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目的探讨过表达整合素连接激酶(ILK)与胶质瘤细胞对替莫唑胺(TMZ)敏感性的影响。方法本研究前期已经构建了重组质粒pEGFP-C1-ILK,并将其转染给SHG-44胶质瘤细胞,筛选出了稳定转染的细胞株。本实验分组如下:SHG-44组、pEGFP-C1+DMSO组、pEGFP-C1-ILK+DMSO组、pEGFP-C1+TMZ组、pEGFP-C1-ILK+TMZ组,应用试剂盒法检测凋亡关键酶Caspase-3的活性,并应用Hoechst法和流式细胞术检测各组细胞的凋亡。结果 Caspase-3的活性在pEGFP-C1-ILK+TMZ组明显低于pEGFPC1+TMZ组(P=0.007 8),pEGFP-C1-ILK+DMSO组中Caspase-3活性与pEGFP-C1+DMSO组比较同样明显降低(P=0.008 3),pEGFP-C1+DMSO组与SHG44组Caspase-3活性比较,差异无统计学意义(P=0.845)。Hoechst法结果:SHG44组与pEGFP-C1+DMSO组中的凋亡细胞,差异无统计学意义(P=0.87),而pEGFP-C1-ILK+TMZ组凋亡细胞明显少于pEGFP-C1+TMZ组(P=0.003),pEGFP-C1-ILK+DMSO组与pEGFP-C1+DMSO组比较,凋亡细胞差异无统计学意义(P=0.92)。流式细胞术结果:pEGFP-C1-ILK+TMZ组与pEGFP-C1+TMZ组比较,早期凋亡细胞百分比明显减少(P=0.001),pEGFP-C1+ILK+DMSO组与pEGFP-C1+DMSO组比较,早期凋亡细胞百分比差异有统计学意义(P=0.005 2),pEGFP-C1+DMSO组与SHG44组早期凋亡细胞百分比相比较,差异无统计学意义(P=0.74)。结论过表达ILK通过降低Caspase-3的酶活性减少胶质瘤细胞的凋亡,从而降低胶质瘤细胞对TMZ的敏感性。
Objective To investigate the effect of overexpressing integrin-linked kinase (ILK) and glioma cells on the sensitivity of temozolomide (TMZ). Methods The recombinant plasmid pEGFP-C1-ILK was constructed in the early stage of this study and transfected into SHG-44 glioma cells. Stably transfected cell lines were screened out. The experiment was divided into the following groups: SHG-44 group, pEGFP-C1 + DMSO group, pEGFP-C1-ILK + DMSO group, pEGFP-C1 + TMZ group and pEGFP-C1-ILK + TMZ group The activity of Caspase-3 was detected by flow cytometry. Hoechst method and flow cytometry were used to detect the apoptosis of each group. Results The activity of Caspase-3 in pEGFP-C1-ILK + TMZ group was significantly lower than that in pEGFPC1 + TMZ group (P = 0.007 8). Caspase-3 activity in pEGFP-C1-ILK + (P = 0.008 3). There was no significant difference in Caspase-3 activity between pEGFP-C1 + DMSO group and SHG44 group (P = 0.845). Hoechst method: There was no significant difference in apoptotic cells between SHG44 group and pEGFP-C1 + DMSO group (P = 0.87), while apoptotic cells in pEGFP-C1-ILK + TMZ group were significantly less than those in pEGFP-C1 + (P = 0.003). There was no significant difference in apoptotic cells between pEGFP-C1-ILK + DMSO group and pEGFP-C1 + DMSO group (P = 0.92). Flow cytometry showed that the percentage of early apoptotic cells was significantly decreased in pEGFP-C1-ILK + TMZ group compared with pEGFP-C1 + TMZ group (P = 0.001) (P = 0.005 2). There was no significant difference in the percentages of apoptotic cells between pEGFP-C1 + DMSO group and SHG44 group (P = 0.74) .Conclusion: 1. Conclusion Overexpression of ILK decreased the apoptosis of glioma cells by decreasing the activity of Caspase-3, thereby reducing the sensitivity of glioma cells to TMZ.