CAFs调控MAPK/ERK5通路抑制结直肠癌HT-29细胞凋亡

CAFs Inhibit Apoptosis of Colorectal Cancer HT-29 Cells via MAPK/ERK5 Pathway

为探讨肿瘤相关成纤维细胞(CAFs)对结直肠癌细胞增殖和凋亡的影响及分子机制,本研究通过组织贴壁法从结直肠癌组织中分离CAFs,并验证CAFs中α-SMA的表达;通过Transwell建立CAFs和结直肠癌细胞系HT-29共培养系统,CCK8检测结直肠癌HT-29细胞活力;流式细胞术检测HT-29细胞凋亡率;通过实时荧光定量PCR和Western blotting检测结直肠癌细胞中VEGF的mRNA和蛋白表达以及ERK5磷酸化水平。与对照NFs相比,α-SMA在结CAFs中表达显著增加(p<0.01)。CCK8结果表明CAFs促进结直肠癌细胞的增殖;流式结果显示CAFs能抑制细胞凋亡;Real-time RT-PCR和Western blotting结果显示CAFs促进结直肠癌细胞内VEGF的mRNA和蛋白表达,并促进ERK5磷酸化。本研究初步表明,CAFs激活MAPK/ERK5通路和VEGF的表达,可促进结直肠癌HT-29细胞增殖。

To investigate the effect of carcinoma associated fibroblasts(CAFs)on the proliferation and apoptosis of colorectal cancer cells and its molecular mechanism.CAFs were isolated from colorectal cancer tissues by tissue adherence method,and the expression ofα-SMA in CAFs was verified;CAFs and colorectal cancer cell line HT-29 co-culture system were established by transwell.The proliferation of the HT-29 cells was assessed using CCK-8 assay.The apoptosis of HT-29 cells was analyzed by flow cytometry.The m RNA and protein expressions of VEGF were measured by real-time RT-PCR and western blotting.The phosphorylation of ERK5 was measured by western blotting.Compared with the NFs,CAFs expressed significantly higher levels ofα-SMA(p<0.01).CCK8 results indicated that CAFs promote the proliferation of colorectal cancer cells.Flow cytometry results showed that CAFs can inhibit the apoptosis of HT-29 cells.Real-time RT-PCR and Western blot showed that CAFs promoted the expression of VEGF m RNA and protein in colorectal cancer cells.The results of western blot showed that CAFs could promote the phosphorylation of ERK5.CAFs promote proliferation of colorectal cancer HT-29 cells by activating MAPK/ERK5 pathway and VEGF expression.