红景天苷缓解高糖合并乙醇暴露加剧的胰岛细胞瘤细胞焦亡  

Salidroside alleviates high glucose and ethanol-induced pyroptosis in insulinoma cells

作　　者：李晓晗 郭旭 陈仕琦 高艳果 周俊 张永红 王启斌 陈黎[1,2,3] 郑涛 LI Xiao-han;CUO Xu;CHEN Shi-qi;GAO Yan-guo;ZHOU Jun;ZHANG Yong-hong;WANG Qi-bin;CHEN Li;ZHENG Tao(School of Pharmaceutical Sciences,Hubei University of Medicine,Shiyan 442000,China;Institute of Wudang Traditional Chinese Medicine,Taihe Hospital,Hubei University of Medicine,Shiyan 442000,China;Department of Pharmacy,Taihe Hospital,Hubei University of Medicine,Shiyan 442000,China)

机构地区：[1]湖北医药学院药学院,湖北十堰442000 [2]湖北医药学院附属太和医院武当中医药研究所,湖北十堰442000 [3]湖北医药学院附属太和医院药学部,湖北十堰442000

出　　处：《中国中药杂志》2024年第22期6181-6189,共9页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金青年科学基金项目(81703582);湖北省自然科学基金项目(2020CFB713,2016CFB153);湖北省卫生健康科研基金项目(WJ2021ZH0017,WJ2019M057);“十四五”湖北省高等学校优势特色学科群(生物与医药)项目(2024BMXKQT3,2023BMXKQT3,2022BMXKQT3);湖北医药学院研究生科技创新项目(YC2022056,YC2023068,YC2023069);吴阶平医学基金会临床科研专项(320.6750.2023-25-7)。

摘　　要：该研究以高糖(HG)刺激大鼠胰岛细胞瘤细胞(INS-1)建立焦亡损伤模型,观察额外的乙醇(ET)暴露对细胞焦亡的影响,并对红景天苷(SAL)的干预作用进行研究。培养INS-1,分别设立正常对照(NG)组、HG组、HG+ET(100 mmol·L^(-1))组和HG+ET+SAL(1~100μmol·L^(-1))组。各组处理72 h后,利用cell counting kit-8(CCK-8)法检测细胞活力变化;使用显微镜观察焦亡小体形成数量变化;采用Western blot法检测细胞内Nod样受体蛋白3(NLRP3)/消皮素D(GSDMD)信号和腺苷酸活化蛋白激酶(AMPK)活性的变化;利用荧光探针检测细胞内活性氧簇(ROS)水平变化;使用时间分辨-荧光共振能量转移(TR-FRET)技术在体外观察SAL对重组AMPK蛋白激酶活性的影响。结果显示同NG相比,HG暴露可诱导INS-1焦亡小体数量增多、ROS水平上升和NLRP3/GSDMD信号激活;同HG组相比,HG+ET暴露可进一步加剧上述变化;同HG+ET组相比,SAL可浓度依赖性增加细胞活力、减少INS-1焦亡小体形成,并抑制ROS生成、NLRP3/GSDMD信号过度活化及AMPK活性下降;TR-FRET实验结果表明SAL可直接激活AMPK;使用AMPK抑制剂预处理INS-1后,SAL增加细胞活力、缓解INS-1焦亡小体形成和抑制ROS过度产生的作用消失。上述结果表明SAL可通过激活AMPK缓解HG合并ET暴露加剧的INS-1焦亡。This study established a pyroptosis injury model by stimulating insulinoma cells(INS-1)of rats with high glucose(HG)and observed the impact of additional ethanol(ET)exposure on cell pyroptosis,as well as the intervention effect of salidroside(SAL).INS-1 cells were cultured and divided into a normal control group(NG),an HG group,an HG+ET(100 mmol·L^(-1))group,and an HG+ET+SAL(1-100μmol·L^(-1))group.After 72 hours of treatment,cell viability was assessed using the cell counting kit-8(CCK-8)assay.The number of pyroptotic bodies was observed under a microscope.Western blot was used to detect changes in the intracellular Nod-like receptor protein 3(NLRP3)/gasdermin D(GSDMD)signaling pathway and adenosine monophosphate-activated protein kinase(AMPK)activity.A fluorescence probe was used to detect changes in intracellular reactive oxygen species(ROS)levels.Time-resolved fluorescence resonance energy transfer(TR-FRET)technology was employed to observe the effect of SAL on recombinant AMPK protein kinase activity in vitro.The results showed that compared to the NG group,HG exposure induced an increase in the number of pyroptotic bodies,elevated ROS levels,and activation of the NLRP3/GSDMD signaling pathway in INS-1 cells.Compared to the HG group,HG+ET exposure further exacerbated these changes.Compared to the HG+ET group,SAL dose-dependently increased cell viability,reduced the formation of pyroptotic bodies in INS-1 cells,and inhibited excessive ROS production,overactivation of the NLRP3/GSDMD signaling pathway,and the decrease in AMPK activity.TR-FRET experiments indicated that SAL could directly activate AMPK.When INS-1 cells were pretreated with an AMPK inhibitor,the effects of SAL on increasing cell viability,alleviating the formation of pyroptotic bodies,and inhibiting excessive ROS production were abolished.These results suggest that SAL can alleviate HG combined with ET-induced exacerbation of INS-1 pyroptosis by activating AMPK.

关 键 词：红景天苷 糖尿病 乙醇暴露 INS-1 焦亡 

分 类 号：R285[医药卫生—中药学]