黄连碱对慢性萎缩性胃炎大鼠PI3K/Akt/mTOR信号通路的影响  

Effect of Coptisine on PI3K/Akt/mTOR Signaling Pathway in Chronic Atrophic Gastritis Rats

作　　者：王杰 杜朋丽 董佳琪 杨玥玮 高云霄 马虹宇[3] 贾雪梅 郭榆西 李博林[1] 杨倩[1] WANG Jie;DU Pengli;DONG Jiaqi;YANG Yuewei;GAO Yunxiao;MA Hongyu;JIA Xuemei;GUO Yuxi;LI Bolin;YANG Qian(Hebei Key Laboratory of Turbidity Toxin Syndrome,Key Laboratory of Integrated Chinese and Western Medicine for Gastroenterology Research(Hebei),Hebei Provincial Hospital of Traditional Chinese Medicine,Shijiazhuang 050011,China;Hebei University of Chinese Medicine,Shijiazhuang 050091,China;Hebei General Hospital,Shijiazhuang 050051,China)

机构地区：[1]河北省中医院,河北省浊毒证重点实验室,河北省中西医结合胃肠病研究重点实验室,石家庄050011 [2]河北中医药大学,石家庄050091 [3]河北省人民医院,石家庄050051

出　　处：《中国实验方剂学杂志》2024年第18期117-124,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家中医临床研究基地建设项目(国中医药办科技函[2018]131号);国家科技部重点研发项目(2018YFC1704100,2018YFC1704102);河北省省级科技计划资助项目(21377724D,21377740D);河北省中医药管理局科研计划项目(2021034,2022026,2022032,2023022);政府资助临床医学优秀人才培养项目(ZF2023162);河北省博士后科研项目择优资助项目(B2023003037)。

摘　　要：目的:基于磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/哺乳动物雷帕霉素靶蛋白(mTOR)信号通路探讨黄连碱对慢性萎缩性胃炎(CAG)大鼠的治疗作用及其作用机制。方法:采用水杨酸钠、N-甲基-N′-硝基-N-亚硝基胍(MNNG)及饥饱失常多因素诱导CAG大鼠模型,将造模成功的大鼠随机分为模型组,叶酸组,黄连碱高、低剂量组,黄连碱高、低剂量组分别灌以黄连碱(50、10 mg·kg^(-1)),叶酸组灌以叶酸2 mg·kg^(-1),干预60 d。苏木素-伊红(HE)染色法检测病理学变化;电镜观察胃黏膜细胞超微结构;免疫比浊法检测大鼠血清胃蛋白酶原Ⅰ(PGⅠ)、胃蛋白酶原Ⅱ(PGⅡ)及计算PGⅠ/PGⅡ(PGR);放射免疫法检测大鼠血清胃泌素-17(G-17)水平;酶联免疫吸附测定法(ELSIA)检测大鼠血清中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)的含量;蛋白免疫印迹法(Western blot)检测胃黏膜转化生长因子-β_(1)(TGF-β_(1))、PI3K、p-Akt、mTOR和人第10号染色体缺失的磷酸酶及张力蛋白同源基因(PTEN)蛋白表达含量;实时荧光定量聚合酶链式反应(Realtime PCR)检测TGF-β_(1)、PI3K、Akt、mTOR、PTEN、微管相关蛋白轻链3Ⅱ(LC3Ⅱ)和酵母Atg6同源物-1(Beclin-1)mRNA水平。结果:与正常组比较,模型组胃黏膜组织固有腺体萎缩,数量减少,炎性细胞浸润;模型组胃黏膜细胞超微结构细胞核固缩,线粒体数量少,肿胀,结构异常;模型组血清G-17、PGⅠ、PGR及胃组织PTEN蛋白及mRNA显著降低(P<0.01),血清IL-6、IL-1β、TNF-α及胃组织TGF-β_(1)、PI3K、Akt和mTOR蛋白及mRNA显著升高(P<0.01)。与模型组比较,各药物组可以不同程度改善CAG大鼠病理损伤和胃黏膜细胞超微结构,明显升高血清G-17、PGⅠ、PGR表达水平(P<0.05,P<0.01),明显降低IL-6、IL-1β、TNF-α水平(P<0.05,P<0.01),黄连碱高剂量组可以明显下调TGF-β_(1)、PI3K、Akt和mTOR蛋白及mRNA(P<0.05,P<0.01)。结论:黄连碱对CAG大鼠�Objective To investigate the therapeutic effect and mechanism of coptisine on chronic atrophic gastritis(CAG)in rats based on the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapamycin(mTOR)signaling pathway.Method A CAG rat model was induced by multiple factors,including sodium salicylate,N-methyl-N′-nitro-N-nitrosoguanidine(MNNG),and irregular feeding.The successfully modeled rats were randomly divided into the model group,folic acid group,and high-and low-dose coptisine groups.The high-and low-dose coptisine groups were given coptisine(50,10 mg·kg^(-1),respectively),and the folic acid group was given folic acid at 2 mg·kg^(-1) for 60 days.The pathological changes were detected by hematoxylin-eosin(HE)staining.The ultrastructure of gastric mucosal cells was observed by electron microscopy.Serum pepsinogenⅠ(PGⅠ),pepsinogenⅡ(PGⅡ),and PGⅠ/PGⅡratio(PGR)were detected by immunoturbidimetry.Serum gastrin-17(G-17)level was detected by radioimmunoassay.The content of interleukin-6(IL-6),tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)in serum of rats was detected by enzyme-linked immunosorbent assay(ELSIA).Western blot analysis was used to detect the expression levels of TGF-β_(1),PI3K,phosphorylated-Akt(p-Akt),mTOR,and phosphatase and tensin homolog deleted on chromosome 10(PTEN)in gastric mucosa.The mRNA levels of TGF-β_(1),PI3K,Akt,mTOR,PTEN,microtubule-associated protein light chain 3Ⅱ(LC3Ⅱ),and Beclin-1 were detected by real-time quantitative polymerase chain reaction(Real-time PCR).Result Compared with the normal group,the model group showed atrophy and reduced number of intrinsic glands in the gastric mucosal tissues,as well as inflammatory cell infiltration.The ultrastructure of gastric mucosal cells in the model group displayed nuclear condensation,reduced and swollen mitochondria,and abnormal structure.The serum levels of G-17,PGⅠ,PGR,and the protein and mRNA levels of PTEN in gastric tissues were significantly lower in the model group(P<0.01)

关 键 词：慢性萎缩性胃炎 黄连碱 磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/哺乳动物雷帕霉素靶蛋白(mTOR) 炎症 胃蛋白酶原 

分 类 号：R22[医药卫生—中医基础理论] R242[医药卫生—中医学] R285.5R573.32