GSDME对SKOV-3卵巢癌细胞化疗耐药的影响

doi:10.20223/j.cnki.1000-8535.2025.10.006

摘要/Abstract

摘要： 目的探讨卵巢癌化学治疗（化疗）耐药与焦孔素E（GSDME）基因的甲基化是否有关,以及地西他滨是否可以通过去甲基化使GSDME蛋白表达水平升高从而逆转卵巢癌化疗耐药。方法顺铂逆浓度梯度构建SKOV-3卵巢癌耐顺铂细胞株（SKOV-3/DDP）;CCK8法检测耐药前后细胞株的半抑制浓度（IC₅₀）;实时荧光定量甲基化特异性PCR法检测两组细胞中GSDME基因的甲基化水平;Wetern Blot检测两组细胞中GSDME的表达水平。将耐药细胞株用不同质量浓度的地西他滨处理,重复上述实验,检测地西他滨处理前后细胞的IC₅₀、GSDME基因的甲基化水平及GSDME蛋白的表达程度。结果与SKOV-3细胞相比,SKOV-3/DDP中GSDME基因的甲基化水平升高（P<0.01）,同时GSDME蛋白的表达水平降低（P<0.001）;随着地西他滨作用浓度的升高,耐药细胞中GSDME基因的甲基化程度逐渐降低,蛋白的表达水平逐渐升高,细胞的IC₅₀逐渐降低：在用0.5 μg/mL地西他滨处理耐药细胞后GSDME基因的甲基化水平虽然降低（P<0.01）,但是此时蛋白的表达水平及耐药细胞的IC₅₀均无明显改变（P>0.05）;当地西他滨的浓度增加到1.0 μg/mL时蛋白的表达水平才明显升高（P<0.05）,而此时细胞的IC₅₀仍未见明显降低（P>0.05）;待药物浓度达到1.5 μg/mL时,细胞的IC₅₀才表现出明显的下降趋势（P<0.05）。结论 GSDME的表达与卵巢癌的化疗耐药密切相关,GSDME的高甲基化水平致使其低表达可促进卵巢癌的化疗耐药。但地西他滨可以通过去甲基化使卵巢癌耐药细胞中GSDME的表达水平升高,从而增加卵巢癌细胞对化疗药物的敏感性,进而逆转卵巢癌化疗耐药。

关键词: 卵巢癌, 焦孔素E, 化疗耐药, 地西他滨, 甲基化

Abstract: Objective To explore whether drug resistance in ovarian cancer is associated with gasdermin E（GSDME） methylation,and to explore whether decitabine can reverse ovarian cancer chemoresistance by increasing GSDME protein expression levels through demethylation．Methods The cisplatin-resistant cell line（SKOV-3/DDP）was constructed by inverse concentration gradient of cisplatin．Semi-inhibitory concentration（IC₅₀）of cell lines after drug resistance was detected using the CCK8 assay．Real-time fluorescence quantitative methylation-specific PCR was used to detect the methylation level of GSDME gene in the two groups of cells．Wetern Blot was used to detect the expression level of GSDME in the two groups of cells．Drug-resistant cell lines were treated with different concentrations of the demethylating drug decitabine．Experiments above were repeated to detect the methylation degree of IC₅₀ and GSDME genes and the expression level of GSDME protein in drug-resistant cells before and after decitabine treatment．Results Compared with SKOV-3 cells,the methylation level of GSDME gene in SKOV-3/DDP was significantly increased（P<0.01）,while the expression level of GSDME protein was significantly decreased（P<0.001）．With the increase of decitabine concentration,the methylation degree of GSDME gene in drug-resistant cells was gradually decreased,the expression level of protein was gradually increased,and the IC₅₀ of cells was gradually decreased：the methylation level of GSDME gene was decreased after 0.05 μg/mL decitabine treatment（P<0.01）,but there were no significant changes in protein expression level and IC₅₀ of drug-resistant cells（P>0.05）．The protein expression level was significantly increased when the concentration of local citabine was increased to 0.10 μg/mL（P<0.05）,while the IC₅₀ of the cells was not significantly decreased（P>0.05）．When the drug concentration reached 0.15 μg/mL,the IC₅₀ of the cells showed a significant downward trend（P<0.05）．Conclusions The expression of GSDME is closely related to chemoresistance in ovarian cancer,and the low expression of GSDME due to its high methylation level can promote chemoresistance in ovarian cancer．However,decitabine can increase the expression level of GSDME in drug-resistant ovarian cancer cells through demethylation,thereby increasing the sensitivity of ovarian cancer cells to chemotherapeutic drugs,and then reversing the chemoresistance of ovarian cancer．

Key words: ovarian cancer, GSDME, chemoresistant, cisplatin, methylation

张渊, 白芷玉, 李琪, 闫俐嘉, 贾娟蓉, 冯勤梅, 尹海珍. GSDME对SKOV-3卵巢癌细胞化疗耐药的影响[J]. 广州医药, 2025, 56(10): 1363-1371.

ZHANG Yuan, BAI Zhiyu, LI Qi, YAN Lijia, JIA Juanrong, FENG Qinmei, YIN Haizhen. Effect of GSDME on chemoresistance in ovarian cancer[J]. Guangzhou Medical Journal, 2025, 56(10): 1363-1371.

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