黑色素瘤分化相关基因-7/白细胞介素-24促进阿霉素杀伤肝癌细胞MHCC-97L机制的研究

目的 探讨黑色紊瘤分化相关基因-7/白细胞介素-24(MDA-7/IL-24)基因促进阿霉素(ADM)杀伤肝癌细胞,逆转肝癌细胞多药耐药(MDR)的机制.方法 以人肝癌细胞株MHCC-97L为实验对象,使用噻唑蓝(MTT)比色法和流式细胞仪比较Ad. MDA-7联合ADM处理组与ADM组、Ad. MDA-7组对肝癌细胞MHCC-97L和正常肝细胞L02的作用差异.观察MDA-7/IL-24对多药耐药的逆转作用.荧光定量聚合酶链反应(PCR)检测MDR-1、STAT-3、bcl-2、bax mRNA的变化.Western blot检测gp-170、STAT-3、bcl-2、bax蛋白的表达的变化.结果 MTT表明Ad. MDA-7对正常肝细胞LO2无生长抑制作用(P>0.05).LO2细胞Ad. MDA-7联合ADM组与ADM组细胞生存率差异无统计学意义(P>0.05).低浓度(100 VP/cell)的Ad. MDA-7联合正常肝细胞的IC50浓度的ADM(1.5 mg/L)使得细胞抑制率从ADM组的17.46%上升到79.50%,生长抑制逆转4.55倍(P<0.05).MDR-1mRNA相对表达量从(16.49±0.11)下降至(5.48±0.05).STAT-3 mRNA相对表达量从(13.17±0.08)上升至(21.57±0.11).bcl-2及BAX表达与其他实验组比较差异均有统计学意义(P<0.05).联合实验组P-170蛋白的表达量较其他组明显降低,而磷酸化STAT-3蛋白的表达量亦增加.结论 Ad. MDA-7具有逆转肝癌细胞MHCC-97L多药耐药的作用,其下调MDR-1 mRNA的表达的同时,并通过活化STAT-3信号通路的表达促进肝癌细胞凋亡.

The MDA-7/IL-24 promote ADM to arrest the growth of hepatoma carcinoma cell lines MHCC-97L

Objective To explore the mechanism of the melanoma differentiation associated gene-7 (MDA-7/IL-24) promoting adriamycin (ADM) to kill the hepatoma carcinoma cell and reversing multidrug resistance (MDR). Methods By using the human hepatoma carcinoma cell line MHCC-97L and normal hepatic cell line LO2, MTT assay and flow cytometry (FCM) were applied to compare the cell growth among the combined ( Ad. mda-7 + ADM ) group, ADM group and Ad. mda-7 group. The expression levels of MDR-1, STAT-3, bcl-2, and bax mRNA were examined by using real-time polymerase chain reaction (PCR). Western blotting was performed to observe the change in the expression of gp-170, STAT-3, bcl-2, and bax among those groups. Results MTT showed that Ad. mda-7 had no toxic effect on LO2 cells, and there was no significant difference in growth rate between adriamycin group and combined group (P>0.05). The ratio of growth suppression in MHCC-97L cells only treated with ADM (1.5 mg/L) was 17.46%, but in combined group subject to treatment of 100 VP/cell Ad. mda-7 and ADM (1.5 mg/L) it was increased to 79.50% (P<0.05 ). Real-time PCR revealed the expression of MDR-1 mRNA was decreased from (16.49±0.11) in ADM group to (5.48±0.05) in combined group, and that of STAT-3 mRNA increased from (13.17±0.08) to (21.57±0.11) correspondingly (P<0.05). Western blotting also demonstrated that the expression of P-170 in combined group was decreased and that of phosphorylated STAT-3 was increased as compared with other two groups. Conclusion Ad. mda-7 can reverse the MDR of ADM to MHCC-97L cells, inhibit the expression of MDR-1 mRNA and activate the signaling of STAT-3 to induce apoptosis of hepatoma carcinoma cell MHCC-97L.