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目的:研究雷丸蛋白对人胃癌细胞MC-4凋亡的诱导作用。方法:以聚乙烯吡咯烷酮(PVP)为溶剂提取雷丸饮片蛋白质,BCA法测定蛋白浓度后以DMEM培养液稀释到180mg.mL-1,120mg.mL-1,90mg.mL-1,60mg.mL-1和45 mg.mL-1,体外作用于胃癌细胞MC-4,37℃、5%CO2条件下培养18h后,光镜检查作用结果。以Hoechst 33258染色后于荧光显微镜下观察细胞凋亡情况,以Annexin V-PI双染色后以流式细胞仪检查细胞的死亡和凋亡情况。结果:随着蛋白质用量的增加,MC-4细胞死亡率与剂量间呈现量效关系,180mg.mL-1时达最高值29.6%。凋亡率和剂量间呈现较好的剂量-效应曲线,120mg.mL-1时凋亡率最高,可达29.4%,聚乙烯吡咯烷酮无作用。结论:PVP荷载雷丸蛋白对人胃癌细胞MC-4有很好的杀伤作用和诱导凋亡作用,杀伤机理和凋亡机理有待深入研究。
Objective: To study the induction of Rayleigh protein on apoptosis of human gastric cancer cell line MC-4. Methods: The protein of Leiwan Decoction was extracted with polyvinylpyrrolidone (PVP) as solvent, the protein concentration was determined by BCA method and diluted to 180mg.mL-1, 120mg.mL-1, 90mg.mL-1 and 60mg.mL- 1 and 45 mg.mL-1, respectively, in vitro effects on gastric cancer cells MC-4, 37 ℃, 5% CO2 for 18h, the results of light microscopy. Cell apoptosis was observed under a fluorescence microscope with Hoechst 33258 staining. Cell death and apoptosis were examined by flow cytometry after Annexin V-PI staining. Results: With the increase of protein dosage, there was a dose-response relationship between the MC-4 cell death rate and the dose, reaching the highest value of 29.6% at 180mg.mL-1. The apoptosis rate and dose showed a good dose-response curve. The highest apoptosis rate was 120 mg.mL-1, up to 29.4%, with no effect of polyvinylpyrrolidone. CONCLUSION: PVP-loaded Leiwan protein has good cytotoxicity and apoptosis-inducing effect on human gastric cancer cell line MC-4, and its killing mechanism and apoptosis mechanism need further study.