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Microporation is a valuable transfection method for gene expression in human adipose tissue-derived stem cells.

ABSTRACT: Stem cells are a promising resource for gene therapy. Adipose tissue-derived stem cells (ADSCs) offer advantages because of their abundance and ease of isolation. However, it is difficult to transduce genes into ADSCs by common transfection methods, especially nonviral methods. We report here the use of a new electroporation method, termed "microporation," to transduce plasmids into human ADSCs (hADSCs). We determined optimal conditions that led to efficient transfection of >76.1% of the microporated hADSCs with only minimal cell damage or cytotoxicity. We demonstrated the expression of both enhanced green fluorescent protein (EGFP) and luciferase from different promoters in microporated hADSCs. More important, the microporated hADSCs retained their multipotency and reporter gene expression was maintained for >2 weeks in vitro and in vivo. We further showed that a Tet-ON-inducible gene expression system could be microporated into hADSCs and that this system was functional following transplantation of the microporated cells into nude mice. Taken together, our data demonstrate that microporation allows a highly efficient transfection of hADSCs, without impairing their stem cell properties.

SUBMITTER: Wang YH

PROVIDER: S-EPMC2835049 | biostudies-literature | 2009 Feb

REPOSITORIES: biostudies-literature

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Microporation is a valuable transfection method for gene expression in human adipose tissue-derived stem cells.

Wang Yan-Hsiung YH Ho Mei-Ling ML Chang Je-Ken JK Chu Hai-Chang HC Lai Shu-Chin SC Wang Gwo-Jaw GJ

Molecular therapy : the journal of the American Society of Gene Therapy 20081209 2

Stem cells are a promising resource for gene therapy. Adipose tissue-derived stem cells (ADSCs) offer advantages because of their abundance and ease of isolation. However, it is difficult to transduce genes into ADSCs by common transfection methods, especially nonviral methods. We report here the use of a new electroporation method, termed "microporation," to transduce plasmids into human ADSCs (hADSCs). We determined optimal conditions that led to efficient transfection of >76.1% of the micropo ...[more]

PMID: 19066595

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Project description:Differences in the inherent properties of adipose tissue-derived stem cells (ASC) may contribute to the biological specificity of the subcutaneous (Sc) and visceral (V) adipose tissue depots. In this study, three distinct subpopulations of ASC, i.e. ASCSVF, ASCBottom, and ASCCeiling, were isolated from Sc and V fat biopsies of non-obese subjects, and their gene expression and functional characteristics were investigated. Genome-wide mRNA expression profiles of ASCSVF, ASCBottom and ASCCeiling from Sc fat were significantly different as compared to their homologous subsets of V-ASCs. Furthermore, ASCSVF, ASCCeiling and ASCBottom from the same fat depot were also distinct from each other. In this respect, both principal component analysis and hierarchical clusters analysis showed that ASCCeiling and ASCSVF shared a similar pattern of closely related genes, which was highly different when compared to that of ASCBottom. However, larger variations in gene expression were found in inter-depot than in intra-depot comparisons. The analysis of connectivity of genes differently expressed in each ASC subset demonstrated that, although there was some overlap, there was also a clear distinction between each Sc-ASC and their corresponding V-ASC subsets, and among ASCSVF, ASCBottom, and ASCCeiling of Sc or V fat depots in regard to networks associated with regulation of cell cycle, cell organization and development, inflammation and metabolic responses. Finally, the release of several cytokines and growth factors in the ASC cultured medium also showed both inter- and intra-depot differences. Thus, ASCCeiling and ASCBottom can be identified as two genetically and functionally heterogeneous ASC populations in addition to the ASCSVF, with ASCBottom showing the highest degree of unmatched gene expression. On the other hand, inter-depot seem to prevail over intra-depot differences in the ASC gene expression assets and network functions, contributing to the high degree of specificity of Sc and V adipose tissue in humans.

Dataset Information

Microporation is a valuable transfection method for gene expression in human adipose tissue-derived stem cells.

Publications

Microporation is a valuable transfection method for gene expression in human adipose tissue-derived stem cells.

Similar Datasets

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