摘要
目的:通过体外培养人Tenon’s囊成纤维细胞(human tenon’s fibroblasts, HTFs)并经TGF-β_1诱导,探讨青光安颗粒含药血清对青光眼滤过性手术后细胞模型自噬的影响。方法:(1)取人的Tenon’s囊组织培养HTFs并鉴定,加入TGF-β_1诱导分化为肌成纤维细胞,构建GFS术后细胞模型。(2)细胞模型中加入最适浓度的青光安含药血清,设立正常组(空白组)、TGF-β_1组(模型组)、TGF-β_1+含药血清组(治疗组)、TGF-β_1+雷帕霉素组(阳性对照组),用Cyto-ID自噬检测试剂盒在荧光显微镜下及Cytation5细胞成像多功能仪器检测,观察核周围和细胞质的自噬小体及自噬溶酶体成分,以自噬阳性细胞数所占百分比表示HTFs的自噬水平,同时检测自噬阳性细胞的平均荧光强度。结果:治疗组自噬阳性细胞百分比明显高于空白组与模型组,差异有统计学意义(P<0.01);且治疗组自噬程度呈含药血清干预时间依赖型即正相关;其自噬阳性细胞平均荧光强度呈含药血清干预时间依赖型即正相关。结论:青光安颗粒含药血清能增强青光眼滤过性手术后细胞模型的自噬表达。
Objective:To investigate the efficacy of Qing Guang An on autophagy in the TGF-β1-activated human tenon’s fibroblasts.Methods:(1)Primary HTFs was stimulated by TGF-β1and underwent immunohistochemistry identification,to establish an cell model after GFS.(2)The cell models were divided into 4 group:blank group(nomal cells),model group(TGF-β1),treatment group(TGF-β1+medicated serum)and positive control group(TGF-β1+RAPA).Then Qing Guang An medicated serum with optimum concentrations were added to the corresponding group.Through the Cyto-ID autophagy detection kits,by fluorescent microscope and Cytation5multifunctional instrument for cell imaging,we observed the autophagosome and autophagy-lysosome around the nucleus and cytoplasm,and assessed the autophagy level of different groups.Results:The treatment group showed higher autophagy than the blank group,and the difference was statistically significant(P<0.01);The results showed that the degree of autophagy in the treatment group was positively correlated with the time-dependent intervention of Qing Guang An medicated serum;The mean fluorescence intensity of the autophagy positive cells was positively correlated with the time-dependent intervention of medicated serum.Conclusion:The Qing Guang An medicated serum can enhance the autophagy expression in the TGF-β1-activated human tenon’s fibroblasts.
作者
喻娟
罗向艳
彭清华
YU Juan;LUO Xiang-yan;PENG Qing-hua(The First Affiliated Hospital,Hu'nan University of TCM,Changsha Hu'nan 410007,China)
出处
《中医药导报》
2019年第5期34-39,共6页
Guiding Journal of Traditional Chinese Medicine and Pharmacy
基金
国家自然科学基金资助项目(No.81603665)
湖南省自然科学基金重点资助项目(No.2017JJ3238)
中国博士后基金资助项目(No.2017M612565)
湖南中医药大学中医学国内一流建设学科
湖南省中医药防治眼耳鼻咽喉疾病与视功能保护工程技术研究中心
中医药防治眼耳鼻咽喉疾病湖南省重点实验室项目
