Human mesenchymal stem cells (HMSCs) have emerged as a promising cell type for tissue repopulat-ing and for use as ex vivo gene delivery vehicles. Herpes simplex virus-1 (HSV-1) possesses many vector features, which make it especially suitable for HMSC applications. Here we performed real-time RT-PCR to detect the level of mature microRNA encoded by the HSV-1 latency-associated transcript (microRNA-LAT) in HMSCs cytoplasm with a specific stem loop reverse primer. Three small interfering RNAs (siRNAs) were chemically synthesized towards microRNA-LAT, TGF-β1 and SMAD3. The results demonstrate that HSV-1 and microRNA-LAT prevented HMSCs from undergoing cisplatin-induced apoptosis. In comparison with cells only treated with cisplatin, the apoptosis phenomenon with HSV-1 and microRNA-LAT were markedly reduced. The apoptosis rates of these two groups were both lower (P<0.05) as determined by flow cytometry analysis. The results of RT-PCR and western blotting analysis confirmed that the mRNA and protein levels of TGF-β1 and SMAD3 were significantly decreased with treatment of HSV-1 and microRNA-LAT. Integral TGF-β signalling pathway components were by these means knocked down. Moreover, the levels of the mature microRNA-LAT were decreased in cis-platin-treated HMSCs. In conclusion, HSV-1 and microRNA-LAT exert their anti-apoptotic effect on HMSCs by down-regulation of the TGF-β signalling pathway. Thus HSV-1, having anti-apoptotic effect naturally encoded in its microRNA-LAT, is a good candidate to be developed for HMSC vector.
ZHANG Jing LI YongMei DU WenCai LI GuangMing LI WeiXia ZHU Ze
