Transgelin 2过表达诱导人乳腺癌细胞MCF-7对紫杉醇耐药作用研究

Effect of Transgelin 2 Overexpression on Paclitaxel Resistance in Human Breast Cancer MCF-7 Cells

目的探讨Transgelin 2过表达对人乳腺癌细胞MCF-7紫杉醇耐药性影响及相关机制。方法运用质粒转染技术升高MCF-7中Transgelin 2的表达,并根据是否转染Transgelin 2的基因(TAGLN2)分为3组:空白对照组(control),阴性对照组(pcDNA3.1)、Transgelin 2转染组(High-TAGLN2);利用Real-time PCR和Western blot检测各组细胞中Transgelin 2和耐药因子的mRNA和蛋白表达;采用四甲基偶氮唑蓝(MTT)法测定细胞活性;通过流式细胞术检测细胞凋亡情况;利用Western blot测定PI3K/Akt通路相关因子的表达。结果PI3K/Akt通路在MCF-7/PTX中激活;与空白对照组和pcDNA3.1组相比,High-TAGLN2组细胞对紫杉醇的敏感性降低,凋亡细胞减少,Transgelin 2和P-gp、MRP1、BCRP的mRNA和蛋白表达均明显增加,PI3K/Akt通路随着Transgelin 2升高而激活,Bcl-2表达升高,Bax表达降低。结论应用质粒转染技术升高Transgelin 2表达降低了MCF-7对紫杉醇的敏感性,诱导了乳腺癌耐药,提示Transgelin 2在乳腺癌紫杉醇耐药中发挥着重要作用,有望成为乳腺癌耐药治疗的新分子标志物。

OBJECTIVE To investigate the effect of Transgelin 2 overexpression on paclitaxel resistance in human breast cancer MCF-7 cells.METHODS Transgelin 2 was increased in MCF-7 cells via plasmid transient transfection.MCF-7 cells were divided into three groups:the high-TAGLN2 group in which MCF-7 cells were infected by plasmid vectors carrying TAGLN2(encoding Transgelin 2)gene,the blank control group and the negative control(pcDNA3.1)group.Real-time PCR and Western blot assay were performed to measure the expression of Transgelin 2 and factors related to drug resistance.Cell viability was determined using MTT method.The apoptosis was detected by flow cytometry.PI3K/Akt signal pathway and its downstream apoptosis molecules including Bcl-2 and Bax were tested by Western blot.RESULTS PI3K/Akt pathway was activated in MCF-7/PTX cells.The mRNA and protein levels of Transgelin 2,P-gp,MRP1 and BCRP in High-TAGLN2 group were higher than those in the control group and pcDNA3.1 group.Transgelin 2 overexpression decreased the sensitivity of breast cancer cells to paclitaxel.After treatment with 20 nmol·L-1 paclitaxel,the cell apoptosis rate in High-TAGLN2 group(23.82%)was lower than that in the pcDNA3.1 group(44.19%).Moreover,overexpression of Transgelin 2 was able to activate PI3K/Akt pathway in MCF-7 cells.On the other hand,the expression of Bcl-2 was aggrandized while Bax was decreased.CONCLUSION Transgelin 2 overexpression desensitizes the MCF-7 cells to paclitaxel and induces the formation of breast cancer drug resistance,suggesting that Transgelin 2 plays a critical role in paclitaxel resistance of breast cancer and could be a novel molecular target for breast cancer treatment.