黄芩素对糖尿病小鼠全层皮肤缺损创面愈合的影响及其机制  

Effects and mechanism of baicalin on wound healing of full-thickness skin defects in diabetic mice

作　　者：施彦 易亮 张伟强 刘妮可 文辉才[1] 杨荣华 Shi Yan;Yi Liang;Zhang Weiqiang;Liu Nike;Wen Huicai;Yang Ronghua(Department of Plastic,Medical Center of Burn Plastic and Wound Repair,the First Affiliated Hospital,Jiangxi Medical College,Nanchang University,Nanchang330006,China;Department of Burn,Plastic Surgery and Wound Repair,the Second Affiliated Hospital(Guangzhou First People's Hospital),South China University of Technology,Guangzhou510180,China)

机构地区：[1]南昌大学第一附属医院烧伤整形与创面修复医学中心,南昌330006 [2]华南理工大学附属第二医院(广州市第一人民医院)烧伤整形美容与创面修复科,广州510180

出　　处：《中华烧伤与创面修复杂志》2024年第11期1085-1094,共10页Chinese Journal of Burns And Wounds

基　　金：国家自然科学基金面上项目(82060350,82272276);中国高校产学研创新基金(2021JH028);广东省基础与应用基础研究基金(2022A1515110490,2022A1515012160)深圳市自然科学基金(JCYJ20220530152015036)。

摘　　要：目的探讨黄芩素对糖尿病小鼠全层皮肤缺损创面愈合的影响及其机制。方法该研究为实验研究。从5只8~12周龄雄性C57BL/6J小鼠中分离单核细胞并诱导分化为巨噬细胞,进行后续实验。按照随机数字表法(分组方法下同),将高糖环境中的巨噬细胞分为采用1μg/mL内毒素/脂多糖(LPS)和相应终物质的量浓度黄芩素处理的0μmol/L黄芩素组(不加黄芩素)、5μmol/L黄芩素组、15μmol/L黄芩素组、25μmol/L黄芩素组、50μmol/L黄芩素组、75μmol/L黄芩素组,处理48 h后,用酶标仪检测细胞增殖活性。将高糖环境中的巨噬细胞分为采用1μg/mL LPS处理的LPS组和用50μmol/L黄芩素+1μg/mL LPS处理的LPS+黄芩素组,处理48 h后,采用免疫荧光法检测细胞中诱导型一氧化氮合酶(iNOS)与CD80双阳性细胞百分比、精氨酸酶1(Arg1)与CD206双阳性细胞百分比,用酶联免疫吸附测定法检测细胞的白细胞介素1β(IL-1β)、IL-6、IL-23、IL-10、胰岛素样生长因子(IGF)、转化生长因子β1(TGF-β1)分泌水平,用荧光探针法检测细胞中活性氧表达,用蛋白质印迹法检测细胞中核因子κB蛋白表达,采用免疫荧光法观察细胞中核因子2表达。细胞实验样本数均为3。取24只8周龄雄性db/db小鼠,于其背部制备全层皮肤缺损创面后将其分为黄芩素组和生理盐水组(每组12只小鼠),伤后3 d分别向小鼠创面注射50μmol/L黄芩素和生理盐水。于伤后4、8、12 d观察创面愈合情况并计算残余创面面积百分比;取伤后8 d创面组织,行苏木精-伊红染色观察表皮新生情况及炎症细胞浸润情况,采用蛋白质印迹法检测CD31的蛋白表达,采用酶标仪检测活性氧表达。动物实验样本数均为6。结果处理48 h后,仅50μmol/L黄芩素组巨噬细胞增殖活性明显高于0μmol/L黄芩素组(P<0.05)。处理48 h后,LPS+黄芩素组巨噬细胞中iNOS与CD80双阳性细胞百分比[(21.0±2.4)%]明显低于LPS组[(66.6±4.5)%,t=15.63,P<ObjectiveTo investigate the effects and mechanism of baicalin on the wound healing of full-thickness skin defects in diabetic mice.MethodsThis study was an experimental research.Mononuclear cells were isolated from five male C57BL/6J mice aged 8-12 weeks and induced to differentiate into macrophages for conducting the subsequent experiments.According to the random number table(the same grouping method below),macrophages in a high-glucose environment were divided into 0μmol/L baicalin group(no baicalin was added),5μmol/L baicalin group,15μmol/L baicalin group,25μmol/L baicalin group,50μmol/L baicalin group,and 75μmol/L baicalin group treated with the corresponding final molarity of baicalin and 1μg/mL endotoxin/lipopolysaccharide(LPS).After treatment for 48 hours,the cell proliferation activity was detected using a microplate reader.Macrophages in a high-glucose environment were divided into LPS group treated with 1μg/mL LPS and LPS+baicalin group treated with 50μmol/L baicalin+1μg/mL LPS.After treatment for 48 hours,the percentage of double-positive cells for inducible nitric oxide synthase(iNOS)and CD80,as well as that for arginase 1(Arg1)and CD206 among the cells,were detected using immunofluorescence method,the secretion levels of interleukin 1β(IL-1β),IL-6,IL-23,IL-10,insulin-like growth factor(IGF),and transforming growth factorβ1(TGF-β1)by the cells were detected using enzyme-linked immunosorbent assay,the expression of reactive oxygen species in the cells was detected using a fluorescent probe method,the protein expression of nuclear factorκB in the cells were detected using Western blotting,and the expression of nuclear factor 2 in the cells was observed using immunofluorescence method.The number of cell experimental samples was 3.Twenty-four 8-week-old male db/db mice were selected.After preparing full-thickness skin defect wounds on their backs,they were divided into baicalin group and normal saline group(with 12 mice in each group).On the third day after injury,50μmol/L baicalin and nor

关 键 词：糖尿病 巨噬细胞 活性氧 黄芩素 创面修复 炎症反应 

分 类 号：R285.5[医药卫生—中药学]