论文部分内容阅读
目的通过构建基因真核表达载体,探讨人2型大麻素受体(h CB2R)对人宫颈癌He La细胞体外凋亡的作用及机制。方法构建GV230-h CB2R质粒,经双酶切、测序鉴定后,转染He La细胞,Western blot法及免疫荧光细胞化学染色联合激光扫描共聚焦显微镜技术检测h CB2R表达及细胞定位;流式细胞术检测细胞凋亡率,Western blot法及实时荧光定量PCR检测h CB2R、Bcl-2、Bax、Bad的表达。结果 h CB2R转染He La细胞后表达相对分子质量(Mr)40 000的h CB2R蛋白,细胞膜和细胞质中均有h CB2R表达;过表达的h CB2R,可上调Bax、Bad的表达,抑制Bcl-2的表达,流式细胞术检测结果显示细胞株呈现凋亡,h CB2R对宫颈癌He La细胞株的生长表现出明显的抑制作用,促进宫颈癌He La细胞凋亡。结论上调He La细胞h CB2R表达可增强Bax、Bad表达,抑制Bcl-2表达,诱导细胞凋亡。
OBJECTIVE: To investigate the effect and mechanism of human CB-2R on human cervical cancer HeLa cells in vitro by constructing eukaryotic expression vector. Methods GV230-h CB2R plasmid was constructed and identified by restriction enzyme digestion and sequencing. HeLa cells were transfected into HeLa cells. Western blot and immunofluorescence staining combined with laser scanning confocal microscopy were used to detect hBB2R expression and cell localization. Flow cytometry The rate of apoptosis, the expression of hBB2R, Bcl-2, Bax and Bad were detected by Western blot and real-time fluorescence quantitative PCR. Results The h CB2R protein expressed by h CB2R transfected He La cells with a relative molecular mass of 40 000 showed h CB2R expression in cell membrane and cytoplasm. Overexpression of h CB2R up-regulated the expression of Bax and Bad and inhibited the expression of Bcl- The results of flow cytometry showed that the cell lines showed apoptosis. H CB2R could inhibit the growth of cervical cancer HeLa cells and promote the apoptosis of cervical cancer HeLa cells. Conclusion Upregulation of h CB2R expression in HeLa cells can enhance the expression of Bax and Bad, inhibit the expression of Bcl-2 and induce apoptosis.