蟾毒灵通过NLRP3/CASP1/GSDMD信号通路诱导卵巢癌细胞焦亡并增强其对顺铂的敏感性OA
Bufalin induces pyroptosis in ovarian cancer cells via the NLRP3/CASP1/GSDMD signaling pathway and enhances their sensitivity to cisplatin
目的:探究蟾毒灵(Buf)对卵巢癌细胞A2780和SKOV3的毒性作用及其机制,评估其与顺铂(DDP)联合应用的协同毒性.方法:常规培养A2780和SKOV3细胞.用CCK-8法、克隆形成实验检测Buf对A2780和SKOV3细胞增殖的影响及半数抑制浓度(IC50).用不同浓度的Buf处理细胞,显微镜下观察细胞形态变化,比色法检测乳酸脱氢酶(LDH)释放水平,流式细胞术分析检测Annexin Ⅴ+PI+细胞比例,qPCR和WB法检测细胞焦亡相关分子mRNA及蛋白表达水平,通过小干扰RNA敲减A2780和SKOV3细胞中的GSDMD并进行后续功能实验进行验证.通过SynergyFinder分析Buf与DDP对于杀伤A2780和SKOV3细胞的协同效应,并通过流式细胞术、qPCR、WB进行机制探索,进一步利用流式细胞术检测Buf与DDP联用对DDP耐药A2780和SKOV3细胞的影响.结果:与对照组相比,Buf显著抑制卵巢癌细胞增殖及迁移能力(均P<0.05),诱导LDH释放和细胞焦亡(均P<0.05),上调核苷酸结合寡聚化结构样受体热蛋白结构域相关蛋白3(NLRP3)、半胱氨酸天冬氨酸蛋白水解酶1(CASP1)、gasdermin家族成员D(GSDMD)、白细胞介素-1β(IL-1β)及IL-18的mRNA及蛋白的表达(均P<0.05),Buf与DDP联用具有协同抑制A2780和SKOV3细胞增殖的作用,并可进一步增强DDP耐药的A2780和SKOV3细胞对DDP的敏感性(P<0.01或P<0.001).结 论:Buf可通过激活NLRP3/CASP1/GSDMD信号通路诱导A2780和SKOV3细胞焦亡,并增强A2780和SKOV3细胞及DDP耐药A2780和SKOV3细胞对DDP的敏感性.
Objective:To investigate the cytotoxic effects and underlying mechanisms of Bufalin(Buf)on ovarian cancer cells(A2780 and SKOV3)and to evaluate its synergistic cytotoxicity with cisplatin(DDP).Methods:A2780 and SKOV3 cells were routinely cultured.The effect of Buf on cell proliferation and its half-maximal inhibitory concentration(IC50)were determined using CCK-8 assay and colony formation assay.Following treatments with various concentrations of Buf,morphological changes of tumor cells were observed under a microscope,lactate dehydrogenase(LDH)release was measured using colorimetric assay,and the proportion of Annexin Ⅴ⁺PI⁺ cells was analyzed using flow cytometry.The mRNA and protein expression levels of pyroptosis-related molecules were detected using qPCR and WB,respectively.Small interfering RNA was used to knock down gasdermin D(GSDMD)in A2780 and SKOV3 cells to validate its functional role.The synergistic cytotoxic effects of Buf and DDP against A2780 and SKOV3 cells were analyzed using the SynergyFinder platform,and the underlying mechanisms were further explored using flow cytometry,qPCR,and WB.Additionally,the synergistic effect of Buf and DDP on DDP-resistant A2780 and SKOV3 cells was evaluated using flow cytometry.Results:Compared with the control group,Buf significantly inhibited proliferation and migration of ovarian cancer cells(both P<0.05),induced LDH release and cell pyroptosis(both P<0.05),and upregulated the mRNA and protein expression levels of nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3(NLRP3),cysteine-aspartic acid protease 1(CASP1),gasdermin D(GSDMD),interleukin(IL)-1β,and IL-18(all P<0.05).The combination of Buf and DDP synergistically inhibited the proliferation of A2780 and SKOV3 cells and further enhanced the sensitivity of DDP-resistant A2780 and SKOV3 cells to DDP.Conclusion:Buf induces pyroptosis in A2780 and SKOV3 cells by activating the NLRP3/CASP1/GSDMD signaling pathway and enhances the sensitivity of DDP-resistant A2780 and SKOV3 cells to DDP.
李婕;奚玲;赵璨
华中科技大学同济医学院附属同济医院 妇产科,湖北 武汉 430030||华中科技大学同济医学院附属同济医院 肿瘤侵袭与转移教育部重点实验室,湖北 武汉 430030华中科技大学同济医学院附属同济医院 妇产科,湖北 武汉 430030||华中科技大学同济医学院附属同济医院 肿瘤侵袭与转移教育部重点实验室,湖北 武汉 430030华中科技大学同济医学院附属同济医院 病理研究所,湖北 武汉 430030
医药卫生
卵巢癌细胞焦亡蟾毒灵NLRP3/CASP1/GSDMD信号通路
ovarian cancerpyroptosisbufalin(Buf)NLRP3/CASP1/GSDMD signaling pathway
《中国肿瘤生物治疗杂志》 2026 (3)
252-261,10
国家自然科学基金(82172717)
评论