Effect of heat shock protein family A member 5(HSPA5) on
artesunateinduced ferroptosis in hepatic carcinoma
WANG Kang1, ZHANG Zheng-yang2, SONG Lian2, GONG Ai-hua1, WANG Dong-qing2, ZHU Hai-tao2
(1. School of Medicine, Jiangsu University, Zhenjiang Jiangsu 212013; 2. Central Laboratory of Radiology, Affiliated Hospital of Jiangsu University, Zhenjiang Jiangsu 212001, China)
摘要 目的: 探讨热休克蛋白5(heat shock protein family A member 5，HSPA5)对青蒿琥酯诱导人肝癌SMMC7721细胞株化疗敏感性的影响。方法: 用不同浓度青蒿琥酯处理SMMC-7721细胞，CCK-8法检测细胞活性，筛选最佳实验浓度。将SMMC-7721细胞按以下分组处理：对照组、青蒿琥酯组、青蒿琥酯+去铁胺组，用流式细胞术检测细胞内脂质来源活性氧水平；试剂盒检测细胞内丙二醛水平。用包装HSPA5干扰或过表达质粒的慢病毒感染SMMC7721细胞，qRTPCR和蛋白质印迹法分别测定转染后HSPA5 mRNA和蛋白表达；CCK-8法检测细胞活性，试剂盒检测细胞内丙二醛水平。结果： 青蒿琥酯浓度为20 μmol/L时，SMMC-7721细胞达到半数致死量，为最佳实验浓度；流式细胞术结果显示，青蒿琥酯处理的细胞内脂质来源活性氧水平、丙二醛水平明显升高，去铁胺可抑制青蒿琥酯导致的细胞内脂质来源活性氧和丙二醛升高(均P<0.05)；经过青蒿琥酯处理的HSPA5干扰组细胞活性水平明显低于未干扰组，丙二醛水平明显高于未干扰组(均P<0.05)。结论： 干扰HSPA5可能增强人肝癌SMMC-7721细胞株对青蒿琥酯的化疗敏感性。
Abstract：Objective: To investigate the effect of heat shock protein family A member 5(HSPA5) on the chemosensitivity of artesunate in human hepatic carcinoma SMMC7721 cells. Methods: SMMC7721 cells were treated with different concentrations of artesunate, CCK8 assay was used to detect the cell viability, and to screen the best conecntration. SMMC-7721 cells were divided into control group, artesunate group, artesunate+deferaxamine group. CCK8 assay was used to test the cell viability. Flow cytometry was used to detect the level of lipid reactive oxygen species(ROS). The malondialdehyde detection kit was used to detect the level of malondialdehyde. qRT-PCR and Western blotting were used to detect the mRNA and protein expression level of HSPA5 in SMMC-7721 cells treated with Vector, HSPA5-shRNA or Flag-HSPA5 plasmid. The CCK-8 assay kit was used to test the cell viability. The malondialdehyde detection kit was used to detect the level of malondialdehyde. Results: The cells showed the modest survival rate when the concentration of artesunate was 20 μmol/L. Flow cytometry showed the levels of lipid ROS and malondialdehyde in cells treated with artesunate increased(P<0.05), which could be reversed by deferoxamine; cell viability in the HSPA5-shRNA group treated with artesunate were lower than those in the Vector group(P<0.05). The level of malondialdehyde in the HSPA5shRNA group treated with artesunate were higher than those in the Vector group(P<0.05). Conclusion: Knockdown of HSPA5 may enhance the chemosensitivity of artesunate in human hepatic carcinoma SMMC7721 cells.
王康1， 张正阳2， 宋廉2， 龚爱华1， 王冬青2， 朱海涛2. 沉默热休克蛋白5可增敏青蒿琥酯诱导肝癌细胞铁死亡[J]. 江苏大学学报:医学版, 2019, 29(04): 316-321.
WANG Kang1, ZHANG Zheng-yang2, SONG Lian2, GONG Ai-hua1, WANG Dong-qing2, ZHU Hai-tao2. Effect of heat shock protein family A member 5(HSPA5) on
artesunateinduced ferroptosis in hepatic carcinoma. Journal of Jiangsu University(Medicine Edition), 2019, 29(04): 316-321.
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2019, Vol. 29 
