摘要
目的基于TGF-β1/Smads信号通路探讨松果菊苷(echinacoside,ECH)对人增生性瘢痕成纤维细胞(HSFbs)的作用机制。方法自2019年3月至2020年2月新疆医科大学第一附属医院整形外科体外培养人增生性瘢痕成纤维细胞;通过CCK-8法分析ECH作用24 h和48 h细胞的抑制活性。采用划痕试验检测HSFbs的迁移能力;流式细胞术检测ECH作用24 h后细胞周期的影响;实时荧光定量聚合酶链反应(RT-q PCR)检测Smad2、Smad3、Smad7、Col1、Col3、Fibronectin和α-SMA的m RNA的表达;蛋白印迹法(Western blot)检测Smad2/3、P-Smad2/3、Smad7、Col1、Col3、Fibronectin和α-SMA蛋白的表达。结果随着ECH质量浓度的增高,HSFbs的细胞存活率逐渐降低,且呈现浓度依赖性,其IC50值为52.26μg/ml;ECH作用于HSFbs 24 h后,可将HSFbs阻滞在G0-G1期;不同浓度的ECH均可抑制HSFbs的迁移;相比模型组,实验组25、50μg/ml ECH均能够降低TGF-β1诱导的Col1、Col3、Smad2、Smad3、Fibronectin和α-SMA m RNA表达,50μg/ml ECH可促进Smad7m RNA表达;在蛋白表达水平上25、50μg/ml ECH均可使Col3、Fibronectin蛋白表达明显降低,Smad7蛋白表达明显增加,50μg/ml ECH组可降低P-Smad2/3、Col1和α-SMA的蛋白表达。结论ECH的可通过阻断TGF-β1/Smad信号通路进而抑制HSFbs活化、增殖与迁移,并减少胶原的分泌。
Objective To investigate the effect of echinacoside(ECH)on human hypertrophic scar fibroblasts(HSFbs)and to elucidate the possible mechanism based on the TGF-β1/Smads signaling pathway.Methods From March 2019 to February 2020,the human hypertrophic scar fibroblasts were cultured in vitro.The inhibitory effect of ECH on HSFbs was analyzed by CCK-8.The migration ability of HSFbs was assessed by the scratch test.The flow cytometry was used to detect the effect of ECH on cell cycle of HSFbs.The expression of Smad2,Smad3,Smad7,Col1,Col3,Fibronectin and m RNA ofα-SMA was detected by RT-q PCR.The expression of Smad2/3,phosphorylated Smad2/3(P-Smad2/3),Smad7,Col1,Col3,fibronectin andα-SMA proteins was detected by Western blot.Results With the increase of mass concentration of ECH,the cell survival rate of HSFbs was gradually decreased and a concentration dependence was showed.The IC50 value was 52.26μg/ml.After treatment with ECH for 24 h,the cell cycle of HSFbs was blocked in the G0-G1 phase.Different concentrations of ECH could inhibit the migration of HSFbs.Compared with the model group,the expression of Col1,Col3,Smad2,Smad3,Fibronectin and m RNA ofα-SMA induced by TGF-β1 could be downregulated by ECH of 25μg/ml and 50μg/ml,and the expression of m RNA of Smad7 could be promoted by ECH of 50μg/ml.At protein level,the expression of Col3 and Fibronectin was significantly inhibited by ECH of 25μg/ml and 50μg/ml.The expression of Smad7 was significantly increased.The expression of P-Smad2/3,Col1 andα-SMA was inhibited by ECH of 50μg/ml.Conclusion ECH can inhibit the activation,proliferation and migration of HSFbs and reduce collagen secretion by blocking the TGF-β1/Smads signaling pathway.
作者
艾江
余扬
费蒙辉
陈朝昱
马少林
AI Jiang;YU Yang;FEI Meng-hui;CHEN Zhao-yu;MA Shao-lin(Department of Burn and Plastic Surgery,The First Affiliated Hospital of Xinjiang Medical University,Urumqi 830034,China)
出处
《中国美容整形外科杂志》
CAS
2021年第3期187-189,I0008,I0009,共5页
Chinese Journal of Aesthetic and Plastic Surgery
基金
国家自然科学基金(81760345)
自治区研究生创新项目(XJ2919G202)
新疆医科大学研究生创新创业启动基金(CXCY2018023)。
