安胃汤含药血清对胃黏膜肠上皮化生模型细胞自噬及凋亡的影响  

Effect of Anwei Decoction-Containing Serum on Autophagy and Apoptosis of Gastric Intestinal Metaplasia Model Cells

作　　者：胡鑫 唐友明[2] 郑景辉[1] 莫云秋[2] 梁柳观 莫少丹 HU Xin;TANG Youming;ZHENG Jinghui;MO Yunqiu;LIANG Liuguan;MO Shaodan(Guangxi University of Chinese Medicine,Nanning 530200 Guangxi,China;Ruikang Hospital Affiliated to Guangxi University of Chinese Medicine,Nanning 530011 Guangxi,China)

机构地区：[1]广西中医药大学,广西南宁530200 [2]广西中医药大学附属瑞康医院,广西南宁530011

出　　处：《中药新药与临床药理》2024年第4期469-476,共8页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金项目(8186150436)。

摘　　要：目的探讨安胃汤(半夏、黄连、干姜、乌药、百合等)含药血清对胃黏膜肠上皮化生(Gastric intestinalmetaplasia,GIM)模型细胞自噬及凋亡的影响。方法采用鹅去氧胆酸(CDCA)诱导人胃黏膜上皮细胞GES-1,制备GIM细胞模型。实验分为正常组(正常胃黏膜上皮细胞GES-1常规培养)、模型组(经CDCA复制成GIM模型后常规培养)、空白对照组(GIM模型细胞以等量空白血清培养)、安胃汤组(GIM模型细胞+7%安胃汤含药血清),各组细胞干预24 h后检测。采用流式细胞术(Annexin V-PE/7-AAD双染法)检测细胞凋亡;免疫荧光双标法检测肠上皮化生相关蛋白SOX2、CDX2的表达情况;透射电镜观察细胞超微结构及自噬情况;qRT-PCR法检测细胞LC3、Bax mRNA表达情况;Western Blot法检测细胞MUC2、KLF4、LC3、Bax蛋白表达情况。结果与正常组比较,模型组细胞的MUC2、KLF4蛋白表达显著上调(P<0.01),细胞的凋亡率明显降低(P<0.05);SOX2荧光强度明显降低(P<0.05),CDX2荧光强度显著增高(P<0.01);细胞结构紊乱,细胞核萎缩畸形,内质网和线粒体等细胞器肿胀破裂及空泡化严重,自噬小体及自噬溶酶体明显增多;Bax mRNA及蛋白表达明显下调(P<0.05),LC3 mRNA表达显著上调(P<0.01),LC3Ⅱ/LC3Ⅰ蛋白表达比值显著升高(P<0.01)。与空白对照组比较,安胃汤组细胞的凋亡率显著增高(P<0.01);SOX2荧光强度显著增高(P<0.01),CDX2荧光强度明显降低(P<0.05);细胞结构较完整,镜下可见自噬小体和自噬溶酶体略有减少;Bax mRNA及蛋白表达显著上调(P<0.01),LC3 mRNA表达显著下调(P<0.01),LC3Ⅱ/LC3Ⅰ蛋白表达比值显著降低(P<0.01)。结论安胃汤可能通过调节GIM模型细胞自噬,并促进其凋亡,从而改善胃黏膜上皮细胞的肠上皮化生情况。To investigate the effects of Anwei Decoction-containing serum(Pinelliae Rhizoma,Coptidis Rhizoma,Zingiberis Rhizoma Recens,Linderae Radix,Lilii Bulbus,etc.)on autophagy and apoptosis of gastric intestinal metaplasia(GIM)model cells.Methods GIM cell model was prepared by inducing human gastric mucosal epithelial cell GES-1 with chenodeoxycholic acid(CDCA).The experiment was divided into normal group(normal gastric mucosal epithelial GES-1 cells were cultured routinely),model group(GIM model was replicated by CDCA and cultured routinely),blank control group(GIM model cells were cultured with the same amount of blank serum),and Anwei Decoction group(GIM model cells+7%Anwei Decoction-containing serum).The cells in each group were detected after 24 hours of intervention.Apoptosis was detected by flow cytometry(Annexin V-PE/7-AAD double staining).The expressions of intestinal metaplasia-related proteins SOX2 and CDX2 were detected by immunofluorescence double labeling method.The ultrastructure and autophagy of cells were observed by transmission electron microscopy.The mRNA expressions of LC3 and Bax were detected by qRT-PCR.The protein expressions of MCU2,KLF4,LC3 and Bax were detected by Western Blot.Results Compared with the normal group,the protein expressions of MUC2 and KLF4 in the model group were significantly up-regulated(P<0.01),and the apoptosis rate of the cells was significantly decreased(P<0.05).The fluorescence intensity of SOX2 was significantly decreased(P<0.05),and the fluorescence intensity of CDX2 was significantly increased(P<0.01).The cell structure was disordered,the nucleus was atrophic and deformed,the organelles such as endoplasmic reticulum and mitochondria were swollen,ruptured and vacuolated,and the number of autophagosomes and autolysosomes were significantly increased.The mRNA and protein expressions of Bax were significantly down-regulated(P<0.05),the mRNA expression of LC3 was significantly up-regulated(P<0.01),and the protein expression ratio of LC3 II/LC3 I was significantly increase

关 键 词：安胃汤含药血清 胃黏膜肠上皮化生 GES-1细胞 自噬 凋亡 慢性萎缩性胃炎 胃癌前病变 

分 类 号：R285.5[医药卫生—中药学]