乳源六肽增加人卵巢癌细胞对顺铂的敏感性

PGPIPN increases the sensitivity of ovarian cancer cells to cisplatin

目的研究乳源六肽(PGPIPN)促进人卵巢癌细胞对顺铂(DDP)的敏感性及其机制。方法体外培养人卵巢癌敏感细胞株COC1和耐DDP的耐药细胞株COC1/DDP,用CCK8法检测其在DDP作用下的细胞增殖抑制率,计算DDP在作用24、48、72 h的半抑制浓度(IC_(50))。不同浓度的PGPIPN联合DDP(IC_(50))分别作用COC1和COC1/DDP 24、48、72 h,CCK8法检测其对生长增殖的影响,用流式细胞术检测其对细胞凋亡的影响。用RT-PCR和Western blot检测人卵巢癌细胞中切除修复交叉互补基因1(ERCC1)的表达情况。结果 PGPIPN联合DDP组同单独使用DDP组相比,人卵巢癌细胞敏感株COC1和人卵巢癌耐药细胞株COC1/DDP的抑制率与凋亡率均增加,增加的幅度与PGPIPN浓度在一定范围内成正比,PGPIPN对耐药株COC1/DDP效果高于敏感株COC1。PGPIPN增加人卵巢癌细胞对DDP敏感性在作用48 h时效果最好,对COC1/DDP和COC1的抑制率最高分别为76.8%和62.3%,凋亡率最高分别为67.4%和45.4%,显著高于单独DDP作用(P<0.01)。PCR和Western blot检测COC1和COC1/DDP的ERCC1基因的表达,结果显示在DDP联合不同浓度PGPIPN作用下ERCC1基因的表达均低于单独使用DDP组,COC1/DDP中ERCC1基因表达量随着PGPIPN浓度的升高而显著降低,尤其是对耐药细胞株COC1/DDP的ERCC1基因表达最为显著,显著高于单独DDP作用(P<0.05,P<0.01)。敏感细胞株COC1细胞中ERCC1基因的表达受PGPIPN的影响不如COC1/DDP明显,只是在高剂量(1×10^(-2)g/L)时,ERCC1基因表达明显下降。结论 PGPIPN能增加人卵巢癌细胞对DDP敏感性,其可能是通过调低ERCC1的表达来实现的。

Objective To explore on the sensitivity and mechanism of immunomodulating peptide（ PGPIPN） promoting human ovarian cancer cells to cisplatin（ DDP）. Methods Human ovarian cancer cell line COC1 and DDP resistant cell line COC1/DDP were cultured in vitro. The inhibition rate of cell proliferation under the action of DDP was detected by CCK8 method,and the concentration of DDP in the half inhibitory concentration of 24 h,48 h and 72 h was calculated. PGPIPN at different concentrations combined with DDP（ IC_（50） concentration） acted on COC1 and COC1/DDP for 24 h,48 h and 72 h respectively. CCK8 method was used to detect its effect on growth and proliferation of ovarian cancer cells,and their apoptosis were detected by flow cytometry. RT-PCR and Western blot were used to detect the expression of excision repair cross complementing 1（ ERCC1） in human ovarian cancer cells. Results Compared with the group treated only with DDP,combination groups were significantly inhibited the proliferation and apoptosis of human ovarian cancer cell both sensitive strain COC1 and resistant strain COC1/DDP,and the inhibition effect showed dose-dependent manor. The effect of PGPIPN on COC1/DDP was higher than that of COC1. PGPIPN increased the sensitivity of human ovarian cancer cells to DDP,and the best was on 48 h,in which the inhibition rates of COC1/DDP and COC1 were 76. 8% and 62. 3% respectively,and the highest apoptosis rates were 67. 4% and 45. 4%,respectively,which was significantly higher than that of individual DDP group（ P〈0. 01）. PCR and Western blot were used to assay the expressions of ERCC1 gene in COC1 and COC1/DDP,the results showed that the expression of ERCC1 gene in combined groups were significantly lower than that of the group treated only with DDP,which showed dose-dependent manor in DDP-resistant cell line COC1/DDP（ P〈0. 05,P〈0. 01）. The expression of ERCC1 gene in COC1 cells was less affected by PGPIPN than that of COC1/DDP,in which the expression of ERCC1 gene decreased obviously only at high dose（1 × 10^（-2） g/L）. Conclusion PGPIPN can increase the sensitivity of human ovarian cancer cells to DDP,which may be achieved by decreasing the expression of ERCC1.