LINC 01118通过调控vimentin蛋白影响卵巢癌细胞紫杉醇耐药

LINC 01118 affects paclitaxel resistance in ovarian cancer cells by regulating vimentin protein

目的探讨LINC 01118对人卵巢癌细胞紫杉醇耐药的影响,并研究其调控机制。方法利用RT-qPCR测定两组卵巢癌紫杉醇耐药细胞系及亲本细胞系中LINC 01118的表达水平;利用Lipofectamine TM2000将siRNA-LINC 01118、siRNA-LINC 01118+GV 230-vim以及相关对照转染至卵巢癌紫杉醇耐药细胞SKOV 3-TR 30中,RT-qPCR验证siRNA对LINC 01118的靶向沉默效率;利用MTT法测定转染后SKOV 3-TR 30细胞对紫杉醇的敏感性;利用流式细胞仪测定转染后SKOV 3-TR 30细胞的凋亡率;利用免疫印迹实验(Western blot)测定转染后vimentin蛋白,耐药蛋白MRP 1、MDR 1及ABCG 2的表达水平。结果RT-q PCR实验结果显示,LINC 01118在卵巢癌紫杉醇耐药细胞中的表达水平显著高于其亲本细胞(P<0.05);转染siRNA-LINC 01118可明显抑制SKOV 3-TR 30细胞中LINC 01118的表达(P<0.05);MTT实验结果显示,抑制LINC 01118的表达显著增加了SKOV 3-TR 30细胞对紫杉醇的敏感性(P<0.05),过表达vimentin能逆转抑制LINC 01118表达时卵巢癌耐药细胞对紫杉醇的敏感性增加现象(P<0.05);流式细胞术结果显示,抑制LINC 01118表达显著增加了SKOV 3-TR 30细胞的凋亡率(P<0.05);Western-blot实验结果显示,抑制LINC 01118表达后,vimentin蛋白的表达水平明显降低(P<0.05),耐药蛋白MRP 1、MDR 1及ABCG 2的表达均显著减少(P<0.05)。结论LINC 01118在卵巢癌紫杉醇耐药细胞中异常高表达;抑制LINC 01118表达可以逆转卵巢癌细胞紫杉醇耐药,诱导细胞凋亡;LINC 01118可以通过正向调控vimentin蛋白影响卵巢癌细胞紫杉醇耐药。

Objective To investigate the effect of LINC01118 on paclitaxel resistance in human ovarian cancer cells and its regulatory mechanism.Methods RT-qPCR was used to determine the expression of LINC01118 in the two groups of paclitaxel resistant ovarian cell lines and its parent cell lines.SiRNA-LINC01118,siRNA-LINC 01118+GV 230-vim and related controls were transfected into paclitaxel resistant ovarian cell SKOV 3一TR 3 using Lipofectamine TM 2000,and the targeted silencing efficiency of siRNA on LINC 01118 was verified by RT-qPCR.The sensitivity of SKOV 3-TR 3 cells to paclitaxel after transfection was determined by MTT assay.Flow cytometry was used to detect the apoptosis rate of SKOV 3-TR 3 after transfection.Western一blot assay was used to detect the expression levels of vimentin protein and drug resistance proteins MRP 1,MDR 1 and ABCG 2 after transfection.Results The results of RT-qPCR showed that the expression level of LINC01118 in paclitaxel resistant ovarian cells was significantly higher than that of parental cells(P<0.05).Transfection of siRNA-LINC 01118 significantly inhibited the expression of LINC 01118 in SKOV 3-TR 30 cells(P<0.05).MTT assay showed that inhibition of LINC 01118 significantly increased the sensitivity of SKOV 3-TR 30 cells to paclitaxel(P<0.05),overexpression of vimentin can reverse increased sensitivity of paclitaxel resistant ovarian cell when inhibiting the expression of LINC 01118(P<0.05).The results of flow cytometry showed that inhibition of LINC 01118 expression significantly increased the apoptosis rate of SKOV 3-TR 30 cells(P<0.05).The results of Western-blot showed that the expression of vimentin protein was significantly decreased after inhibition of LINC 01118 expression(P<0.05),and the expressions of drug resistance proteins MRP 1,MDR 1 and ABCG2 were significantly decreased(P<0.05).Conclusion LINC 01118 is highly expressed in paclitaxel resistant ovarian cells;inhibition of LINC 01118 expression can reverse paclitaxel resistance and induce apoptosis in paclitaxel resistant ovarian cells;LINC 01118 can affect paclitaxel resistance of ovarian cancer cells by positive regulation of vimentin protein.