热疗联合人肿瘤坏死因子对TNFR1高表达胶质瘤的细胞周期、F-肌动蛋白及其侵袭性的影响

Effect of Hyperthermia Combined with rhTNF on Cell Cycle,F-actin and Invasiveness to Over-expressed TNFR1 Glioma Cells

目的探讨热疗联合重组人肿瘤坏死因子(recombinant human tumor necrosis factor,rhTNF)对肿瘤坏死因子受体1(tumornecrosis factor receptor 1,TNFR1)高表达的胶质瘤细胞的细胞周期和F-肌动蛋白(F-actin)的影响及其与胶质瘤侵袭性的关系。方法建立TNFR1高表达胶质瘤细胞株,RT-PCR和Western blot法检测胶质瘤细胞TNFR1的表达水平;碘化丙啶染色后用流式细胞术检测胶质瘤细胞细胞周期的变化;WST-8法检测细胞增殖;免疫荧光技术检测胶质瘤细胞内F-actin的表达水平;Transwell小室法检测胶质瘤细胞侵袭性改变。结果与对照组相比,TNFR1高表达胶质瘤细胞株的TNFR1 mRNA水平增加了78.5%,其蛋白质的表达水平增加了89.7%(P<0.05);经热疗联合rhT-NF处理后细胞增殖受抑制,S和G2/M期的TNFR1高表达胶质瘤细胞数之和明显增多,而F-actin的荧光强度和胶质瘤侵袭性分别降低了72.3%和83.10%。结论热疗联合rhTNF可能是通过阻滞TNFR1高表达胶质瘤细胞的细胞周期和降低F-actin的表达来实现降低胶质瘤侵袭性的作用。

Objective The study objective was to investigate the effect of hyperthermia combined with rhTNF on cell cycle and F aetin of TNFR1 in over-expressed glioma, as well as invasiveness in vitro. Methods C6 cell Line of over-expressed TNFR1 （C6/TNFR1） was constructed. , The mRNA and pro- tein of TNFR1 were measured by reverse transcription-polymerase chain reaction （RT-PCR） and Western blot respectively, and the cell cycle and cell proliferation were determined by flow cytometry（stained by propidium iodide） and WST-8 respectively. The invasiveness was measured by transwell assay and immu- nofluorescence technique was used to measure F-actin protein expression. Results Compared with the control group, the mRNA and protein levels of TNFR1 in c6/TNFR1 cell was increased, by 78. 5 % and 89.7% （P〈0. 05）, respectively. The cell proliferation was inhibited and most of e6/TNFR1 cells were arrested in S ＋ G2/M phase compared with the control group cells after hyperthermia combined with rhT- NF treatment （P〈0. 05）. The fluorescence intensity of F-aetin and the average number of C6/TNFR1 cells passing through the inserted filter were decreased by 72. 3% and 83.10〈respectively, compared to the control group cells after hyperthermia combined with rhTNF treatment （P〈0. 01）. Conclusion Hy- perthermia combined with rhTNF might reduce glioma of C6/TNFR1 invasiveness through blocking cell cycle and reducing the expression of F- action.