清热化瘀健脾方抑制小鼠结肠炎相关性结肠癌的作用机制  

Mechanism of Qingre Huayu Jianpi Prescription Inhibiting Development of Colitis-associated Colorectal Cancer in Mice

作　　者：郑燕秋 游弋晖 柯俊羽 宋锦彬 吴永强 刘昌辉[3] 李燕舞[1] ZHENG Yanqiu;YOU Yihui;KE Junyu;SONG Jinbin;WU Yongqiang;LIU Changhui;LI Yanwu(Science and Technology Innovation Center,Guangzhou University of Chinese Medicine,Guangzhou 510405,China;Gaozhou Hospital of Traditional Chinese Medicine Affiliated to Guangzhou University of Chinese Medicine,Gaozhou 525200,China;School of Pharmaceutical Sciences,Guangzhou University of Chinese Medicine,Guangzhou 510006,China)

机构地区：[1]广州中医药大学科技创新中心,广州510405 [2]广州中医药大学附属高州中医院,广东高州525200 [3]广州中医药大学中药学院,广州510006

出　　处：《中国实验方剂学杂志》2024年第8期83-90,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(82274600);广州市科学技术局民生科技项目(202002020032);茂名市科技创新专题(2022S014)。

摘　　要：目的:探讨清热化瘀健脾方(QHJ)对小鼠结肠炎相关性结肠癌(CAC)的干预作用及相关机制。方法:将C57BL/6小鼠随机分为4组:正常组、模型组、QHJ低剂量(QHJ-L,10 g·kg^(-1))组、QHJ高剂量(QHJ-H,40 g·kg^(-1))组,除正常组外,采用氧化偶氮甲烷(AOM)和葡聚糖硫酸钠(DSS)联合化学诱导CAC小鼠模型,造模时间14周。各药物组于第5周开始按剂量灌胃给药,每天1次至14周结束,正常及模型组平行给予等体积水。评估各组小鼠的生存率、结肠长度、质量及病理形态改变;采用蛋白免疫印迹法(Western blot)检测Wnt信号蛋白3a(Wnt3a)、β-连环蛋白(β-catenin)、非磷酸化β-连环蛋白(Non-p-β-catenin)、胆固醇结合糖蛋白133(CD133)表达;免疫组化(IHC)检测分化簇80(CD80)、CD11抗原样家族成员B(CD11b)蛋白的定位及表达。分离培养CAC小鼠来源的结肠类器官,检测Wnt信号通路相关蛋白表达。结果:QHJ可明显改善CAC小鼠生存率,抑制结肠瘤状体个数。与正常组比较,模型组小鼠结肠组织中Wnt3a、β-catenin、p-β-catenin、CD133蛋白表达水平均明显升高(P<0.05,P<0.01);与模型组比较,QHJ-L组Wnt3a、β-catenin水平显著降低(P<0.01),QHJ-H组Wnt3a、β-catenin、Non-p-β-catenin和CD133蛋白水平明显下降(P<0.05,P<0.01)。与正常组比较,模型组小鼠结肠中CD11b表达水平明显升高,而CD80水平明显降低(P<0.05,P<0.01);与模型组比较,QHJ-L、QHJ-H组CD11b均明显降低,CD80均明显升高(P<0.05,P<0.01)。CAC模型小鼠结肠类器官中Non-p-β-catenin、CD133蛋白的表达均显著升高;而QHJ干预后可抑制模型小鼠结肠类器官Non-p-β-catenin、CD133蛋白的表达(P<0.01)。结论:QHJ可抑制CAC模型小鼠结肠炎-癌进程,其机制可能与调节微环境并抑制Wnt信号过度激活有关。Objective:To explore the effect of Qingre Huayu Jianpi prescription(QHJ)on colitisassociated colorectal cancer(CAC)in mice,and its related mechanism.Method:C57BL/6 mice were randomly divided into four groups including the normal,model,QHJ low-dose(QHJ-L,10 g·kg^(-1)),and QHJ high-dose(QHJ-H,40 g·kg^(-1))groups.Azoxymethane(AOM)and dextran sodium sulfate(DSS)were combined to chemically build a CAC mouse model for 14 weeks.Each drug group was given intragastrically from the 5th week to the 14th week,once per day.An equal volume of water was fed to the normal and model groups.The mouse survival rate,colon length,weight,and pathological alterations were assessed.The protein expressions of Wnt-3a protein signaling(Wnt3a),β-catenin,Non-phosphor-β-catenin(Non-p-β-catenin),and cholesterol-binding glycoproteins 133(CD133)were detected by Western blot.The localization and expression of the cluster of differentiation(CD)80 and CD11 antigen-like family member B(CD11b)were detected by immunohistochemistry(IHC).The colon organoids derived from CAC mice were isolated and cultured to detect the expression of Wnt signaling pathway-related proteins.Result:The survival rate of the CAC mice was improved by QHJ treatment and the number of colon tumors was inhibited significantly.Compared with those in the normal group,the expression levels of Wnt3a,β-catenin,Non-p-β-catenin,and CD133 in colon tissues in the model group were significantly increased(P<0.05,P<0.01).Compared with those in the model group,the levels of Wnt3a andβ-catenin in the QHJ-L group were significantly decreased(P<0.01),and the protein levels of Wnt3a,β-catenin,Non-p-β-catenin,and CD133 in the QHJ-H group were significantly decreased(P<0.05,P<0.01).Meanwhile,the expression level of CD11b in the model group was significantly increased compared with that in the normal group while the CD80 level was significantly decreased(P<0.05,P<0.01).Compared with those in the model group,CD11b in QHJ-L and QHJ-H groups was significantly decreased,and CD80 was significant

关 键 词：结肠炎相关性结肠癌 清热化瘀健脾方 WNT信号通路 结肠类器官 

分 类 号：R2-0[医药卫生—中医学] R22R242R285.5R735