ERCC1基因与卵巢上皮癌细胞铂类耐药相关性探讨

发布时间：2018-10-14 18:34

【摘要】：目的检测切除修复交叉互补1(ERCC1)基因在人卵巢上皮癌顺铂(DDP)耐药细胞株SKOV3/DDP及其亲本细胞株SKOV3中的表达,探讨ERCC1基因在卵巢上皮癌顺铂耐药中的意义以及有效沉默ERCC1基因能否有效逆转SKOV3/DDP细胞对DDP耐药。方法于2013—2014年在南方医科大学采用实时荧光定量PCR(RT-PCR)和Western印迹法检测ERCC1 m RNA和蛋白在SKOV3/DDP及SKOV3细胞中的表达。人工构建3个靶向沉默ERCC1基因的短发夹状RNA(sh RNA)质粒表达载体,慢病毒载体法转染至SKOV3/DDP细胞,RT-PCR和Western印迹法检测ERCC1基因水平并挑选沉默效果最佳的表达载体。RT-PCR和Western印迹法检测稳定转染后SKOV3/DDP细胞中ERCC1基因的表达,CCK-8法检测转染细胞对DDP的耐药性。结果 SKOV3/DDP细胞中ERCC1 m RNA和蛋白的表达量明显高于SKOV3,差异有统计学意义(P0.01)。稳定转染ERCC1 sh RNA后的SKOV3/DDP细胞,ERCC1的m RNA及蛋白的表达量明显下降,差异有统计学意义(P0.01)。细胞活性检测提示特异性沉默ERCC1基因能增加SKOV3/DDP细胞对顺铂的敏感性(P0.05)。结论 ERCC1基因在SKOV3/DDP细胞中的表达明显升高,有效沉默ERCC1基因能有效逆转SKOV3/DDP细胞对DDP耐药,增加卵巢上皮癌DDP耐药细胞对DDP的敏感性,为卵巢上皮癌耐药的治疗提供新靶点。
[Abstract]:Objective to detect the expression of excision repair cross complementary 1 (ERCC1) gene in cisplatin (DDP) resistant human ovarian carcinoma (OECC) cell line SKOV3/DDP and its parent cell line SKOV3. To investigate the significance of ERCC1 gene in cisplatin resistance in ovarian epithelial carcinoma and whether the effective silencing of ERCC1 gene can effectively reverse the resistance of SKOV3/DDP cells to DDP. Methods Real-time fluorescence quantitative PCR (RT-PCR) and Western blot were used to detect the expression of ERCC1 m RNA and protein in SKOV3/DDP and SKOV3 cells from 2013 to 2014. Three short hairpin RNA (sh RNA) expression vectors targeting silencing ERCC1 gene were constructed. Lentivirus vector was transfected into SKOV3/DDP cells, RT-PCR and Western blotting were used to detect the level of ERCC1 gene and the best expression vector was selected. RT-PCR and Western blotting were used to detect the expression of ERCC1 gene in SKOV3/DDP cells after stable transfection, and CCK-8 assay was used to detect the resistance of transfected cells to DDP. Results the expression of ERCC1 m RNA and protein in SKOV3/DDP cells was significantly higher than that in SKOV3, cells (P0.01). After stable transfection of ERCC1 sh RNA, the expression of m RNA and protein of ERCC1 in SKOV3/DDP cells decreased significantly (P0.01). Specific silencing of ERCC1 gene increased the sensitivity of SKOV3/DDP cells to cisplatin (P0.05). Conclusion the expression of ERCC1 gene in SKOV3/DDP cells was significantly increased. Effectively silencing ERCC1 gene could effectively reverse the drug resistance of SKOV3/DDP cells to DDP, increase the sensitivity of DDP resistant cells to DDP, and provide a new target for the treatment of ovarian epithelial carcinoma.
【作者单位】：南方医科大学;南方医科大学珠江医院;广州医科大学附属第一医院妇产科;
【分类号】：R737.31

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