摘要
背景:缺血缺氧的心肌微环境导致植入的细胞存活率低。目的:观察沉默caspase-3基因对大鼠骨髓间充质干细胞增殖和体外缺血缺氧环境下凋亡的影响。方法:构建靶向caspase-3的shRNA重组慢病毒并转染骨髓间充质干细胞为转基因组,以正常细胞组和空载体组做对照,采用MTS法检测各组细胞增殖情况。建立缺血缺氧模型,real-time PCR和免疫组织化学分别检测缺血缺氧环境各组细胞的caspase-3mRNA和蛋白表达水平,应用流式细胞术检测不同缺血缺氧时间点(0,6,12,24,48h)各组细胞的凋亡率。结果与结论:重组慢病毒成功转染骨髓间充质干细胞,且细胞增殖活性升高(P<0.05)。缺血缺氧环境下,转基因组细胞caspase-3在mRNA和蛋白表达水平相比对照组下降(P<0.05)。沉默caspase-3能显著降低骨髓间充质干细胞的凋亡率(P<0.05),且随着缺血缺氧时间的延长凋亡率缓慢升高。结果提示,沉默caspase-3能加快骨髓间充质干细胞的生长速度和提高在体外缺血缺氧环境下的抗凋亡能力。
BACKGROUND： Ischemia and hypoxia myocardial microenvironment leads to poor survival of transplanted cells. OBJECTIVE： To investigate the effects of silencing caspase-3 gene on proliferation and apoptosis of rat bone marrow mesenchymal stem cells under ischemia and hypoxia in vitro. METHODS： Lentiviral short hairpin RNA interference vector targeting caspase-3 was constructed and transfected into mesenchymal stem cells as the transgene group. The normal cell group and the empty vector group were as controls. MTS assay was applied to examine the proliferation of cells in each group. The ischemia and hypoxia model was established. The expressions of caspase-3 mRNA and protein were detected by real-time PCR and immunohistochemistry. The apoptotic rates of the cells at different time points （0, 6, 12, 24 and 48 hours） in each group were evaluated by flow cytometry. RESULTS AND CONCLUSION： Recombinant lentivirus was transfected into mesenchymal stern cells successfully and the proliferation activity of the cells was increased （P 〈 0.05）. Compared with control groups, the levels of caspase-3 mRNA and protein in the transgene group were decreased （P 〈 0.05） under ischemia and hypoxia. Silencing caspase-3 could reduce the apoptotic rate of mesenchymal stem cells （P 〈 0.05）, and the apoptotic rate was increased slowly as ischemia and hypoxia time prolonging. Silencing caspase-3 can increase the growth speed and the anti-apoptosis ability of mesenchymal stem cells under ischemia and hypoxia in vitro.
出处
《中国组织工程研究》
CAS
CSCD
2013年第14期2480-2487,共8页
Chinese Journal of Tissue Engineering Research
基金
广东省自然科学基金资助项目(7001619)
广东省科技计划项目(2007B050200011)~~
关键词
干细胞
骨髓干细胞
骨髓间充质干细胞
基因沉默
CASPASE-3
基因
细胞增殖
凋亡
慢病毒
缺血缺氧
shRNA
省级基金
干细胞图片文章
stem cells
bone marrow-derived stem cells
bone marrow mesenchymal stem cells
gene silencingCaspase-3
gene
cell proliferation
apoptosis
lentivirus
ischemia and hypoxia
short hairpin RNA
provincialgrants-supported paper
stem cell photographs-containing paper
