摘要
【目的】探讨沉默caspase-3对大鼠骨髓间充质干细胞(MSC)在体外缺血缺氧环境下凋亡的影响。【方法】构建靶向caspase-3的shRNA重组慢病毒并转染MSC。建立缺血缺氧模型,流式细胞术和Hoechst荧光染色法检测细胞的凋亡情况。Western blot检测caspase-3的蛋白表达,Real-time PCR检测caspase-3、Bcl-2和Bax的mRNA表达。【结果】重组慢病毒成功转染MSC。缺血缺氧环境下,转染组的细胞凋亡率为(13.66±0.20)%,低于空载体组的(21.86±0.43)%和空白对照组的(22.28±0.48)%(P<0.01)。沉默caspase-3后caspase-3表达下调,Bcl-2表达上调,Bax表达下调,Bcl-2/Bax比值升高(P<0.05)。【结论】沉默caspase-3能显著提高MSC在缺血缺氧环境下的存活能力。
[Objective] To investigate the effect of silencing caspase-3 gene on apoptosis of rat MSC under hypoxia and serum deprivation in vitro.[Methods] Lentiviral shRNA interference vector targeting caspase-3 was constructed and transfected into MSC.Cells were treated with hypoxia and serum deprivation.The apoptosis were evaluated by flow cytometry and Hoechst fluorochrome staining.The expression of caspase-3 protein was detected by Western blot analysis.The expression of caspase-3,Bcl-2 and Bax mRNA were examined by Real-time PCR.[Results] Recombinant lentivirus was transfected into MSC successfully.The apoptotic rate in MSC-shRNA,MSC-vector and MSC were （13.66 ± 0.20）%,（21.86 ± 0.43）% and （22.28 ± 0.48）% （P 〈 0.01）,respectively.Compared with control groups,silencing caspase-3 down-regulated the mRNA level of caspase-3,up-regulated the mRNA level of Bcl-2 and down-regulated the mRNA level of Bax and the ratio of Bcl-2 to Bax increased （P 〈 0.05）.[Conclusions] Silencing caspase-3 improves the ability of survival of MSC under hypoxia and serum deprivation in vitro.
出处
《中山大学学报（医学科学版）》
CAS
CSCD
北大核心
2013年第6期819-824,共6页
Journal of Sun Yat-Sen University:Medical Sciences
基金
广东省科技计划项目(2012B031800258)
中山大学青年教师培育项目(11ykpy30)
