摘要
目的研究替莫唑胺对U373胶质瘤细胞增殖和侵袭能力的抑制作用及其分子调控机制。方法将U373胶质瘤细胞分别用0μmol/L、100μmol/L、200μmol/L和400μmol/L替莫唑胺处理,培养48 h。用control siRNA、siPTPN1、pcDNA3.1空载体及pcDNA3.1/PTPN1,分别转染用400μmol/L替莫唑胺处理的细胞。用CCK-8法和Transwell实验检测替莫唑胺对U373细胞增殖、侵袭能力的影响。用Western blot检测各组细胞的PTPN1、C-myc、Cyclin D1、PI3K、p-mTOR和p-AKT蛋白的表达水平。结果与空白对照组相比,各替莫唑胺处理组U373细胞的增殖和侵袭能力均显著下降(P<0.05~0.01),PTPN1的表达水平显著降低(P<0.05~0.01);其中以400μmol/L替莫唑胺处理组细胞的增殖、侵袭能力及PTPN1表达水平最低(均P<0.01)。与pcDNA3.1空载体转染组相比,过表达PTPN1组U373细胞的PTPN1蛋白表达水平,以及增殖和侵袭能力均显著增高(均P<0.01)。与control siRNA转染组相比,siPTPN1转染组U373细胞的PTPN1蛋白表达水平及增殖和侵袭能力均显著下降(均P<0.01)。替莫唑胺处理细胞的C-myc、Cyclin D1、PI3K、p-mTOR和p-AKT表达水平均显著降低(P<0.05~0.01)。PTPN1过表达细胞的C-myc、Cyclin D1、PI3K、p-mTOR和p-AKT表达水平显著升高(均P<0.01);而PTPN1表达沉默细胞的C-myc、Cyclin D1、PI3K、p-mTOR和p-AKT的表达水平则明显降低(均P<0.05)。结论替莫唑胺可通过下调胶质瘤细胞的PTPN1表达影响mTOR信号通路,抑制胶质瘤细胞的增殖和侵袭能力。
Objective To study the inhibitory effect of temozolomide on the proliferation and invasion of glioma cells and its molecular regulation mechanism.Methods U373 glioma cells were treated with 0μmol/L,100μmol/L,200μmol/L and 400μmol/L temozolomide,respectively,then the cells were cultured for 48 h.The cells that treated with 400μmol/L temozolomide were transfected with control siRNA,siPTPN1,pcDNA3.1 empty vector,pcDNA3.1/PTPN1,respectively.The effect of temozolomide on the proliferation and invasion of glioma cells was detected by CCK-8 and transwell assay,respectively.The effects of temozolomide on the expression of PTPN1,C-myc,Cyclin D1 and p-mTOR in glioma cells was detected by Western blot.Results Compared with the control group,the proliferation and invasion ability of U373 cells were significantly decreased after temozolomide treated(P<0.05-0.01),and the expression levels of PTPN1 were significantly decreased(P<0.05-0.01).When the concentration of temozolomide was 400μmol/L,the cell proliferation and invasion ability,and expression level of PTPN1 were the lowest(P<0.01).The expression of PTPN1 protein,the cell proliferation and invasion ability were significantly up-regulated in pcDNA3.1/PTPN1 transfected U373 cells compared with pcDNA3.1 empty vector transfection group(P<0.01).The expression of PTPN1 protein,and cell proliferation and invasion ability in U373 cells of siPTPN1 transfected group were significantly down-regulated compared with control siRNA transfection group(P<0.01).The expression levels of C-myc,Cyclin D1,PI3K,p-mTOR and p-AKT were significantly down-regulated when treated with temozolomide(P<0.05-0.01).When PTPN1 was overexpressed,the expression levels of C-myc,Cyclin D1,PI3K,p-mTOR and p-AKT were significantly up-regulated(P<0.05-0.01).When PTPN1 was silenced,the expression levels of C-myc,Cyclin D1,PI3K,p-mTOR and p-AKT were significantly down-regulated(P<0.05).Conclusion Temozolomide can affect the mTOR signaling pathway by down-regulating the expression of PTPN1 in glioma cells,and inhibit the proliferation and invasion of glioma cells.
作者
赵海康
王凤鹿
杨磊
袁致海
陈明生
赵瑛
解杨
张亮
ZHAO Hai-kang;WANG Feng-lu;YANG Lei(Department of Neurosurgery, Second Affiliated Hospital of Xi'an Medical College, Xi'an 710038, China)
出处
《临床神经外科杂志》
CAS
2020年第6期638-644,共7页
Journal of Clinical Neurosurgery
基金
国家自然科学基金(81870172)。
