蟾蜍灵通过程序性坏死途径诱导人胃癌细胞SGC-7901死亡

目的 探讨蟾蜍灵（Bufalin）诱导人胃癌SGC-7901细胞死亡过程中程序性坏死途径的发生机制。方法 体外培养SGC-7901细胞，于对照组加入RPMI 1640培养液，实验组加入不同浓度的蟾蜍灵（50、100、150、200 nmol/L），对照组与实验组细胞继续培养48 h。MTT法检测细胞活性，DAPI染色法观察细胞核形态改变，透射电镜观察细胞膜及细胞核改变，流式细胞术检测细胞坏死率，Western blotting法检测程序性坏死关键蛋白受体相互作用蛋白激酶1（receptor interacting protein kinase 1，RIP1）的表达。结果 MTT实验结果表明，蟾蜍灵对SGC-7901细胞的生长具有显著的抑制作用（P<0.05）。通过透射电镜可以观察到细胞坏死时的特征性改变，如细胞膜破裂、细胞内空泡的产生以及细胞器的崩解。DAPI染色显示，蟾蜍灵（100 nmol/L）处理细胞48 h后，无明显细胞凋亡特征性改变，如细胞核固缩、核碎裂、凋亡小体形成。与对照组相比细胞坏死率升高（P<0.05）。蟾蜍灵对程序性坏死途径关键蛋白RIP1的表达有促进作用（P<0.05）。结论 蟾蜍灵通过程序性坏死途径诱导SGC-7901细胞死亡。

Bufalin induces necroptosis in human gastric cancer SGC-7901 cells

Objective To investigate the necroptosis mechanism induced by Bufalin in human gastric cancer SGC-7901 cells. Methods SGC-7901 cells were cultured in vitro. The cells in the control group were cultured in RPMI 1640 medium, while those in the experimental group were exposed to various concentrations of Bufalin(50,100,150,200 nmol/L). After treatment for 48 h, the cell survival rate was assessed by MTT assay. Cell ultrastructures were observed by transmission electron microscope. Morphological changes in the nuclei were observed by DAPI fluorescence staining. Cellular necroptosis was assayed by flow cytometry. Western blotting was used to analyze the expression of receptor interacting protein kinase 1 (RIP1) in the necroptosis pathway. Results MTT assay showed that Bufalin had a significant effect on the growth of SGC-7901 cells(P<0.05). Transmission electron microscopy can be used to observe characteristic changes in cell necrosis, such as disruption of cell membranes, production of intracellular vacuoles, and disintegration of organelles. DAPI staining showed that after treatment with 100 nmol/L for 48 h, there were no obvious changes in cell apoptosis, such as nuclear condensation, nuclear fragmentation and formation of apoptotic bodies. The cell necrosis rate was higher than control group(P<0.05). Bufalin promotes the expression of RIP1, a key protein of necroptosis pathway(P<0.05). Conclusion Bufalin efficiently induced cell death through necroptosis pathway in SGC-7901 cells.