Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord

Mesenchymal stem cells (MSC) are multipotent stem cells that possess the ability to self-renew, capable of differentiating into mesodermal lineage and exert an immunomodulatory activity. These qualities grant MSC great potential in regenerative medicine, immunotherapy and gene therapy. The increasin...

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Main Author:	Tong, Chih Kong
Format:	Thesis
Language:	English English
Published:	2010
Subjects:	Mesenchymal Stem Cells - cytology
Online Access:	http://psasir.upm.edu.my/id/eprint/21036/1/FPSK%28m%29_2010_9_IR.pdf
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spelling	my-upm-ir.210362013-05-27T08:14:35Z Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord 2010-01 Tong, Chih Kong Mesenchymal stem cells (MSC) are multipotent stem cells that possess the ability to self-renew, capable of differentiating into mesodermal lineage and exert an immunomodulatory activity. These qualities grant MSC great potential in regenerative medicine, immunotherapy and gene therapy. The increasing demand of tissue regeneration and allogenic transplantation necessitate the need for a readily available source of MSC as an ‘off-the-shelf’ product for quick and effective treatment. To date, MSC can be generated from human umbilical cord which was once considered clinical waste. The aim of this study is to establish an optimal method to generate and characterise MSC from human umbilical cord samples. In this study, a novel generation method by combining mild enzymatic digestion and mechanical dissociation is established. Briefly, the outer layer of umbilical cord is separated from the blood vessels and remnant cord blood prior to processing. The umbilical cord samples are subjected to explants culture, enzymatic degradation or a combination of enzymatic and mechanical dissociation to obtain a single cell suspension for the generation of umbilical cord MSC (UC-MSC). Once UC-MSC expand, they are characterised by immunophenotyping and differentiation assays. The data show that UC-MSC express common MSC surface markers (CD105+, CD73+, CD29+, CD90+, CD34-, CD45-, MHC class I+, MHC class II-, CD80-,CD86-) and are capable of differentiating into osteoblasts and adipocytes. Simultaneously, the properties of UC-MSC are evaluated in the presence and absence of basic fibroblast growth factor (bFGF). The results show that bFGF supplementation significantly affects the morphology, growth kinetics, cell cycle and cellular functions of UC-MSC. Furthermore, bFGF enhances the growth rate of UCMSC by reducing the doubling time (about 3-4 fold) and skews the cytokine secretion profile (decreases MMP3 and VEGF production). Similar to bone marrow MSC, UC-MSC also exert an immunomodulatory effect on T cells. In the presence of UC-MSC, T cell activation is preserved. However their proliferation is profoundly inhibited in a dose-dependent manner. T cells are arrested anergy in G0 phase of the cell cycle and this inhibitory activity requires cell-to-cell contact. This study reveals that UC-MSC share similar characteristics with BM-MSC and potentially serve as a future source of MSC for clinical use. Mesenchymal Stem Cells - cytology 2010-01 Thesis http://psasir.upm.edu.my/id/eprint/21036/ http://psasir.upm.edu.my/id/eprint/21036/1/FPSK%28m%29_2010_9_IR.pdf application/pdf en public masters Universiti Putra Malaysia Mesenchymal Stem Cells - cytology Faculty of Medicine and Health Sciences English
institution	Universiti Putra Malaysia
collection	PSAS Institutional Repository
language	English English
topic	Mesenchymal Stem Cells - cytology
spellingShingle	Mesenchymal Stem Cells - cytology Tong, Chih Kong Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
description	Mesenchymal stem cells (MSC) are multipotent stem cells that possess the ability to self-renew, capable of differentiating into mesodermal lineage and exert an immunomodulatory activity. These qualities grant MSC great potential in regenerative medicine, immunotherapy and gene therapy. The increasing demand of tissue regeneration and allogenic transplantation necessitate the need for a readily available source of MSC as an ‘off-the-shelf’ product for quick and effective treatment. To date, MSC can be generated from human umbilical cord which was once considered clinical waste. The aim of this study is to establish an optimal method to generate and characterise MSC from human umbilical cord samples. In this study, a novel generation method by combining mild enzymatic digestion and mechanical dissociation is established. Briefly, the outer layer of umbilical cord is separated from the blood vessels and remnant cord blood prior to processing. The umbilical cord samples are subjected to explants culture, enzymatic degradation or a combination of enzymatic and mechanical dissociation to obtain a single cell suspension for the generation of umbilical cord MSC (UC-MSC). Once UC-MSC expand, they are characterised by immunophenotyping and differentiation assays. The data show that UC-MSC express common MSC surface markers (CD105+, CD73+, CD29+, CD90+, CD34-, CD45-, MHC class I+, MHC class II-, CD80-,CD86-) and are capable of differentiating into osteoblasts and adipocytes. Simultaneously, the properties of UC-MSC are evaluated in the presence and absence of basic fibroblast growth factor (bFGF). The results show that bFGF supplementation significantly affects the morphology, growth kinetics, cell cycle and cellular functions of UC-MSC. Furthermore, bFGF enhances the growth rate of UCMSC by reducing the doubling time (about 3-4 fold) and skews the cytokine secretion profile (decreases MMP3 and VEGF production). Similar to bone marrow MSC, UC-MSC also exert an immunomodulatory effect on T cells. In the presence of UC-MSC, T cell activation is preserved. However their proliferation is profoundly inhibited in a dose-dependent manner. T cells are arrested anergy in G0 phase of the cell cycle and this inhibitory activity requires cell-to-cell contact. This study reveals that UC-MSC share similar characteristics with BM-MSC and potentially serve as a future source of MSC for clinical use.
format	Thesis
qualification_level	Master's degree
author	Tong, Chih Kong
author_facet	Tong, Chih Kong
author_sort	Tong, Chih Kong
title	Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
title_short	Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
title_full	Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
title_fullStr	Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
title_full_unstemmed	Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord
title_sort	generation and characterisation of human mesenchymal stem cells from human umbilical cord
granting_institution	Universiti Putra Malaysia
granting_department	Faculty of Medicine and Health Sciences
publishDate	2010
url	http://psasir.upm.edu.my/id/eprint/21036/1/FPSK%28m%29_2010_9_IR.pdf
_version_	1747811480278925312

Generation and Characterisation of Human Mesenchymal Stem Cells from Human Umbilical Cord

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