甲基莲心碱对耐长春新碱人胃癌细胞多药耐药性的逆转

目的:研究甲基莲心碱(Nef)逆转耐长春新碱人胃癌细胞多药耐药性(MDR)的作用及机制。方法:采用噻唑蓝(MTT)比色法检测长春新碱(VCR)的细胞毒性;PI染色流式细胞计数测定VCR诱导细胞凋亡;间接免疫荧光流式细胞术检测细胞P-gp和MRP的表达。结果:Nef(5、10μmol·L^-1)对人胃癌细胞(SGC7901)和耐长春新碱人胃癌细胞(SGC7901/VCR)无显著毒性作用,VCR对敏感株SGC7901的IC₅₀为0.06mg·L^-1,而对MDR细胞株SGC7901/VCR的IC₅₀为2.32mg·L^-1,SGC7901/VCR较SGC7901对VCR耐药39倍,Nef(2.5、5、10μmol·L^-1)能使VCR对SGC7901/VCR细胞的IC₅₀从2.32mg·L^-1依次下降至0.34、0.12、0.05mg·L^-1,逆转倍数分别为6.8、18.1、43.8。Nef(2.5、5、10μmol·L^-1)能降低SGC7901/VCR细胞对VCR的凋亡抗性,其作用强于维拉帕米(VRP)。SGC7901/VCR细胞较SGC7901细胞高表达P-gp、MRP,Nef(10μmol·L^-1)处理24h后,SGC7901/VCR细胞P-gp、MRP的表达明显低下。结论:Nef具有逆转耐长春新碱人胃癌细胞的MDR作用,其作用机理与下调P-pg和MRP表达有关。

Reversal effect of neferine on resistance to vincristine in human multidrug-resistant gastric carcinoma cell line SGC7901/VCR

AIM: To investigate the effect of neferine (Nef) on human gastric carcinoma cell line with multidrug resistance (MDR). METHODS: The cytotoxic effect of vincristine (VCR) was evaluated by MTT assay. The cell apoptosis induced by VCR was determined by flow cytometry, and the expression of P-glycoprotein (P-gp) and multidrug-resistance-associated protein (MRP) in cells was examined by immunofluorescence flow cytometry. RESULTS: MTT assay showed that Nef at the concentration of 5 μmol·L^-1 to 10 μmol·L^-1 have no cytotoxicity to parent human gastric carcinoma cell line (SGC7901) and its VCR-resistant variant cell line (SGC7901/VCR). The IC₅₀ value of VCR to SGC7901 cell line was 0.06 mg·L^-1and that of to SGC7901/VCR cell line was 2.32 mg·L^-1, which indicated SGC7901/VCR cell line were 39 times more resistant to VCR in comparison with the parent SGC7901 cell line. After treatment with Nef at the concentrations of 2.5, 5 and 10 μmol·L^-1, the IC₅₀ value of VCR to SGC7901/VCR cell line decreased to 0.34, 0.12 and 0.05 mg·L^-1, respectively and those increased by 6.8-, 18.1- and 43.8- fold in the chemosensitivity, respectively. Flow cytometry showed that SGC7901/VCR cells were resistant to apoptosis induced by VCR. After 24 h treatment with Nef (2.5, 5 and 10 μmol·L^-1) and VCR, the apoptosis of SGC7901/VCR cells increased, which indicated Nef could abolish resistance of SGC7901/VCR cells to VCR-induced apoptosis. Furthermore, the action of Nef was more potent than verapamil. The expression of P-glycoprotein and multidrug resistance associated protein was strongly positive in SGC7901/VCR cells, and the expression level of P-gp and MRP in SGC701/VCR cells was significantly down-regulated at 24 h after treatment with Nef (10 μmol·L^-1). CONCLUSIONS: Nef can reverse MDR in multidrug-resistant human gastric carcinoma SGC7901/VCR cell line. Its mechanism might be associated with down-regulation of expression of P-gp and MRP in SGC7901/VCR cells.