• 3D bioprinting
• Bioink
• Gel
• Genetics
• Tissue engineering

• DBT-RA/2024/January/N/5132 Department of Biotechnology, Ministry of Science and Technology, India
• TN02000025 NCEII, Czech republic

• 3D bioprinting
• bioreactors
• molecules
• tissue engineering
• vascularization control

• Tissue Engineering/methods
• Humans
• Neovascularization, Physiologic/drug effects
• Animals
• Biocompatible Materials/chemistry
• Bioprinting/methods
• Tissue Scaffolds/chemistry
• Printing, Three-Dimensional
• Bioreactors

• doi.org/10.54499/2022.02781.CEECIND/CP1745/CT0001 FCT-Fundação para a Ciência e a Tecnologia

• bone
• microvascular fragments
• osteoblasts
• spheroid
• tissue engineering
• vascularization

• Deutsche Forschungsgemeinschaft

• pluripotent stem cells
• tissue engineering
• immune evasion
• applied immunology
• regenerative medicine

• Animals
• Mice
• Humans
• Mouse Embryonic Stem Cells/immunology
• Mouse Embryonic Stem Cells/metabolism
• Transplantation, Homologous
• Transgenes
• Mice, Inbred C57BL
• Leukocytes, Mononuclear/immunology
• Cell Survival

• Lunenfeld-Tanenbaum Research Institute
• Medicine by Design, C1TPA-2016-20
• CIHR Foundation Grant, FDN-143231
• Fighting Blindness Canada
• Not Related
• Canada Research Chair, 950-230422
• Ontario Research Fund; RE08-064
• Canadian Research Chair; 950-230422
• Medicine by Design C1TPA-2016-20
• Canada Research Chair; 950-230422

• port wine birthmark
• induced pluripotent stem cells
• mesenchymal stem cells
• endothelial cells
• hippo pathway
• wnt pathway
• zinc-finger proteins

• R01 AR073172 NIAMS NIH HHS
• R21 AR083066 NIAMS NIH HHS

• endothelial cell
• extracellular matrix
• induced pluripotent stem cell
• mechanical cues
• stem cell

• cardiology
• regenerative medicine
• tissue engineering

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.

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.

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.

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.

The online version contains supplementary material available at 10.1186/s13287-022-02925-w.

• Endothelial differentiation
• Endothelium
• Hematopoietic cells
• Hematopoietic differentiation
• Hemogenic endothelium
• hESCs
• hPSC
• hiPSC

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.

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.

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).

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.

The online version contains supplementary material available at 10.1186/s13287-022-02924-x.

• 3D scalable bioreactor suspension culture
• Angiogenesis
• Arterial endothelium
• Differentiation
• Endothelial cells
• Human induced pluripotent stem cells
• LDL uptake
• Regenerative medicine, 2D monolayer culture
• Vascular endothelium
• Wnt signaling
• eNOS
• hiPSCs
• iPS cells

• 3d reconstruction of protein
• Biomaterials
• Materials science

• cell signalling
• medical research

• Antigens, CD34
• Cell Differentiation
• Colony-Forming Units Assay
• Endothelial Cells
• Humans
• Niacinamide/pharmacology
• Stem Cells

• R01 DK074720 NIDDK NIH HHS
• T32 HL134644 NHLBI NIH HHS
• U.S. Department of Health & Human Services | NIH | National Institute of Diabetes and Digestive and Kidney Diseases (National Institute of Diabetes & Digestive & Kidney Diseases)

• angiogenesis
• cell therapy
• endothelial dysfunction
• stem cells
• vascularization

• Animals
• Endothelium, Vascular/metabolism
• Microvessels
• Oxidative Stress
• Regeneration
• Vascular Diseases/metabolism

• P30 ES030283 NIEHS NIH HHS
• R01 AG049821 NIA NIH HHS
• R01 AG053585 NIA NIH HHS
• Gheen's Foundation
• Canadian Network for Research and Innovation in Machining Technology, Natural Sciences and Engineering Research Council of Canada (CRSNG)
• Gouvernement du Canada | Canadian Institutes of Health Research (CIHR)
• HHS | NIH | National Institute on Aging (NIA)
• Ministère du Développement économique, de la Création d'emplois et du Commerce | Ontario Ministry of Research and Innovation (MRI)
• U.S. Department of Defense (DOD)

The prognosis of rheumatic diseases is generally better than that of malignant diseases. However, some cases with poor prognoses resist conventional therapies and cause irreversible functional and organ damage. In recent years, there has been much research on regenerative medicine, which uses stem cells to restore the function of missing or dysfunctional tissues and organs. The development of regenerative medicine is also being attempted in rheumatic diseases. In diseases such as systemic sclerosis (SSc), systemic lupus erythematosus (SLE), and rheumatoid arthritis, hematopoietic stem cell transplantation has been attempted to correct and reconstruct abnormalities in the immune system. Mesenchymal stem cells (MSCs) have also been tried for the treatment of refractory skin ulcers in SSc using the ability of MSCs to differentiate into vascular endothelial cells and for the treatment of systemic lupus erythematosus SLE using the immunosuppressive effect of MSCs. CD34-positive endothelial progenitor cells (EPCs), which are found in the mononuclear cell fraction of bone marrow and peripheral blood, can differentiate into vascular endothelial cells at the site of ischemia. Therefore, EPCs have been used in research on vascular regeneration therapy for patients with severe lower limb ischemia caused by rheumatic diseases such as SSc. Since the first report of induced pluripotent stem cells (iPSCs) in 2007, research on regenerative medicine using iPSCs has been actively conducted, and their application to rheumatic diseases is expected. However, there are many safety issues and bioethical issues involved in regenerative medicine research, and it is essential to resolve these issues for practical application and spread of regenerative medicine in the future. The environment surrounding regenerative medicine research is changing drastically, and the required expertise is becoming higher. This paper outlines the current status and challenges of regenerative medicine in rheumatic diseases.

• Japan Society for the Promotion of Science
• Japan Agency for Medical Research and Development
• SENSHIN Medical Research Foundation

• Differentiation
• Endothelial cells
• Induced pluripotent stem cells
• WNT pathway
• iPSCs

• disease models, drug screening
• induced pluripotent stem cells
• lung cell differentiation
• lung-on-chip
• organoids models, toxicity screening

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.

• cardiovascular disease
• endothelial cell
• human induced pluripotent stem cell
• regenerative medicine
• smooth muscle cell
• stem cell

Severe acute respiratory syndrome (SARS)-CoV-2 infection leads to severe disease associated with cytokine storm, vascular dysfunction, coagulation, and progressive lung damage. It affects several vital organs, seemingly through a pathological effect on endothelial cells. The SARS-CoV-2 genome encodes 29 proteins, whose contribution to the disease manifestations, and especially endothelial complications, is unknown. We cloned and expressed 26 of these proteins in human cells and characterized the endothelial response to overexpression of each, individually. Whereas most proteins induced significant changes in endothelial permeability, nsp2, nsp5_c145a (catalytic dead mutant of nsp5), and nsp7 also reduced CD31, and increased von Willebrand factor expression and IL-6, suggesting endothelial dysfunction. Using propagation-based analysis of a protein–protein interaction (PPI) network, we predicted the endothelial proteins affected by the viral proteins that potentially mediate these effects. We further applied our PPI model to identify the role of each SARS-CoV-2 protein in other tissues affected by coronavirus disease (COVID-19). While validating the PPI network model, we found that the tight junction (TJ) proteins cadherin-5, ZO-1, and β-catenin are affected by nsp2, nsp5_c145a, and nsp7 consistent with the model prediction. Overall, this work identifies the SARS-CoV-2 proteins that might be most detrimental in terms of endothelial dysfunction, thereby shedding light on vascular aspects of COVID-19.

• SARS-COV-2
• endothelium
• human
• infectious disease
• microbiology
• protein interactions
• vasculature

Intravenous thrombolysis using recombinant tissue plasminogen activator (rt-PA) is the standard treatment for acute ischemic stroke. Standard-dose rt-PA (0.9 mg/kg) is known to achieve good recanalization but carries a high bleeding risk. Lower dose of rt-PA has less bleeding risk but carries a high re-occlusion rate. We investigate if induced pluripotent stem cells (iPSCs) can improve the thrombolytic effect of low-dose rt-PA (0.45 mg/kg).

Single irradiation with 6 mW/cm² light-emitting diode (LED) for 4 h at rat common carotid artery was used as thrombosis model according to our previous report. Endothelin-1 (ET-1), intercellular adhesion molecule-1 (ICAM-1), and interleukin 1 beta (IL-1 beta) were used as the inflammatory markers for artery endothelial injury. Angiopoietin-2 (AP-2), brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) were examined in artery wall and iPSCs culture. Animal ultrasound was used to evaluate the stenosis degree of common carotid artery before and at 2 h, 24 h, 4 days and 7 days after LED irradiation.

After LED irradiation alone, there was a persistent occlusion from 2 h to 7 days. Standard-dose rt-PA alone could recanalize the occluded artery from 24 h to 7 days to stenotic degree ≤ 50%. Low-dose rt-PA or 1 × 10⁶ mouse iPSCs alone could not recanalize the occluded arteries from 2 h to 7 days. Combination use of low-dose rt-PA plus 1 × 10⁶ mouse iPSCs caused better recanalization from 24 h to 7 days. ET-1, ICAM-1 and IL-1 beta were strongly expressed after LED irradiation but reduced after iPSCs treatment. AP-2, BDNF and VEGF were rarely induced after LED irradiation but strongly expressed after iPSCs treatment. In vitro study showed iPSCs could express AP-2, BDNF and VEGF.

The adjuvant use of iPSCs may help improving the thrombolytic effect of low-dose rt-PA by suppressing inflammatory factors and inducing angiogenic trophic factors. Stem cells could be a potential regimen in acute thrombolytic therapy to improve recanalization and reduce complications.

The online version contains supplementary material available at 10.1186/s13287-021-02615-z.

• Carotid thrombosis
• Endothelium
• Stem cell
• Thrombolysis
• Tissue plasminogen activator

Cell-cell interactions are crucial for organ development and function. In the heart, endothelial cells engage in bidirectional communication with cardiomyocytes regulating cardiac development and growth. We aimed to elucidate the organotypic development of cardiac endothelial cells and cardiomyocyte and endothelial cell crosstalk using human induced pluripotent stem cells (hiPSC). Single-cell RNA sequencing was performed with hiPSC-derived cardiomyocytes (hiPS-CMs) and endothelial cells (hiPS-ECs) in mono- and co-culture. The presence of hiPS-CMs led to increased expression of transcripts related to vascular development and maturation, cardiac development, as well as cardiac endothelial cell and endocardium-specific genes in hiPS-ECs. Interestingly, co-culture induced the expression of cardiomyocyte myofibrillar genes and MYL7 and MYL4 protein expression was detected in hiPS-ECs. Major regulators of BMP- and Notch-signaling pathways were induced in both cell types in co-culture. These results reflect the findings from animal studies and extend them to human endothelial cells, demonstrating the importance of EC-CM interactions during development.

• cardiomyocytes
• co-culture
• endothelial cells
• human induced pluripotent stem cells
• single cell RNA sequencing

• Cell differentiation
• Endothelial cells
• Mechanotransduction
• Shear stress
• Stem cells

• co-culture
• contact differentiation
• differentiation
• embryonic stem cells
• endothelial differentiation
• hepatic differentiation

• R01 GM061731 NIGMS NIH HHS

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.

• cardiovascular cells
• differentiation
• human pluripotent stem cells (hPSCs)
• large animal
• therapeutic application

• pluripotent human stem cells
• human endothelial cells
• hemogenic endothelial cells
• arterial endothelial cells
• cell culture protocol
• directed differentiation

• Cell Differentiation
• Endothelial Cells/metabolism
• Humans
• Pluripotent Stem Cells/metabolism

• R01 HL128064 NHLBI NIH HHS
• R01 HL146056 NHLBI NIH HHS
• UH2 EB017103 NIBIB NIH HHS

Stem cell technology has been around for almost 30 years and in that time has grown into an enormous field. The stem cell technique progressed from the first successful isolation of mammalian embryonic stem cells (ESCs) in the 1990s, to the production of human induced-pluripotent stem cells (iPSCs) in the early 2000s, to finally culminate in the differentiation of pluripotent cells into highly specialized cell types, such as neurons, endothelial cells (ECs), cardiomyocytes, fibroblasts, and lung and intestinal cells, in the last decades. In recent times, we have attained a new height in stem cell research whereby we can produce 3D organoids derived from stem cells that more accurately mimic the in vivo environment. This review summarizes the development of stem cell research in the context of vascular research ranging from differentiation techniques of ECs and smooth muscle cells (SMCs) to the generation of vascularized 3D organoids. Furthermore, the different techniques are critically reviewed, and future applications of current 3D models are reported.

• Differentiation
• Endothelial cells
• Induced pluripotent stem cells
• Organoids
• Smooth muscle cells
• Vasculature

• diabetes
• insulin
• islet transplantation
• microvascular fragments
• vascularization

• Animals
• Diabetes Mellitus, Experimental
• Endothelial Cells
• Insulin
• Insulin-Secreting Cells
• Islets of Langerhans
• Islets of Langerhans Transplantation
• Mice

• Biocompatible Materials/chemistry
• Biomechanical Phenomena
• Cells, Cultured
• Collagen/chemistry
• Diabetes Complications/etiology
• Diabetes Complications/pathology
• Diabetes Complications/therapy
• Diabetes Mellitus, Type 2/complications
• Diabetes Mellitus, Type 2/pathology
• Diabetes Mellitus, Type 2/therapy
• Fibroblasts/cytology
• Gelatin/chemistry
• Human Umbilical Vein Endothelial Cells
• Humans
• Hydrogels/chemistry
• Imaging, Three-Dimensional
• In Vitro Techniques
• Keratinocytes/pathology
• Materials Testing
• Methacrylates/chemistry
• Models, Biological
• Skin/blood supply
• Skin/pathology
• Skin Diseases/etiology
• Skin Diseases/pathology
• Skin Diseases/therapy
• Tissue Scaffolds/chemistry
• Wound Healing/physiology

• Cell Culture Techniques/methods
• Cell Differentiation/genetics
• Cell Lineage/genetics
• Electrophysiological Phenomena
• Heart Ventricles/cytology
• Humans
• Induced Pluripotent Stem Cells/cytology
• Mesoderm/cytology
• Mesoderm/growth & development
• Myocytes, Cardiac/cytology
• Receptor, Platelet-Derived Growth Factor alpha/genetics
• Wnt Proteins/antagonists & inhibitors

• Japan Agency for Medical Research and Development

• RNA sequencing
• endothelial cells
• flow
• induced pluripotent stem cells
• shear stress
• single-cell RNA sequencing

• Dental stem cells
• endothelial cells
• growth factors
• reprogramming
• small molecules

Regenerative therapy holds great promise in the development of cures of some untreatable diseases such as cardiovascular diseases, and pluripotent stem cells (PSCs) including induced PSCs (iPSCs) are the most important regenerative seed cells. Recently, differentiation of human PSCs into functional tissues and cells in vitro has been widely reported. However, although porcine reports are rare they are quite essential, as the pig is an important animal model for the in vitro generation of human organs. In this study, we reprogramed porcine embryonic fibroblasts into porcine iPSCs (piPSCs), and differentiated them into cluster of differentiation 31 (CD31)-positive endothelial cells (ECs) (piPSC-derived ECs, piPS-ECs) using an optimized single-layer culture method. During differentiation, we observed that a combination of GSK3β inhibitor (CHIR99021) and bone morphogenetic protein 4 (BMP4) promoted mesodermal differentiation, resulting in higher proportions of CD31-positive cells than those from separate CHIR99021 or BMP4 treatment. Importantly, the piPS-ECs showed comparable morphological and functional properties to immortalized porcine aortic ECs, which are capable of taking up low-density lipoprotein and forming network structures on Matrigel. Our study, which is the first trial on a species other than human and mouse, has provided an optimized single-layer culture method for obtaining ECs from porcine PSCs. Our approach can be beneficial when evaluating autologous EC transplantation in pig models.

• endothelial cell
• iPS cell
• mesoderm
• swine

• ADSCs
• Adipose tissue
• Adult stem cells
• Angiogenesis
• CD34+
• EPCs
• Endothelial progenitor cells
• MSCs
• Microvessel fragments
• Vascularization
• Vasculogenesis
• iPSCs

• 3-D printing
• anatomy
• microenvironment
• microphysiological systems
• organ chips
• organoids
• physiology

• Animals
• Drug Discovery
• Humans
• Lab-On-A-Chip Devices

• R15 GM122953 NIGMS NIH HHS

• Neuropilins
• cell migration
• endothelium
• integrins
• protein trafficking

Endothelial cells (ECs) and endothelial progenitor cells (EPCs) play crucial roles in maintaining vascular health and homeostasis. Both cell types have been used in regenerative therapy as well as in various in vitro models; however, the properties of primary human ECs and EPCs are dissimilar owing to differences in genetic backgrounds and sampling techniques. Human induced pluripotent stem cells (hiPSCs) are an alternative cell source of ECs and EPCs. However, owing to the low purity of differentiated cells from hiPSCs, purification via an antigen–antibody reaction, which damages the cells, is indispensable. Besides, owing to limited expandability, it is difficult to produce these cells in large numbers. Here we report the development of relatively simple differentiation and purification methods for hiPSC-derived EPCs (iEPCs). Furthermore, we discovered that a combination of three small molecules, that is, Y-27632 (a selective inhibitor of Rho-associated, coiled-coil containing protein kinase [ROCK]), A 83–01 (a receptor-like kinase inhibitor of transforming growth factor beta [TGF-β]), and CHIR-99021 (a selective inhibitor of glycogen synthase kinase-3β [GSK3β] that also activates Wnt), dramatically stimulated protein synthesis-related pathways and enhanced the proliferative capacity of iEPCs. These findings will help to establish a supply system of EPCs at an industrial scale.

• Cell biology
• Cell culture
• Cell differentiation
• Differentiation
• Endothelial progenitor cell
• Expansion
• Human induced pluripotent stem cell
• Pharmaceutical science
• Purification
• Stem cell research

Vascular endothelial growth factors (VEGFs) are hypoxia-inducible secreted proteins to promote angiogenesis, in which VEGF-A is an important molecule that binds and activates VEGF receptor-1 (VEGFR-1) and VEGFR-2. In this study, two DNA aptamers, Apt01 and Apt02, were successfully isolated by alternating consecutive systematic evolution of ligands by exponential enrichment (SELEX) against VEGFR-1 and -2 using deep sequencing analysis in an early selection round. Their binding affinities for VEGFR-2 were lower than that of VEGFR-1, which is similar to that of VEGF-A. Structural analyses with the measurements of circular dichroism spectra and ultraviolet melting curve showed that Apt01 possessed the stem-loop structure in the molecule, whereas Apt02 formed G-quadruplex structures. In addition, Apt02 accelerated a tube formation of human umbilical vein endothelial cells faster than Apt01, which was affected by difference of binding affinity and nuclease resistance due to G-quadruplex structures. These results demonstrated that Apt02 might have a potential to function as an alternative to VEGF-A.

• G-quadruplex
• VEGF
• VEGF receptor
• angiogenesis
• aptamer
• vascular endothelial growth factor

• CD82
• Wnt inhibition
• cardiomyocytes
• exosome
• iPSCs
• progenitors

Endothelial cells (ECs) are required for a multitude of cardiovascular clinical applications, such as revascularization of ischemic tissues or endothelialization of tissue engineered grafts. Patient derived primary ECs are limited in number, have donor variabilities and their in vitro phenotypes and functions can deteriorate over time. This necessitates the exploration of alternative EC sources. Although there has been a recent surge in the use of pluripotent stem cell derived endothelial cells (PSC-ECs) for various cardiovascular clinical applications, current differentiation protocols yield a heterogeneous EC population, where their specification into arterial or venous subtypes is undefined. Since arterial and venous ECs are phenotypically and functionally different, inappropriate matching of exogenous ECs to host sites can potentially affect clinical efficacy, as exemplified by venous graft mismatch when placed into an arterial environment. Therefore, there is a need to design and employ environmental cues that can effectively modulate PSC-ECs into a more homogeneous arterial or venous phenotype for better adaptation to the host environment, which will in turn contribute to better application efficacy. In this review, we will first give an overview of the developmental and functional differences between arterial and venous ECs. This provides the foundation for our subsequent discussion on the different bioengineering strategies that have been investigated to varying extent in providing biochemical and biophysical environmental cues to mature PSC-ECs into arterial or venous subtypes. The ability to efficiently leverage on a combination of biochemical and biophysical environmental cues to modulate intrinsic arterio-venous specification programs in ECs will greatly facilitate future translational applications of PSC-ECs. Since the development and maintenance of arterial and venous ECs in vivo occur in disparate physio-chemical microenvironments, it is conceivable that the application of these environmental factors in customized combinations or magnitudes can be used to selectively mature PSC-ECs into an arterial or venous subtype.

• arterial specification
• endothelial cells
• environmental cues
• functional maturation
• human pluripotent stem cells
• shear stress
• substrate topography

• Cyclic adenosine monophosphate
• differentiation
• stem cell
• tissue engineering

The current work reports the functional characterization of human induced pluripotent stem cells (iPSCs)- arterial and venous-like endothelial cells (ECs), derived in chemically defined conditions, either in monoculture or seeded in a scaffold with mechanical properties similar to blood vessels. iPSC-derived arterial- and venous-like endothelial cells were obtained in two steps: differentiation of iPSCs into endothelial precursor cells (CD31^pos/KDR^pos/VE-Cad^med/EphB2^neg/COUP-TF^neg) followed by their differentiation into arterial and venous-like ECs using a high and low vascular endothelial growth factor (VEGF) concentration. Cells were characterized at gene, protein and functional levels. Functionally, both arterial and venous-like iPSC-derived ECs responded to vasoactive agonists such as thrombin and prostaglandin E2 (PGE₂), similar to somatic ECs; however, arterial-like iPSC-derived ECs produced higher nitric oxide (NO) and elongation to shear stress than venous-like iPSC-derived ECs. Both cells adhered, proliferated and prevented platelet activation when seeded in poly(caprolactone) scaffolds. Interestingly, both iPSC-derived ECs cultured in monoculture or in a scaffold showed a different inflammatory profile than somatic ECs. Although both somatic and iPSC-derived ECs responded to tumor necrosis factor-α (TNF-α) by an increase in the expression of intercellular adhesion molecule 1 (ICAM-1), only somatic ECs showed an upregulation in the expression of E-selectin or vascular cell adhesion molecule 1 (VCAM-1).

• Extracellular matrix
• Shear stress
• Stem cells
• Stiffness

• Animals
• Cardiovascular Diseases/therapy
• Cardiovascular System/cytology
• Cell Differentiation
• Humans
• Mechanotransduction, Cellular
• Stem Cells/cytology
• Stress, Mechanical

• R01 AG045428 NIA NIH HHS
• T32 HL105373 NHLBI NIH HHS

• decellularization
• endothelialization
• gelatin
• liver scaffold
• transplantation

• Animals
• Biocompatible Materials/chemistry
• Cell Proliferation
• Endothelial Cells/cytology
• Gelatin/chemistry
• Liver/blood supply
• Liver/chemistry
• Liver/ultrastructure
• Liver Transplantation
• Neovascularization, Physiologic
• Rats
• Rats, Sprague-Dawley
• Tissue Engineering/methods
• Tissue Scaffolds/chemistry