Effect of the human insulin-like growth factor 1 gene transfection to human umbilical cord blood mesenchymal stem cells.
OBJECTIVE
To observe the effect of transfecting the gene human insulin-like growth factor (hIGF)-1 into human umbilical cord blood mesenchymal stem cells (hUCB-MSCs) via non-viral vector.


METHODS
This study was performed in the Affiliated Hospital of Qingdao University, Qingdao, China from June 2012 to May 2013. Twelve hUCB samples were harvested, and isolated in lymphocyte separation medium, and then cultured. Surface antigen expression in MSCs was detected by flow cytometry. Recombinant plasmid pIRES2-enhanced green fluorescent protein (EGFP)-hIGF-1 was transfected into MSCs by X-treme GENE HP DNA transfection reagent. Then, EGFP was observed with reverse fluorescent microscope at different time points. Enzyme-linked immunosorbent assay was used to determine the hIGF-1 protein concentration in supernatants. Immunofluorescence microscopy and reverse transcription polymerase chain reaction were used to detect the expression of hIGF-1 in the hUCB-MSCs. Expression of type II collagen was detected by immunohistochemistry staining.


RESULTS
Transfection efficiency was 28.74 +/- 7.31%. The cluster of differentiation (CD)90, CD105, and CD146 expression increased CD34, CD45, and anti-HLA-DR expression decreased. Results of immunofluorescence microscopy and RT-PCR confirmed expression of the hIGF-1 gene. The hIGF-1 protein concentration in the supernatants showed a peak level at 34.63 +/- 1.61 ng/ml 48 hours after transfection. Immunohistochemical analysis of transfected hUCB-MSCs proved that type II collagen could be expressed positively.


CONCLUSIONS
Human IGF-1 gene can be transfected into hUCB-MSCs, and expressed at a high level with subsequent expression of type II collagen.
Department of Surgery, The Qilu Hospital, Shandong University, Jinan, China.
Zeng-Chi
ZC
Han
Hai-Ning
HN
Zhang
Ying-Zhen
YZ
Wang
Cheng-Yu
CY
Lv
Zong-Yao
ZY
Xu
D001483:Base Sequence; D017931:DNA Primers; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D014162:Transfection; D014470:Umbilical Cord / Q000166:cytology / Q000378:metabolism*
D017931:DNA Primers; D007334:Insulin-Like Growth Factor I