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1
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Chung H, Rahmani W, Sinha S, Imanzadeh A, Pun A, Arora R, Jaffer A, Biernaskie J, Chun J. Nephron progenitor fate is modulated by angiotensin type 1 receptor signaling in human kidney organoids. Stem Cells 2025; 43:sxaf012. [PMID: 40111092 PMCID: PMC12080355 DOI: 10.1093/stmcls/sxaf012] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2024] [Accepted: 03/03/2025] [Indexed: 03/22/2025]
Abstract
The renin-angiotensin system (RAS) is essential for normal kidney development. Dysregulation of the RAS during embryogenesis can result in kidney abnormalities. To explore how angiotensin type 1 receptor (AT1R) signaling modulates nephron progenitor (NP) fate specification, we used induced pluripotent stem cell (iPSC) derived human kidney organoids treated with angiotensin II (Ang II) or the AT1R blocker losartan during differentiation. Ang II promoted NP proliferation and differentiation preferentially toward a podocyte fate, depleted the podocyte precursor population, and accelerated glomerular maturation. By contrast, losartan expanded the podocyte precursor population, delayed podocyte differentiation, and regressed the transcriptional signature to a more immature fetal state. Overall, using various in silico approaches with validation by RNAscope, we identified a role for AT1R signaling in regulating NP fate during nephrogenesis in kidney organoids. Our work supports the use of RAS modulators to improve organoid maturation and suggests that RAS may be a determinant of nephron endowment in vivo.
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Affiliation(s)
- Hyunjae Chung
- Department of Medicine, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Waleed Rahmani
- Department of Medicine, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Sarthak Sinha
- Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Aysa Imanzadeh
- Department of Medicine, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Alexander Pun
- Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Rohit Arora
- Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Arzina Jaffer
- Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Jeff Biernaskie
- Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
- Hotchkiss Brain Institute, University of Calgary, Calgary, AB, Canada T2N 4N1
- Alberta Children’s Hospital Research Institute, University of Calgary, Calgary, AB, Canada T2N 4N1
| | - Justin Chun
- Department of Medicine, Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1
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2
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Zhong J, Gao RR, Zhang X, Yang JX, Liu Y, Ma J, Chen Q. Dissecting endothelial cell heterogeneity with new tools. CELL REGENERATION (LONDON, ENGLAND) 2025; 14:10. [PMID: 40121354 PMCID: PMC11929667 DOI: 10.1186/s13619-025-00223-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/30/2024] [Revised: 02/20/2025] [Accepted: 02/22/2025] [Indexed: 03/25/2025]
Abstract
The formation of a blood vessel network is crucial for organ development and regeneration. Over the past three decades, the central molecular mechanisms governing blood vessel growth have been extensively studied. Recent evidence indicates that vascular endothelial cells-the specialized cells lining the inner surface of blood vessels-exhibit significant heterogeneity to meet the specific needs of different organs. This review focuses on the current understanding of endothelial cell heterogeneity, which includes both intra-organ and inter-organ heterogeneity. Intra-organ heterogeneity encompasses arterio-venous and tip-stalk endothelial cell specialization, while inter-organ heterogeneity refers to organ-specific transcriptomic profiles and functions. Advances in single-cell RNA sequencing (scRNA-seq) have enabled the identification of new endothelial subpopulations and the comparison of gene expression patterns across different subsets of endothelial cells. Integrating scRNA-seq with other high-throughput sequencing technologies promises to deepen our understanding of endothelial cell heterogeneity at the epigenetic level and in a spatially resolved context. To further explore human endothelial cell heterogeneity, vascular organoids offer powerful tools for studying gene function in three-dimensional culture systems and for investigating endothelial-tissue interactions using human cells. Developing organ-specific vascular organoids presents unique opportunities to unravel inter-organ endothelial cell heterogeneity and its implications for human disease. Emerging technologies, such as scRNA-seq and vascular organoids, are poised to transform our understanding of endothelial cell heterogeneity and pave the way for innovative therapeutic strategies to address human vascular diseases.
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Affiliation(s)
- Jing Zhong
- Center for Cell Lineage Atlas, CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 510530, China
- University of Chinese Academy of Sciences, Beijing, China
- China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangdong-Hong Kong Joint Laboratory for Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
- Center for Cell Lineage Atlas, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Rong-Rong Gao
- Biomedical Sciences College & Shandong Medicinal Biotechnology Centre, Shandong First Medical University & Shandong Academy of Medical Sciences, NHC Key Laboratory of Biotechnology Drugs (Shandong Academy of Medical Sciences); Key Lab for Rare & Uncommon Diseases of Shandong Province, Ji'nan 250117, Shandong, China
| | - Xin Zhang
- Center for Cell Lineage Atlas, CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 510530, China
- University of Chinese Academy of Sciences, Beijing, China
- China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangdong-Hong Kong Joint Laboratory for Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
- Center for Cell Lineage Atlas, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Jia-Xin Yang
- The Innovation Centre of Ministry of Education for Development and Diseases, School of Medicine, South China University of Technology, Guangzhou, 510006, China
| | - Yang Liu
- The Innovation Centre of Ministry of Education for Development and Diseases, School of Medicine, South China University of Technology, Guangzhou, 510006, China.
| | - Jinjin Ma
- The Innovation Centre of Ministry of Education for Development and Diseases, School of Medicine, South China University of Technology, Guangzhou, 510006, China.
- The Institute of Future Health, South China of Technology, Guangzhou International Campus, Guangzhou, 511442, China.
| | - Qi Chen
- Center for Cell Lineage Atlas, CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 510530, China.
- China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangdong-Hong Kong Joint Laboratory for Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
- Center for Cell Lineage Atlas, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
- Biomedical Sciences College & Shandong Medicinal Biotechnology Centre, Shandong First Medical University & Shandong Academy of Medical Sciences, NHC Key Laboratory of Biotechnology Drugs (Shandong Academy of Medical Sciences); Key Lab for Rare & Uncommon Diseases of Shandong Province, Ji'nan 250117, Shandong, China.
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3
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Mitani S, Hosoda C, Onodera Y, Takabayashi Y, Sakata A, Shima M, Tatsumi K. Efficient generation of liver sinusoidal endothelial-like cells secreting coagulation factor VIII from human induced pluripotent stem cells. Mol Ther Methods Clin Dev 2024; 32:101355. [PMID: 39559558 PMCID: PMC11570519 DOI: 10.1016/j.omtm.2024.101355] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2023] [Accepted: 10/15/2024] [Indexed: 11/20/2024]
Abstract
Liver sinusoidal endothelial cells (LSECs) and LSEC progenitor cells (LPCs) derived from human pluripotent stem cells (PSCs) are expected as valuable cell sources for the development of cell therapy for hemophilia A, a congenital deficiency of coagulation factor VIII (FVIII), as LSECs are responsible for FVIII production. However, there is room for improvement in the efficiency of LSEC and LPC differentiation from human PSCs. In this study, we sought to optimize the method of mesoderm differentiation induction, the initial step of LSEC differentiation from human PSCs, to efficiently induce LSEC-like cells capable of secreting FVIII from human induced pluripotent stem cells (iPSCs). Following optimization of the concentration and stimulation period of CHIR99021 (glycogen synthase kinase 3β inhibitor), bone morphogenetic protein 4, fibroblast growth factor 2, and Activin A in the mesoderm induction step, approximately 65% and 54% of cells differentiated into LPCs and LSEC-like cells, respectively. Furthermore, we observed substantial FVIII protein secretion from LSEC-like cells in vitro. In conclusion, we established an efficient method for obtaining LPCs and functional LSEC-like cells from human iPSCs in vitro.
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Affiliation(s)
- Seiji Mitani
- Advanced Medical Science of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Chihiro Hosoda
- Advanced Medical Science of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Yu Onodera
- Advanced Medical Science of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Yoko Takabayashi
- Advanced Medical Science of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Asuka Sakata
- Medicinal Biology of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Midori Shima
- Medicinal Biology of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
| | - Kohei Tatsumi
- Advanced Medical Science of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
- Medicinal Biology of Thrombosis and Hemostasis, Nara Medical University, Kashihara, Nara 634-8521, Japan
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4
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Jia H, Moore M, Wadhwa M, Burns C. Human iPSC-Derived Endothelial Cells Exhibit Reduced Immunogenicity in Comparison With Human Primary Endothelial Cells. Stem Cells Int 2024; 2024:6153235. [PMID: 39687754 PMCID: PMC11649354 DOI: 10.1155/sci/6153235] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/20/2024] [Accepted: 11/07/2024] [Indexed: 12/18/2024] Open
Abstract
Human induced pluripotent stem cell (iPSC)-derived endothelial cells (ECs) have emerged as a promising source of autologous cells with great potential to produce novel cell therapy for ischemic vascular diseases. However, their clinical application still faces numerous challenges including safety concerns such as the potential aberrant immunogenicity derived from the reprogramming process. This study investigated immunological phenotypes of iPSC-ECs by a side-by-side comparison with primary human umbilical vein ECs (HUVECs). Three types of human iPSC-ECs, NIBSC8-EC generated in house and two commercial iPSC-ECs, alongside HUVECs, were examined for surface expression of proteins of immune relevance under resting conditions and after cytokine activation. All iPSC-EC populations failed to express major histocompatibility complex (MHC) Class II on their surface following interferon-gamma (IFN-γ) treatment but showed similar basal and IFN-γ-stimulated expression levels of MHC Class I of HUVECs. Multiple iPSC-ECs also retained constitutive and tumor necrosis factor-alpha (TNF-α)-stimulated expression levels of intercellular adhesion molecule-1 (ICAM-1) like HUVECs. However, TNF-α induced a differential expression of E-selectin and vascular cell adhesion molecule-1 (VCAM-1) on iPSC-ECs. Furthermore, real-time monitoring of proliferation of human peripheral blood mononuclear cells (PBMCs) cocultured on an endothelial monolayer over 5 days showed that iPSC-ECs provoked distinct dynamics of PBMC proliferation, which was generally decreased in alloreactivity and IFN-γ-stimulated proliferation of PBMCs compared with HUVECs. Consistently, in the conventional mixed lymphocyte reaction (MLR), the proliferation of total CD3+ and CD4+ T cells after 5-day cocultures with multiple iPSC-EC populations was largely reduced compared to HUVECs. Last, multiple iPSC-EC cocultures secreted lower levels of proinflammatory cytokines than HUVEC cocultures. Collectively, iPSC-ECs manifested many similarities, but also some disparities with a generally weaker inflammatory immune response than primary ECs, indicating that iPSC-ECs may possibly exhibit hypoimmunogenicity corresponding with less risk of immune rejection in a transplant setting, which is important for safe and effective cell therapies.
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Affiliation(s)
- Haiyan Jia
- Biotherapeutics and Advanced Therapies, Research and Development, Science and Research Group, Medicines and Healthcare Products Regulatory Agency, Blanche Lane, South Mimms, Potters Bar EN6 3QG, Hertfordshire, UK
| | - Melanie Moore
- Therapeutic Reference Materials, Standards Lifecycle, Science and Research Group, Medicines and Healthcare Products Regulatory Agency, Blanche Lane, South Mimms, Potters Bar EN6 3QG, Hertfordshire, UK
| | - Meenu Wadhwa
- Biotherapeutics and Advanced Therapies, Research and Development, Science and Research Group, Medicines and Healthcare Products Regulatory Agency, Blanche Lane, South Mimms, Potters Bar EN6 3QG, Hertfordshire, UK
| | - Chris Burns
- Biotherapeutics and Advanced Therapies, Research and Development, Science and Research Group, Medicines and Healthcare Products Regulatory Agency, Blanche Lane, South Mimms, Potters Bar EN6 3QG, Hertfordshire, UK
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5
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Naderi-Meshkin H, Wahyu Setyaningsih WA, Yacoub A, Carney G, Cornelius VA, Nelson CA, Kelaini S, Donaghy C, Dunne PD, Amirkhah R, Zampetaki A, Zeng L, Stitt AW, Lois N, Grieve DJ, Margariti A. Unveiling impaired vascular function and cellular heterogeneity in diabetic donor-derived vascular organoids. Stem Cells 2024; 42:791-808. [PMID: 39049437 PMCID: PMC11384901 DOI: 10.1093/stmcls/sxae043] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2023] [Accepted: 06/06/2024] [Indexed: 07/27/2024]
Abstract
Vascular organoids (VOs), derived from induced pluripotent stem cells (iPSCs), hold promise as in vitro disease models and drug screening platforms. However, their ability to faithfully recapitulate human vascular disease and cellular composition remains unclear. In this study, we demonstrate that VOs derived from iPSCs of donors with diabetes (DB-VOs) exhibit impaired vascular function compared to non-diabetic VOs (ND-VOs). DB-VOs display elevated levels of reactive oxygen species (ROS), heightened mitochondrial content and activity, increased proinflammatory cytokines, and reduced blood perfusion recovery in vivo. Through comprehensive single-cell RNA sequencing, we uncover molecular and functional differences, as well as signaling networks, between vascular cell types and clusters within DB-VOs. Our analysis identifies major vascular cell types (endothelial cells [ECs], pericytes, and vascular smooth muscle cells) within VOs, highlighting the dichotomy between ECs and mural cells. We also demonstrate the potential need for additional inductions using organ-specific differentiation factors to promote organ-specific identity in VOs. Furthermore, we observe basal heterogeneity within VOs and significant differences between DB-VOs and ND-VOs. Notably, we identify a subpopulation of ECs specific to DB-VOs, showing overrepresentation in the ROS pathway and underrepresentation in the angiogenesis hallmark, indicating signs of aberrant angiogenesis in diabetes. Our findings underscore the potential of VOs for modeling diabetic vasculopathy, emphasize the importance of investigating cellular heterogeneity within VOs for disease modeling and drug discovery, and provide evidence of GAP43 (neuromodulin) expression in ECs, particularly in DB-VOs, with implications for vascular development and disease.
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Affiliation(s)
- Hojjat Naderi-Meshkin
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Wiwit A Wahyu Setyaningsih
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
- Department of Anatomy, Faculty of Medicine, Public Health, and Nursing, Universitas Gadjah Mada, Sleman, D.I. Yogyakarta, 55281, Indonesia
| | - Andrew Yacoub
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Garrett Carney
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Victoria A Cornelius
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Clare-Ann Nelson
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Sophia Kelaini
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Clare Donaghy
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Philip D Dunne
- The Patrick G Johnston Centre for Cancer Research, Queen's University Belfast, Belfast, BT9 7AE, United Kingdom
| | - Raheleh Amirkhah
- The Patrick G Johnston Centre for Cancer Research, Queen's University Belfast, Belfast, BT9 7AE, United Kingdom
| | - Anna Zampetaki
- School of Cardiovascular Medicine and Sciences, BHF Centre of Research Excellence, King's College London, London, SE5 9NU, United Kingdom
| | - Lingfang Zeng
- School of Cardiovascular Medicine and Sciences, BHF Centre of Research Excellence, King's College London, London, SE5 9NU, United Kingdom
| | - Alan W Stitt
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Noemi Lois
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - David J Grieve
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
| | - Andriana Margariti
- The Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, BT9 7BL, United Kingdom
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6
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Ng ES, Sarila G, Li JY, Edirisinghe HS, Saxena R, Sun S, Bruveris FF, Labonne T, Sleebs N, Maytum A, Yow RY, Inguanti C, Motazedian A, Calvanese V, Capellera-Garcia S, Ma F, Nim HT, Ramialison M, Bonifer C, Mikkola HKA, Stanley EG, Elefanty AG. Long-term engrafting multilineage hematopoietic cells differentiated from human induced pluripotent stem cells. Nat Biotechnol 2024:10.1038/s41587-024-02360-7. [PMID: 39223325 DOI: 10.1038/s41587-024-02360-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2023] [Accepted: 07/20/2024] [Indexed: 09/04/2024]
Abstract
Hematopoietic stem cells (HSCs) derived from human induced pluripotent stem cells (iPS cells) have important biomedical applications. We identified differentiation conditions that generate HSCs defined by robust long-term multilineage engraftment in immune-deficient NOD,B6.Prkdcscid Il2rgtm1Wjl/SzJ KitW41/W41 mice. We guided differentiating iPS cells, as embryoid bodies in a defined culture medium supplemented with retinyl acetate, through HOXA-patterned mesoderm to hemogenic endothelium specified by bone morphogenetic protein 4 and vascular endothelial growth factor (VEGF). Removal of VEGF facilitated an efficient endothelial-to-hematopoietic transition, evidenced by release into the culture medium of CD34+ blood cells, which were cryopreserved. Intravenous transplantation of two million thawed CD34+ cells differentiated from four independent iPS cell lines produced multilineage bone marrow engraftment in 25-50% of immune-deficient recipient mice. These functionally defined, multipotent CD34+ hematopoietic cells, designated iPS cell-derived HSCs (iHSCs), produced levels of engraftment similar to those achieved following umbilical cord blood transplantation. Our study provides a step toward the goal of generating HSCs for clinical translation.
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Grants
- GNT1117596,GNT1068866, GNT1129861,GNT2012535 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT1164577, GNT2012936 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT2012535 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT1186019 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT1068866, GNT1129861, GNT2012535 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT1068866, GNT1129861, GNT1186019 Department of Health | National Health and Medical Research Council (NHMRC)
- GNT1079004, GNT1068866, GNT1129861, GNT1186019 Department of Health | National Health and Medical Research Council (NHMRC)
- RT3-07763 California Institute for Regenerative Medicine (CIRM)
- NNF21CC0073729 Novo Nordisk Fonden (Novo Nordisk Foundation)
- NIH 1RO1DK125097-01 Foundation for the National Institutes of Health (Foundation for the National Institutes of Health, Inc.)
- IPD2 2018-06635 Vetenskapsrådet (Swedish Research Council)
- BB/R014809/1 RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
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Affiliation(s)
- Elizabeth S Ng
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia.
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia.
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia.
| | - Gulcan Sarila
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Jacky Y Li
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Hasindu S Edirisinghe
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Ritika Saxena
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Shicheng Sun
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
- Changping Laboratory, Beijing, China
| | - Freya F Bruveris
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Tanya Labonne
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Nerida Sleebs
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Alexander Maytum
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
- Institute for Cancer and Genomic Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
| | - Raymond Y Yow
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Chantelle Inguanti
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Ali Motazedian
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
- Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia
| | - Vincenzo Calvanese
- Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, Los Angeles, CA, USA
- Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA
- Laboratory for Molecular Cell Biology, University College London, London, UK
| | - Sandra Capellera-Garcia
- Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, Los Angeles, CA, USA
- Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA
| | - Feiyang Ma
- Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, Los Angeles, CA, USA
- Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA
| | - Hieu T Nim
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
- Australian Regenerative Medicine Institute, Monash University, Clayton, Victoria, Australia
| | - Mirana Ramialison
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
- Australian Regenerative Medicine Institute, Monash University, Clayton, Victoria, Australia
| | - Constanze Bonifer
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
- Institute for Cancer and Genomic Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
| | - Hanna K A Mikkola
- Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, Los Angeles, CA, USA
- Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA
| | - Edouard G Stanley
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia
| | - Andrew G Elefanty
- Murdoch Children's Research Institute, The Royal Children's Hospital, Parkville, Victoria, Australia.
- Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Parkville, Victoria, Australia.
- The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Murdoch Children's Research Institute, Parkville, Victoria, Australia.
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Lu H, Zuo X, Yuan J, Xie Z, Yin L, Pu Y, Chen Z, Zhang J. Research progress in the development of 3D skin models and their application to in vitro skin irritation testing. J Appl Toxicol 2024; 44:1302-1316. [PMID: 38711121 DOI: 10.1002/jat.4618] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2024] [Revised: 04/07/2024] [Accepted: 04/11/2024] [Indexed: 05/08/2024]
Abstract
Toxicological assessment of chemicals is crucial for safeguarding human health and the environment. However, traditional animal experiments are associated with ethical, technical, and predictive limitations in assessing the toxicity of chemicals to the skin. With the recent development of bioengineering and tissue engineering, three-dimensional (3D) skin models have been commonly used as an alternative for toxicological studies. The skin consists of the subcutaneous, dermis, and epidermis. All these layers have crucial functions such as physical and biological protection and thermoregulation. The epidermis is the shallowest layer protecting against external substances and media. Because the skin is the first contact point for many substances, this organ is very significant for assessing local toxicity following skin exposure. According to the classification of the United Nations Global Harmonized System, skin irritation is a major potentially hazardous characteristic of chemicals, and this characteristic must be accurately assessed and classified for enhancing chemical safety management and preventing and reducing chemical accidents. This review discusses the research progress of 3D skin models and introduces their application in assessing chemical skin irritation.
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Affiliation(s)
- Hongxia Lu
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
| | - Xulei Zuo
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
| | - Jiayu Yuan
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
| | - Zhuoying Xie
- State Key Laboratory of Digital Medical Engineering, School of Biological Science and Medical Engineering, Southeast University, Nanjing, P. R. China
| | - Lihong Yin
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
| | - Yuepu Pu
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
| | - Zaozao Chen
- State Key Laboratory of Digital Medical Engineering, School of Biological Science and Medical Engineering, Southeast University, Nanjing, P. R. China
| | - Juan Zhang
- Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing, P. R. China
- Jiangsu Institute for Sport and Health (JISH), Nanjing, P. R. China
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8
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Zamora Alvarado JE, McCloskey KE, Gopinathan A. Migration and proliferation drive the emergence of patterns in co-cultures of differentiating vascular progenitor cells. MATHEMATICAL BIOSCIENCES AND ENGINEERING : MBE 2024; 21:6731-6757. [PMID: 39483091 PMCID: PMC11556463 DOI: 10.3934/mbe.2024295] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/03/2024]
Abstract
Vascular cells self-organize into unique structures guided by cell proliferation, migration, and/or differentiation from neighboring cells, mechanical factors, and/or soluble signals. However, the relative contribution of each of these factors remains unclear. Our objective was to develop a computational model to explore the different factors affecting the emerging micropatterns in 2D. This was accomplished by developing a stochastic on-lattice population-based model starting with vascular progenitor cells with the potential to proliferate, migrate, and/or differentiate into either endothelial cells or smooth muscle cells. The simulation results yielded patterns that were qualitatively and quantitatively consistent with experimental observations. Our results suggested that post-differentiation cell migration and proliferation when balanced could generate between 30-70% of each cell type enabling the formation of vascular patterns. Moreover, the cell-to-cell sensing could enhance the robustness of this patterning. These findings computationally supported that 2D patterning is mechanistically similar to current microfluidic platforms that take advantage of the migration-directed self-assembly of mature endothelial and mural cells to generate perfusable 3D vasculature in permissible hydrogel environments and suggest that stem or progenitor cells may not be fully necessary components in many tissue formations like those formed by vasculogenesis.
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Affiliation(s)
- Jose E. Zamora Alvarado
- School of Engineering, University of California Merced, Merced, CA 95343, USA
- Graduate Program in Materials and Biomaterials Science and Engineering, University of California Merced, Merced, CA 95343, USA
| | - Kara E. McCloskey
- School of Engineering, University of California Merced, Merced, CA 95343, USA
- Graduate Program in Materials and Biomaterials Science and Engineering, University of California Merced, Merced, CA 95343, USA
| | - Ajay Gopinathan
- Graduate Program in Materials and Biomaterials Science and Engineering, University of California Merced, Merced, CA 95343, USA
- Department of Physics, University of California Merced, Merced, CA 95343, USA
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9
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Kiskin FN, Yang Y, Yang H, Zhang JZ. Cracking the code of the cardiovascular enigma: hPSC-derived endothelial cells unveil the secrets of endothelial dysfunction. J Mol Cell Cardiol 2024; 192:65-78. [PMID: 38761989 DOI: 10.1016/j.yjmcc.2024.05.005] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/13/2024] [Revised: 05/08/2024] [Accepted: 05/10/2024] [Indexed: 05/20/2024]
Abstract
Endothelial dysfunction is a central contributor to the development of most cardiovascular diseases and is characterised by the reduced synthesis or bioavailability of the vasodilator nitric oxide together with other abnormalities such as inflammation, senescence, and oxidative stress. The use of patient-specific and genome-edited human pluripotent stem cell-derived endothelial cells (hPSC-ECs) has shed novel insights into the role of endothelial dysfunction in cardiovascular diseases with strong genetic components such as genetic cardiomyopathies and pulmonary arterial hypertension. However, their utility in studying complex multifactorial diseases such as atherosclerosis, metabolic syndrome and heart failure poses notable challenges. In this review, we provide an overview of the different methods used to generate and characterise hPSC-ECs before comprehensively assessing their effectiveness in cardiovascular disease modelling and high-throughput drug screening. Furthermore, we explore current obstacles that will need to be overcome to unleash the full potential of hPSC-ECs in facilitating patient-specific precision medicine. Addressing these challenges holds great promise in advancing our understanding of intricate cardiovascular diseases and in tailoring personalised therapeutic strategies.
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Affiliation(s)
- Fedir N Kiskin
- Institute of Neurological and Psychiatric Disorders, Shenzhen Bay Laboratory, Shenzhen 518132, China.
| | - Yuan Yang
- Institute of Neurological and Psychiatric Disorders, Shenzhen Bay Laboratory, Shenzhen 518132, China.
| | - Hao Yang
- Institute of Neurological and Psychiatric Disorders, Shenzhen Bay Laboratory, Shenzhen 518132, China.
| | - Joe Z Zhang
- Institute of Neurological and Psychiatric Disorders, Shenzhen Bay Laboratory, Shenzhen 518132, China.
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10
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Saha S, Haynes WJ, Del Rio NM, Young EE, Zhang J, Seo J, Huang L, Holm AM, Blashka W, Murphy L, Scholz MJ, Henrichs A, Suresh Babu J, Steill J, Stewart R, Kamp TJ, Brown ME. Diminished Immune Cell Adhesion in Hypoimmune ICAM-1 Knockout Pluripotent Stem Cells. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.06.07.597791. [PMID: 38895244 PMCID: PMC11185752 DOI: 10.1101/2024.06.07.597791] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/21/2024]
Abstract
Hypoimmune gene edited human pluripotent stem cells (hPSCs) are a promising platform for developing reparative cellular therapies that evade immune rejection. Existing first-generation hypoimmune strategies have used CRISPR/Cas9 editing to modulate genes associated with adaptive (e.g., T cell) immune responses, but have largely not addressed the innate immune cells (e.g., monocytes, neutrophils) that mediate inflammation and rejection processes occurring early after graft transplantation. We identified the adhesion molecule ICAM-1 as a novel hypoimmune target that plays multiple critical roles in both adaptive and innate immune responses post-transplantation. In a series of studies, we found that ICAM-1 blocking or knock-out (KO) in hPSC-derived cardiovascular therapies imparted significantly diminished binding of multiple immune cell types. ICAM-1 KO resulted in diminished T cell proliferation responses in vitro and in longer in vivo retention/protection of KO grafts following immune cell encounter in NeoThy humanized mice. The ICAM-1 KO edit was also introduced into existing first-generation hypoimmune hPSCs and prevented immune cell binding, thereby enhancing the overall hypoimmune capacity of the cells. This novel hypoimmune editing strategy has the potential to improve the long-term efficacy and safety profiles of regenerative therapies for cardiovascular pathologies and a number of other diseases.
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Affiliation(s)
- Sayandeep Saha
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - W. John Haynes
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Natalia M. Del Rio
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Elizabeth E. Young
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Jue Zhang
- Morgridge Institute for Research, Madison, WI
| | - Jiwon Seo
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Liupei Huang
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Alexis M. Holm
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Wesley Blashka
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Lydia Murphy
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Merrick J. Scholz
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | - Abigale Henrichs
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
| | | | - John Steill
- Morgridge Institute for Research, Madison, WI
| | - Ron Stewart
- Morgridge Institute for Research, Madison, WI
| | - Timothy J. Kamp
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Medicine, Madison, WI
| | - Matthew E. Brown
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Madison, WI
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11
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Cai Z, Zhu M, Xu L, Wang Y, Xu Y, Yim WY, Cao H, Guo R, Qiu X, He X, Shi J, Qiao W, Dong N. Directed Differentiation of Human Induced Pluripotent Stem Cells to Heart Valve Cells. Circulation 2024; 149:1435-1456. [PMID: 38357822 PMCID: PMC11062615 DOI: 10.1161/circulationaha.123.065143] [Citation(s) in RCA: 10] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/12/2023] [Accepted: 01/19/2024] [Indexed: 02/16/2024]
Abstract
BACKGROUND A main obstacle in current valvular heart disease research is the lack of high-quality homogeneous functional heart valve cells. Human induced pluripotent stem cells (hiPSCs)-derived heart valve cells may help with this dilemma. However, there are no well-established protocols to induce hiPSCs to differentiate into functional heart valve cells, and the networks that mediate the differentiation have not been fully elucidated. METHODS To generate heart valve cells from hiPSCs, we sequentially activated the Wnt, BMP4, VEGF (vascular endothelial growth factor), and NFATc1 signaling pathways using CHIR-99021, BMP4, VEGF-165, and forskolin, respectively. The transcriptional and functional similarity of hiPSC-derived heart valve cells compared with primary heart valve cells were characterized. Longitudinal single-cell RNA sequencing was used to uncover the trajectory, switch genes, pathways, and transcription factors of the differentiation. RESULTS An efficient protocol was developed to induce hiPSCs to differentiate into functional hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells. After 6-day differentiation and CD144 magnetic bead sorting, ≈70% CD144+ cells and 30% CD144- cells were obtained. On the basis of single-cell RNA sequencing data, the CD144+ cells and CD144- cells were found to be highly similar to primary heart valve endothelial cells and primary heart valve interstitial cells in gene expression profile. Furthermore, CD144+ cells had the typical function of primary heart valve endothelial cells, including tube formation, uptake of low-density lipoprotein, generation of endothelial nitric oxide synthase, and response to shear stress. Meanwhile, CD144- cells could secret collagen and matrix metalloproteinases, and differentiate into osteogenic or adipogenic lineages like primary heart valve interstitial cells. Therefore, we identified CD144+ cells and CD144- cells as hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells, respectively. Using single-cell RNA sequencing analysis, we demonstrated that the trajectory of heart valve cell differentiation was consistent with embryonic valve development. We identified the main switch genes (NOTCH1, HEY1, and MEF2C), signaling pathways (TGF-β, Wnt, and NOTCH), and transcription factors (MSX1, SP5, and MECOM) that mediated the differentiation. Finally, we found that hiPSC-derived valve interstitial-like cells might derive from hiPSC-derived valve endothelial-like cells undergoing endocardial-mesenchymal transition. CONCLUSIONS In summary, this is the first study to report an efficient strategy to generate functional hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells from hiPSCs, as well as to elucidate the differentiation trajectory and transcriptional dynamics of hiPSCs differentiated into heart valve cells.
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Affiliation(s)
- Ziwen Cai
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
- Department of Cardiovascular Surgery, Union Hospital, Fujian Medical University, Fuzhou, China (Z.C.)
| | - Miaomiao Zhu
- Department of Cardiovascular Surgery, Union Hospital, Fujian Medical University, Fuzhou, China (Z.C.)
- Institute of Maternal and Children Health, Wuhan Children’s Hospital (Wuhan Maternal and Child Healthcare Hospital), Tongji medical College, Huazhong University of Science & Technology, Hubei, China (M.Z.)
| | - Li Xu
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Yue Wang
- Department of Anesthesiology, Union Hospital, Fujian Medical University, Fuzhou, China (Y.W.)
| | - Yin Xu
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Wai Yen Yim
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Hong Cao
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Ruikang Guo
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Xiang Qiu
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Ximiao He
- Department of Physiology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (M.Z., X.H.)
| | - Jiawei Shi
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Weihua Qiao
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
| | - Nianguo Dong
- Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China (Z.C., L.X., Y.X., W.Y.Y., H.C., R.G., X.Q, J.S., W.Q., N.D.)
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12
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Yan W, Xia Y, Zhao H, Xu X, Ma X, Tao L. Stem cell-based therapy in cardiac repair after myocardial infarction: Promise, challenges, and future directions. J Mol Cell Cardiol 2024; 188:1-14. [PMID: 38246086 DOI: 10.1016/j.yjmcc.2023.12.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/10/2023] [Revised: 12/09/2023] [Accepted: 12/22/2023] [Indexed: 01/23/2024]
Abstract
Stem cells represent an attractive resource for cardiac regeneration. However, the survival and function of transplanted stem cells is poor and remains a major challenge for the development of effective therapies. As two main cell types currently under investigation in heart repair, mesenchymal stromal cells (MSCs) indirectly support endogenous regenerative capacities after transplantation, while induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) functionally integrate into the damaged myocardium and directly contribute to the restoration of its pump function. These two cell types are exposed to a common microenvironment with many stressors in ischemic heart tissue. This review summarizes the research progress on the mechanisms and challenges of MSCs and iPSC-CMs in post-MI heart repair, introduces several randomized clinical trials with 3D-mapping-guided cell therapy, and outlines recent findings related to the factors that affect the survival and function of stem cells. We also discuss the future directions for optimization such as biomaterial utilization, cell combinations, and intravenous injection of engineered nucleus-free MSCs.
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Affiliation(s)
- Wenjun Yan
- Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China
| | - Yunlong Xia
- Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China
| | - Huishou Zhao
- Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China
| | - Xiaoming Xu
- Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China
| | - Xinliang Ma
- Department of Emergency Medicine, Thomas Jefferson University, Philadelphia, PA 19107, United States of America
| | - Ling Tao
- Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
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13
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Jiao YC, Wang YX, Liu WZ, Xu JW, Zhao YY, Yan CZ, Liu FC. Advances in the differentiation of pluripotent stem cells into vascular cells. World J Stem Cells 2024; 16:137-150. [PMID: 38455095 PMCID: PMC10915963 DOI: 10.4252/wjsc.v16.i2.137] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/21/2023] [Revised: 12/20/2023] [Accepted: 01/16/2024] [Indexed: 02/26/2024] Open
Abstract
Blood vessels constitute a closed pipe system distributed throughout the body, transporting blood from the heart to other organs and delivering metabolic waste products back to the lungs and kidneys. Changes in blood vessels are related to many disorders like stroke, myocardial infarction, aneurysm, and diabetes, which are important causes of death worldwide. Translational research for new approaches to disease modeling and effective treatment is needed due to the huge socio-economic burden on healthcare systems. Although mice or rats have been widely used, applying data from animal studies to human-specific vascular physiology and pathology is difficult. The rise of induced pluripotent stem cells (iPSCs) provides a reliable in vitro resource for disease modeling, regenerative medicine, and drug discovery because they carry all human genetic information and have the ability to directionally differentiate into any type of human cells. This review summarizes the latest progress from the establishment of iPSCs, the strategies for differentiating iPSCs into vascular cells, and the in vivo transplantation of these vascular derivatives. It also introduces the application of these technologies in disease modeling, drug screening, and regenerative medicine. Additionally, the application of high-tech tools, such as omics analysis and high-throughput sequencing, in this field is reviewed.
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Affiliation(s)
- Yi-Chang Jiao
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
| | - Ying-Xin Wang
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
| | - Wen-Zhu Liu
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
| | - Jing-Wen Xu
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
| | - Yu-Ying Zhao
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
| | - Chuan-Zhu Yan
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Mitochondrial Medicine Laboratory, Qilu Hospital (Qingdao) of Shandong University, Qingdao 266103, Shandong Province, China
- Brain Science Research Institute, Shandong University, Jinan 250012, Shandong Province, China
| | - Fu-Chen Liu
- Department of Neurology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Research Institute of Neuromuscular and Neurodegenerative Diseases, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
- Brain Science Research Institute, Shandong University, Jinan 250012, Shandong Province, China.
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14
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Esparza A, Jimenez N, Borrego EA, Browne S, Natividad-Diaz SL. Review: Human stem cell-based 3D in vitro angiogenesis models for preclinical drug screening applications. Mol Biol Rep 2024; 51:260. [PMID: 38302762 PMCID: PMC10834608 DOI: 10.1007/s11033-023-09048-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2023] [Accepted: 11/06/2023] [Indexed: 02/03/2024]
Abstract
Vascular diseases are the underlying pathology in many life-threatening illnesses. Human cellular and molecular mechanisms involved in angiogenesis are complex and difficult to study in current 2D in vitro and in vivo animal models. Engineered 3D in vitro models that incorporate human pluripotent stem cell (hPSC) derived endothelial cells (ECs) and supportive biomaterials within a dynamic microfluidic platform provide a less expensive, more controlled, and reproducible platform to better study angiogenic processes in response to external chemical or physical stimulus. Current studies to develop 3D in vitro angiogenesis models aim to establish single-source systems by incorporating hPSC-ECs into biomimetic extracellular matrices (ECM) and microfluidic devices to create a patient-specific, physiologically relevant platform that facilitates preclinical study of endothelial cell-ECM interactions, vascular disease pathology, and drug treatment pharmacokinetics. This review provides a detailed description of the current methods used for the directed differentiation of human stem cells to endothelial cells and their use in engineered 3D in vitro angiogenesis models that have been developed within the last 10 years.
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Affiliation(s)
- Aibhlin Esparza
- Department of Metallurgical, Materials, and Biomedical Engineering (MMBME), The University of Texas at El Paso (UTEP), El Paso, TX, USA
- 3D Printed Microphysiological Systems Laboratory, The University of Texas at El Paso, El Paso, TX, USA
| | - Nicole Jimenez
- Department of Metallurgical, Materials, and Biomedical Engineering (MMBME), The University of Texas at El Paso (UTEP), El Paso, TX, USA
- 3D Printed Microphysiological Systems Laboratory, The University of Texas at El Paso, El Paso, TX, USA
| | - Edgar A Borrego
- Department of Metallurgical, Materials, and Biomedical Engineering (MMBME), The University of Texas at El Paso (UTEP), El Paso, TX, USA
- 3D Printed Microphysiological Systems Laboratory, The University of Texas at El Paso, El Paso, TX, USA
| | - Shane Browne
- Department of Anatomy and Regenerative Medicine, Tissue Engineering Research Group, Royal College of Surgeons, Dublin, Ireland
- CÚRAM, Centre for Research in Medical Devices, University of Galway, Galway, H91 W2TY, Ireland
| | - Sylvia L Natividad-Diaz
- Department of Metallurgical, Materials, and Biomedical Engineering (MMBME), The University of Texas at El Paso (UTEP), El Paso, TX, USA.
- 3D Printed Microphysiological Systems Laboratory, The University of Texas at El Paso, El Paso, TX, USA.
- Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX, USA.
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15
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Seo J, Saha S, Brown ME. The past, present, and future promise of pluripotent stem cells. JOURNAL OF IMMUNOLOGY AND REGENERATIVE MEDICINE 2024; 22-23:100077. [PMID: 38706532 PMCID: PMC11065261 DOI: 10.1016/j.regen.2024.100077] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/07/2024]
Affiliation(s)
| | | | - Matthew E. Brown
- University of Wisconsin-Madison, School of Medicine and Public Health, Department of Surgery, Division of Transplantation, 600 Highland Avenue, Madison, WI, 53792, United States
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16
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Sullivan MA, Lane S, Volkerling A, Engel M, Werry EL, Kassiou M. Three-dimensional bioprinting of stem cell-derived central nervous system cells enables astrocyte growth, vasculogenesis, and enhances neural differentiation/function. Biotechnol Bioeng 2023; 120:3079-3091. [PMID: 37395340 PMCID: PMC10953436 DOI: 10.1002/bit.28470] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2023] [Revised: 05/16/2023] [Accepted: 06/05/2023] [Indexed: 07/04/2023]
Abstract
Current research tools for preclinical drug development such as rodent models and two-dimensional immortalized monocultures have failed to serve as effective translational models for human central nervous system (CNS) disorders. Recent advancements in the development of induced pluripotent stem cells (iPSCs) and three-dimensional (3D) culturing can improve the in vivo-relevance of preclinical models, while generating 3D cultures though novel bioprinting technologies can offer increased scalability and replicability. As such, there is a need to develop platforms that combine iPSC-derived cells with 3D bioprinting to produce scalable, tunable, and biomimetic cultures for preclinical drug discovery applications. We report a biocompatible poly(ethylene glycol)-based matrix which incorporates Arg-Gly-Asp and Tyr-Ile-Gly-Ser-Arg peptide motifs and full-length collagen IV at a stiffness similar to the human brain (1.5 kPa). Using a high-throughput commercial bioprinter we report the viable culture and morphological development of monocultured iPSC-derived astrocytes, brain microvascular endothelial-like cells, neural progenitors, and neurons in our novel matrix. We also show that this system supports endothelial-like vasculogenesis and enhances neural differentiation and spontaneous activity. This platform forms a foundation for more complex, multicellular models to facilitate high-throughput translational drug discovery for CNS disorders.
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Affiliation(s)
- Michael A. Sullivan
- School of Medical Sciences, The Faculty of Medicine and HealthThe University of SydneySydneyNew South WalesAustralia
| | - Samuel Lane
- School of Chemistry, The Faculty of ScienceThe University of SydneySydneyNew South WalesAustralia
| | | | - Martin Engel
- Inventia Life Science Operations Pty Ltd.AlexandriaNew South WalesAustralia
| | - Eryn L. Werry
- School of Chemistry, The Faculty of ScienceThe University of SydneySydneyNew South WalesAustralia
- Central Clinical School, Faculty of Medicine and HealthThe University of SydneySydneyNew South WalesAustralia
| | - Michael Kassiou
- School of Chemistry, The Faculty of ScienceThe University of SydneySydneyNew South WalesAustralia
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Bailin SS, Kropski JA, Gangula RD, Hannah L, Simmons JD, Mashayekhi M, Ye F, Fan R, Mallal S, Warren CM, Kalams SA, Gabriel CL, Wanjalla CN, Koethe JR. Changes in subcutaneous white adipose tissue cellular composition and molecular programs underlie glucose intolerance in persons with HIV. Front Immunol 2023; 14:1152003. [PMID: 37711619 PMCID: PMC10499182 DOI: 10.3389/fimmu.2023.1152003] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2023] [Accepted: 08/07/2023] [Indexed: 09/16/2023] Open
Abstract
Introduction Subcutaneous adipose tissue (SAT) is a critical regulator of systemic metabolic homeostasis. Persons with HIV (PWH) have an increased risk of metabolic diseases and significant alterations in the SAT immune environment compared with the general population. Methods We generated a comprehensive single-cell multi-omic SAT atlas to characterize cellular compositional and transcriptional changes in 59 PWH across a spectrum of metabolic health. Results Glucose intolerance was associated with increased lipid-associated macrophages, CD4+ and CD8+ T effector memory cells, and decreased perivascular macrophages. We observed a coordinated intercellular regulatory program which enriched for genes related to inflammation and lipid-processing across multiple cell types as glucose intolerance increased. Increased CD4+ effector memory tissue-resident cells most strongly associated with altered expression of adipocyte genes critical for lipid metabolism and cellular regulation. Intercellular communication analysis demonstrated enhanced pro-inflammatory and pro-fibrotic signaling between immune cells and stromal cells in PWH with glucose intolerance compared with non-diabetic PWH. Lastly, while cell type-specific gene expression among PWH with diabetes was globally similar to HIV-negative individuals with diabetes, we observed substantially divergent intercellular communication pathways. Discussion These findings suggest a central role of tissue-resident immune cells in regulating SAT inflammation among PWH with metabolic disease, and underscore unique mechanisms that may converge to promote metabolic disease.
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Affiliation(s)
- Samuel S. Bailin
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Jonathan A. Kropski
- Department of Medicine, Division of Allergy, Pulmonary, and Critical Care Medicine, Vanderbilt University Medical Center, Nashville, TN, United States
- Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN, United States
- Deparment of Cell and Developmental Biology, Vanderbilt University, Nashville, TN, United States
| | - Rama D. Gangula
- Tennessee Center for AIDS Research, Vanderbilt University Medical Center, Nashville, TN, United States
| | - LaToya Hannah
- Department of Medicine, Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Joshua D. Simmons
- Tennessee Center for AIDS Research, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Mona Mashayekhi
- Department of Medicine, Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Fei Ye
- Department of Biostatics, Division of Epidemiology, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Run Fan
- Department of Biostatistics, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Simon Mallal
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
- Tennessee Center for AIDS Research, Vanderbilt University Medical Center, Nashville, TN, United States
- Insitute for Immunology and Infectious Diseases, Murdoch University, Perth, WA, Australia
- Vanderbilt Technologies for Advanced Genomics, Vanderbilt University Medical Center, Nashville, TN, United States
- Center for Translational Immunology and Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Christian M. Warren
- Tennessee Center for AIDS Research, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Spyros A. Kalams
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
- Tennessee Center for AIDS Research, Vanderbilt University Medical Center, Nashville, TN, United States
- Center for Translational Immunology and Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Curtis L. Gabriel
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Nashville, TN, United States
| | - Celestine N. Wanjalla
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
- Center for Translational Immunology and Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
| | - John R. Koethe
- Department of Medicine, Division of Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
- Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN, United States
- Center for Translational Immunology and Infectious Diseases, Vanderbilt University Medical Center, Nashville, TN, United States
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18
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Tattersall MC, Esnault S, Stewart R, Vereide DT, Swanson S, Zhang J, Steill J, Jarjour N, Hansen KM, Korcarz CE, Baker TB, Stein JH. Effects of an In Vivo Vaping Challenge on In Vitro Interleukin-6 Biosynthesis Pathways in Arterial Endothelial Cells Derived From Human Embryonic Stem Cells. J Am Heart Assoc 2023:e030139. [PMID: 37449574 PMCID: PMC10382107 DOI: 10.1161/jaha.123.030139] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/15/2023] [Accepted: 06/27/2023] [Indexed: 07/18/2023]
Affiliation(s)
- Matthew C Tattersall
- Department of Medicine, Division of Cardiovascular Medicine University of Wisconsin School of Medicine and Public Health Madison WI
| | - Stephane Esnault
- Department of Medicine, Division of Allergy, Pulmonary and Critical Care Medicine The University of Wisconsin-Madison School of Medicine and Public Health Madison WI
| | | | | | | | - Jue Zhang
- Morgridge Institute for Research Madison WI
| | | | - Nizar Jarjour
- Department of Medicine, Division of Allergy, Pulmonary and Critical Care Medicine The University of Wisconsin-Madison School of Medicine and Public Health Madison WI
| | - Kristin M Hansen
- Department of Medicine, Division of Cardiovascular Medicine University of Wisconsin School of Medicine and Public Health Madison WI
| | - Claudia E Korcarz
- Department of Medicine, Division of Cardiovascular Medicine University of Wisconsin School of Medicine and Public Health Madison WI
| | - Timothy B Baker
- University of Wisconsin Center for Tobacco Research and Intervention (UW-CTRI) University of Wisconsin School of Medicine and Public Health Madison WI
| | - James H Stein
- Department of Medicine, Division of Cardiovascular Medicine University of Wisconsin School of Medicine and Public Health Madison WI
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19
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Cho S, Aakash P, Lee S, Yoon YS. Endothelial cell direct reprogramming: Past, present, and future. J Mol Cell Cardiol 2023; 180:22-32. [PMID: 37080451 PMCID: PMC10330356 DOI: 10.1016/j.yjmcc.2023.04.006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/03/2023] [Revised: 04/04/2023] [Accepted: 04/17/2023] [Indexed: 04/22/2023]
Abstract
Ischemic cardiovascular disease still remains as a leading cause of morbidity and mortality despite various medical, surgical, and interventional therapy. As such, cell therapy has emerged as an attractive option because it tackles underlying problem of the diseases by inducing neovascularization in ischemic tissue. After overall failure of adult stem or progenitor cells, studies attempted to generate endothelial cells (ECs) from pluripotent stem cells (PSCs). While endothelial cells (ECs) differentiated from PSCs successfully induced vascular regeneration, differentiating volatility and tumorigenic potential is a concern for their clinical applications. Alternatively, direct reprogramming strategies employ lineage-specific factors to change cell fate without achieving pluripotency. ECs have been successfully reprogrammed via ectopic expression of transcription factors (TFs) from endothelial lineage. The reprogrammed ECs induced neovascularization in vitro and in vivo and thus demonstrated their therapeutic value in animal models of vascular insufficiency. Methods of delivering reprogramming factors include lentiviral or retroviral vectors and more clinically relevant, non-integrative adenoviral and episomal vectors. Most studies made use of fibroblast as a source cell for reprogramming, but reprogrammability of other clinically relevant source cell types has to be evaluated. Specific mechanisms and small molecules that are involved in the aforementioned processes tackles challenges associated with direct reprogramming efficiency and maintenance of reprogrammed EC characteristics. After all, this review provides summary of past and contemporary methods of direct endothelial reprogramming and discusses the future direction to overcome these challenges to acquire clinically applicable reprogrammed ECs.
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Affiliation(s)
- Seonggeon Cho
- Division of Cardiology, Department of Medicine, Emory University School of Medicine, Atlanta, GA, USA
| | - Parthasarathy Aakash
- Division of Cardiology, Department of Medicine, Emory University School of Medicine, Atlanta, GA, USA
| | - Sangho Lee
- Division of Cardiology, Department of Medicine, Emory University School of Medicine, Atlanta, GA, USA.
| | - Young-Sup Yoon
- Division of Cardiology, Department of Medicine, Emory University School of Medicine, Atlanta, GA, USA; Severance Biomedical Science Institute, Yonsei University College of Medicine, Seoul, Republic of Korea.
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20
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Noh KM, Park SJ, Moon SH, Jung SY. Extracellular matrix cues regulate the differentiation of pluripotent stem cell-derived endothelial cells. Front Cardiovasc Med 2023; 10:1169331. [PMID: 37435057 PMCID: PMC10330705 DOI: 10.3389/fcvm.2023.1169331] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2023] [Accepted: 05/23/2023] [Indexed: 07/13/2023] Open
Abstract
The generation of endothelial cells (ECs) from human pluripotent stem cells (PSCs) has been a promising approach for treating cardiovascular diseases for several years. Human PSCs, particularly induced pluripotent stem cells (iPSCs), are an attractive source of ECs for cell therapy. Although there is a diversity of methods for endothelial cell differentiation using biochemical factors, such as small molecules and cytokines, the efficiency of EC production varies depending on the type and dose of biochemical factors. Moreover, the protocols in which most EC differentiation studies have been performed were in very unphysiological conditions that do not reflect the microenvironment of native tissue. The microenvironment surrounding stem cells exerts variable biochemical and biomechanical stimuli that can affect stem cell differentiation and behavior. The stiffness and components of the extracellular microenvironment are critical inducers of stem cell behavior and fate specification by sensing the extracellular matrix (ECM) cues, adjusting the cytoskeleton tension, and delivering external signals to the nucleus. Differentiation of stem cells into ECs using a cocktail of biochemical factors has been performed for decades. However, the effects of mechanical stimuli on endothelial cell differentiation remain poorly understood. This review provides an overview of the methods used to differentiate ECs from stem cells by chemical and mechanical stimuli. We also propose the possibility of a novel EC differentiation strategy using a synthetic and natural extracellular matrix.
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Affiliation(s)
- Kyung Mu Noh
- Stem Cell Research Institute, T&R Biofab Co. Ltd., Seongnam-si, Republic of Korea
| | - Soon-Jung Park
- Stem Cell Research Institute, T&R Biofab Co. Ltd., Seongnam-si, Republic of Korea
| | - Sung-Hwan Moon
- Department of Animal Science and Technology, College of Biotechnology and Natural Resources, Chung-Ang University, Anseong-si, Republic of Korea
| | - Seok Yun Jung
- Stem Cell Research Institute, T&R Biofab Co. Ltd., Seongnam-si, Republic of Korea
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21
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Lou X, Tang Y, Ye L, Pretorius D, Fast VG, Kahn-Krell AM, Zhang J, Zhang J, Qiao A, Qin G, Kamp T, Thomson JA, Zhang J. Cardiac muscle patches containing four types of cardiac cells derived from human pluripotent stem cells improve recovery from cardiac injury in mice. Cardiovasc Res 2023; 119:1062-1076. [PMID: 36647784 PMCID: PMC10153642 DOI: 10.1093/cvr/cvad004] [Citation(s) in RCA: 24] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/11/2022] [Revised: 10/24/2022] [Accepted: 11/04/2022] [Indexed: 01/18/2023] Open
Abstract
AIMS We have shown that human cardiac muscle patches (hCMPs) containing three different types of cardiac cells-cardiomyocytes (CMs), smooth muscle cells (SMCs), and endothelial cells (ECs), all of which were differentiated from human pluripotent stem cells (hPSCs)-significantly improved cardiac function, infarct size, and hypertrophy in a pig model of myocardial infarction (MI). However, hPSC-derived CMs (hPSC-CMs) are phenotypically immature, which may lead to arrhythmogenic concerns; thus, since hPSC-derived cardiac fibroblasts (hPSC-CFs) appear to enhance the maturity of hPSC-CMs, we compared hCMPs containing hPSC-CMs, -SMCs, -ECs, and -CFs (4TCC-hCMPs) with a second hCMP construct that lacked hPSC-CFs but was otherwise identical [hCMP containing hPSC-CMs, -AECs, and -SMCs (3TCC-hCMPs)]. METHODS AND RESULTS hCMPs were generated in a fibrin scaffold. MI was induced in severe combined immunodeficiency (SCID) mice through permanent coronary artery (left anterior descending) ligation, followed by treatment with cardiac muscle patches. Animal groups included: MI heart treated with 3TCC-hCMP; with 4TCC-hCMP; MI heart treated with no patch (MI group) and sham group. Cardiac function was evaluated using echocardiography, and cell engraftment rate and infarct size were evaluated histologically at 4 weeks after patch transplantation. The results from experiments in cultured hCMPs demonstrate that the inclusion of cardiac fibroblast in 4TCC-hCMPs had (i) better organized sarcomeres; (ii) abundant structural, metabolic, and ion-channel markers of CM maturation; and (iii) greater conduction velocities (31 ± 3.23 cm/s, P < 0.005) and action-potential durations (APD50 = 365 ms ± 2.649, P < 0.0001; APD = 408 ms ± 2.757, P < 0.0001) than those (velocity and APD time) in 3TCC-hCMPs. Furthermore, 4TCC-hCMPs transplantation resulted in better cardiac function [ejection fraction (EF) = 49.18% ± 0.86, P < 0.05], reduced infarct size (22.72% ± 0.98, P < 0.05), and better engraftment (15.99% ± 1.56, P < 0.05) when compared with 3TCC-hCMPs (EF = 41.55 ± 0.92%, infarct size = 39.23 ± 4.28%, and engraftment = 8.56 ± 1.79%, respectively). CONCLUSION Collectively, these observations suggest that the inclusion of hPSC-CFs during hCMP manufacture promotes hPSC-CM maturation and increases the potency of implanted hCMPs for improving cardiac recovery in mice model of MI.
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Affiliation(s)
- Xi Lou
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Yawen Tang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Lei Ye
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Danielle Pretorius
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Vladimir G Fast
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Asher M Kahn-Krell
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Jue Zhang
- Morgridge Institute for Research, Madison, WI 53715, USA
| | - Jianhua Zhang
- Division of Cardiovascular Medicine, Department of Medicine, University of Wisconsin-Madison, Madison, WI, USA
- Department of Cell and Regenerative Biology, University of Wisconsin-Madison, Madison, WI, USA
| | - Aijun Qiao
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Gangjian Qin
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
| | - Timothy Kamp
- Division of Cardiovascular Medicine, Department of Medicine, University of Wisconsin-Madison, Madison, WI, USA
- Department of Cell and Regenerative Biology, University of Wisconsin-Madison, Madison, WI, USA
| | - James A Thomson
- Morgridge Institute for Research, Madison, WI 53715, USA
- Department of Cell and Regenerative Biology, University of Wisconsin-Madison, Madison, WI, USA
| | - Jianyi Zhang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, 1670 University Boulevard, Volker Hall G094J, Birmingham, AL 35294, USA
- Department of Medicine, School of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, USA
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22
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Liver X receptor controls follicular helper T cell differentiation via repression of TCF-1. Proc Natl Acad Sci U S A 2023; 120:e2213793120. [PMID: 36802434 PMCID: PMC9992818 DOI: 10.1073/pnas.2213793120] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/23/2023] Open
Abstract
Liver X receptor (LXR) is a critical regulator of cholesterol homeostasis that inhibits T cell receptor (TCR)-induced proliferation by altering intracellular sterol metabolism. However, the mechanisms by which LXR regulates helper T cell subset differentiation remain unclear. Here, we demonstrate that LXR is a crucial negative regulator of follicular helper T (Tfh) cells in vivo. Both mixed bone marrow chimera and antigen-specific T cell adoptive cotransfer studies show a specific increase in Tfh cells among LXRβ-deficient CD4+ T cell population in response to immunization and lymphocytic choriomeningitis mammarenavirus (LCMV) infection. Mechanistically, LXRβ-deficient Tfh cells express augmented levels of T cell factor 1 (TCF-1) but comparable levels of Bcl6, CXCR5, and PD-1 in comparison with those of LXRβ-sufficient Tfh cells. Loss of LXRβ confers inactivation of GSK3β induced by either AKT/Extracellular signal-regulated kinase (ERK) activation or Wnt/β-catenin pathway, leading to elevated TCF-1 expression in CD4+ T cells. Conversely, ligation of LXR represses TCF-1 expression and Tfh cell differentiation in both murine and human CD4+ T cells. LXR agonist significantly diminishes Tfh cells and the levels of antigen-specific IgG upon immunization. These findings unveil a cell-intrinsic regulatory function of LXR in Tfh cell differentiation via the GSK3β-TCF1 pathway, which may serve as a promising target for pharmacological intervention in Tfh-mediated diseases.
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23
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Bulut M, Vila Cuenca M, de Graaf M, van den Hil FE, Mummery CL, Orlova VV. Three-Dimensional Vessels-on-a-Chip Based on hiPSC-derived Vascular Endothelial and Smooth Muscle Cells. Curr Protoc 2022; 2:e564. [PMID: 36250774 PMCID: PMC11648816 DOI: 10.1002/cpz1.564] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/16/2023]
Abstract
Blood vessels are composed of endothelial cells (ECs) that form the inner vessel wall and mural cells that cover the ECs to mediate their stabilization. Crosstalk between ECs and VSMCs while the ECs undergo microfluidic flow is vital for the function and integrity of blood vessels. Here, we describe a protocol to generate three-dimensional (3D) engineered vessels-on-chip (VoCs) composed of vascular cells derived from human induced pluripotent stem cells (hiPSCs). We first describe protocols for robust differentiation of vascular smooth muscle cells (hiPSC-VSMCs) from hiPSCs that are effective across multiple hiPSC lines. Second, we describe the fabrication of a simple microfluidic device consisting of a single collagen lumen that can act as a cell scaffold and support fluid flow using the viscous finger patterning (VFP) technique. After the channel is seeded sequentially with hiPSC-derived ECs (hiPSC-ECs) and hiPSC-VSMCs, a stable EC barrier covered by VSMCs lines the collagen lumen. We demonstrate that this 3D VoC model can recapitulate physiological cell-cell interaction and can be perfused under physiological shear stress using a microfluidic pump. The uniform geometry of the vessel lumens allows precise control of flow dynamics. We have thus developed a robust protocol to generate an entirely isogenic hiPSC-derived 3D VoC model, which could be valuable for studying vessel barrier function and physiology in healthy or disease states. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Differentiation of hiPSC-VSMCs Support Protocol 1: Characterization of hiPSC-NCCs and hiPSC-VSMCs Support Protocol 2: Preparation of cryopreserved hiPSC-VSMCs and hiPSC-ECs for VoC culture Basic Protocol 2: Generation of 3D VoC model composed of hiPSC-ECs and hiPSC-VSMCs Support Protocol 3: Structural characterization of 3D VoC model.
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Affiliation(s)
- Merve Bulut
- Department of Anatomy and EmbryologyLeiden University Medical CenterLeidenThe Netherlands
| | - Marc Vila Cuenca
- Department of Anatomy and EmbryologyLeiden University Medical CenterLeidenThe Netherlands
| | - Mees de Graaf
- Department of Anatomy and EmbryologyLeiden University Medical CenterLeidenThe Netherlands
| | | | - Christine L. Mummery
- Department of Anatomy and EmbryologyLeiden University Medical CenterLeidenThe Netherlands
- Department of Applied Stem Cell TechnologiesUniversity of TwenteEnschedeThe Netherlands
| | - Valeria V. Orlova
- Department of Anatomy and EmbryologyLeiden University Medical CenterLeidenThe Netherlands
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Functional Characterization of Human Induced Pluripotent Stem Cell-Derived Endothelial Cells. Int J Mol Sci 2022; 23:ijms23158507. [PMID: 35955642 PMCID: PMC9368986 DOI: 10.3390/ijms23158507] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2022] [Revised: 07/27/2022] [Accepted: 07/29/2022] [Indexed: 12/03/2022] Open
Abstract
Endothelial cells derived from human induced pluripotent stem cells (hiPSC-ECs) provide a new opportunity for mechanistic research on vascular regeneration and drug screening. However, functions of hiPSC-ECs still need to be characterized. The objective of this study was to investigate electrophysiological and functional properties of hiPSC-ECs compared with primary human cardiac microvascular endothelial cells (HCMECs), mainly focusing on ion channels and membrane receptor signaling, as well as specific cell functions. HiPSC-ECs were derived from hiPS cells that were generated from human skin fibroblasts of three independent healthy donors. Phenotypic and functional comparison to HCMECs was performed by flow cytometry, immunofluorescence staining, quantitative reverse-transcription polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA), tube formation, LDL uptake, exosome release assays and, importantly, patch clamp techniques. HiPSC-ECs were successfully generated from hiPS cells and were identified by endothelial markers. The mRNA levels of KCNN2, KCNN4, KCNMA1, TRPV2, and SLC8A1 in hiPSC-ECs were significantly higher than HCMECs. AT1 receptor mRNA level in hiPSC-ECs was higher than in HCMECs. AT2 receptor mRNA level was the highest among all receptors. Adrenoceptor ADRA2 expression in hiPSC-ECs was lower than in HCMECs, while ADRA1, ADRB1, ADRB2, and G-protein GNA11 and Gai expression were similar in both cell types. The expression level of muscarinic and dopamine receptors CHRM3, DRD2, DRD3, and DRD4 in hiPSC-ECs were significantly lower than in HCMECs. The functional characteristics of endothelial cells, such as tube formation and LDL uptake assay, were not statistically different between hiPSC-ECs and HCMECs. Phenylephrine similarly increased the release of the vasoconstrictor endothelin-1 (ET-1) in hiPSC-ECs and HCMECs. Acetylcholine also similarly increased nitric oxide generation in hiPSC-ECs and HCMECs. The resting potentials (RPs), ISK1–3, ISK4 and IK1 were similar in hiPSC-ECs and HCMECs. IBK was larger and IKATP was smaller in hiPSC-ECs. In addition, we also noted a higher expression level of exosomes marker CD81 in hiPSC-ECs and a higher expression of CD9 and CD63 in HCMECs. However, the numbers of exosomes extracted from both types of cells did not differ significantly. The study demonstrates that hiPSC-ECs are similar to native endothelial cells in ion channel function and membrane receptor-coupled signaling and physiological cell functions, although some differences exist. This information may be helpful for research using hiPSC-ECs.
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25
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Vargas-Valderrama A, Ponsen AC, Le Gall M, Clay D, Jacques S, Manoliu T, Rouffiac V, Ser-le-Roux K, Quivoron C, Louache F, Uzan G, Mitjavila-Garcia MT, Oberlin E, Guenou H. Endothelial and hematopoietic hPSCs differentiation via a hematoendothelial progenitor. Stem Cell Res Ther 2022; 13:254. [PMID: 35715824 PMCID: PMC9205076 DOI: 10.1186/s13287-022-02925-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2022] [Accepted: 05/29/2022] [Indexed: 11/10/2022] Open
Abstract
Background hPSC-derived endothelial and hematopoietic cells (ECs and HCs) are an interesting source of cells for tissue engineering. Despite their close spatial and temporal embryonic development, current hPSC differentiation protocols are specialized in only one of these lineages. In this study, we generated a hematoendothelial population that could be further differentiated in vitro to both lineages.
Methods Two hESCs and one hiPSC lines were differentiated into a hematoendothelial population, hPSC-ECs and blast colonies (hPSC-BCs) via CD144+-embryoid bodies (hPSC-EBs). hPSC-ECs were characterized by endothelial colony-forming assay, LDL uptake assay, endothelial activation by TNF-α, nitric oxide detection and Matrigel-based tube formation. Hematopoietic colony-forming cell assay was performed from hPSC-BCs. Interestingly, we identified a hPSC-BC population characterized by the expression of both CD144 and CD45. hPSC-ECs and hPSC-BCs were analyzed by flow cytometry and RT-qPCR; in vivo experiments have been realized by ischemic tissue injury model on a mouse dorsal skinfold chamber and hematopoietic reconstitution in irradiated immunosuppressed mouse from hPSC-ECs and hPSC-EB-CD144+, respectively. Transcriptomic analyses were performed to confirm the endothelial and hematopoietic identity of hESC-derived cell populations by comparing them against undifferentiated hESC, among each other’s (e.g. hPSC-ECs vs. hPSC-EB-CD144+) and against human embryonic liver (EL) endothelial, hematoendothelial and hematopoietic cell subpopulations.
Results A hematoendothelial population was obtained after 84 h of hPSC-EBs formation under serum-free conditions and isolated based on CD144 expression. Intrafemorally injection of hPSC-EB-CD144+ contributed to the generation of CD45+ human cells in immunodeficient mice suggesting the existence of hemogenic ECs within hPSC-EB-CD144+. Endothelial differentiation of hPSC-EB-CD144+ yields a population of > 95% functional ECs in vitro. hPSC-ECs derived through this protocol participated at the formation of new vessels in vivo in a mouse ischemia model. In vitro, hematopoietic differentiation of hPSC-EB-CD144+ generated an intermediate population of > 90% CD43+ hPSC-BCs capable to generate myeloid and erythroid colonies. Finally, the transcriptomic analyses confirmed the hematoendothelial, endothelial and hematopoietic identity of hPSC-EB-CD144+, hPSC-ECs and hPSC-BCs, respectively, and the similarities between hPSC-BC-CD144+CD45+, a subpopulation of hPSC-BCs, and human EL hematopoietic stem cells/hematopoietic progenitors.
Conclusion The present work reports a hPSC differentiation protocol into functional hematopoietic and endothelial cells through a hematoendothelial population. Both lineages were proven to display characteristics of physiological human cells, and therefore, they represent an interesting rapid source of cells for future cell therapy and tissue engineering. Supplementary Information The online version contains supplementary material available at 10.1186/s13287-022-02925-w.
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Affiliation(s)
| | - Anne-Charlotte Ponsen
- INSERM UMRS-MD 1197, Hôpital Paul Brousse, Université Paris-Saclay, 94807, Villejuif, France
| | - Morgane Le Gall
- Plateforme Protéomique 3P5-Proteom'IC, Institut Cochin, INSERM U1016, CNRS UMR8104, Université de Paris, 75014, Paris, France
| | - Denis Clay
- INSERM UMS-44, Hôpital Paul Brousse, Université Paris Sud-Université Paris-Saclay, 94807, Villejuif, France
| | - Sébastien Jacques
- Plateforme de Génomique- GENOM'IC, Institut Cochin, INSERM U1016, CNRS UMR8104, Université de Paris, 75014, Paris, France
| | - Tudor Manoliu
- Plate-forme Imagerie et Cytométrie, UMS AMMICa, Gustave Roussy, Université Paris-Saclay, 94805, Villejuif, France
| | - Valérie Rouffiac
- Plate-forme Imagerie et Cytométrie, UMS AMMICa, Gustave Roussy, Université Paris-Saclay, 94805, Villejuif, France
| | - Karine Ser-le-Roux
- INSERM, UMS AMMICa, Plate-forme d'Evaluation Préclinique, Gustave Roussy, 94807, Villejuif, France
| | - Cyril Quivoron
- Laboratoire d'Hématologie Translationnelle, Gustave Roussy, 94805, Villejuif, France
| | - Fawzia Louache
- INSERM UMRS-MD 1197, Hôpital Paul Brousse, Université Paris-Saclay, 94807, Villejuif, France
| | - Georges Uzan
- INSERM UMRS-MD 1197, Hôpital Paul Brousse, Université Paris-Saclay, 94807, Villejuif, France
| | | | - Estelle Oberlin
- INSERM UMRS-MD 1197, Hôpital Paul Brousse, Université Paris-Saclay, 94807, Villejuif, France
| | - Hind Guenou
- INSERM UMRS-MD 1197, Hôpital Paul Brousse, Université Paris-Saclay, 94807, Villejuif, France. .,Université d'Evry-Val-d'Essonne, Université Paris-Saclay, 91000, Evry, France.
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Hamad S, Derichsweiler D, Gaspar JA, Brockmeier K, Hescheler J, Sachinidis A, Pfannkuche KP. High-efficient serum-free differentiation of endothelial cells from human iPS cells. Stem Cell Res Ther 2022; 13:251. [PMID: 35690874 PMCID: PMC9188069 DOI: 10.1186/s13287-022-02924-x] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2022] [Accepted: 05/29/2022] [Indexed: 11/10/2022] Open
Abstract
Introduction Endothelial cells (ECs) form the inner lining of all blood vessels of the body play important roles in vascular tone regulation, hormone secretion, anticoagulation, regulation of blood cell adhesion and immune cell extravasation. Limitless ECs sources are required to further in vitro investigations of ECs’ physiology and pathophysiology as well as for tissue engineering approaches. Ideally, the differentiation protocol avoids animal-derived components such as fetal serum and yields ECs at efficiencies that make further sorting obsolete for most applications.
Method Human induced pluripotent stem cells (hiPSCs) are cultured under serum-free conditions and induced into mesodermal progenitor cells via stimulation of Wnt signaling for 24 h. Mesodermal progenitor cells are further differentiated into ECs by utilizing a combination of human vascular endothelial growth factor A165 (VEGF), basic fibroblast growth factor (bFGF), 8-Bromoadenosine 3′,5′-cyclic monophosphate sodium salt monohydrate (8Bro) and melatonin (Mel) for 48 h.
Result This combination generates hiPSC derived ECs (hiPSC-ECs) at a fraction of 90.9 ± 1.5% and is easily transferable from the two-dimensional (2D) monolayer into three-dimensional (3D) scalable bioreactor suspension cultures. hiPSC-ECs are positive for CD31, VE-Cadherin, von Willebrand factor and CD34. Furthermore, the majority of hiPSC-ECs express the vascular endothelial marker CD184 (CXCR4).
Conclusion The differentiation method presented here generates hiPSC-ECs in only 6 days, without addition of animal sera and at high efficiency, hence providing a scalable source of hiPSC-ECs.
Supplementary Information The online version contains supplementary material available at 10.1186/s13287-022-02924-x.
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Affiliation(s)
- Sarkawt Hamad
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany.,Biology Department, Faculty of Science, Soran University, Kurdistan Region, Soran, Iraq
| | - Daniel Derichsweiler
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany
| | - John Antonydas Gaspar
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany
| | - Konrad Brockmeier
- Department of Pediatric Cardiology, University Hospital of Cologne, Cologne, Germany
| | - Jürgen Hescheler
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany
| | - Agapios Sachinidis
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany.,Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany
| | - Kurt Paul Pfannkuche
- Medical Faculty, Center for Physiology and Pathophysiology, Institute for Neurophysiology, University of Cologne, Robert Koch Str. 39, 50931, Cologne, Germany. .,Department of Pediatric Cardiology, University Hospital of Cologne, Cologne, Germany. .,Marga-and-Walter-Boll Laboratory for Cardiac Tissue Engineering, University of Cologne, Cologne, Germany. .,Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany.
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Integrative epigenomic and transcriptomic analysis reveals the requirement of JUNB for hematopoietic fate induction. Nat Commun 2022; 13:3131. [PMID: 35668082 PMCID: PMC9170695 DOI: 10.1038/s41467-022-30789-4] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2021] [Accepted: 05/18/2022] [Indexed: 11/08/2022] Open
Abstract
Human pluripotent stem cell differentiation towards hematopoietic progenitor cell can serve as an in vitro model for human embryonic hematopoiesis, but the dynamic change of epigenome and transcriptome remains elusive. Here, we systematically profile the chromatin accessibility, H3K4me3 and H3K27me3 modifications, and the transcriptome of intermediate progenitors during hematopoietic progenitor cell differentiation in vitro. The integrative analyses reveal sequential opening-up of regions for the binding of hematopoietic transcription factors and stepwise epigenetic reprogramming of bivalent genes. Single-cell analysis of cells undergoing the endothelial-to-hematopoietic transition and comparison with in vivo hemogenic endothelial cells reveal important features of in vitro and in vivo hematopoiesis. We find that JUNB is an essential regulator for hemogenic endothelium specialization and endothelial-to-hematopoietic transition. These studies depict an epigenomic roadmap from human pluripotent stem cells to hematopoietic progenitor cells, which may pave the way to generate hematopoietic progenitor cells with improved developmental potentials.
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Macklin BL, Lin YY, Emmerich K, Wisniewski E, Polster BM, Konstantopoulos K, Mumm JS, Gerecht S. Intrinsic epigenetic control of angiogenesis in induced pluripotent stem cell-derived endothelium regulates vascular regeneration. NPJ Regen Med 2022; 7:28. [PMID: 35551465 PMCID: PMC9098630 DOI: 10.1038/s41536-022-00223-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/21/2021] [Accepted: 04/14/2022] [Indexed: 11/17/2022] Open
Abstract
Human-induced pluripotent stem cell-derived endothelial cells (iECs) provide opportunities to study vascular development and regeneration, develop cardiovascular therapeutics, and engineer model systems for drug screening. The differentiation and characterization of iECs are well established; however, the mechanisms governing their angiogenic phenotype remain unknown. Here, we aimed to determine the angiogenic phenotype of iECs and the regulatory mechanism controlling their regenerative capacity. In a comparative study with HUVECs, we show that iECs increased expression of vascular endothelial growth factor receptor 2 (VEGFR2) mediates their highly angiogenic phenotype via regulation of glycolysis enzymes, filopodia formation, VEGF mediated migration, and robust sprouting. We find that the elevated expression of VEGFR2 is epigenetically regulated via intrinsic acetylation of histone 3 at lysine 27 by histone acetyltransferase P300. Utilizing a zebrafish xenograft model, we demonstrate that the ability of iECs to promote the regeneration of the amputated fin can be modulated by P300 activity. These findings demonstrate how the innate epigenetic status of iECs regulates their phenotype with implications for their therapeutic potential.
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Affiliation(s)
- Bria L Macklin
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD, 21218, USA
| | - Ying-Yu Lin
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD, 21218, USA
| | - Kevin Emmerich
- Department of Ophthalmology, Wilmer Eye Institute and McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA
| | - Emily Wisniewski
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD, 21218, USA
| | - Brian M Polster
- Department of Anesthesiology and Center for Shock, Trauma, and Anesthesiology Research, University of Maryland School of Medicine, Baltimore, MD, 21201, USA
| | - Konstantinos Konstantopoulos
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD, 21218, USA
| | - Jeff S Mumm
- Department of Ophthalmology, Wilmer Eye Institute and McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA
| | - Sharon Gerecht
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD, 21218, USA. .,Department of Biomedical Engineering, Duke University, Durham, NC, 27708, USA.
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29
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Vivas A, Mikhal J, Ong GM, Eigenbrodt A, van der Meer AD, Aquarius R, Geurts BJ, Boogaarts HD. Aneurysm-on-a-Chip: Setting Flow Parameters for Microfluidic Endothelial Cultures Based on Computational Fluid Dynamics Modeling of Intracranial Aneurysms. Brain Sci 2022; 12:603. [PMID: 35624990 PMCID: PMC9139202 DOI: 10.3390/brainsci12050603] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2022] [Revised: 04/20/2022] [Accepted: 04/27/2022] [Indexed: 11/30/2022] Open
Abstract
Intracranial aneurysms are pouch-like extrusions from the vessels at the base of the brain which can rupture and cause a subarachnoid hemorrhage. The pathophysiological mechanism of aneurysm formation is thought to be a consequence of blood flow (hemodynamic) induced changes on the endothelium. In this study, the results of a personalized aneurysm-on-a-chip model using patient-specific flow parameters and patient-specific cells are presented. CT imaging was used to calculate CFD parameters using an immersed boundary method. A microfluidic device either cultured with human umbilical vein endothelial cells (HUVECs) or human induced pluripotent stem cell-derived endothelial cells (hiPSC-EC) was used. Both types of endothelial cells were exposed for 24 h to either 0.03 Pa or 1.5 Pa shear stress, corresponding to regions of low shear and high shear in the computational aneurysm model, respectively. As a control, both cell types were also cultured under static conditions for 24 h as a control. Both HUVEC and hiPSC-EC cultures presented as confluent monolayers with no particular cell alignment in static or low shear conditions. Under high shear conditions HUVEC elongated and aligned in the direction of the flow. HiPSC-EC exhibited reduced cell numbers, monolayer gap formation and cells with aberrant, spread-out morphology. Future research should focus on hiPSC-EC stabilization to allow personalized intracranial aneurysm models.
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Affiliation(s)
- Aisen Vivas
- Applied Stem Cell Technologies, University of Twente, 7522 NB Enschede, The Netherlands; (A.V.); (A.E.); (A.D.v.d.M.)
| | - Julia Mikhal
- Multiscale Modeling and Simulation Group, Department of Applied Mathematics, University of Twente, 7522 NB Enschede, The Netherlands; (J.M.); (G.M.O.); (B.J.G.)
| | - Gabriela M. Ong
- Multiscale Modeling and Simulation Group, Department of Applied Mathematics, University of Twente, 7522 NB Enschede, The Netherlands; (J.M.); (G.M.O.); (B.J.G.)
| | - Anna Eigenbrodt
- Applied Stem Cell Technologies, University of Twente, 7522 NB Enschede, The Netherlands; (A.V.); (A.E.); (A.D.v.d.M.)
| | - Andries D. van der Meer
- Applied Stem Cell Technologies, University of Twente, 7522 NB Enschede, The Netherlands; (A.V.); (A.E.); (A.D.v.d.M.)
| | - Rene Aquarius
- Department of Neurosurgery, Radboud University Medical Center, 6525 XZ Nijmegen, The Netherlands;
| | - Bernard J. Geurts
- Multiscale Modeling and Simulation Group, Department of Applied Mathematics, University of Twente, 7522 NB Enschede, The Netherlands; (J.M.); (G.M.O.); (B.J.G.)
| | - Hieronymus D. Boogaarts
- Department of Neurosurgery, Radboud University Medical Center, 6525 XZ Nijmegen, The Netherlands;
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Seymour AJ, Westerfield AD, Cornelius VC, Skylar-Scott MA, Heilshorn SC. Bioprinted microvasculature: progressing from structure to function. Biofabrication 2022; 14:10.1088/1758-5090/ac4fb5. [PMID: 35086069 PMCID: PMC8988885 DOI: 10.1088/1758-5090/ac4fb5] [Citation(s) in RCA: 24] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Accepted: 01/27/2022] [Indexed: 11/12/2022]
Abstract
Three-dimensional (3D) bioprinting seeks to unlock the rapid generation of complex tissue constructs, but long-standing challenges with efficientin vitromicrovascularization must be solved before this can become a reality. Microvasculature is particularly challenging to biofabricate due to the presence of a hollow lumen, a hierarchically branched network topology, and a complex signaling milieu. All of these characteristics are required for proper microvascular-and, thus, tissue-function. While several techniques have been developed to address distinct portions of this microvascularization challenge, no single approach is capable of simultaneously recreating all three microvascular characteristics. In this review, we present a three-part framework that proposes integration of existing techniques to generate mature microvascular constructs. First, extrusion-based 3D bioprinting creates a mesoscale foundation of hollow, endothelialized channels. Second, biochemical and biophysical cues induce endothelial sprouting to create a capillary-mimetic network. Third, the construct is conditioned to enhance network maturity. Across all three of these stages, we highlight the potential for extrusion-based bioprinting to become a central technique for engineering hierarchical microvasculature. We envision that the successful biofabrication of functionally engineered microvasculature will address a critical need in tissue engineering, and propel further advances in regenerative medicine andex vivohuman tissue modeling.
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Affiliation(s)
- Alexis J. Seymour
- Department of Bioengineering, Stanford University, 443 Via Ortega, Shriram Center Room 119, Stanford, CA 94305, USA
| | - Ashley D. Westerfield
- Department of Bioengineering, Stanford University, 443 Via Ortega, Shriram Center Room 119, Stanford, CA 94305, USA
| | - Vincent C. Cornelius
- Department of Bioengineering, Stanford University, 443 Via Ortega, Shriram Center Room 119, Stanford, CA 94305, USA
| | - Mark A. Skylar-Scott
- Department of Bioengineering, Stanford University, 443 Via Ortega, Shriram Center Room 119, Stanford, CA 94305, USA
| | - Sarah C. Heilshorn
- Department of Materials Science & Engineering, Stanford University, 476 Lomita Mall, McCullough Room 246, Stanford, CA 94305, USA
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31
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Stem cells, organoids, and organ-on-a-chip models for personalized in vitro drug testing. CURRENT OPINION IN TOXICOLOGY 2021. [DOI: 10.1016/j.cotox.2021.08.006] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
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Zhang J, Bolli R, Garry DJ, Marbán E, Menasché P, Zimmermann WH, Kamp TJ, Wu JC, Dzau VJ. Basic and Translational Research in Cardiac Repair and Regeneration: JACC State-of-the-Art Review. J Am Coll Cardiol 2021; 78:2092-2105. [PMID: 34794691 PMCID: PMC9116459 DOI: 10.1016/j.jacc.2021.09.019] [Citation(s) in RCA: 64] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/02/2021] [Revised: 09/13/2021] [Accepted: 09/13/2021] [Indexed: 12/25/2022]
Abstract
This paper aims to provide an important update on the recent preclinical and clinical trials using cell therapy strategies and engineered heart tissues for the treatment of postinfarction left ventricular remodeling and heart failure. In addition to the authors’ own works and opinions on the roadblocks of the field, they discuss novel approaches for cardiac remuscularization via the activation of proliferative mechanisms in resident cardiomyocytes or direct reprogramming of somatic cells into cardiomyocytes. This paper’s main mindset is to present current and future strategies in light of their implications for the design of future patient trials with the ultimate objective of facilitating the translation of discoveries in regenerative myocardial therapies to the clinic.
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Affiliation(s)
- Jianyi Zhang
- Department of Biomedical Engineering, School of Medicine, School of Engineering, The University of Alabama at Birmingham, Birmingham, Alabama, USA.
| | - Roberto Bolli
- Institute of Molecular Cardiology, University of Louisville, Louisville, Kentucky, USA
| | - Daniel J Garry
- Department of Medicine, Lillehei Heart Institute, University of Minnesota, Minneapolis, Minnesota, USA
| | - Eduardo Marbán
- Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles California, USA
| | - Philippe Menasché
- Department of Cardiovascular Surgery, Hôpital Européen Georges Pompidou, University of Paris, PARCC, INSERM, F-75015, Paris, France
| | - Wolfram-Hubertus Zimmermann
- Institute of Pharmacology and Toxicology, University Medical Center Göttingen, and DZHK (German Center for Cardiovascular Research), partner site Göttingen, Göttingen, Germany
| | - Timothy J Kamp
- Department of Medicine, School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Joseph C Wu
- Stanford Cardiovascular Institute, Stanford University School of Medicine, Stanford, California, USA
| | - Victor J Dzau
- Mandel Center for Hypertension Research, Duke Cardiovascular Center, Duke University School of Medicine, Durham, North Carolina, USA
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Sabatier P, Beusch CM, Saei AA, Aoun M, Moruzzi N, Coelho A, Leijten N, Nordenskjöld M, Micke P, Maltseva D, Tonevitsky AG, Millischer V, Carlos Villaescusa J, Kadekar S, Gaetani M, Altynbekova K, Kel A, Berggren PO, Simonson O, Grinnemo KH, Holmdahl R, Rodin S, Zubarev RA. An integrative proteomics method identifies a regulator of translation during stem cell maintenance and differentiation. Nat Commun 2021; 12:6558. [PMID: 34772928 PMCID: PMC8590018 DOI: 10.1038/s41467-021-26879-4] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2020] [Accepted: 10/25/2021] [Indexed: 12/21/2022] Open
Abstract
Detailed characterization of cell type transitions is essential for cell biology in general and particularly for the development of stem cell-based therapies in regenerative medicine. To systematically study such transitions, we introduce a method that simultaneously measures protein expression and thermal stability changes in cells and provide the web-based visualization tool ProteoTracker. We apply our method to study differences between human pluripotent stem cells and several cell types including their parental cell line and differentiated progeny. We detect alterations of protein properties in numerous cellular pathways and components including ribosome biogenesis and demonstrate that modulation of ribosome maturation through SBDS protein can be helpful for manipulating cell stemness in vitro. Using our integrative proteomics approach and the web-based tool, we uncover a molecular basis for the uncoupling of robust transcription from parsimonious translation in stem cells and propose a method for maintaining pluripotency in vitro.
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Affiliation(s)
- Pierre Sabatier
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
| | - Christian M Beusch
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
| | - Amir A Saei
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
- Department of Cell Biology, Harvard Medical School, Boston, MA, USA
| | - Mike Aoun
- Division of Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
| | - Noah Moruzzi
- The Rolf Luft Research Center for Diabetes and Endocrinology, Department of Molecular Medicine and Surgery, Karolinska Institute, Stockholm, 17176, Sweden
| | - Ana Coelho
- Division of Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
| | - Niels Leijten
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, Utrecht, 3584 CH, The Netherlands
| | - Magnus Nordenskjöld
- Center for Molecular Medicine, Karolinska University Hospital, Stockholm, 171 76, Sweden
- Department of Molecular Medicine and Surgery, Karolinska Institute, Stockholm, 17177, Sweden
- Department of Clinical Genetics, Karolinska University Hospital, Stockholm, 171 76, Sweden
| | - Patrick Micke
- Immunology, Genetics and Pathology, Rudbecklaboratoriet, Uppsala University, Uppsala, 751 85, Sweden
| | - Diana Maltseva
- Faculty of biology and biotechnology, National Research University Higher School of Economics, Myasnitskaya Street, 13/4, Moscow, 117997, Russia
| | - Alexander G Tonevitsky
- Faculty of biology and biotechnology, National Research University Higher School of Economics, Myasnitskaya Street, 13/4, Moscow, 117997, Russia
- Scientific Research Center Bioclinicum, Ugreshskaya str. 2/85, Moscow, 115088, Russia
| | - Vincent Millischer
- Department of Molecular Medicine and Surgery, Karolinska Institute, Stockholm, 17177, Sweden
- Translational Psychiatry, Center for Molecular Medicine, Karolinska University Hospital, Stockholm, 171 76, Sweden
- Department of Psychiatry and Psychotherapy, Medical University of Vienna, Vienna, 1090, Austria
| | - J Carlos Villaescusa
- Neurogenetic Unit, Department of Molecular Medicine and Surgery, Karolinska University Hospital, Stockholm, 171 76, Sweden
- Stem Cell R&D-TRU, Novo Nordisk A/S, Måløv, Denmark
| | - Sandeep Kadekar
- Department of Surgical Sciences, Uppsala University, Uppsala, 752 37, Sweden
| | - Massimiliano Gaetani
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
- Chemical Proteomics Core Facility, Division of Physiological Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, 171 77, Sweden
- Chemical Proteomics, Science for Life Laboratory (SciLifeLab), Stockholm, 17 177, Sweden
| | | | - Alexander Kel
- geneXplain GmbH, Am Exer 19B, 38302, Wolfenbuettel, Germany
| | - Per-Olof Berggren
- The Rolf Luft Research Center for Diabetes and Endocrinology, Department of Molecular Medicine and Surgery, Karolinska Institute, Stockholm, 17176, Sweden
| | - Oscar Simonson
- Department of Surgical Sciences, Uppsala University, Uppsala, 752 37, Sweden
- Department of Cardio-thoracic Surgery and Anesthesiology, Uppsala University Hospital, Uppsala, 751 85, Sweden
| | - Karl-Henrik Grinnemo
- Department of Surgical Sciences, Uppsala University, Uppsala, 752 37, Sweden
- Department of Cardio-thoracic Surgery and Anesthesiology, Uppsala University Hospital, Uppsala, 751 85, Sweden
| | - Rikard Holmdahl
- Division of Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden
| | - Sergey Rodin
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden.
- Department of Surgical Sciences, Uppsala University, Uppsala, 752 37, Sweden.
- Department of Cardio-thoracic Surgery and Anesthesiology, Uppsala University Hospital, Uppsala, 751 85, Sweden.
| | - Roman A Zubarev
- Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, 17177, Sweden.
- Department of Pharmacological & Technological Chemistry, I.M. Sechenov First Moscow State Medical University, Moscow, 119146, Russia.
- The National Medical Research Center for Endocrinology, Moscow, 115478, Russia.
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Human Induced Pluripotent Stem Cell-Derived Vascular Cells: Recent Progress and Future Directions. J Cardiovasc Dev Dis 2021; 8:jcdd8110148. [PMID: 34821701 PMCID: PMC8622843 DOI: 10.3390/jcdd8110148] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2021] [Revised: 11/01/2021] [Accepted: 11/02/2021] [Indexed: 12/12/2022] Open
Abstract
Human induced pluripotent stem cells (hiPSCs) hold great promise for cardiovascular regeneration following ischemic injury. Considerable effort has been made toward the development and optimization of methods to differentiate hiPSCs into vascular cells, such as endothelial and smooth muscle cells (ECs and SMCs). In particular, hiPSC-derived ECs have shown robust potential for promoting neovascularization in animal models of cardiovascular diseases, potentially achieving significant and sustained therapeutic benefits. However, the use of hiPSC-derived SMCs that possess high therapeutic relevance is a relatively new area of investigation, still in the earlier investigational stages. In this review, we first discuss different methodologies to derive vascular cells from hiPSCs with a particular emphasis on the role of key developmental signals. Furthermore, we propose a standardized framework for assessing and defining the EC and SMC identity that might be suitable for inducing tissue repair and regeneration. We then highlight the regenerative effects of hiPSC-derived vascular cells on animal models of myocardial infarction and hindlimb ischemia. Finally, we address several obstacles that need to be overcome to fully implement the use of hiPSC-derived vascular cells for clinical application.
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Qin K, Lei J, Yang J. The Differentiation of Pluripotent Stem Cells towards Endothelial Progenitor Cells - Potential Application in Pulmonary Arterial Hypertension. Int J Stem Cells 2021; 15:122-135. [PMID: 34711697 PMCID: PMC9148829 DOI: 10.15283/ijsc21044] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2021] [Revised: 08/26/2021] [Accepted: 09/02/2021] [Indexed: 11/22/2022] Open
Abstract
Background and Objectives Endothelial progenitor cells (EPCs) and endothelial cells (ECs) have been applied in the clinic to treat pulmonary arterial hypertension (PAH), a disease characterized by disordered pulmonary vasculature. However, the lack of sufficient transplantable cells before the deterioration of disease condition is a current limitation to apply cell therapy in patients. It is necessary to differentiate pluripotent stem cells (PSCs) into EPCs and identify their characteristics. Methods and Results Comparing previously reported methods of human PSCs-derived ECs, we optimized a highly efficient differentiation protocol to obtain cells that match the phenotype of isolated EPCs from healthy donors. The protocol is compatible with chemically defined medium (CDM), it could produce a large number of clinically applicable cells with low cost. Moreover, we also found PSCs-derived EPCs express CD133, have some characteristics of mesenchymal stem cells and are capable of homing to repair blood vessels in zebrafish xenograft assays. In addition, we further revealed that IPAH PSCs-derived EPCs have higher expression of proliferation-related genes and lower expression of immune-related genes than normal EPCs and PSCs-derived EPCs through microarray analysis. Conclusions In conclusion, we optimized a highly efficient differentiation protocol to obtain PSCs-derived EPCs with the phenotypic and molecular characteristics of EPCs from healthy donors which distinguished them from EPCs from PAH.
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Affiliation(s)
- Kezhou Qin
- Department of Cell Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China
| | - Jia Lei
- Department of Physiology, and Department of Cardiology of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China
| | - Jun Yang
- Department of Cell Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.,Department of Physiology, and Department of Cardiology of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China
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Pappalardo A, Herron L, Alvarez Cespedes DE, Abaci HE. Quantitative Evaluation of Human Umbilical Vein and Induced Pluripotent Stem Cell-Derived Endothelial Cells as an Alternative Cell Source to Skin-Specific Endothelial Cells in Engineered Skin Grafts. Adv Wound Care (New Rochelle) 2021; 10:490-502. [PMID: 32870778 DOI: 10.1089/wound.2020.1163] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
Objective: We compared the capability of human umbilical vein endothelial cells (HUVECs), induced pluripotent stem cell (iPSC)-derived endothelial cells (iECs), and human dermal blood endothelial cells (HDBECs) to effectively vascularize engineered human skin constructs (HSCs) in vitro and on immunodeficient mice. Approach: We quantified the angiogenesis within HSCs both in vitro and in vivo through computational analyses of immunofluorescent (IF) staining. We assayed with real-time quantitative PCR (RT-qPCR) the expression of key endothelial, dermal, and epidermal genes in 2D culture and HSCs. Epidermal integrity and proliferation were also evaluated through haematoxylin and eosin staining, and IF staining. Results: IF confirmed iEC commitment to endothelial phenotype. RT-qPCR showed HUVECs and iECs immaturity compared with HDBECs. In vitro, the vascular network extension was comparable for HDBECs and HUVECs despite differences in vascular diameter, whereas iECs formed unorganized rudimentary tubular structures. In vivo, all ECs produced discrete vascular networks of varying dimensions. HUVECs and HDBECs maintained a higher proliferation of basal keratinocytes. HDBECs had the best impact on extracellular matrix expression, and epidermal proliferation and differentiation. Innovation: To our knowledge, this study represents the first direct and quantitative comparison of HDBECs, HUVECs, and iECs angiogenic performance in HSCs. Conclusions: Our data indicate that HUVECs and iECs can be an alternative cell source to HDBEC to promote the short-term viability of prevascularized engineered grafts. Nevertheless, HDBECs maintain their capillary identity and outperform other EC types in promoting the maturation of the dermis and epidermis. These intrinsic characteristics of HDBECs may influence the long-term function of skin grafts.
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Affiliation(s)
- Alberto Pappalardo
- Dermatology Department, Columbia University Medical Center, New York, New York, USA
| | - Lauren Herron
- Dermatology Department, Columbia University Medical Center, New York, New York, USA
| | | | - Hasan Erbil Abaci
- Dermatology Department, Columbia University Medical Center, New York, New York, USA
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37
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Huang Y, Qian JY, Cheng H, Li XM. Effects of shear stress on differentiation of stem cells into endothelial cells. World J Stem Cells 2021; 13:894-913. [PMID: 34367483 PMCID: PMC8316872 DOI: 10.4252/wjsc.v13.i7.894] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/27/2021] [Revised: 04/20/2021] [Accepted: 06/22/2021] [Indexed: 02/06/2023] Open
Abstract
Stem cell transplantation is an appealing potential therapy for vascular diseases and an indispensable key step in vascular tissue engineering. Substantial effort has been made to differentiate stem cells toward vascular cell phenotypes, including endothelial cells (ECs) and smooth muscle cells. The microenvironment of vascular cells not only contains biochemical factors that influence differentiation but also exerts hemodynamic forces, such as shear stress and cyclic strain. More recently, studies have shown that shear stress can influence the differentiation of stem cells toward ECs. A deep understanding of the responses and underlying mechanisms involved in this process is essential for clinical translation. This review highlights current data supporting the role of shear stress in stem cell differentiation into ECs. Potential mechanisms and signaling cascades for transducing shear stress into a biological signal are proposed. Further study of stem cell responses to shear stress will be necessary to apply stem cells for pharmacological applications and cardiovascular implants in the realm of regenerative medicine.
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Affiliation(s)
- Yan Huang
- Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, Beijing Advanced Innovation Center for Biomedical Engineering, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China
| | - Jia-Yi Qian
- Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, Beijing Advanced Innovation Center for Biomedical Engineering, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China
| | - Hong Cheng
- Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, Beijing Advanced Innovation Center for Biomedical Engineering, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China
| | - Xiao-Ming Li
- Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, Beijing Advanced Innovation Center for Biomedical Engineering, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China.
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38
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Fu M, Song J. Single-Cell Transcriptomics Reveals the Cellular Heterogeneity of Cardiovascular Diseases. Front Cardiovasc Med 2021; 8:643519. [PMID: 34179129 PMCID: PMC8225933 DOI: 10.3389/fcvm.2021.643519] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2020] [Accepted: 05/13/2021] [Indexed: 12/23/2022] Open
Abstract
"A world in a wild flower, and a bodhi in a leaf," small cells contain huge secrets. The vasculature is composed of many multifunctional cell subpopulations, each of which is involved in the occurrence and development of cardiovascular diseases. Single-cell transcriptomics captures the full picture of genes expressed within individual cells, identifies rare or de novo cell subpopulations, analyzes single-cell trajectory and stem cell or progenitor cell lineage conversion, and compares healthy tissue and disease-related tissue at single-cell resolution. Single-cell transcriptomics has had a profound effect on the field of cardiovascular research over the past decade, as evidenced by the construction of cardiovascular cell landscape, as well as the clarification of cardiovascular diseases and the mechanism of stem cell or progenitor cell differentiation. The classification and proportion of cell subpopulations in vasculature vary with species, location, genotype, and disease, exhibiting unique gene expression characteristics in organ development, disease progression, and regression. Specific gene markers are expected to be the diagnostic criteria, therapeutic targets, or prognostic indicators of diseases. Therefore, treatment of vascular disease still has lots of potentials to develop. Herein, we summarize the cell clusters and gene expression patterns in normal vasculature and atherosclerosis, aortic aneurysm, and pulmonary hypertension to reveal vascular heterogeneity and new regulatory factors of cardiovascular disease in the use of single-cell transcriptomics and discuss its current limitations and promising clinical potential.
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Affiliation(s)
- Mengxia Fu
- State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
- The Cardiomyopathy Research Group at Fuwai Hospital, Beijing, China
| | - Jiangping Song
- State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
- The Cardiomyopathy Research Group at Fuwai Hospital, Beijing, China
- Department of Cardiovascular Surgery, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
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39
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Hermsen J, Brown ME. Humanized Mouse Models for Evaluation of PSC Immunogenicity. ACTA ACUST UNITED AC 2021; 54:e113. [PMID: 32588980 DOI: 10.1002/cpsc.113] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
New human pluripotent stem cell (hPSC)-derived therapies are advancing to clinical trials at an increasingly rapid pace. In addition to ensuring that the therapies function properly, there is a critical need to investigate the human immune response to these cell products. A robust allogeneic (or autologous) immune response could swiftly eliminate an otherwise promising cell therapy, even in immunosuppressed patients. In coming years, researchers in the regenerative medicine field will need to utilize a number of in vitro and in vivo assays and models to evaluate and better understand hPSC immunogenicity. Humanized mouse models-mice engrafted with functional human immune cell types-are an important research tool for investigating the mechanisms of the adaptive immune response to hPSC therapies. This article provides an overview of humanized mouse models relevant to the study of hPSC immunogenicity and explores central considerations for investigators seeking to utilize these powerful models in their research. © 2020 Wiley Periodicals LLC.
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Affiliation(s)
- Jack Hermsen
- University of Wisconsin School of Medicine and Public Health Western Clinical Campus, Madison, Wisconsin
| | - Matthew E Brown
- University of Wisconsin School of Medicine and Public Health Western Clinical Campus, Madison, Wisconsin
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40
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Nguyen J, Lin YY, Gerecht S. The next generation of endothelial differentiation: Tissue-specific ECs. Cell Stem Cell 2021; 28:1188-1204. [PMID: 34081899 DOI: 10.1016/j.stem.2021.05.002] [Citation(s) in RCA: 32] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
Endothelial cells (ECs) sense and respond to fluid flow and regulate immune cell trafficking in all organs. Despite sharing the same mesodermal origin, ECs exhibit heterogeneous tissue-specific characteristics. Human pluripotent stem cells (hPSCs) can potentially be harnessed to capture this heterogeneity and further elucidate endothelium behavior to satisfy the need for increased accuracy and breadth of disease models and therapeutics. Here, we review current strategies for hPSC differentiation to blood vascular ECs and their maturation into continuous, fenestrated, and sinusoidal tissues. We then discuss the contribution of hPSC-derived ECs to recent advances in organoid development and organ-on-chip approaches.
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Affiliation(s)
- Jane Nguyen
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD 21218, USA
| | - Ying-Yu Lin
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD 21218, USA
| | - Sharon Gerecht
- Department of Chemical and Biomolecular Engineering, The Institute for NanoBioTechnology, Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, MD 21218, USA; Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, MD 21218, USA; Department of Biomedical Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
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41
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Su L, Kong X, Loo SJ, Gao Y, Kovalik JP, Su X, Ma J, Ye L. Diabetic Endothelial Cells Differentiated From Patient iPSCs Show Dysregulated Glycine Homeostasis and Senescence Associated Phenotypes. Front Cell Dev Biol 2021; 9:667252. [PMID: 34136485 PMCID: PMC8201091 DOI: 10.3389/fcell.2021.667252] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2021] [Accepted: 03/30/2021] [Indexed: 11/13/2022] Open
Abstract
Induced pluripotent stem cells derived cells (iPSCs) not only can be used for personalized cell transfer therapy, but also can be used for modeling diseases for drug screening and discovery in vitro. Although prior studies have characterized the function of rodent iPSCs derived endothelial cells (ECs) in diabetes or metabolic syndrome, feature phenotypes are largely unknown in hiPSC-ECs from patients with diabetes. Here, we used hiPSC lines from patients with type 2 diabetes mellitus (T2DM) and differentiated them into ECs (dia-hiPSC-ECs). We found that dia-hiPSC-ECs had disrupted glycine homeostasis, increased senescence, and impaired mitochondrial function and angiogenic potential as compared with healthy hiPSC-ECs. These signature phenotypes will be helpful to establish dia-hiPSC-ECs as models of endothelial dysfunction for understanding molecular mechanisms of disease and for identifying and testing new targets for the treatment of endothelial dysfunction in diabetes.
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Affiliation(s)
- Liping Su
- National Heart Centre Singapore, National Heart Research Institute Singapore, Singapore, Singapore
| | - Xiaocen Kong
- Department of Endocrinology, Nanjing First Hospital, Nanjing Medical University, Nanjing, China
| | - Sze Jie Loo
- National Heart Centre Singapore, National Heart Research Institute Singapore, Singapore, Singapore
| | - Yu Gao
- Department of Cardiology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
| | - Jean-Paul Kovalik
- Programme in Cardiovascular & Metabolic Disorders, Duke-National University of Singapore, Singapore, Singapore
| | - Xiaofei Su
- Department of Endocrinology, Nanjing First Hospital, Nanjing Medical University, Nanjing, China
| | - Jianhua Ma
- Department of Endocrinology, Nanjing First Hospital, Nanjing Medical University, Nanjing, China
| | - Lei Ye
- National Heart Centre Singapore, National Heart Research Institute Singapore, Singapore, Singapore
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42
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Gao Y, Pu J. Differentiation and Application of Human Pluripotent Stem Cells Derived Cardiovascular Cells for Treatment of Heart Diseases: Promises and Challenges. Front Cell Dev Biol 2021; 9:658088. [PMID: 34055788 PMCID: PMC8149736 DOI: 10.3389/fcell.2021.658088] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2021] [Accepted: 03/25/2021] [Indexed: 12/15/2022] Open
Abstract
Human pluripotent stem cells (hPSCs) are derived from human embryos (human embryonic stem cells) or reprogrammed from human somatic cells (human induced pluripotent stem cells). They can differentiate into cardiovascular cells, which have great potential as exogenous cell resources for restoring cardiac structure and function in patients with heart disease or heart failure. A variety of protocols have been developed to generate and expand cardiovascular cells derived from hPSCs in vitro. Precisely and spatiotemporally activating or inhibiting various pathways in hPSCs is required to obtain cardiovascular lineages with high differentiation efficiency. In this concise review, we summarize the protocols of differentiating hPSCs into cardiovascular cells, highlight their therapeutic application for treatment of cardiac diseases in large animal models, and discuss the challenges and limitations in the use of cardiac cells generated from hPSCs for a better clinical application of hPSC-based cardiac cell therapy.
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Affiliation(s)
- Yu Gao
- Department of Cardiology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
| | - Jun Pu
- Department of Cardiology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
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43
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Guan G, Huo D, Li Y, Zhao X, Li Y, Qin Z, Sun D, Yang G, Yang M, Tan J, Zeng W, Zhu C. Engineering hiPSC-CM and hiPSC-EC laden 3D nanofibrous splenic hydrogel for improving cardiac function through revascularization and remuscularization in infarcted heart. Bioact Mater 2021; 6:4415-4429. [PMID: 33997517 PMCID: PMC8113784 DOI: 10.1016/j.bioactmat.2021.04.010] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2021] [Revised: 04/06/2021] [Accepted: 04/07/2021] [Indexed: 12/13/2022] Open
Abstract
Cell therapy has been a promising strategy for cardiac repair after myocardial infarction (MI), but a poor ischemic environment and low cell delivery efficiency remain significant challenges. The spleen serves as a hematopoietic stem cell niche and secretes cardioprotective factors after MI, but it is unclear whether it could be used for human pluripotent stem cell (hiPSC) cultivation and provide a proper microenvironment for cell grafts against the ischemic environment. Herein, we developed a splenic extracellular matrix derived thermoresponsive hydrogel (SpGel). Proteomics analysis indicated that SpGel is enriched with proteins known to modulate the Wnt signaling pathway, cell-substrate adhesion, cardiac muscle contraction and oxidation-reduction processes. In vitro studies demonstrated that hiPSCs could be efficiently induced into endothelial cells (iECs) and cardiomyocytes (iCMs) with enhanced function on SpGel. The cytoprotective effect of SpGel on iECs/iCMs against oxidative stress damage was also proven. Furthermore, in vivo studies revealed that iEC/iCM-laden SpGel improved cardiac function and inhibited cardiac fibrosis of infarcted hearts by improving cell survival, revascularization and remuscularization. In conclusion, we successfully established a novel platform for the efficient generation and delivery of autologous cell grafts, which could be a promising clinical therapeutic strategy for cardiac repair and regeneration after MI.
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Affiliation(s)
- Ge Guan
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Da Huo
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Yanzhao Li
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Xiaolin Zhao
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Yinghao Li
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Zhongliang Qin
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China.,Chongqing Institute of Zhong Zhi Yi Gu, Shapingba District, Chongqing, 400030, China
| | - Dayu Sun
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Guanyuan Yang
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Mingcan Yang
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Ju Tan
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
| | - Wen Zeng
- Department of Cell Biology, Third Military Army Medical University, Chongqing, 400038, China
| | - Chuhong Zhu
- Department of Anatomy, State Key Laboratory of Trauma, Burn and Combined Injury, Key Lab for Biomechanics and Tissue Engineering of Chongqing, Army Medical University (Third Military Medical University), Chongqing, 400038, China
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44
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Browne S, Gill EL, Schultheiss P, Goswami I, Healy KE. Stem cell-based vascularization of microphysiological systems. Stem Cell Reports 2021; 16:2058-2075. [PMID: 33836144 PMCID: PMC8452487 DOI: 10.1016/j.stemcr.2021.03.015] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2020] [Revised: 03/11/2021] [Accepted: 03/15/2021] [Indexed: 12/27/2022] Open
Abstract
Microphysiological systems (MPSs) (i.e., tissue or organ chips) exploit microfluidics and 3D cell culture to mimic tissue and organ-level physiology. The advent of human induced pluripotent stem cell (hiPSC) technology has accelerated the use of MPSs to study human disease in a range of organ systems. However, in the reduction of system complexity, the intricacies of vasculature are an often-overlooked aspect of MPS design. The growing library of pluripotent stem cell-derived endothelial cell and perivascular cell protocols have great potential to improve the physiological relevance of vasculature within MPS, specifically for in vitro disease modeling. Three strategic categories of vascular MPS are outlined: self-assembled, interface focused, and 3D biofabricated. This review discusses key features and development of the native vasculature, linking that to how hiPSC-derived vascular cells have been generated, the state of the art in vascular MPSs, and opportunities arising from interdisciplinary thinking.
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Affiliation(s)
- Shane Browne
- Department of Bioengineering and California Institute for Quantitative Biosciences (QB3), University of California at Berkeley, Berkeley, CA 94720, USA
| | - Elisabeth L Gill
- Department of Bioengineering and California Institute for Quantitative Biosciences (QB3), University of California at Berkeley, Berkeley, CA 94720, USA
| | - Paula Schultheiss
- Department of Bioengineering and California Institute for Quantitative Biosciences (QB3), University of California at Berkeley, Berkeley, CA 94720, USA
| | - Ishan Goswami
- Department of Bioengineering and California Institute for Quantitative Biosciences (QB3), University of California at Berkeley, Berkeley, CA 94720, USA; Department of Materials Science and Engineering, University of California, Berkeley, CA 94720, USA
| | - Kevin E Healy
- Department of Bioengineering and California Institute for Quantitative Biosciences (QB3), University of California at Berkeley, Berkeley, CA 94720, USA; Department of Materials Science and Engineering, University of California, Berkeley, CA 94720, USA.
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45
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Guo Z, Mo Z. Regulation of endothelial cell differentiation in embryonic vascular development and its therapeutic potential in cardiovascular diseases. Life Sci 2021; 276:119406. [PMID: 33785330 DOI: 10.1016/j.lfs.2021.119406] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2020] [Revised: 03/05/2021] [Accepted: 03/14/2021] [Indexed: 12/17/2022]
Abstract
During vertebrate development, the cardiovascular system begins operating earlier than any other organ in the embryo. Endothelial cell (EC) forms the inner lining of blood vessels, and its extensive proliferation and migration are requisite for vasculogenesis and angiogenesis. Many aspects of cellular biology are involved in vasculogenesis and angiogenesis, including the tip versus stalk cell specification. Recently, epigenetics has attracted growing attention in regulating embryonic vascular development and controlling EC differentiation. Some proteins that regulate chromatin structure have been shown to be directly implicated in human cardiovascular diseases. Additionally, the roles of important EC signaling such as vascular endothelial growth factor and its receptors, angiopoietin-1 and tyrosine kinase containing immunoglobulin and epidermal growth factor homology domain-2, and transforming growth factor-β in EC differentiation during embryonic vasculature development are briefly discussed in this review. Recently, the transplantation of human induced pluripotent stem cell (iPSC)-ECs are promising approaches for the treatment of ischemic cardiovascular disease including myocardial infarction. Patient-specific iPSC-derived EC is a potential new target to study differences in gene expression or response to drugs. However, clinical application of the iPSC-ECs in regenerative medicine is often limited by the challenges of maintaining cell viability and function. Therefore, novel insights into the molecular mechanisms underlying EC differentiation might provide a better understanding of embryonic vascular development and bring out more effective EC-based therapeutic strategies for cardiovascular diseases.
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Affiliation(s)
- Zi Guo
- Department of Endocrinology, The Third Xiangya Hospital, Central South University, Changsha, Hunan, China
| | - Zhaohui Mo
- Department of Endocrinology, The Third Xiangya Hospital, Central South University, Changsha, Hunan, China.
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46
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Wang Z, Zuo F, Liu Q, Wu X, Du Q, Lei Y, Wu Z, Lin H. Comparative Study of Human Pluripotent Stem Cell-Derived Endothelial Cells in Hydrogel-Based Culture Systems. ACS OMEGA 2021; 6:6942-6952. [PMID: 33748608 PMCID: PMC7970572 DOI: 10.1021/acsomega.0c06187] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/19/2020] [Accepted: 02/19/2021] [Indexed: 06/12/2023]
Abstract
Human pluripotent stem cell (hPSC)-derived endothelial cells (ECs) are promising cell sources for drug discovery, tissue engineering, and studying or treating vascular diseases. However, hPSC-ECs derived from different culture methods display different phenotypes. Herein, we made a detailed comparative study of hPSC-ECs from three different culture systems (e.g., 2D, 3D PNIPAAm-PEG hydrogel, and 3D alginate hydrogel cultures) based on our previous reports. We expanded hPSCs and differentiated them into ECs in three culture systems. Both 3D hydrogel systems could mimic an in vivo physiologically relevant microenvironment to protect cells from shear force and prevent cell agglomeration, leading to a high culture efficiency and a high volumetric yield. We demonstrated that hPSC-ECs produced from both hydrogel systems had similar results as 2D-ECs. The transcriptome analysis showed that PEG-ECs and alginate-ECs displayed a functional phenotype due to their higher gene expressions in vasculature development, extracellular matrix, angiogenesis, and glycolysis, while 2D-ECs showed a proliferative phenotype due to their higher gene expressions in cell proliferation. Taken together, both PEG- and alginate-hydrogel systems will significantly advance the applications of hPSC-ECs in various biomedical fields.
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Affiliation(s)
- Zhanqi Wang
- Department
of Vascular Surgery, Beijing Anzhen Hospital of Capital Medical University, Beijing Institute of Heart Lung and Blood Vessel Diseases, Beijing 100029, China
| | - Fuxing Zuo
- Department
of Neurosurgery, National Cancer Center/National Clinical Research
Center for Cancer/Cancer Hospital, Chinese
Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
| | - Qing Liu
- Department
of Obstetrics, Beijing Obstetrics and Gynecology
Hospital Capital Medical University, Beijing 100006, China
| | - Xuesheng Wu
- Department
of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China
| | - Qian Du
- Department
of Biological Systems Engineering, University
of Nebraska-Lincoln, Lincoln, Nebraska 68588, United States
| | - Yuguo Lei
- Department
of Chemical and Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, Nebraska 68588, United States
| | - Zhangmin Wu
- Department
of Vascular Surgery, Beijing Anzhen Hospital of Capital Medical University, Beijing Institute of Heart Lung and Blood Vessel Diseases, Beijing 100029, China
| | - Haishuang Lin
- Department
of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China
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47
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Wang L, Serpooshan V, Zhang J. Engineering Human Cardiac Muscle Patch Constructs for Prevention of Post-infarction LV Remodeling. Front Cardiovasc Med 2021; 8:621781. [PMID: 33718449 PMCID: PMC7952323 DOI: 10.3389/fcvm.2021.621781] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2020] [Accepted: 02/04/2021] [Indexed: 12/20/2022] Open
Abstract
Tissue engineering combines principles of engineering and biology to generate living tissue equivalents for drug testing, disease modeling, and regenerative medicine. As techniques for reprogramming human somatic cells into induced pluripotent stem cells (iPSCs) and subsequently differentiating them into cardiomyocytes and other cardiac cells have become increasingly efficient, progress toward the development of engineered human cardiac muscle patch (hCMP) and heart tissue analogs has accelerated. A few pilot clinical studies in patients with post-infarction LV remodeling have been already approved. Conventional methods for hCMP fabrication include suspending cells within scaffolds, consisting of biocompatible materials, or growing two-dimensional sheets that can be stacked to form multilayered constructs. More recently, advanced technologies, such as micropatterning and three-dimensional bioprinting, have enabled fabrication of hCMP architectures at unprecedented spatiotemporal resolution. However, the studies working on various hCMP-based strategies for in vivo tissue repair face several major obstacles, including the inadequate scalability for clinical applications, poor integration and engraftment rate, and the lack of functional vasculature. Here, we review many of the recent advancements and key concerns in cardiac tissue engineering, focusing primarily on the production of hCMPs at clinical/industrial scales that are suitable for administration to patients with myocardial disease. The wide variety of cardiac cell types and sources that are applicable to hCMP biomanufacturing are elaborated. Finally, some of the key challenges remaining in the field and potential future directions to address these obstacles are discussed.
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Affiliation(s)
- Lu Wang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, Birmingham, AL, United States
| | - Vahid Serpooshan
- Department of Biomedical Engineering, Emory University School of Medicine and Georgia Institute of Technology, Atlanta, GA, United States
- Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, United States
- Children's Healthcare of Atlanta, Atlanta, GA, United States
| | - Jianyi Zhang
- Department of Biomedical Engineering, School of Medicine and School of Engineering, University of Alabama at Birmingham, Birmingham, AL, United States
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48
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Huang Y, Wu H, Han X, Wu J, Yu M, Zhao ZA, Shen Z, Hu S, Lei W. Generation of an EFNB2-2A-mCherry reporter human embryonic stem cell line using CRISPR/Cas9-mediated site-specific homologous recombination. Stem Cell Res 2021; 52:102241. [PMID: 33611045 DOI: 10.1016/j.scr.2021.102241] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/28/2020] [Revised: 01/20/2021] [Accepted: 02/05/2021] [Indexed: 10/22/2022] Open
Abstract
Ephrin B2 (EFNB2) is the first identified and most widely used marker for arterial endothelial cells (AECs). We generated a heterozygous EFNB2-2A-mCherry reporter H1 cell line, H1-EFNB2-2A-mCherry+/- (WAe001-A-57), by CRISPR/Cas9-mediated insertion of 2A-mCherry cassette into the EFNB2 gene locus, immediately before the translation stop codon. The H1-EFNB2-2A-mCherry reporter cells were pluripotent and could differentiate into all three germ layer lineages. Simultaneous expression of mCherry was observed when expression of EFNB2 was increased during endothelial cell differentiation. Thus, the generated reporter cells enable live identification of EFNB2-positive AECs, and screening of small molecule compound and target genes that promote AEC differentiation.
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Affiliation(s)
- Ying Huang
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Hongchun Wu
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Xinglong Han
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Jie Wu
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Miao Yu
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Zhen-Ao Zhao
- Institute of Microcirculation & Department of Pathophysiology of Basic Medical College, Hebei North University, Zhangjiakou 075000, China; Hebei Key Laboratory of Critical Disease Mechanism and Intervention, Zhangjiakou 075000, China
| | - Zhenya Shen
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China
| | - Shijun Hu
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China.
| | - Wei Lei
- Department of Cardiovascular Surgery of the First Affiliated Hospital & Institute for Cardiovascular Science, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Protection, Medical College, Soochow University, Suzhou 215000, China.
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Veschini L, Sailem H, Malani D, Pietiäinen V, Stojiljkovic A, Wiseman E, Danovi D. High-Content Imaging to Phenotype Human Primary and iPSC-Derived Cells. Methods Mol Biol 2021; 2185:423-445. [PMID: 33165865 DOI: 10.1007/978-1-0716-0810-4_27] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Increasingly powerful microscopy, liquid handling, and computational techniques have enabled cell imaging in high throughput. Microscopy images are quantified using high-content analysis platforms linking object features to cell behavior. This can be attempted on physiologically relevant cell models, including stem cells and primary cells, in complex environments, and conceivably in the presence of perturbations. Recently, substantial focus has been devoted to cell profiling for cell therapy, assays for drug discovery or biomarker identification for clinical decision-making protocols, bringing this wealth of information into translational applications. In this chapter, we focus on two protocols enabling to (1) benchmark human cells, in particular human endothelial cells as a case study and (2) extract cells from blood for follow-up experiments including image-based drug testing. We also present concepts of high-content imaging and discuss the benefits and challenges, with the aim of enabling readers to tailor existing pipelines and bring such approaches closer to translational research and the clinic.
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Affiliation(s)
- Lorenzo Veschini
- Academic Centre of Reconstructive Science, Faculty of Dentistry, Oral & Craniofacial Sciences, King's College London, London, UK
| | - Heba Sailem
- The Institute of Biomedical Engineering, Oxford, UK
| | - Disha Malani
- Institute for Molecular Medicine Finland-FIMM, Helsinki Institute of Life Science-HiLIFE, University of Helsinki, Helsinki, Finland
| | - Vilja Pietiäinen
- Institute for Molecular Medicine Finland-FIMM, Helsinki Institute of Life Science-HiLIFE, University of Helsinki, Helsinki, Finland
| | - Ana Stojiljkovic
- Division of Veterinary Anatomy, Vetsuisse Faculty, University of Bern, Bern, Switzerland
| | - Erika Wiseman
- Stem Cell Hotel, Centre for Stem Cells and Regenerative Medicine, King's College London, London, UK
| | - Davide Danovi
- Stem Cell Hotel, Centre for Stem Cells and Regenerative Medicine, King's College London, London, UK.
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50
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Ramakrishna RR, Abd Hamid Z, Wan Zaki WMD, Huddin AB, Mathialagan R. Stem cell imaging through convolutional neural networks: current issues and future directions in artificial intelligence technology. PeerJ 2020; 8:e10346. [PMID: 33240655 PMCID: PMC7680049 DOI: 10.7717/peerj.10346] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2020] [Accepted: 10/21/2020] [Indexed: 12/12/2022] Open
Abstract
Stem cells are primitive and precursor cells with the potential to reproduce into diverse mature and functional cell types in the body throughout the developmental stages of life. Their remarkable potential has led to numerous medical discoveries and breakthroughs in science. As a result, stem cell-based therapy has emerged as a new subspecialty in medicine. One promising stem cell being investigated is the induced pluripotent stem cell (iPSC), which is obtained by genetically reprogramming mature cells to convert them into embryonic-like stem cells. These iPSCs are used to study the onset of disease, drug development, and medical therapies. However, functional studies on iPSCs involve the analysis of iPSC-derived colonies through manual identification, which is time-consuming, error-prone, and training-dependent. Thus, an automated instrument for the analysis of iPSC colonies is needed. Recently, artificial intelligence (AI) has emerged as a novel technology to tackle this challenge. In particular, deep learning, a subfield of AI, offers an automated platform for analyzing iPSC colonies and other colony-forming stem cells. Deep learning rectifies data features using a convolutional neural network (CNN), a type of multi-layered neural network that can play an innovative role in image recognition. CNNs are able to distinguish cells with high accuracy based on morphologic and textural changes. Therefore, CNNs have the potential to create a future field of deep learning tasks aimed at solving various challenges in stem cell studies. This review discusses the progress and future of CNNs in stem cell imaging for therapy and research.
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Affiliation(s)
- Ramanaesh Rao Ramakrishna
- Biomedical Science Programme and Centre for Diagnostic, Therapeutic and Investigative Science, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia
| | - Zariyantey Abd Hamid
- Biomedical Science Programme and Centre for Diagnostic, Therapeutic and Investigative Science, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia
| | - Wan Mimi Diyana Wan Zaki
- Department of Electrical, Electronic & Systems Engineering, Faculty of Engineering & Built Environment, Universiti Kebangsaan Malaysia, Bangi, Selangor, Malaysia
| | - Aqilah Baseri Huddin
- Department of Electrical, Electronic & Systems Engineering, Faculty of Engineering & Built Environment, Universiti Kebangsaan Malaysia, Bangi, Selangor, Malaysia
| | - Ramya Mathialagan
- Biomedical Science Programme and Centre for Diagnostic, Therapeutic and Investigative Science, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia
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