IL-17A和IFN-γ对甲状腺相关眼病患者CD34-眼眶成纤维细胞纤维化的作用  

Effects of IL-17A and IFN-γon the fibrosis of CD34-orbital fibroblasts in Graves orbitopathy

作　　者：鲁奕 黄雅琢 李寅炜[1] 吴钰 方思捷 孙静[1] 周慧芳[1] Lu Yi;Huang Yazhuo;Li Yinwei;Wu Yu;Fang Sijie;Sun Jing;Zhou Huifang(Department of Ophthalmology,Shanghai Ninth People's Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200011,China)

机构地区：[1]上海交通大学医学院附属第九人民医院眼科,200011

出　　处：《中华实验眼科杂志》2020年第11期923-928,共6页Chinese Journal Of Experimental Ophthalmology

基　　金：国家重点研发计划项目(2018YFC1106100);国家自然科学基金面上项目(81770960);国家自然科学基金国际(地区)合作与交流项目(81761168037);上海市科学技术委员会基金项目(17DZ2260100);上海市教委“高水平地方高校建设创新团队”眼肿瘤与眼眶修复协同创新团队(SSMV-ZDCX20180401);上海交通大学医学院附属第九人民医院院级基金项目(JYHL2019MS04)。

摘　　要：目的观察白细胞介素-17A(IL-17A)与γ干扰素(IFN-γ)对Graves眼病(GO)患者CD34-眼眶成纤维细胞(OFs)的作用,探讨GO的发病机制。方法收集5例GO患者眼眶减压术中获取的眼眶脂肪结缔组织以及泪腺组织,采用组织块消化法进行培养和传代。采用免疫磁珠富集其中的CD34-OFs。将培养的OFs分为转化生长因子-β(TGF-β)组、TGF-β+10 ng/ml IL-17A组和TGF-β+100 ng/ml IL-17A组、TGF-β+1 ng/ml IFN-γ组和TGF-β+5 ng/ml IFN-γ组,在各组细胞培养液中均添加5 ng/ml TGF-β以诱导细胞发生纤维化,按照分组方法分别加入不同质量浓度的IL-17A或IFN-γ。采用Western blot法检测各组眼眶脂肪结缔组织和泪腺组织来源的CD34-OFs中纤连蛋白、Ⅰ型胶原、ɑ-平滑肌肌动蛋白(ɑ-SMA)和组织金属基质蛋白酶抑制剂1(TIMP-1)等纤维化相关蛋白表达。结果在眼眶脂肪结缔组织来源的CD34-OFs中,TGF-β+100 ng/ml IL-17A组细胞中纤连蛋白、Ⅰ型胶原、α-SMA和TIMP-1蛋白相对表达量明显高于TGF-β组和TGF-β+10 ng/ml IL-17A组,差异均有统计学意义(均P<0.05);TGF-β+5 ng/ml IFN-γ组细胞中纤连蛋白、Ⅰ型胶原和α-SMA相对表达量均明显低于TGF-β组和TGF-β+1 ng/ml IFN-γ组,差异均有统计学意义(均P<0.05)。在泪腺组织来源的CD34-OFs中,TGF-β+100 ng/ml IL-17A组细胞中纤连蛋白、Ⅰ型胶原、α-SMA和TIMP-1蛋白相对表达量明显高于TGF-β组和TGF-β+10 ng/ml IL-17A组,差异均有统计学意义(均P<0.05);TGF-β+1 ng/ml IFN-γ组细胞中纤连蛋白、Ⅰ型胶原和TIMP-1蛋白相对表达量明显高于TGF-β组和TGF-β+5 ng/ml IFN-γ组,而α-SMA蛋白相对表达量明显高于TGF-β+5 ng/ml IFN-γ组,差异均有统计学意义(均P<0.05)。结论高质量浓度IL-17A处理可促进TGF-β诱导的CD34-OFs纤维化,而高质量浓度的IFN-γ则抑制TGF-β诱导的CD34-OFs纤维化。Objective To observe the effects of interleukin-17A(IL-17A)and interferon-γ(IFN-γ)on CD34-orbital fibroblasts(OFs)in patients with Graves ophthalmopathy(GO),and to explore the pathogenesis of GO.Methods Orbital adipose connective tissue and lacrimal gland tissue of 5 patients with GO were collected during orbital decompression surgery.These tissue was cultured by tissue explant culture method.CD34-OFs were enriched by immunomagnetic beads after passage and expansion.The cultured cells were divided into transforming growth factor-β(TGF-β)group,TGF-β+10 ng/ml IL-17A group,TGF-β+100 ng/ml IL-17A group,TGF-β+1 ng/ml IFN-γgroup and TGF-β+5 ng/ml IFN-γgroup.The fibrosis of the cells was induced with 5 ng/ml TGF-βand then was treated with different concentrations of IL-17A or IFN-γaccording to grouping.The expression of fibronectin,collagen typeⅠ,α-smooth muscle actin(α-SMA)and tissue metalloproteinase inhibitor-1(TIMP-1)in CD34-OFs derived from both orbital adipose connective tissue and lacrimal gland tissue were detected by Western blot.This study protocol was approved by an Ethic Committee of Shanghai Ninth People's Hospital(SH9H-2020-TK195-1)and written informed consent was obtained from each patient.Results For CD34-OFs derived from orbital adipose connective tissue,the relative expressions of fibronectin,typeⅠcollagen,α-SMA and TIMP-1 protein were significantly higher in the TGF-β+100 ng/ml IL-17A group than those in the TGF-βgroup and TGF-β+10 ng/ml IL-17A group(all at P<0.05);the relative expressions of fibronectin,typeⅠcollagen andα-SMA in the cells in the TGF-β+5 ng/ml IFN-γgroup were significantly lower than those in the TGF-βgroup and the TGF-β+1 ng/ml IFN-γgroup(all at P<0.05).For CD34-OFs derived from lacrimal gland,the relative expressions of fibronectin,typeⅠcollagen,α-SMA and TIMP-1 protein in the TGF-β+100 ng/ml IL-17A group were significantly higher than those in the TGF-βgroup and the TGF-β+10 ng/ml IL-17A group(all at P<0.05);the relative expressions of fibronecti

关 键 词：GRAVES眼病 成纤维细胞 转化生长因子-β 白细胞介素-17A Γ干扰素 纤维化 

分 类 号：R581[医药卫生—内分泌] R771.3[医药卫生—内科学]