中文摘要：

目的：研究caspase-8小发夹RNA（Cap8 shRNA）对人骨髓间充质干细胞（hMSCs）凋亡的调控作用。方法：构建2个靶向人caspase-8基因的穿梭质粒pAd-Cap8 shRNA，并鉴定可有效抑制人HEK293细胞中caspase-8表达的pAd-Cap8 shRNA质粒。构建表达Cap8 shRNA的重组腺病毒质粒，并在HEK293细胞中包装、扩增重组腺病毒rAd-Cap8 shRNA。检测rAd-Cap8 shRNA感染的hMSCs中caspase-8 mRNA和蛋白表达。利用anne-xin V/PI双染色法和caspase-8活性检测分析去血清/缺氧诱导的hMSCs凋亡变化，利用荧光定量PCR检测hMSCs中VEGF、肝细胞生长因子1（HGF）、胰岛素样生长因子（IGF-1）、Bcl-2和Bcl-xL mRNA表达。结果：通过定量PCR筛选到可有效抑制caspase-8表达的pAd-Cap8 shRNA质粒。成功构建Cap8 shRNA的重组腺病毒质粒，并包装、扩增重组腺病毒rAd-Cap8 shRNA。荧光定量PCR和Western blotting结果证实rAd-Cap8 shRNA可有效抑制hMSCs中caspase-8表达。 rAd-Cap8 shRNA能有效抑制去血清/缺氧诱导的hMSCs凋亡，降低caspase-8活性，增强hMSCs中HGF、IGF-1和Bcl-2表达。结论：Caspase-8 shRNA可以抑制去血清/缺氧诱导的hMSCs凋亡。

英文摘要：

AIM：To investigate the effect of caspase-8 small hairpin RNA （ shRNA） on attenuating apoptosis of human mesenchymal stem cells （ hMSCs ） .METHODS： Two recombinant plasmids for over-expression of caspase-8 shRNA, pAd-Cap8 shRNA1 and pAd-Cap8 shRNA2, were constructed.Caspase-8 mRNA was determined in pAd-Cap8 shRNA-transfected human HEK293 cells by Q-PCR.The screened pAd-Cap8 shRNA was used to construct the recombinant adenovirus plasmid , which was linearized and transfected into HEK 293 cells for packaging and amplification of the recombi-nant adenovirus rAd-Cap8 shRNA.The expression of caspase-8 at mRNA and protein levels was determined by Q-PCR and Western blotting .Annexin V/PI staining and determination of caspase-8 activity were performed to assess apoptosis of hM-SCs under the conditions of serum deprivation and hypoxia .The mRNA expression of vascular endothelial growth factor （VEGF）, hepatocyte growth factor （HGF）, insulin-like growth factor 1 （IGF-1）, Bcl-2 and Bcl-xL was analyzed by Q-PCR.RESULTS：The pAd-Cap8 shRNA, which efficiently inhibited caspase-8 expression, was screened by Q-PCR.The recombinant adenovirus plasmid for caspase-8 shRNA was constructed and used to package and amplify the recombinant ad-enovirus （ rAd）-Cap8 shRNA successfully .rAd-Cap8 shRNA-mediated caspase-8 shRNA markedly inhibited caspase-8 ex-pression in hMSCs .Over-expression of caspase-8 shRNA by infection of rAd-Cap8 shRNA also efficiently decreased the ap-optotic rate and caspase-8 activity in hMSCs under the conditions of serum deprivation and hypoxia , with up-regulation of the mRNA expression of HGF, IGF-1 and Bcl-2.CONCLUSION：Caspase-8 shRNA attenuates hMSC apoptosis under the conditions of serum deprivation and hypoxia .