枫杨总黄酮调控类风湿关节炎成纤维样滑膜细胞增殖、迁移及凋亡的作用及机制  

Effect and mechanism by which Pterocarya hupehensis skan total flavonoids regulates the proliferation,migration and apoptosis of fibroblast-like synoviocytes

作　　者：包卓玛 侯孜明 江露 李玮怡 张宗星 刘道忠 袁林 Bao Zhuoma;Hou Ziming;Jiang Lu;Li Weiyi;Zhang Zongxing;Liu Daozhong;Yuan Lin(Hubei Provincial Key Laboratory of Rheumatic Diseases,Hubei Minzu University,Enshi 445000,Hubei Province,China;Department of Medicine,Hubei Minzu University,Enshi 445000,Hubei Province,China;The First Affiliated Hospital of Anhui University of Chinese Medicine,Hefei 230031,Anhui Province,China;Shizhu Tujiazu Autonomous County Hospital,Chongqing 409100,China;Hubei Provincial Clinical Research Center for Nephrology,Enshi 445000,Hubei Province,China)

机构地区：[1]湖北民族大学风湿性疾病发生与干预湖北省重点实验室,湖北省恩施州445000 [2]湖北民族大学医学部,湖北省恩施州445000 [3]安徽中医药大学第一附属医院,安徽省合肥市230031 [4]石柱土家族自治县中医院,重庆市409100 [5]湖北省肾脏病临床医学研究中心,湖北省恩施州445000

出　　处：《中国组织工程研究》2026年第4期816-823,共8页Chinese Journal of Tissue Engineering Research

基　　金：恩施土家族苗族自治州科技局指导计划项目(恩施科业[2019]15号),项目负责人:袁林;2023年湖北民族大学研究生教育创新项目(MYK2023064),项目负责人:包卓玛。

摘　　要：背景:研究证实枫杨总黄酮能够改善胶原诱导的大鼠关节炎水平,但它对成纤维样滑膜细胞Wnt/β-catenin信号通路的调控作用及对相关细胞功能的影响尚缺少研究。目的:基于Wnt/β-catenin信号通路探究枫杨总黄酮对脂多糖诱导的成纤维样滑膜细胞增殖、迁移及凋亡的作用及机制。方法:成纤维样滑膜细胞分为对照组、脂多糖组、脂多糖+枫杨总黄酮低、中、高剂量组(10,20,40μg/mL)、脂多糖+Wnt通路抑制剂Dickkopf相关蛋白1(DKK1)组、脂多糖+DKK1+枫杨总黄酮高剂量组(40μg/mL)。采用CCK-8法检测枫杨总黄酮对成纤维样滑膜细胞活力的影响并筛选最终用药浓度和时间;流式细胞术检测成纤维样滑膜细胞的凋亡情况;细胞划痕实验、EDU染色和细胞克隆实验检测成纤维样滑膜细胞的迁移和增殖能力;蛋白质免疫印迹检测成纤维样滑膜细胞Wnt3a、β-catenin、细胞致瘤基因、基质金属蛋白酶2、基质金属蛋白酶9、Bax、Bcl-2蛋白表达水平。结果与结论:(1)与对照组相比,当枫杨总黄酮质量浓度>40μg/mL时细胞活力明显下降(P<0.01),故选择药物剂量≤40μg/mL进行后续实验;(2)与脂多糖组相比,枫杨总黄酮低、中、高剂量组细胞划痕愈合率、细胞克隆形成率、EDU阳性细胞数均显著降低,细胞凋亡率明显升高(P<0.05-0.01);(3)蛋白质免疫印迹结果显示,与脂多糖组相比,枫杨总黄酮低、中、高剂量组显著抑制细胞Wnt3a、β-catenin、细胞致瘤基因、基质金属蛋白酶2、基质金属蛋白酶9、Bcl-2蛋白表达,并促进Bax蛋白表达(P<0.01);(4)与单用Wnt通路抑制剂DKK1组相比,DKK1+枫杨总黄酮高剂量联合干预后,能够明显增强DKK1对细胞Wnt3a、β-catenin、基质金属蛋白酶2、基质金属蛋白酶9、Bcl-2蛋白表达的抑制作用和对Bax蛋白表达的促进作用(P<0.01);(5)结果表明,枫杨总黄酮可能通过抑制Wnt/β-catenin信号通路下调脂多糖诱导的成纤维BACKGROUND:Studies have confirmed that Pterocarya hupehensis skan total flavonoids(PHSTF)can improve the level of collagen-induced arthritis in rats,but there is still a lack of research on the regulation of Wnt/β-catenin signaling pathway in fibroblast-like synoviocytes and its effect on related cell functions.OBJECTIVE:To investigate the effect and mechanism of PHSTF on lipopolysaccharide-induced proliferation,migration and apoptosis of fibroblast-like synoviocytes based on the Wnt/β-catenin signaling pathway METHODS:Fibroblast-like synoviocytes were divided into control group,lipopolysaccharide group,lipopolysaccharide+low-,medium-,and high-dose PHSTF groups(10,20,and 40μg/mL),lipopolysaccharide+Wnt pathway inhibitor DKK1 group,and lipopolysaccharide+Wnt pathway inhibitor DKK1+high-dose PHSTF group(40μg/mL).The cell counting kit-8 method was used to detect the effect of PHSTF on the viability of fibroblast-like synoviocytes,and the final drug concentration and time were screened.Flow cytometry was used to detect the apoptosis of fibroblast-like synoviocytes.Cell scratch assay,EDU staining and cell cloning assay were used to detect the migration and proliferation of fibroblast-like synoviocytes.Western blot assay was used to detect the protein expression levels of Wnt3a,β-catenin,tumorigenic genes,matrix metalloproteinase 2,matrix metalloproteinase 9,Bax and Bcl-2 in fibroblast-like synoviocytes.RESULTS AND CONCLUSION:(1)Compared with the control group,the cell viability decreased significantly when the concentration of PHSTF was>40μg/mL(P<0.01).Therefore,the drug concentration of≤40μg/mL was selected for subsequent experiments.(2)Compared with the lipopolysaccharide group,the wound healing rate,cell clone formation rate and the number of EDU-positive cells in the low-,medium-and high-dose PHSTF groups were significantly reduced,while the apoptosis rate was significantly increased(P<0.05-0.01).(3)Western blot results showed that compared with the lipopolysaccharide group,low-,medium-and high-dose PHSTF

关 键 词：枫杨总黄酮 成纤维样滑膜细胞 类风湿关节炎 Wnt/β-catenin 凋亡 增殖 

分 类 号：R459.9[医药卫生—治疗学] R329.21[医药卫生—临床医学] R593.22