紫草素逆转绒癌细胞JAR/MTX耐药与survivin和Bcl-2表达关系研究

Relationship between the reverse effect of shikonin on methotrexate-resistant human choriocarcinoma cell line JAR/MTX and expressions of survivin and Bcl-2

目的探讨紫草素逆转绒癌细胞耐药效应在凋亡方面的机制。方法体外培养细胞并将其分为4组,各组加药后观察每组细胞24、48、72h生长情况并计算细胞贴壁率,电化学发光法检测β-HCG值,S-P免疫组织化学及RT-PCR法检测survivin、Bcl-2基因表达。结果MTX对细胞JAR/MTX毒性作用、细胞贴壁率、β-HCG下降程度与对照相比差异无统计学意义(P>0.05);紫草素作用细胞后与对照相比差异有统计学意义(P<0.05);二者合用作用于JAR/MTX细胞与对照相比差异显著(P<0.01),survivin基因的mRNA和蛋白表达与对照相比差异显著(P<0.01),Bcl-2基因的mRNA表达与对照相比差异显著(P<0.01),蛋白表达与对照相比差异有统计学意义(P<0.05)。结论紫草素可逆转JAR/MTX细胞对MTX耐药是通过下调survivin、Bcl-2基因表达增加细胞凋亡实现的;紫草素有可能成为绒癌化疗的新型增敏剂。

Objective To explore the mechanism of how shikonin to reverse the muhi-drug resistance of human choriocarcinoma ceils. Methods JAR/MTX ceils were divided into 4 groups. The cell growth condition of each group at 24, 48, and 72 h was observed and cell adhesion rate was calculated. β-HCG values at 24 and 48 h after adding drugs were assayed. Cytomorphologic changes were observed by H-E staining. The gene expression of survivin and Bcl-2 was detected by S-P immunocytochemistry and RT-PCR. Results There was no significant difference in cell adhesion rate and secretion of β-HCG between methotrexate treatment group and control group （ P 〉 0.05 ）, but there was difference between shikonin group and control one （ P 〈 0.05 ）. The strong effect was noted after combining use of methotrexate and shikonin as compared with control group （P 〈 0. 01 ）. There were significant differences in mRNA and protein expression of survivin and Bcl-2 between shikonin group and control one （ P 〈 0.01 ）. Conclusion Shikonin can reverse drug-resistance of JAR/MTX to methotrexate by down-regulating the expression of survivin and Bcl-2 and accelerating apoptosis. Shikonin is promising as a synergist of chemotherapy used for choriocarcinoma.