Expression of lentiviral vectors mediated NEP1-40 in mesenchymal stem cells fromhuman umbilical cord blood

doi:10.3969/j.issn.2095-4344.2013.01.005

Abstract

Abstract:

BACKGROUND: In recent years, studies have shown that mesenchymal stem cells are ideal seed cells used fortissue engineering because of their strong proliferation and multi-differentiation potential. Mesenchymal stem cells can be efficiently transfected and expressed exogenous gene, and therefore they have broad application prospects in gene therapy.
OBJECTIVE: NEP1-40 gene-containing lentiviral vectors were transfected into umbilical cord blood-derived mesenchymal stem cells to evaluate the biological function changes of mesenchymal stem cells and detect NEP1-40 expression in umbilical cord blood-derived mesenchymal stem cells.
METHODS: Human umbilical cord blood-derived mesenchymal stem cells were isolated and cultured in vitro. Cell surface markers were detected by flow cytometry, and their biological characteristics were identified. NEP1-40 gene was cloned into the lentiviral vector and lentiviral supernatant was packaged. Then umbilical cord blood-derived mesenchymal stem cells were transfected with different multiplicities of infection.
RESULTS AND CONCLUSIONS: We successfully isolated and cultured human umbilical cord blood-derived mesenchymal stem cells in vitro by density gradient centrifugation method, and the cells could be induced to differentiate into adipocytes. Flow cytometry results showed that umbilical cord blood-derived mesenchymal stem cells were positive for CD90, CD73 and CD105 protein, but they were negative for CD14, CD34, CD45, CD19, HLA-DR, Stro-1 and CD106 protein. Real-time PCR detection showed that the NEP1-40 mRNA expression level was positively correlated with multiplicity of infection. Higher multiplicity of infection yielded higher NEP1-40 expression. In addition, NEP1-40 protein expression could be seen in human umbilical cord blood-derived mesenchymal stem cells after transfected with NEP1-40. These findings suggest that the transfection of NEP1-40 gene has little impact on biological function of human umbilical cord blood-derived mesenchymal stem cells.

Key words: stem cells, umbilical cord blood-derived stem cells, cord blood, mesenchymal stem cells, lentiviral vectors, transfection, NEP1-40, 863 program, stem cell photographs-containing paper

CLC Number:

R394.2

Huang Ai-hua, Zhou Yi-dan, Zhang Ping-ping, Wang Sai-ying, Lin Yuan-yuan, Ge Ting-ai, Chen Zan. Expression of lentiviral vectors mediated NEP1-40 in mesenchymal stem cells fromhuman umbilical cord blood[J]. Chinese Journal of Tissue Engineering Research, 2013, 17(1): 31-37.

Figures/Tables 7

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