归芪白术方联合奥沙利铂通过调节VIP/cAMP/PKA/AQPs信号通路保护胃癌荷瘤小鼠肠道屏障  

Guiqi Baizhu Prescription Combined with Oxaliplatin Protects Intestinal Barrier of Tumor-bearing Mice with Gastric Cancer by Regulation of VIP/cAMP/PKA/AQP Signaling Pathway

作　　者：董焕成 苏韫 龚红霞 曹旺杰 和建政 刘永琦 张晗 曾元丁 李从艺 康倩 DONG Huancheng;SU Yun;GONG Hongxia;CAO Wangjie;HE Jianzheng;LIU Yongqi;ZHANG Han;ZENG Yuanding;LI Congyi;KANG Qian(School of Basic Medicine,Gansu University of Chinese Medicine,Lanzhou 730000,China;Provincial Key Laboratory of Molecular Medicine and Traditional Chinese Medicine(TCM)Prevention and Treatment of Major Diseases in Colleges and Universities of Gansu Province,Lanzhou 730000,China;Key Laboratory of TCM Prevention and Treatment of Chronic Diseases in Gansu Province,Lanzhou 730000,China;NHC Key Laboratory of Diagnosis and Therapy of Gastrointestinal Tumor,Gansu Provincial Hospital,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学基础医学院,兰州730000 [2]甘肃省高校重大疾病分子医学与中医药防治研究省级重点实验室,兰州730000 [3]甘肃省中医药防治慢性疾病重点实验室,兰州730000 [4]国家卫生健康委胃肠肿瘤诊治重点实验室,甘肃省人民医院,兰州730000

出　　处：《中国实验方剂学杂志》2023年第5期129-136,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：甘肃省“双一流”科研重点项目(GSSYLXM-05);中国医学科学院中央级公益性科研院所基本科研业务费专项资金项目(2019PT320005);甘肃省高效中(藏)药化学与质量研究重点实验室开放基金项目(zzy-2022-06);甘肃省中医药研究中心专项(zyzx-2020-zx18)。

摘　　要：目的:探讨归芪白术方联合奥沙利铂通过血管活性肠肽(VIP)/环磷酸腺苷(cAMP)/蛋白激酶A(PKA)信号通路调控下游水通道蛋白3(AQP3)和水通道蛋白4(AQP4)从而保护胃癌荷瘤小鼠肠道屏障的作用。方法:将密度为1×10^(7)个/mL的胃癌细胞株MFC制成细胞悬液,经荷瘤小鼠右腋下接种细胞悬液0.2 mL,构建胃癌荷瘤小鼠模型,造模成功后将小鼠随机分为5组,模型组、奥沙利铂组(10mg·kg^(-1))、奥沙利铂加归芪白术方高、中、低剂量组(17.68、8.84、4.42 g·kg^(-1)),每组10只,另外余10只作为空白组。各组小鼠经灌胃或腹腔注射中药、奥沙利铂或生理盐水,治疗14 d。末次给药后,次日眼球取血分离血清并取其结肠样本。苏木素-伊红(HE)染色观察组织形态的变化。酶联免疫吸附测定法(ELISA)检测血清中D-乳酸(D-LA)、二胺氧化酶(DAO)含量;实时荧光定量聚合酶链式反应(Real-time PCR)及蛋白免疫印迹法(Western blot)分别检测各组小鼠结肠组织中VIP、cAMP、PKA、AQP3和AQP4 mRNA及蛋白的表达。结果:与正常组比较,模型组黏膜下层水肿,黏膜层中肠腺排列紊乱,杯状细胞缺失,可见大量炎性细胞浸润及绒毛脱落的现象。而各给药组均有不同程度的改善;与正常组比较,模型组血清中DAO和D-LA水平均显著上升(P<0.01),与模型组比较,联合用药高剂量组DAO和D-LA水平及联合用药中剂量组D-LA水平均有所下降(P<0.05,P<0.01),与奥沙利铂组比较,联合用药高、中剂量组D-LA水平有所下降(P<0.05),其余组DAO和D-LA表达水平也有所下降,但差异无统计学意义;与正常组比较,模型组小鼠的VIP、cAMP、PKA、AQP3和AQP4 mRNA及蛋白表达水平明显降低(P<0.05,P<0.01);与模型组比较,各给药组小鼠的VIP、cAMP、PKA、AQP3和AQP4 mRNA及蛋白表达水平均有所升高,其中联合用药高剂量组VIP、cAMP、PKA、AQP3和AQP4 mRNA及蛋白表达水平明显升高(P<0.05,P<0.01),联合用药中剂量组cAMPObjective:To investigate the protective effect of Guiqi Baizhu prescription combined with oxaliplatin on the intestinal barrier of tumor-bearing mice with gastric cancer by regulating downstream aquaporin 3(AQP3) and aquaporin 4(AQP4) through the vasoactive intestinal peptide(VIP)/cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA) signaling pathway. Method:The gastric cancer cell lines MFC with a density of 1×10^(7)/mL were prepared into cell suspension. The tumor-bearing mouse model of gastric cancer was established by inoculating 0.2 mL cell suspension under the right axilla of mice. After successful modeling, mice were randomly divided into 5 groups, namely, model group, oxaliplatin group(10 mg·kg^(-1)), and high, medium, and low-dose oxaliplatin + Guiqi Baizhu prescription groups(17.68, 8.84, 4.42 g·kg^(-1)), with 10 mice in each group, and the remaining 10 mice were set as a blank group. Mice in each group were treated with Chinese medicine, oxaliplatin, or normal saline by gavage or intraperitoneal injection for 14 d. The next day after the last dose, blood was taken from the eyeball to separate serum and take colonic samples.Hematoxylin-eosin(HE) staining was used to observe the changes in tissue morphology. The content of Dlactate acid(D-LA) and diamine oxidase(DAO) in the serum was determined by enzyme-linked immunosorbent assay(ELISA). The mRNA and protein expressions of VIP, cAMP, PKA, AQP3, and AQP4 were detected by Real-time quantitative polymerase chain reaction(Real-time PCR) and Western blot, respectively. Result:Compared with the blank group, the model group showed edema in the colonic submucosa, disordered arrangement of intestinal glands in the mucosal layer, loss of goblet cells, infiltration of inflammatory cells, and villus shedding. However, there were different degrees of improvement in each administration group. As compared with the blank group, the serum levels of DAO and D-LA in the model group were significantly increased(P<0.01). As compared with the model group, the levels

关 键 词：归芪白术方 胃癌 肠道屏障损伤 血管活性肠肽(VIP)/环磷酸腺苷(cAMP)/蛋白激酶A(PKA)信号通路 水通道蛋白3(AQP3) AQP4 

分 类 号：R2-0[医药卫生—中医学] R22R285.5R289R33