苦参碱调控抑制肠上皮细胞凋亡影响脓毒症诱导的肠损伤研究  

Matrine Influences Sepsis-Induced Intestinal Injury by Regulating Intestinal Epithelial Cells Apoptosis

作　　者：陈林[1] 徐玉峰 范永强[1] 樊均明[1] 陈朝晖[1] 冷潇 Chen Lin;Xu Yufeng;Fan Yongqiang;Fan Junming;Chen Zhaohui;Leng Xiao(The First Affiliated Hospital of Chengdu Medical College,Chengdu 610500,China;Experimental Center,Chengdu Medical College,Chengdu 610500,China)

机构地区：[1]成都医学院第一附属医院,成都610500 [2]成都医学院实验中心,成都610500

出　　处：《成都医学院学报》2025年第2期202-206,共5页Journal of Chengdu Medical College

基　　金：四川省中医药管理局一般项目(No:2020LC0074);第七批全国老中医药学术继承项目(No:YS000126)。

摘　　要：目的 探究苦参碱能否改善脓毒症相关的肠损伤。方法 将Caco-2细胞分为对照组(Con)、脂多糖刺激组(LPS)和苦参碱低、中、高剂量组(50、100、200μmol/L)。相应刺激各组细胞24 h后,采用Hoechst 33258染色观察细胞凋亡并计算细胞凋亡率;采用蛋白质印迹法检测BAX、BCL2、剪切型caspase-3和β-actin的表达情况;采用ROS活性探针染色检测细胞内活性氧(ROS)水平。50只雄性小鼠随机分为假伤组(Sham)、模型组(CLP)和苦参碱低、中、高剂量组(25、50、100 mg/kg),每组各10只,采用盲肠结扎穿孔手术(CLP)构建小鼠脓毒症模型,造模成功,按分组给予相应药物连续治疗24 h后,采用流式细胞术检测小肠细胞凋亡情况;采用ELISA法测定肠脂肪酸结合蛋白(I-FABP)、D-乳酸、白介素-6(IL-6)、肿瘤坏死因子α(TNF-α)与IL-1β的含量。60只雄性小鼠随机分为Sham、CLP和大剂量苦参碱治疗组(CLP+苦参碱100 mg/kg),每组各20只,造模成功后给予相应药物连续治疗,记录小鼠死亡情况并计算各组小鼠的7 d死亡率。结果 与Con组相比,LPS组细胞凋亡率增高(P<0.05),BAX与剪切型caspase-3表达增多(P<0.05),而BCL2表达下调(P<0.05),苦参碱组可逆转上述改变,并呈现出浓度依赖性(P<0.05);同时,与LPS组相比,苦参碱组细胞内ROS水平也随给药浓度的提高而逐渐降低;与Sham组相比,CLP组血清I-FABP、D-乳酸、IL-6、TNF-α与IL-1β的浓度增高且小肠细胞凋亡增多(P<0.05),而苦参碱组的治疗可减少血浆中上述因子含量并减少细胞凋亡(P<0.05),同样呈现出浓度依赖性;与Sham组相比,CLP组死亡率提高(P<0.001),而给予大剂量的苦参碱治疗可提高脓毒症小鼠的存活率(P<0.05)。结论 苦参碱可通过缓和脓毒症时小肠细胞凋亡,进而改善脓毒症相关的肠损伤。Objective To investigate whether matrine can improve sepsis-related intestinal injury.Methods Caco-2 cells were divided into Con group,lipopolysaccharide(LPS)group and Matrine low-,medium-,and high-dose(50,100,200μmol/L)groups.After 24 h of corresponding stimulation,cell apoptosis was visualized by Hoechst 33258 staining and apoptosis rate was calculated.The expression of BAX,B-cell lymphoma 2(BCL2),cleaved caspase-3 andβ-actin were detected by western blot.Intracellular reactive oxygen species(ROS)levels were measured by ROS activity probe staining.Fifty male mice were randomly and averagely divided into Sham group,cecal ligation and puncture(CLP)group and Matrine low-,medium-and high-dose(25,50,100 mg/kg)groups,with 10 mice in each group.And sepsis model was constructed by CLP surgery.After successful modeling,mice in each group were administrated the corresponding drugs for 24 h.And then,the apoptosis of intestinal epithelial cells was detected by flow cytometry.Enzyme-linked immunosorbent assay(ELISA)was used to detect the level of intestinal fatty acid binding protein(I-FABP),D-lactate,interleukin-6(IL-6),tumor necrosis factorα(TNF-α)and interleukin-1β(IL-1β).Sixty male mice were randomly assigned to Sham,CLP and high-dose Matrine treatment(CLP+Matrine 100 mg/kg)groups,with 20 mice in each group.After successful modeling,the mice were treated with corresponding drugs.The death of mice was recorded and the 7-d mortality of each group was calculated.Results Compared with the Con group,the LPS group exhibited higher cell apoptosis rate(P<0.05),increased expression of BAX and cleaved caspase-3(P<0.05),but down-regulated BCL2 expression(P<0.05);the Matrine group could reverse the above changes and presented a concentration-dependence(P<0.05).Meanwhile,the intracellular ROS level also gradually decreased with the increasing administration concentration in the Matrine group compared with the LPS group.Compared with the Sham group,the concentration of serum I-FABP,D-lactate,IL-6,TNF-αand IL-1βand cell apopto

关 键 词：脓毒症 肠损伤 苦参碱 细胞凋亡 活性氧 

分 类 号：R285[医药卫生—中药学]