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Xu B, Liu D, Liu W, Long G, Liu W, Wu Y, He X, Shen Y, Jiang P, Yin M, Fan Y, Shen H, Shi L, Zhang Q, Xue W, Jin C, Chen Z, Chen B, Li J, Hu Y, Li X, Xiao Z, Zhao Y, Dai J. Engineered human spinal cord-like tissues with dorsal and ventral neuronal progenitors for spinal cord injury repair in rats and monkeys. Bioact Mater 2023; 27:125-137. [PMID: 37064803 PMCID: PMC10090126 DOI: 10.1016/j.bioactmat.2023.03.015] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2022] [Revised: 03/05/2023] [Accepted: 03/21/2023] [Indexed: 03/31/2023] Open
Abstract
Transplanting human neural progenitor cells is a promising method of replenishing the lost neurons after spinal cord injury (SCI), but differentiating neural progenitor cells into the diverse types of mature functional spinal cord neurons in vivo is challenging. In this study, engineered human embryonic spinal cord-like tissues with dorsal and ventral neuronal characters (DV-SC) were generated by inducing human neural progenitor cells (hscNPCs) to differentiate into various types of dorsal and ventral neuronal cells on collagen scaffold in vitro. Transplantation of DV-SC into complete SCI models in rats and monkeys showed better therapeutic effects than undifferentiated hscNPCs, including pronounced cell survival and maturation. DV-SC formed a targeted connection with the host's ascending and descending axons, partially restored interrupted neural circuits, and improved motor evoked potentials and the hindlimb function of animals with SCI. This suggests that the transplantation of pre-differentiated hscNPCs with spinal cord dorsal and ventral neuronal characteristics could be a promising strategy for SCI repair.
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2
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Nie L, Yao D, Chen S, Wang J, Pan C, Wu D, Liu N, Tang Z. Directional induction of neural stem cells, a new therapy for neurodegenerative diseases and ischemic stroke. Cell Death Discov 2023; 9:215. [PMID: 37393356 DOI: 10.1038/s41420-023-01532-9] [Citation(s) in RCA: 25] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2023] [Revised: 06/16/2023] [Accepted: 06/22/2023] [Indexed: 07/03/2023] Open
Abstract
Due to the limited capacity of the adult mammalian brain to self-repair and regenerate, neurological diseases, especially neurodegenerative disorders and stroke, characterized by irreversible cellular damage are often considered as refractory diseases. Neural stem cells (NSCs) play a unique role in the treatment of neurological diseases for their abilities to self-renew and form different neural lineage cells, such as neurons and glial cells. With the increasing understanding of neurodevelopment and advances in stem cell technology, NSCs can be obtained from different sources and directed to differentiate into a specific neural lineage cell phenotype purposefully, making it possible to replace specific cells lost in some neurological diseases, which provides new approaches to treat neurodegenerative diseases as well as stroke. In this review, we outline the advances in generating several neuronal lineage subtypes from different sources of NSCs. We further summarize the therapeutic effects and possible therapeutic mechanisms of these fated specific NSCs in neurological disease models, with special emphasis on Parkinson's disease and ischemic stroke. Finally, from the perspective of clinical translation, we compare the strengths and weaknesses of different sources of NSCs and different methods of directed differentiation, and propose future research directions for directed differentiation of NSCs in regenerative medicine.
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Affiliation(s)
- Luwei Nie
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Dabao Yao
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Shiling Chen
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Jingyi Wang
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Chao Pan
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Dongcheng Wu
- Department of Biochemistry and Molecular Biology, Wuhan University School of Basic Medical Sciences, Wuhan, 430030, China
- Wuhan Hamilton Biotechnology Co., Ltd., Wuhan, 430030, China
| | - Na Liu
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China.
| | - Zhouping Tang
- Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China.
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3
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Schaefers C, Rothmiller S, Thiermann H, Rein T, Schmidt A. The Efficiency of Direct Maturation: the Comparison of Two hiPSC Differentiation Approaches into Motor Neurons. Stem Cells Int 2022; 2022:1320950. [PMID: 36530489 PMCID: PMC9757946 DOI: 10.1155/2022/1320950] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2022] [Revised: 11/10/2022] [Accepted: 11/11/2022] [Indexed: 02/23/2025] Open
Abstract
Motor neurons (MNs) derived from human-induced pluripotent stem cells (hiPSC) hold great potential for the treatment of various motor neurodegenerative diseases as transplantations with a low-risk of rejection are made possible. There are many hiPSC differentiation protocols that pursue to imitate the multistep process of motor neurogenesis in vivo. However, these often apply viral vectors, feeder cells, or antibiotics to generate hiPSC and MNs, limiting their translational potential. In this study, a virus-, feeder-, and antibiotic-free method was used for reprogramming hiPSC, which were maintained in culture medium produced under clinical good manufacturing practice. Differentiation into MNs was performed with standardized, chemically defined, and antibiotic-free culture media. The identity of hiPSC, neuronal progenitors, and mature MNs was continuously verified by the detection of specific markers at the genetic and protein level via qRT-PCR, flow cytometry, Western Blot, and immunofluorescence. MNX1- and ChAT-positive motoneuronal progenitor cells were formed after neural induction via dual-SMAD inhibition and expansion. For maturation, an approach aiming to directly mature these progenitors was compared to an approach that included an additional differentiation step for further specification. Although both approaches generated mature MNs expressing characteristic postmitotic markers, the direct maturation approach appeared to be more efficient. These results provide new insights into the suitability of two standardized differentiation approaches for generating mature MNs, which might pave the way for future clinical applications.
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Affiliation(s)
- Catherine Schaefers
- Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstr. 11, 80937 Munich, Germany
| | - Simone Rothmiller
- Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstr. 11, 80937 Munich, Germany
| | - Horst Thiermann
- Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstr. 11, 80937 Munich, Germany
| | - Theo Rein
- Max Planck Institute of Psychiatry, Kraepelinstr. 2-10, 80804 Munich, Germany
| | - Annette Schmidt
- Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstr. 11, 80937 Munich, Germany
- Institute of Sport Science, University of the Bundeswehr Munich, Werner-Heisenberg-Weg 39, 85577 Neubiberg, Germany
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Miceli M, Maruotti GM, Sarno L, Carbone L, Guida M, Pelagalli A. Preliminary Characterization of the Epigenetic Modulation in the Human Mesenchymal Stem Cells during Chondrogenic Process. Int J Mol Sci 2022; 23:9870. [PMID: 36077266 PMCID: PMC9456537 DOI: 10.3390/ijms23179870] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2022] [Revised: 08/23/2022] [Accepted: 08/25/2022] [Indexed: 11/16/2022] Open
Abstract
Regenerative medicine represents a growing hot topic in biomedical sciences, aiming at setting out novel therapeutic strategies to repair or regenerate damaged tissues and organs. For this perspective, human mesenchymal stem cells (hMSCs) play a key role in tissue regeneration, having the potential to differentiate into many cell types, including chondrocytes. Accordingly, in the last few years, researchers have focused on several in vitro strategies to optimize hMSC differentiation protocols, including those relying on epigenetic manipulations that, in turn, lead to the modulation of gene expression patterns. Therefore, in the present study, we investigated the role of the class II histone deacetylase (HDAC) inhibitor, MC1568, in the hMSCs-derived chondrogenesis. The hMSCs we used for this work were the hMSCs obtained from the amniotic fluid, given their greater differentiation capacity. Our preliminary data documented that MC1568 drove both the improvement and acceleration of hMSCs chondrogenic differentiation in vitro, since the differentiation process in MC1568-treated cells took place in about seven days, much less than that normally observed, namely 21 days. Collectively, these preliminary data might shed light on the validity of such a new differentiative protocol, in order to better assess the potential role of the epigenetic modulation in the process of the hypertrophic cartilage formation, which represents the starting point for endochondral ossification.
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Affiliation(s)
- Marco Miceli
- Department of Neuroscience, Reproductive and Odontostomatological Sciences, University of Naples “Federico II”, 80131 Naples, Italy
- CEINGE Biotecnologie Avanzate, 80145 Naples, Italy
| | - Giuseppe Maria Maruotti
- Department of Neuroscience, Reproductive and Odontostomatological Sciences, University of Naples “Federico II”, 80131 Naples, Italy
| | - Laura Sarno
- Department of Neuroscience, Reproductive and Odontostomatological Sciences, University of Naples “Federico II”, 80131 Naples, Italy
| | - Luigi Carbone
- Department of Neuroscience, Reproductive and Odontostomatological Sciences, University of Naples “Federico II”, 80131 Naples, Italy
| | - Maurizio Guida
- Department of Neuroscience, Reproductive and Odontostomatological Sciences, University of Naples “Federico II”, 80131 Naples, Italy
| | - Alessandra Pelagalli
- Department of Advanced Biomedical Sciences, University of Naples “Federico II”, 80131 Naples, Italy
- Institute of Biostructures and Bioimaging (IBB), National Research Council (CNR), 80131 Naples, Italy
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5
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Peli1 signaling blockade attenuates congenital zika syndrome. PLoS Pathog 2020; 16:e1008538. [PMID: 32544190 PMCID: PMC7297310 DOI: 10.1371/journal.ppat.1008538] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2019] [Accepted: 04/13/2020] [Indexed: 12/16/2022] Open
Abstract
Zika virus (ZIKV) infects pregnant women and causes devastating congenital zika syndrome (CZS). How the virus is vertically transmitted to the fetus and induces neuronal loss remains unclear. We previously reported that Pellino (Peli)1, an E3 ubiquitin ligase, promotes p38MAPK activation in microglia and induction of lethal encephalitis by facilitating the replication of West Nile virus (WNV), a closely related flavivirus. Here, we found that Peli1 expression was induced on ZIKV-infected human monocytic cells, peripheral blood mononuclear cells, human first-trimester placental trophoblasts, and neural stem cell (hNSC)s. Peli1 mediates ZIKV cell attachment, entry and viral translation and its expression is confined to the endoplasmic reticulum. Moreover, Peli1 mediated inflammatory cytokine and chemokine responses and induced cell death in placental trophoblasts and hNSCs. ZIKV-infected pregnant mice lacking Peli1 signaling had reduced placental inflammation and tissue damage, which resulted in attenuated congenital abnormalities. Smaducin-6, a membrane-tethered Smad6-derived peptide, blocked Peli1-mediated NF-κB activation but did not have direct effects on ZIKV infection. Smaducin-6 reduced inflammatory responses and cell death in placental trophoblasts and hNSCs, and diminished placental inflammation and damage, leading to attenuated congenital malformations in mice. Collectively, our results reveal a novel role of Peli1 in flavivirus pathogenesis and suggest that Peli1 promotes ZIKV vertical transmission and neuronal loss by mediating inflammatory cytokine responses and induction of cell death. Our results also identify Smaducin-6 as a potential therapeutic candidate for treatment of CZS. We previously reported that Pellino (Peli)1, an E3 ubiquitin ligase mediates p38MAPK activation in microglia and induces lethal encephalitis by facilitating replication of a mosquito -borne flavivirus, West Nile virus (WNV). Zika virus (ZIKV), a closely related flavivirus, causes devastating congenital zika syndrome (CZS) in pregnant women. How ZIKV is vertically transmitted to the fetus and induces neuronal loss remains unclear. Here, we found that Peli1 expression was enhanced in human monocytic cells, peripheral blood mononuclear cells, first-trimester placental trophoblasts and neural stem cell (hNSC)s following ZIKV infection. Peli1 expression colocalized with the endoplasmic reticulum and double-stranded RNA in ZIKV-infected cells and was required for ZIKV cell attachment and replication. Peli1 knockdown in placental trophoblasts inhibited ZIKV replication and decreased inflammatory cytokine responses and cell death. ZIKV-infected pregnant mice lacking Peli1 signaling showed reduced placental inflammation and tissue damage, which resulted in attenuated congenital abnormalities. Furthermore, Smaducin-6, a membrane-tethered Smad6-derived peptide, blocked Peli1-mediated NF-κB activation, but not ZIKV replication. Smaducin 6 inhibited Peli1-mediated inflammatory cytokine responses and cell death in placental trophoblasts and hNSCs, and attenuated congenital malformations in mice. Collectively, our results reveal a novel role of Peli1 in flavivirus pathogenesis and suggest that Peli1 promotes ZIKV vertical transmission and neuronal loss by mediating inflammatory cytokine responses and induction of cell death.
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Liu S, Jia J, Zhou H, Zhang C, Liu L, Liu J, Lu L, Li X, Kang Y, Lou Y, Cai Z, Ren Y, Kong X, Feng S. PTEN modulates neurites outgrowth and neuron apoptosis involving the PI3K/Akt/mTOR signaling pathway. Mol Med Rep 2019; 20:4059-4066. [PMID: 31702028 PMCID: PMC6797942 DOI: 10.3892/mmr.2019.10670] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2018] [Accepted: 07/18/2019] [Indexed: 02/07/2023] Open
Abstract
The present study aimed to explore the role of the PTEN/Akt/mTOR signaling pathway in the neurite outgrowth and apoptosis of cortical neurons. Cortical neurons were seeded on or adjacent to chondroitin sulfate proteoglycans. The length, number and crossing behavior of the neurites were calculated. Immunohistochemical staining and TUNEL data were analyzed. Neurites treated with PTEN inhibitor exhibited significant enhancements in elongation, initiation and crossing abilities when they encountered chondroitin sulfate proteoglycans in vitro. These effects disappeared when the PTEN/Akt/mTOR signaling pathway was blocked. Neurons exhibited significant enhancements in survival ability following PTEN inhibition. The present study demonstrated that PTEN inhibition can promote axonal elongation and initiation in cerebral cortical neurons, as well as the ability to cross the chondroitin sulfate proteoglycan border. In addition, PTEN inhibition is useful for protecting the neuron from apoptosis. The PTEN/Akt/mTOR signaling pathway is an important signaling pathway.
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Affiliation(s)
- Shen Liu
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Jun Jia
- Department of Trauma Orthopedics, Tianjin Hospital, Tianjin 300211, P.R. China
| | - Hengxing Zhou
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Chi Zhang
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Lu Liu
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Jun Liu
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Lu Lu
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Xueying Li
- Key Laboratory of Immuno Microenvironment and Disease of the Educational Ministry of China, Department of Immunology, Tianjin Medical University, Tianjin 300070, P.R. China
| | - Yi Kang
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Yongfu Lou
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Zhiwei Cai
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Yiming Ren
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
| | - Xiaohong Kong
- Laboratory of Medical Molecular Virology, School of Medicine, Nankai University, Tianjin 300071, P.R. China
| | - Shiqing Feng
- Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin 300052, P.R. China
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7
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Miceli M, Baldi D, Cavaliere C, Soricelli A, Salvatore M, Napoli C. Peripheral artery disease: the new frontiers of imaging techniques to evaluate the evolution of regenerative medicine. Expert Rev Cardiovasc Ther 2019; 17:511-532. [PMID: 31220944 DOI: 10.1080/14779072.2019.1635012] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
Introduction: Stem cells (ESC, iPSC, MSC) are known to have intrinsic regenerative properties. In the last decades numerous findings have favored the development of innovative therapeutic protocols based on the use of stem cells (Regenerative Medicine/Cell Therapy) for the treatment of numerous diseases including PAD, with promising results in preclinical studies. So far, several clinical studies have shown a general improvement of the patient's clinical outcome, however they possess many critical issues caused by the non-randomized design of the limited number of patients examined, the type cells to be used, their dosage, the short duration of treatment and also their delivery strategy. Areas covered: In this context, the use of the most advanced molecular imaging techniques will allow the visualization of very important physio-pathological processes otherwise invisible with conventional techniques, such as angiogenesis, also providing important structural and functional data. Expert opinion: The new frontier of cell therapy applied to PAD, potentially able to stop or even the process that causes the disease, with particular emphasis on the clinical aspects that different types of cells involve and on the use of more innovative molecular imaging techniques now available.
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Affiliation(s)
| | | | | | - Andrea Soricelli
- a IRCCS SDN , Naples , Italy.,b Department of Exercise and Wellness Sciences , University of Naples Parthenope , Naples , Italy
| | | | - Claudio Napoli
- a IRCCS SDN , Naples , Italy.,c University Department of Advanced Medical and Surgical Sciences, Clinical Department of Internal Medicine and Specialty Medicine , Università degli Studi della Campania 'Luigi Vanvitelli' , Napes , Italy
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8
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Mitra J, Guerrero EN, Hegde PM, Liachko NF, Wang H, Vasquez V, Gao J, Pandey A, Taylor JP, Kraemer BC, Wu P, Boldogh I, Garruto RM, Mitra S, Rao KS, Hegde ML. Motor neuron disease-associated loss of nuclear TDP-43 is linked to DNA double-strand break repair defects. Proc Natl Acad Sci U S A 2019; 116:4696-4705. [PMID: 30770445 PMCID: PMC6410842 DOI: 10.1073/pnas.1818415116] [Citation(s) in RCA: 199] [Impact Index Per Article: 33.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Genome damage and their defective repair have been etiologically linked to degenerating neurons in many subtypes of amyotrophic lateral sclerosis (ALS) patients; however, the specific mechanisms remain enigmatic. The majority of sporadic ALS patients feature abnormalities in the transactivation response DNA-binding protein of 43 kDa (TDP-43), whose nucleo-cytoplasmic mislocalization is characteristically observed in spinal motor neurons. While emerging evidence suggests involvement of other RNA/DNA binding proteins, like FUS in DNA damage response (DDR), the role of TDP-43 in DDR has not been investigated. Here, we report that TDP-43 is a critical component of the nonhomologous end joining (NHEJ)-mediated DNA double-strand break (DSB) repair pathway. TDP-43 is rapidly recruited at DSB sites to stably interact with DDR and NHEJ factors, specifically acting as a scaffold for the recruitment of break-sealing XRCC4-DNA ligase 4 complex at DSB sites in induced pluripotent stem cell-derived motor neurons. shRNA or CRISPR/Cas9-mediated conditional depletion of TDP-43 markedly increases accumulation of genomic DSBs by impairing NHEJ repair, and thereby, sensitizing neurons to DSB stress. Finally, TDP-43 pathology strongly correlates with DSB repair defects, and damage accumulation in the neuronal genomes of sporadic ALS patients and in Caenorhabditis elegans mutant with TDP-1 loss-of-function. Our findings thus link TDP-43 pathology to impaired DSB repair and persistent DDR signaling in motor neuron disease, and suggest that DSB repair-targeted therapies may ameliorate TDP-43 toxicity-induced genome instability in motor neuron disease.
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Affiliation(s)
- Joy Mitra
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
| | - Erika N Guerrero
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
- Center for Neuroscience, Instituto de Investigaciones Científicas y Servicios de Alta Tecnología, City of Knowledge, Panama, Republic of Panama
- Department of Biotechnology, Acharya Nagarjuna University, Guntur 522510, India
| | - Pavana M Hegde
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
| | - Nicole F Liachko
- Geriatric Research Education and Clinical Center, Veterans Affairs Puget Sound Health Care System, Seattle, WA 98108
- Division of Gerontology and Geriatric Medicine, Department of Medicine, University of Washington, Seattle, WA 98104
| | - Haibo Wang
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
- Institute of Academic Medicine, Houston Methodist Research Institute, Houston, TX 77030
| | - Velmarini Vasquez
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
- Center for Neuroscience, Instituto de Investigaciones Científicas y Servicios de Alta Tecnología, City of Knowledge, Panama, Republic of Panama
- Department of Biotechnology, Acharya Nagarjuna University, Guntur 522510, India
| | - Junling Gao
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555
| | - Arvind Pandey
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
| | - J Paul Taylor
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, TN 38105
- Howard Hughes Medical Institute, St. Jude Children's Research Hospital, Chevy Chase, MD 20815
| | - Brian C Kraemer
- Geriatric Research Education and Clinical Center, Veterans Affairs Puget Sound Health Care System, Seattle, WA 98108
- Division of Gerontology and Geriatric Medicine, Department of Medicine, University of Washington, Seattle, WA 98104
| | - Ping Wu
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555
| | - Istvan Boldogh
- Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555
| | - Ralph M Garruto
- Department of Anthropology, Binghamton University, State University of New York, Binghamton, NY 13902;
- Department of Biological Sciences, Binghamton University, State University of New York, Binghamton, NY 13902
| | - Sankar Mitra
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030
- Department of Radiation Oncology, Weill Medical College, New York, NY 10065
| | - K S Rao
- Center for Neuroscience, Instituto de Investigaciones Científicas y Servicios de Alta Tecnología, City of Knowledge, Panama, Republic of Panama
| | - Muralidhar L Hegde
- Department of Radiation Oncology, Houston Methodist Research Institute, Houston, TX 77030;
- Institute of Academic Medicine, Houston Methodist Research Institute, Houston, TX 77030
- Department of Radiation Oncology, Weill Medical College, New York, NY 10065
- Houston Methodist Neurological Institute, Houston Methodist Research Institute, Houston, TX 77030
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Wang L, Schlagal CR, Gao J, Hao Y, Dunn TJ, McGrath EL, Labastida JA, Yu Y, Feng SQ, Liu SY, Wu P. Oligodendrocyte differentiation from human neural stem cells: A novel role for c-Src. Neurochem Int 2018; 120:21-32. [PMID: 30041015 DOI: 10.1016/j.neuint.2018.07.006] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2018] [Revised: 06/28/2018] [Accepted: 07/18/2018] [Indexed: 01/06/2023]
Abstract
Human neural stem cells (hNSCs) can differentiate into an oligodendrocyte lineage to facilitate remyelination in patients. Molecular mechanisms underlying oligodendrocyte fate specification remains unknown, hindering the development of efficient methods to generate oligodendrocytes from hNSCs. We have found that Neurobasal-A medium (NB) is capable of inducing hNSCs to oligodendrocyte progenitor cells (OPCs). We identified several signaling molecules are altered after cultivation in NB medium, including Akt, ERK1/2 and c-Src. While sustained activation of Akt and ERK1/2 during both NB induction and subsequent differentiation was required for OPC differentiation, c-Src phosphorylation was increased temporally during the period of NB induction. Both pharmacological inhibition and RNA interference confirmed that a transient elevation of phospho-c-Src is critical for OPC induction. Furthermore, inactivation of c-Src inhibited phosphorylation of Akt and ERK1/2. In summary, we identified a novel and critical role of c-Src in guiding hNSC differentiation to an oligodendrocyte lineage.
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Affiliation(s)
- Le Wang
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA; Department of Spine Surgery, The First Affiliated Hospital, Sun Yat-sen University, 58 Zhongshan Er Rd, Yuexiu Qu, Guangzhou Shi, Guangdong Sheng, China
| | - Caitlin R Schlagal
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Junling Gao
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Yan Hao
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA; Department of Orthopedics, Tianjin Medical University General Hospital, 154 Anshan Rd, Heping Qu, 300051, China
| | - Tiffany J Dunn
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Erica L McGrath
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Javier Allende Labastida
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Yongjia Yu
- Department of Radiation Oncology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA
| | - Shi-Qing Feng
- Department of Orthopedics, Tianjin Medical University General Hospital, 154 Anshan Rd, Heping Qu, 300051, China
| | - Shao-Yu Liu
- Department of Spine Surgery, The First Affiliated Hospital, Sun Yat-sen University, 58 Zhongshan Er Rd, Yuexiu Qu, Guangzhou Shi, Guangdong Sheng, China
| | - Ping Wu
- Department of Neuroscience, Cell Biology and Anatomy, University of Texas Medical Branch, 301 University Blvd, Galveston, TX, 77555, USA.
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10
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Marei HES, El-Gamal A, Althani A, Afifi N, Abd-Elmaksoud A, Farag A, Cenciarelli C, Thomas C, Anwarul H. Cholinergic and dopaminergic neuronal differentiation of human adipose tissue derived mesenchymal stem cells. J Cell Physiol 2018; 233:936-945. [PMID: 28369825 DOI: 10.1002/jcp.25937] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2016] [Accepted: 03/27/2017] [Indexed: 02/06/2023]
Abstract
Mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into various cell types such as cartilage, bone, and fat cells. Recent studies have shown that induction of MSCs in vitro by growth factors including epidermal growth factor (EGF) and fibroblast growth factor (FGF2) causes them to differentiate into neural like cells. These cultures also express ChAT, a cholinergic marker; and TH, a dopaminergic marker for neural cells. To establish a protocol with maximum differentiation potential, we examined MSCs under three experimental culture conditions using neural induction media containing FGF2, EGF, BMP-9, retinoic acid, and heparin. Adipose-derived MSCs were extracted and expanded in vitro for 3 passages after reaching >80% confluency, for a total duration of 9 days. Cells were then characterized by flow cytometry for CD markers as CD44 positive and CD45 negative. MSCs were then treated with neural induction media and were characterized by morphological changes and Q-PCR. Differentiated MSCs expressed markers for immature and mature neurons; β Tubulin III (TUBB3) and MAP2, respectively, showing the neural potential of these cells to differentiate into functional neurons. Improved protocols for MSCs induction will facilitate and ensure the reproducibility and standard production of MSCs for therapeutic applications in neurodegenerative diseases.
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Affiliation(s)
| | - Aya El-Gamal
- Faculty of Veterinary Medicine, Department of Cytology and Histology, Mansoura University, Mansoura, Egypt
| | - Asma Althani
- Biomedical Research Center, Qatar University, Doha, Qatar
| | | | - Ahmed Abd-Elmaksoud
- Faculty of Veterinary Medicine, Department of Cytology and Histology, Mansoura University, Mansoura, Egypt
| | - Amany Farag
- Faculty of Veterinary Medicine, Department of Cytology and Histology, Mansoura University, Mansoura, Egypt
| | | | - Caceci Thomas
- Department of Biomedical Sciences, Virginia Tech Carilion School of Medicine, Roanoke, Virginia
| | - Hasan Anwarul
- Department of Mechanical and Industrial Engineering, Qatar University, Doha, Qatar
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11
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Cardozo MJ, Mysiak KS, Becker T, Becker CG. Reduce, reuse, recycle – Developmental signals in spinal cord regeneration. Dev Biol 2017; 432:53-62. [DOI: 10.1016/j.ydbio.2017.05.011] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2016] [Revised: 02/03/2017] [Accepted: 05/11/2017] [Indexed: 02/06/2023]
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12
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Raik S, Kumar A, Bhattacharyya S. Insights into cell-free therapeutic approach: Role of stem cell "soup-ernatant". Biotechnol Appl Biochem 2017; 65:104-118. [PMID: 28321921 DOI: 10.1002/bab.1561] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2016] [Accepted: 03/02/2017] [Indexed: 12/16/2022]
Abstract
Current advances in medicine have revolutionized the field of regenerative medicine dramatically with newly evolved therapies for repair or replacement of degenerating or injured tissues. Stem cells (SCs) can be harvested from different sources for clinical therapeutics, which include fetal tissues, umbilical cord blood, embryos, and adult tissues. SCs can be isolated and differentiated into desired lineages for tissue regeneration and cell replacement therapy. However, several loopholes need to be addressed properly before this can be extended for large-scale therapeutic application. These include a careful approach for patient safety during SC treatments and tolerance of recipients. SC treatments are associated with a number of risk factors and require successful integration and survival of transplanted cells in the desired microenvironment with concurrent tissue regeneration. Recent studies have focused on developing alternatives that can replace the cell-based therapy using paracrine factors. The development of stem "cell free" therapies can be devoted mainly to the use of soluble factors (secretome), extracellular vesicles, and mitochondrial transfer. The present review emphasizes on the paradigms related to the use of SC-based therapeutics and the potential applications of a cell-free approach as an alternative to cell-based therapy in the area of regenerative medicine.
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Affiliation(s)
- Shalini Raik
- Department of Biophysics, PGIMER, Chandigarh, India
| | - Ajay Kumar
- Department of Biophysics, PGIMER, Chandigarh, India
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13
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Abstract
During vertebrate embryonic development, the spinal cord is formed by the neural derivatives of a neuromesodermal population that is specified at early stages of development and which develops in concert with the caudal regression of the primitive streak. Several processes related to spinal cord specification and maturation are coupled to this caudal extension including neurogenesis, ventral patterning and neural crest specification and all of them seem to be crucially regulated by Fibroblast Growth Factor (FGF) signaling, which is prominently active in the neuromesodermal region and transiently in its derivatives. Here we review the role of FGF signaling in those processes, trying to separate its different functions and highlighting the interactions with other signaling pathways. Finally, these early functions of FGF signaling in spinal cord development may underlay partly its ability to promote regeneration in the lesioned spinal cord as well as its action promoting specific fates in neural stem cell cultures that may be used for therapeutical purposes.
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Affiliation(s)
- Ruth Diez Del Corral
- Department of Cellular, Molecular and Developmental Neurobiology, Cajal Institute, Consejo Superior de Investigaciones CientíficasMadrid, Spain.,Champalimaud Research, Champalimaud Centre for the UnknownLisbon, Portugal
| | - Aixa V Morales
- Department of Cellular, Molecular and Developmental Neurobiology, Cajal Institute, Consejo Superior de Investigaciones CientíficasMadrid, Spain
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14
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McGrath EL, Rossi SL, Gao J, Widen SG, Grant AC, Dunn TJ, Azar SR, Roundy CM, Xiong Y, Prusak DJ, Loucas BD, Wood TG, Yu Y, Fernández-Salas I, Weaver SC, Vasilakis N, Wu P. Differential Responses of Human Fetal Brain Neural Stem Cells to Zika Virus Infection. Stem Cell Reports 2017; 8:715-727. [PMID: 28216147 PMCID: PMC5355569 DOI: 10.1016/j.stemcr.2017.01.008] [Citation(s) in RCA: 86] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2016] [Revised: 01/12/2017] [Accepted: 01/12/2017] [Indexed: 11/17/2022] Open
Abstract
Zika virus (ZIKV) infection causes microcephaly in a subset of infants born to infected pregnant mothers. It is unknown whether human individual differences contribute to differential susceptibility of ZIKV-related neuropathology. Here, we use an Asian-lineage ZIKV strain, isolated from the 2015 Mexican outbreak (Mex1-7), to infect primary human neural stem cells (hNSCs) originally derived from three individual fetal brains. All three strains of hNSCs exhibited similar rates of Mex1-7 infection and reduced proliferation. However, Mex1-7 decreased neuronal differentiation in only two of the three stem cell strains. Correspondingly, ZIKA-mediated transcriptome alterations were similar in these two strains but significantly different from that of the third strain with no ZIKV-induced neuronal reduction. This study thus confirms that an Asian-lineage ZIKV strain infects primary hNSCs and demonstrates a cell-strain-dependent response of hNSCs to ZIKV infection.
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Affiliation(s)
- Erica L McGrath
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Shannan L Rossi
- Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Junling Gao
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Steven G Widen
- Sealy Center for Molecular Medicine, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Auston C Grant
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Tiffany J Dunn
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Sasha R Azar
- Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX 77555, USA; Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Christopher M Roundy
- Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX 77555, USA; Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Ying Xiong
- Department of Radiology and Oncology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Deborah J Prusak
- Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Bradford D Loucas
- Department of Radiology and Oncology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Thomas G Wood
- Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Yongjia Yu
- Department of Radiology and Oncology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Ildefonso Fernández-Salas
- Instituto Nacional de Salud Pública, Centro Regional de Salud Pública, Tapachula, Chiapas 30700, Mexico
| | - Scott C Weaver
- Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX 77555, USA; Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA
| | - Nikos Vasilakis
- Department of Pathology, Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX 77555, USA.
| | - Ping Wu
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA; Beijing Institute for Brain Disorders, Capital Medical University, Beijing 100069, China.
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15
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Han Y, Kim KT. Neural Growth Factor Stimulates Proliferation of Spinal Cord Derived-Neural Precursor/Stem Cells. J Korean Neurosurg Soc 2016; 59:437-41. [PMID: 27651860 PMCID: PMC5028602 DOI: 10.3340/jkns.2016.59.5.437] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2016] [Revised: 05/10/2016] [Accepted: 06/08/2016] [Indexed: 01/08/2023] Open
Abstract
Objective Recently, regenerative therapies have been used in clinical trials (heart, cartilage, skeletal). We don't make use of these treatments to spinal cord injury (SCI) patients yet, but regenerative therapies are rising interest in recent study about SCI. Neural precursor/stem cell (NPSC) proliferation is a significant event in functional recovery of the central nervous system (CNS). However, brain NPSCs and spinal cord NPSCs (SC-NPSCs) have many differences including gene expression and proliferation. The purpose of this study was to investigate the influence of neural growth factor (NGF) on the proliferation of SC-NPSCs. Methods NPSCs (2×104) were suspended in 100 µL of neurobasal medium containing NGF-7S (Sigma-Aldrich) and cultured in a 96-well plate for 12 days. NPSC proliferation was analyzed five times for either concentration of NGF (0.02 and 2 ng/mL). Sixteen rats after SCI were randomly allocated into two groups. In group 1 (SCI-vehicle group, n=8), animals received 1.0 mL of the saline vehicle solution. In group 2 (SCI-NGF group, n=8), the animals received single doses of NGF (Sigma-Aldrich). A dose of 0.02 ng/mL of NGF or normal saline as a vehicle control was intra-thecally injected daily at 24 hour intervals for 7 days. For Immunohistochemistry analysis, rats were sacrificed after one week and the spinal cords were obtained. Results The elevation of cell proliferation with 0.02 ng/mL NGF was significant (p<0.05) but was not significant for 2 ng/mL NGF. The optical density was increased in the NGF 0.02 ng/mL group compared to the control group and NGF 2 ng/mL groups. The density of nestin in the SCI-NGF group was significantly increased over the SCI-vehicle group (p<0.05). High power microscopy revealed that the density of nestin in the SCI-NGF group was significantly increased over the SCI-vehicle group. Conclusion SC-NPSC proliferation is an important pathway in the functional recovery of SCI. NGF enhances SC-NPSC proliferation in vitro and in vivo. NGF may be a useful option for treatment of SCI patients pending further studies to verify the clinical applicability.
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Affiliation(s)
- Youngmin Han
- Department of Neurosurgery, Kyungpook National University Hospital, Daegu, Korea
| | - Kyoung-Tae Kim
- Department of Neurosurgery, Kyungpook National University Hospital, Daegu, Korea
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16
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Schubert KO, Weiland F, Baune BT, Hoffmann P. The use of MALDI-MSI in the investigation of psychiatric and neurodegenerative disorders: A review. Proteomics 2016; 16:1747-58. [DOI: 10.1002/pmic.201500460] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2015] [Revised: 02/08/2016] [Accepted: 02/24/2016] [Indexed: 11/11/2022]
Affiliation(s)
| | - Florian Weiland
- Adelaide Proteomics Centre; The University of Adelaide; Adelaide Australia
- Institute for Photonics and Advanced Sensing (IPAS); The University of Adelaide; Adelaide Australia
| | - Bernhard T. Baune
- Discipline of Psychiatry; The University of Adelaide; Adelaide Australia
| | - Peter Hoffmann
- Adelaide Proteomics Centre; The University of Adelaide; Adelaide Australia
- Institute for Photonics and Advanced Sensing (IPAS); The University of Adelaide; Adelaide Australia
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17
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Javanmardy S, Asadi MH, Movahedin M, Moradpour F, Bahadoran H. Derivation of motor neuron-like cells from neonatal mouse testis in a simple culture condition. Andrologia 2016; 48:1100-1107. [PMID: 26892722 DOI: 10.1111/and.12545] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/04/2015] [Indexed: 11/28/2022] Open
Abstract
Embryonic stem cell (ESC) therapy is an exciting way to treat neurodegenerative disease and central nervous system injury. However, many ethical and immunological problems surround the use of embryonic stem cells. Finding an alternative source of stem cells is therefore pertinent. In this study, spermatogonia stem cells (SSCs) were used to generate mature motor neurons. SSCs were extracted from neonatal testes and cultured in DMED/F12 medium for 3 weeks. Characterisation of SSC-derived ESC-like cells was confirmed by RT-qPCR, immunostaining, alkaline phosphatase activity and their ability to form embryoid bodies (EBs). The EBs were induced by retinoic acid and Sonic hedgehog and trypsinised to obtain single induced cells. The single cells were cultured in neural medium for 18 days. Characterisation of neural precursors and motor neuron-like cells was confirmed by RT-qPCR and immunocytochemical analysis at the 7th day (early stage) and 18th day (late stage), respectively, of culturing. The neural precursors were found to be positive for nestin and Sox2, and a small fraction of cells expressed β-tubulin III. Upon further differentiation, multipolar neurons were detected that expressed β-tubulin III and MAP2 markers. Moreover, the expression levels of Olig2 and PAX6 were significantly lower, while HB9, Isl1 and Isl2 expression levels were higher at the late stage when compared to the early stage. These results show that SSCs have the potential to differentiate to motor neuron-like cells and express markers specific for mature motor neurons. However, the functional ability of these cells remains to be evaluated in future studies.
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Affiliation(s)
- S Javanmardy
- Department of Midwifery, School of Nursing & Midwifery, Kermanshah University of Medical Sciences, Kermanshah, Iran
| | - M H Asadi
- Department of Anatomical Sciences, Faculty of medicine, Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - M Movahedin
- Department of Anatomical Sciences, Faculty of medical sciences, Tarbiat Modares University, Tehran, Iran
| | - F Moradpour
- Department of Physiology & Pharmacology, School of Medicine, Kermanshah University of Medical Sciences, Kermanshah, Iran
| | - H Bahadoran
- Department of Anatomical Sciences, Faculty of medicine, Baqiyatallah University of Medical Sciences, Tehran, Iran
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18
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Nam H, Lee KH, Nam DH, Joo KM. Adult human neural stem cell therapeutics: Current developmental status and prospect. World J Stem Cells 2015; 7:126-136. [PMID: 25621112 PMCID: PMC4300923 DOI: 10.4252/wjsc.v7.i1.126] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/28/2014] [Revised: 09/01/2014] [Accepted: 10/16/2014] [Indexed: 02/06/2023] Open
Abstract
Over the past two decades, regenerative therapies using stem cell technologies have been developed for various neurological diseases. Although stem cell therapy is an attractive option to reverse neural tissue damage and to recover neurological deficits, it is still under development so as not to show significant treatment effects in clinical settings. In this review, we discuss the scientific and clinical basics of adult neural stem cells (aNSCs), and their current developmental status as cell therapeutics for neurological disease. Compared with other types of stem cells, aNSCs have clinical advantages, such as limited proliferation, inborn differentiation potential into functional neural cells, and no ethical issues. In spite of the merits of aNSCs, difficulties in the isolation from the normal brain, and in the in vitro expansion, have blocked preclinical and clinical study using aNSCs. However, several groups have recently developed novel techniques to isolate and expand aNSCs from normal adult brains, and showed successful applications of aNSCs to neurological diseases. With new technologies for aNSCs and their clinical strengths, previous hurdles in stem cell therapies for neurological diseases could be overcome, to realize clinically efficacious regenerative stem cell therapeutics.
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19
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Zhou H, Li X, Wu Q, Li F, Fu Z, Liu C, Liang Z, Chu T, Wang T, Lu L, Ning G, Kong X, Feng S. shRNA against PTEN promotes neurite outgrowth of cortical neurons and functional recovery in spinal cord contusion rats. Regen Med 2014; 10:411-29. [PMID: 25495396 DOI: 10.2217/rme.14.88] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
AIM To explore neurite growth/regeneration and spinal cord injury repair after PTEN silencing via lentivirus-mediated RNAi. MATERIALS & METHODS Cortical neurons were seeded on or adjacent to chondroitin sulfate proteoglycans. The length, number and crossing behavior of neurites were calculated. Lentivirus was locally injected into spinal cord contusion rats. The functional recovery and immunohistochemical staining were analyzed. RESULTS Neurites with PTEN silencing exhibited significant enhancements in elongation, initiation and crossing ability when they encountered chondroitin sulfate proteoglycans in vitro. In vivo PTEN silencing improved functional recovery significantly, and promoted axon and synapse formation, but not scar formation. CONCLUSIONS PTEN silencing may be promising for spinal cord injury repair.
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Affiliation(s)
- Hengxing Zhou
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | | | - Qiang Wu
- 3Department of Orthopaedics, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, No. 314 Anshanxi Road, Nankai District, Tianjin 300193, PR China
| | - Fuyuan Li
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | | | - Chang Liu
- 4School of Medicine, Nankai University, No. 94 Weijin Road, Nankai District, Tianjin 300071, PR China
| | - Zhipin Liang
- 4School of Medicine, Nankai University, No. 94 Weijin Road, Nankai District, Tianjin 300071, PR China
| | - Tianci Chu
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | - Tianyi Wang
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | - Lu Lu
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | - Guangzhi Ning
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
| | - Xiaohong Kong
- 4School of Medicine, Nankai University, No. 94 Weijin Road, Nankai District, Tianjin 300071, PR China
| | - Shiqing Feng
- 1Department of Orthopaedics, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, PR China
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20
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Cholinergic differentiation of neural stem cells generated from cell aggregates-derived from Human Bone marrow stromal cells. Tissue Eng Regen Med 2014. [DOI: 10.1007/s13770-014-0019-6] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023] Open
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Liu X, Li D, Jiang D, Fang Y. Acetylcholine secretion by motor neuron-like cells from umbilical cord mesenchymal stem cells. Neural Regen Res 2014; 8:2086-92. [PMID: 25206517 PMCID: PMC4146069 DOI: 10.3969/j.issn.1673-5374.2013.22.008] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2013] [Accepted: 07/25/2013] [Indexed: 11/29/2022] Open
Abstract
Umbilical cord mesenchymal stem cells were isolated by a double enzyme digestion method. The third passage of umbilical cord mesenchymal stem cells was induced with heparin and/or basic fibroblast growth factor. Results confirmed that cell morphology did not change after induction with basic fibroblast growth factor alone. However, neuronal morphology was visible, and microtubule-associated protein-2 expression and acetylcholine levels increased following induction with heparin alone or heparin combined with basic fibroblast growth factor. Hb9 and choline acetyltransferase expression was high following inductive with heparin combined with basic fibroblast growth factor. Results indicate that the inductive effect of basic fibroblast growth factor alone was not obvious. Heparin combined with basic fibroblast growth factor noticeably promoted the differentiation of umbilical cord mesenchymal stem cells into motor neuron-like cells. Simultaneously, umbilical cord mesenchymal stem cells could secrete acetylcholine.
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Affiliation(s)
- Xueyuan Liu
- Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
| | - Dehua Li
- Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
| | - Dong Jiang
- Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
| | - Yan Fang
- Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
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Combined use of NGF/BDNF/bFGF promotes proliferation and differentiation of neural stem cells in vitro. Int J Dev Neurosci 2014; 38:74-8. [DOI: 10.1016/j.ijdevneu.2014.08.002] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2014] [Revised: 08/03/2014] [Accepted: 08/03/2014] [Indexed: 02/06/2023] Open
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23
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Natarajan R, Singal V, Benes R, Gao J, Chan H, Chen H, Yu Y, Zhou J, Wu P. STAT3 modulation to enhance motor neuron differentiation in human neural stem cells. PLoS One 2014; 9:e100405. [PMID: 24945434 PMCID: PMC4063761 DOI: 10.1371/journal.pone.0100405] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2014] [Accepted: 05/27/2014] [Indexed: 11/19/2022] Open
Abstract
Spinal cord injury or amyotrophic lateral sclerosis damages spinal motor neurons and forms a glial scar, which prevents neural regeneration. Signal transducer and activator of transcription 3 (STAT3) plays a critical role in astrogliogenesis and scar formation, and thus a fine modulation of STAT3 signaling may help to control the excessive gliogenic environment and enhance neural repair. The objective of this study was to determine the effect of STAT3 inhibition on human neural stem cells (hNSCs). In vitro hNSCs primed with fibroblast growth factor 2 (FGF2) exhibited a lower level of phosphorylated STAT3 than cells primed by epidermal growth factor (EGF), which correlated with a higher number of motor neurons differentiated from FGF2-primed hNSCs. Treatment with STAT3 inhibitors, Stattic and Niclosamide, enhanced motor neuron differentiation only in FGF2-primed hNSCs, as shown by increased homeobox gene Hb9 mRNA levels as well as HB9+ and microtubule-associated protein 2 (MAP2)+ co-labeled cells. The increased motor neuron differentiation was accompanied by a decrease in the number of glial fibrillary acidic protein (GFAP)-positive astrocytes. Interestingly, Stattic and Niclosamide did not affect the level of STAT3 phosphorylation; rather, they perturbed the nuclear translocation of phosphorylated STAT3. In summary, we demonstrate that FGF2 is required for motor neuron differentiation from hNSCs and that inhibition of STAT3 further increases motor neuron differentiation at the expense of astrogliogenesis. Our study thus suggests a potential benefit of targeting the STAT3 pathway for neurotrauma or neurodegenerative diseases.
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Affiliation(s)
- Rajalaxmi Natarajan
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Vinamrata Singal
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Richard Benes
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Junling Gao
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Hoi Chan
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Haijun Chen
- Department of Pharmacology & Toxicology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Yongjia Yu
- Department of Radiation Oncology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Jia Zhou
- Department of Pharmacology & Toxicology, The University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Ping Wu
- Department of Neuroscience and Cell Biology, The University of Texas Medical Branch, Galveston, Texas, United States of America
- * E-mail:
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24
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Acquadro E, Caron I, Tortarolo M, Bucci EM, Bendotti C, Corpillo D. Human SOD1-G93A specific distribution evidenced in murine brain of a transgenic model for amyotrophic lateral sclerosis by MALDI imaging mass spectrometry. J Proteome Res 2014; 13:1800-9. [PMID: 24579824 DOI: 10.1021/pr400942n] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023]
Abstract
Amyotrophic lateral sclerosis (ALS) is a progressive, fatal neurodegenerative disease caused by the degeneration of motor neurons. The transgenic mouse model carrying the human SOD1G93A mutant gene (hSOD1G93A mouse) represents one of the most reliable and widely used model of this pathology. In the present work, the innovative technique of matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) was applied in the study of pathological alterations at the level of small brain regions such as facial and trigeminal nuclei, which in rodents are extremely small and would be difficult to analyze with classical proteomics approaches. Comparing slices from three mice groups (transgenic hSOD1G93A, transgenic hSOD1WT, and nontransgenic, Ntg), this technique allowed us to evidence the accumulation of hSOD1G93A in the facial and trigeminal nuclei, where it generates aggregates. This phenomenon is likely to be correlated to the degeneration observed in these regions. Moreover, a statistical analysis allowed us to highlight other proteins as differentially expressed among the three mice groups analyzed. Some of them were identified by reverse-phase HPLC fractionation of extracted proteins and mass spectrometric analysis before and after trypsin digestion. In particular, the 40S ribosomal protein S19 (RPS19) was upregulated in the parenkyma and reactive glial cells in facial nuclei of hSOD1G93A mice when compared to transgenic hSOD1WT and nontransgenic ones.
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Affiliation(s)
- Elena Acquadro
- ABLE Bioscences, BioIndustry Park Silvano Fumero S.p.A., Via Ribes 5, 10010 Colleretto Giacosa, TO, Italy
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25
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Small molecules enable neurogenin 2 to efficiently convert human fibroblasts into cholinergic neurons. Nat Commun 2014; 4:2183. [PMID: 23873306 PMCID: PMC3843951 DOI: 10.1038/ncomms3183] [Citation(s) in RCA: 258] [Impact Index Per Article: 23.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2013] [Accepted: 06/24/2013] [Indexed: 12/16/2022] Open
Abstract
Cell fate can be reprogrammed by modifying intrinsic and extrinsic cues. Here, we show that two small molecules (forskolin and dorsomorphin) enable the transcription factor Neurogenin 2 (NGN2) to convert human fetal lung fibroblasts into cholinergic neurons with high purity (>90%) and efficiency (up to 99% of NGN2-expressing cells). The conversion is direct without passing through a proliferative progenitor state. These human induced cholinergic neurons show mature electrophysiological properties and exhibit motor neuron-like features including morphology, gene expression, and the formation of functional neuromuscular junctions. Inclusion of an additional transcription factor, SOX11 also efficiently converts postnatal and adult skin fibroblasts from healthy and diseased human patients to cholinergic neurons. Taken together, this study identifies a simple and highly efficient strategy for reprogramming human fibroblasts to subtype-specific neurons. These findings offer a unique venue for investigating the molecular mechanisms underlying cellular plasticity and human neurodegenerative diseases.
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Li H, Miao J, Zhao G, Wu D, Liu B, Wei X, Cao S, Gu H, Zhang Y, Wang L, Fan Y, Yuan Z. Different expression patterns of growth factors in rat fetuses with spina bifida aperta after in utero mesenchymal stromal cell transplantation. Cytotherapy 2013; 16:319-30. [PMID: 24364908 DOI: 10.1016/j.jcyt.2013.10.005] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2012] [Revised: 10/07/2013] [Accepted: 10/14/2013] [Indexed: 12/14/2022]
Abstract
BACKGROUND AIMS In a previous study, we successfully devised a prenatal surgical approach and transplanted mesenchymal stromal cells (MSCs) to fetal rat spinal column to treat retinoic acid-induced neural tube defects in rat. Our results show that MSCs survived, migrated and differentiated into neural lineage cells. We intended to study various growth factor expressions in rat fetal spinal cords with spina bifida aperta after in utero MSC transplantation and the effect of in vivo growth factor introduction for prenatal spina bifida treatment. METHODS Pregnant rats were treated with retinoic acid on embryonic day 10 and then received fetal surgery for MSC transplantation and/or lentiviral epidermal growth factor (EGF) injection on embryonic day 16; various growth factor expression in spinal cords from embryonic day 20 fetuses were analyzed by means of quantitative reverse transcriptase-polymerase chain reaction. Terminal deoxynucleotidyl transferase dUTP nick end labeling analysis was performed to observe spinal tissue apoptosis. RESULTS Growth factor expression was dysregulated in spinal cords with spina bifida. After MSC transplantation, we observed significantly increased expression of EGF, fibroblast growth factor (FGF)-8, FGF-2 and FGF-20 in the MSC transplantation group compared with blank injection; Furthermore, EGF expression positively correlated with surviving MSC amounts. Expression of other growth factors was not significantly different. In vivo EGF introduction reduced spinal tissue apoptosis. CONCLUSIONS Our results suggest that intrinsic EGF and FGF-2, FGF-8 and FGF-20 might affect the in vivo fate of transplanted MSCs in a fetal rat spina bifida model. In vivo EGF introduction together with MSC transplantation might serve as a new strategy for prenatal spina bifida treatment.
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Affiliation(s)
- Hui Li
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Jianing Miao
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Guifeng Zhao
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Di Wu
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Bo Liu
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Xiaowei Wei
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Songying Cao
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Hui Gu
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Yi Zhang
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Lili Wang
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Yang Fan
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China
| | - Zhengwei Yuan
- Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, Shenyang, China.
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Yoo J, Kim HS, Hwang DY. Stem cells as promising therapeutic options for neurological disorders. J Cell Biochem 2013; 114:743-53. [PMID: 23097262 DOI: 10.1002/jcb.24427] [Citation(s) in RCA: 48] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2012] [Accepted: 10/12/2012] [Indexed: 12/13/2022]
Abstract
Due to the limitations of pharmacological and other current therapeutic strategies, stem cell therapies have emerged as promising options for treating many incurable neurologic diseases. A variety of stem cells including pluripotent stem cells (i.e., embryonic stem cells and induced pluripotent stem cells) and multipotent adult stem cells (i.e., fetal brain tissue, neural stem cells, and mesenchymal stem cells from various sources) have been explored as therapeutic options for treating many neurologic diseases, and it is becoming obvious that each type of stem cell has pros and cons as a source for cell therapy. Wise selection of stem cells with regard to the nature and status of neurologic dysfunctions is required to achieve optimal therapeutic efficacy. To this aim, the stem cell-mediated therapeutic efforts on four major neurological diseases, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and stroke, will be introduced, and current problems and future directions will be discussed.
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Affiliation(s)
- Jongman Yoo
- Department of Biological Science, CHA University, Kyeonggido, Korea
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Morimoto Y, Kato-Negishi M, Onoe H, Takeuchi S. Three-dimensional neuron-muscle constructs with neuromuscular junctions. Biomaterials 2013; 34:9413-9. [PMID: 24041425 DOI: 10.1016/j.biomaterials.2013.08.062] [Citation(s) in RCA: 124] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2013] [Accepted: 08/20/2013] [Indexed: 12/13/2022]
Abstract
This paper describes a fabrication method of muscle tissue constructs driven by neurotransmitters released from activated motor neurons. The constructs consist of three-dimensional (3D) free-standing skeletal muscle fibers co-cultured with motor neurons. We differentiated mouse neural stem cells (mNSCs) cultured on the skeletal muscle fibers into neurons that extend their processes into the muscle fibers. We found that acetylcholine receptors (AChRs) were formed at the connection between the muscle fibers and the neurons. The neuron-muscle constructs consist of highly aligned, long and matured muscle fibers that facilitate wide contractions of muscle fibers in a single direction. The contractions of the neuron-muscle construct were observed after glutamic acid activation of the neurons. The contraction was stopped by treatment with curare, an neuromuscular junction (NMJ) antagonist. These results indicate that our method succeeded in the formation of NMJs in the neuron-muscle constructs. The neuron-muscle construct system can potentially be used in pharmacokinetic assays related to NMJ disease therapies and in soft-robotic actuators.
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Affiliation(s)
- Yuya Morimoto
- Center for International Research on Micronano Mechatronics (CIRMM), Institute of Industrial Science (IIS), The University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8505, Japan
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Miceli M, Franci G, Dell'Aversana C, Ricciardiello F, Petraglia F, Carissimo A, Perone L, Maruotti GM, Savarese M, Martinelli P, Cancemi M, Altucci L. MePR: a novel human mesenchymal progenitor model with characteristics of pluripotency. Stem Cells Dev 2013; 22:2368-83. [PMID: 23597129 DOI: 10.1089/scd.2012.0498] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Human embryo stem cells or adult tissues are excellent models for discovery and characterization of differentiation processes. The aims of regenerative medicine are to define the molecular and physiological mechanisms that govern stem cells and differentiation. Human mesenchymal stem cells (hMSCs) are multipotent adult stem cells that are able to differentiate into a variety of cell types under controlled conditions both in vivo and in vitro, and they have the remarkable ability of self-renewal. hMSCs derived from amniotic fluid and characterized by the expression of Oct-4 and Nanog, typical markers of pluripotent cells, represent an excellent model for studies on stemness. Unfortunately, the limited amount of cells available from each donation and, above all, the limited number of replications do not allow for detailed studies. Here, we report on the immortalization and characterization of novel mesenchymal progenitor (MePR) cell lines from amniotic fluid-derived hMSCs, whose biological properties are similar to primary amniocytes. Our data indicate that MePR cells display the multipotency potential and differentiation rates of hMSCs, thus representing a useful model to study both mechanisms of differentiation and pharmacological approaches to induce selective differentiation. In particular, MePR-2B cells, which carry a bona fide normal karyotype, might be used in basic stem cell research, leading to the development of new approaches for stem cell therapy and tissue engineering.
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Affiliation(s)
- Marco Miceli
- Dipartimento di Biochimica, Biofisica e Patologia Generale, Seconda Università di Napoli , Napoli, Italy
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Liao JM, Hu XX. Transplantation of umbilical cord blood-derived mesenchymal stem cells for treatment of liver cirrhosis: Research progress. Shijie Huaren Xiaohua Zazhi 2013; 21:508-513. [DOI: 10.11569/wcjd.v21.i6.508] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Cirrhosis is a serious threat to human health. Currently, there have been no available radical measures that can effectively block the process of this disease. The research progress in the field of stem cells brings an opportunity for the treatment of cirrhosis. Having a wide variety of sources, weak immunogenicity, and strong proliferation and differentiation ability, human umbilical cord blood-derived mesenchymal stem cells have been demonstrated to be promising in the treatment of liver cirrhosis. This article reviews the biological characteristics of human umbilical cord blood mesenchymal stem cells and their application in the treatment of cirrhosis.
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Luan P, Zhou HH, Zhang B, Liu AM, Yang LH, Weng XL, Tao EX, Liu J. Basic fibroblast growth factor protects C17.2 cells from radiation-induced injury through ERK1/2. CNS Neurosci Ther 2013; 18:767-72. [PMID: 22943143 DOI: 10.1111/j.1755-5949.2012.00365.x] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022] Open
Abstract
AIMS To establish a radiation-induced neural injury model using C17.2 neural stem cells (NSCs) and to investigate whether basic fibroblast growth factor (bFGF) can protect the radiation-induced injury of C17.2 NSCs. Furthermore, we aim to identify the possible mechanisms involved in this model. METHODS C17.2 NSCs received a single exposure (3, 6, and 9 Gy, respectively) at a dose rate of 300 cGy/min with a control group receiving 0 Gy. Different concentrations of bFGF were added for 24 h, 5 min postirradiation. The MTS assay and flow cytometry were used to detect cytotoxicity and apoptosis. Expression of FGFR1, ERK1/2, and p-ERK1/2 proteins was detected with or without U0126 was pretreated prior to C17.2 NSCs receiving irradiation. RESULTS C17.2 NSCs showed a dose-dependent cell death as the dose of radiation was increased. Additionally, the rate of apoptosis in the C17.2 NSCs reached 31.2 ± 1.23% in the 6 Gy irradiation group, which was the most significant when compared to the other irradiation treated groups. bFGF showed protective effect on cell apoptosis in a dose-dependent manner. The mean percentage of apoptotic cells decreased to 7.83 ± 1.75% when 100 ng/mL bFGF was given. Furthermore, U0126 could block the protective effect of bFGF by inhibiting the phosphorylation of ERK1/2. CONCLUSIONS An in vitro cellular model of radiation-induced apoptosis of NSCs, in C17.2 NSCs, was developed successfully. Additionally, bFGF can protect neurons from radiation injury in vitro via the ERK1/2 signal transduction pathway.
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Affiliation(s)
- Ping Luan
- Medical School, Shenzhen University, China
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Abstract
Regeneration of the nervous system requires either the repair or replacement of nerve cells that have been damaged by injury or disease. While lower organisms possess extensive capacity for neural regeneration, evolutionarily higher organisms including humans are limited in their ability to regenerate nerve cells, posing significant issues for the treatment of injury and disease of the nervous system. This chapter focuses on current approaches for neural regeneration, with a discussion of traditional methods to enhance neural regeneration as well as emerging concepts within the field such as stem cells and cellular reprogramming. Stem cells are defined by their ability to self-renew as well as their ability to differentiate into multiple cell types, and hence can serve as a source for cell replacement of damaged neurons. Traditionally, adult stem cells isolated from the hippocampus and subventricular zone have served as a source of neural stem cells for replacement purposes. With the advancement of pluripotent stem cells, including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (iPSCs), new and exciting approaches for neural cell replacement are being developed. Furthermore, with increased understanding of the human genome and epigenetics, scientists have been successful in the direct genetic reprogramming of somatic cells to a neuronal fate, bypassing the intermediary pluripotent stage. Such breakthroughs have accelerated the timing of production of mature neuronal cell types from a patient-specific somatic cell source such as skin fibroblasts or mononuclear blood cells. While extensive hurdles remain to the translational application of such stem cell and reprogramming strategies, these approaches have revolutionized the field of regenerative biology and have provided innovative approaches for the potential regeneration of the nervous system.
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Affiliation(s)
- Melissa M Steward
- Department of Biology, Indiana University Purdue University, Indianapolis, IN, USA
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Pérez López S, Otero Hernández J. Advances in Stem Cell Therapy. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2012; 741:290-313. [DOI: 10.1007/978-1-4614-2098-9_19] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
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Guillemot F, Zimmer C. From cradle to grave: the multiple roles of fibroblast growth factors in neural development. Neuron 2011; 71:574-88. [PMID: 21867876 DOI: 10.1016/j.neuron.2011.08.002] [Citation(s) in RCA: 175] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 08/04/2011] [Indexed: 01/08/2023]
Abstract
The generation of a functional nervous system involves a multitude of steps that are controlled by just a few families of extracellular signaling molecules. Among these, the fibroblast growth factor (FGF) family is particularly prominent for the remarkable diversity of its functions. FGFs are best known for their roles in the early steps of patterning of the neural primordium and proliferation of neural progenitors. However, other equally important functions have emerged more recently, including in the later steps of neuronal migration, axon navigation, and synaptogenesis. We review here these diverse functions and discuss the mechanisms that account for this unusual range of activities. FGFs are essential components of most protocols devised to generate therapeutically important neuronal populations in vitro or to stimulate neuronal repair in vivo. How FGFs promote the development of the nervous system and maintain its integrity will thus remain an important focus of research in the future.
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Affiliation(s)
- François Guillemot
- Division of Molecular Neurobiology, Medical Research Council, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW71AA, UK.
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35
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Ojeda L, Gao J, Hooten KG, Wang E, Thonhoff JR, Dunn TJ, Gao T, Wu P. Critical role of PI3K/Akt/GSK3β in motoneuron specification from human neural stem cells in response to FGF2 and EGF. PLoS One 2011; 6:e23414. [PMID: 21887250 PMCID: PMC3160859 DOI: 10.1371/journal.pone.0023414] [Citation(s) in RCA: 61] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2011] [Accepted: 07/16/2011] [Indexed: 12/17/2022] Open
Abstract
Fibroblast growth factor (FGF) and epidermal growth factor (EGF) are critical for the development of the nervous system. We previously discovered that FGF2 and EGF had opposite effects on motor neuron differentiation from human fetal neural stem cells (hNSCs), but the underlying mechanisms remain unclear. Here, we show that FGF2 and EGF differentially affect the temporal patterns of Akt and glycogen synthase kinase 3 beta (GSK3β) activation. High levels of phosphatidylinositol 3-kinase (PI3K)/Akt activation accompanied with GSK3β inactivation result in reduction of the motor neuron transcription factor HB9. Inhibition of PI3K/Akt by chemical inhibitors or RNA interference or overexpression of a constitutively active form of GSK3β enhances HB9 expression. Consequently, PI3K inhibition increases hNSCs differentiation into HB9+/microtubule-associated protein 2 (MAP2)+ motor neurons in vitro. More importantly, blocking PI3K not only enhances motor neuron differentiation from hNSCs grafted into the ventral horn of adult rat spinal cords, but also permits ectopic generation of motor neurons in the dorsal horn by overriding environmental influences. Our data suggest that FGF2 and EGF affect the motor neuron fate decision in hNSCs differently through a fine tuning of the PI3K/AKT/GSK3β pathway, and that manipulation of this pathway can enhance motor neuron generation.
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Affiliation(s)
- Luis Ojeda
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
| | - Junling Gao
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
| | - Kristopher G. Hooten
- Department of Neurosurgery, University of Florida, Gainesville, Florida, United States of America
| | - Enyin Wang
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
- West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu, Sichuan, China
| | - Jason R. Thonhoff
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
| | - Tiffany J. Dunn
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
| | - Tianyan Gao
- Markey Cancer Center and Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky, United States of America
| | - Ping Wu
- Department of Neuroscience and Cell Biology, University Of Texas Medical Branch at Galveston, Galveston, Texas, United States of America
- * E-mail:
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Thonhoff JR, Gao J, Dunn TJ, Ojeda L, Wu P. Mutant SOD1 microglia-generated nitroxidative stress promotes toxicity to human fetal neural stem cell-derived motor neurons through direct damage and noxious interactions with astrocytes. AMERICAN JOURNAL OF STEM CELLS 2011; 1:2-21. [PMID: 23671793 PMCID: PMC3643388] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Received: 08/10/2011] [Accepted: 08/18/2011] [Indexed: 06/02/2023]
Abstract
Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease. Human neural stem cells (hNSCs) may have the potential to replace lost motor neurons. The therapeutic efficacy of stem cell therapy depends greatly on the survival of grafted stem cell-derived motor neurons in the microenvironment of the spinal cord in ALS. After transplantation of hNSCs into the spinal cords of transgenic ALS rats, morphological analysis reveals that grafted hNSCs differentiate into motor neurons. However, hNSCs degenerate and show signs of nitroxidative damage at the disease end-stage. Using an in vitro coculture system, we systematically assess interactions between microglia and astroglia derived from both nontransgenic rats and transgenic rats expressing human mutant SOD1(G93A) before and after symptomatic disease onset, and determine the effects of such microglia-astroglia interactions on the survival of hNSC-derived motor neurons. We found that ALS microglia, specifically isolated after symptomatic disease onset, are directly toxic to hNSC-derived motor neurons. Furthermore, nontransgenic astrocytes not only lose their protective role in hNSC-derived motor neuron survival in vitro, but also exhibit toxic features when cocultured with mutant SOD1(G93A) microglia. Using inhibitors of inducible nitric oxide synthase and NADPH oxidase, we show that microglia-generated nitric oxide and superoxide partially contribute to motor neuron loss and astrocyte dysfunction in this coculture paradigm. In summary, reactive oxygen/nitrogen species released from overactivated microglia in ALS directly eliminate human neural stem cell-derived motor neurons and reduce the neuroprotective capacities of astrocytes.
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Affiliation(s)
- Jason R Thonhoff
- Department of Neuroscience and Cell Biology, University of Texas Medical BranchGalveston, Texas 77555, USA
| | - Junling Gao
- Department of Neuroscience and Cell Biology, University of Texas Medical BranchGalveston, Texas 77555, USA
| | - Tiffany J Dunn
- Department of Neuroscience and Cell Biology, University of Texas Medical BranchGalveston, Texas 77555, USA
| | - Luis Ojeda
- Department of Biochemistry and Molecular Biology, University of Texas Medical BranchGalveston, Texas 77555, USA
| | - Ping Wu
- Department of Neuroscience and Cell Biology, University of Texas Medical BranchGalveston, Texas 77555, USA
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Lunn JS, Sakowski SA, Federici T, Glass JD, Boulis NM, Feldman EL. Stem cell technology for the study and treatment of motor neuron diseases. Regen Med 2011; 6:201-13. [PMID: 21391854 DOI: 10.2217/rme.11.6] [Citation(s) in RCA: 56] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Amyotrophic lateral sclerosis and spinal muscular atrophy are devastating neurodegenerative diseases that lead to the specific loss of motor neurons. Recently, stem cell technologies have been developed for the investigation and treatment of both diseases. Here we discuss the different stem cells currently being studied for mechanistic discovery and therapeutic development, including embryonic, adult and induced pluripotent stem cells. We also present supporting evidence for the utilization of stem cell technology in the treatment of amyotrophic lateral sclerosis and spinal muscular atrophy, and describe key issues that must be considered for the transition of stem cell therapies for motor neuron diseases from bench to bedside. Finally, we discuss the first-in-human Phase I trial currently underway examining the safety and feasibility of intraspinal stem cell injections in amyotrophic lateral sclerosis patients as a foundation for translating stem cell therapies for various neurological diseases.
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Affiliation(s)
- J Simon Lunn
- University of Michigan Department of Neurology, 109 Zina Pitcher Place, Ann Arbor, MI 48109, USA
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Ernst N, Tiede S, Tronnier V, Kruse C, Zechel C, Paus R. An improved, standardised protocol for the isolation, enrichment and targeted neural differentiation of Nestin+ progenitors from adult human dermis. Exp Dermatol 2010; 19:549-55. [DOI: 10.1111/j.1600-0625.2009.01041.x] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/21/2023]
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Lindvall O, Kokaia Z. Stem cells in human neurodegenerative disorders--time for clinical translation? J Clin Invest 2010; 120:29-40. [PMID: 20051634 PMCID: PMC2798697 DOI: 10.1172/jci40543] [Citation(s) in RCA: 462] [Impact Index Per Article: 30.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Stem cell-based approaches have received much hype as potential treatments for neurodegenerative disorders. Indeed, transplantation of stem cells or their derivatives in animal models of neurodegenerative diseases can improve function by replacing the lost neurons and glial cells and by mediating remyelination, trophic actions, and modulation of inflammation. Endogenous neural stem cells are also potential therapeutic targets because they produce neurons and glial cells in response to injury and could be affected by the degenerative process. As we discuss here, however, significant hurdles remain before these findings can be responsibly translated to novel therapies. In particular, we need to better understand the mechanisms of action of stem cells after transplantation and learn how to control stem cell proliferation, survival, migration, and differentiation in the pathological environment.
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Affiliation(s)
- Olle Lindvall
- Address correspondence to: Olle Lindvall, Laboratory of Neurogenesis and Cell Therapy, Wallenberg Neuroscience Center, University Hospital, SE-221 84, Lund, Sweden. Phone: 46-46-222-0543; Fax: 46-46-222-0560; E-mail:
| | - Zaal Kokaia
- Address correspondence to: Olle Lindvall, Laboratory of Neurogenesis and Cell Therapy, Wallenberg Neuroscience Center, University Hospital, SE-221 84, Lund, Sweden. Phone: 46-46-222-0543; Fax: 46-46-222-0560; E-mail:
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Seo MS, Jeong YH, Park JR, Park SB, Rho KH, Kim HS, Yu KR, Lee SH, Jung JW, Lee YS, Kang KS. Isolation and characterization of canine umbilical cord blood-derived mesenchymal stem cells. J Vet Sci 2009; 10:181-7. [PMID: 19687617 PMCID: PMC2801133 DOI: 10.4142/jvs.2009.10.3.181] [Citation(s) in RCA: 84] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Human umbilical cord blood-derived mesenchymal stem cells (MSCs) are known to possess the potential for multiple differentiations abilities in vitro and in vivo. In canine system, studying stem cell therapy is important, but so far, stem cells from canine were not identified and characterized. In this study, we successfully isolated and characterized MSCs from the canine umbilical cord and its fetal blood. Canine MSCs (cMSCs) were grown in medium containing low glucose DMEM with 20% FBS. The cMSCs have stem cells expression patterns which are concerned with MSCs surface markers by fluorescence- activated cell sorter analysis. The cMSCs had multipotent abilities. In the neuronal differentiation study, the cMSCs expressed the neuronal markers glial fibrillary acidic protein (GFAP), neuronal class III beta tubulin (Tuj-1), neurofilament M (NF160) in the basal culture media. After neuronal differentiation, the cMSCs expressed the neuronal markers Nestin, GFAP, Tuj-1, microtubule-associated protein 2, NF160. In the osteogenic & chondrogenic differentiation studies, cMSCs were stained with alizarin red and toluidine blue staining, respectively. With osteogenic differentiation, the cMSCs presented osteoblastic differentiation genes by RT-PCR. This finding also suggests that cMSCs might have the ability to differentiate multipotentially. It was concluded that isolated MSCs from canine cord blood have multipotential differentiation abilities. Therefore, it is suggested that cMSCs may represent a be a good model system for stem cell biology and could be useful as a therapeutic modality for canine incurable or intractable diseases, including spinal cord injuries in future regenerative medicine studies.
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Affiliation(s)
- Min-Soo Seo
- Adult Stem Cell Research Center, Department of Veterinary Public Health, College of Veterinery Medicine, Seoul National University, Seoul 151-742, Korea
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