摘要 目的探讨人真皮乳头层成纤维细胞(Fp)、网状层成纤维细胞(Fr)和肌成纤维细胞(MFB)在瘢痕疙瘩皮损组织中的表达与分布。方法2019年5-12月在武汉大学人民医院皮肤科门诊确诊的15例瘢痕疙瘩患者,男8例,女7例,年龄20~50岁,取皮损组织,以15例年龄匹配的女性乳房整形术正常皮肤组织为对照。采用双重免疫荧光染色法检测成纤维细胞活化蛋白(FAP)、CD90和α平滑肌肌动蛋白(α-SMA)在瘢痕疙瘩和正常皮肤组织中的分布。从3例正常皮肤和3例瘢痕疙瘩组织中分离成纤维细胞原代培养,采用10 ng/ml转化生长因子β1(TGF-β1)体外处理两组细胞0~48 h,观察细胞表型的变化,荧光定量RT-PCR和Western印迹检测FAP、CD90和α-SMA mRNA和蛋白表达。两组间差异比较采用t检验。结果免疫荧光结果显示,正常皮肤组织中,FAP+/CD90-细胞主要分布在真皮浅层,FAP-/CD90+细胞集中在真皮深层,CD90+细胞几乎不表达α-SMA;瘢痕疙瘩组织深层可见大量FAP+和CD90+细胞,大量CD90+细胞同时表达α-SMA。双重免疫荧光染色显示,正常皮肤成纤维细胞几乎不表达α-SMA,瘢痕疙瘩成纤维细胞表达α-SMA;TGF-β1处理24 h时,正常成纤维细胞和瘢痕疙瘩成纤维细胞α-SMA+细胞荧光强度(21.058±0.709、27.112±0.097)均高于未处理组(11.312±0.636、21.306±0.464),t值为22.430、13.370,P<0.05。RT-PCR和Western印迹显示,TGF-β1处理48 h时,瘢痕疙瘩成纤维细胞FAP、CD90、α-SMA mRNA相对表达水平(92.610±3.667、1.366±0.105、3.240±0.141)与蛋白表达水平(0.652±0.073、1.046±0.119、0.946±0.117)均高于处理前(均P<0.05)。结论瘢痕疙瘩组织真皮深层的CD90+(Fr)细胞异常增生,提示针对真皮深层异常增殖活跃的FAP-/CD90+(Fr)细胞群进行定向干预可能提高瘢痕疙瘩治疗疗效。 Objective To investigate the expression and distribution of human dermal papillary fibroblasts(Fp),reticular fibroblasts(Fr),and myofibroblasts(MFB)in keloid tissues.Methods Keloid tissues were collected from 15 outpatients(including 8 males and 7 females)aged 20-50 years,who were diagnosed in the Department of Dermatology,Renmin Hospital of Wuhan University from May to December 2019.Normal skin tissues were taken from 15 age-matched women who underwent mammoplasty,and served as controls.The distribution of fibroblast activation protein(FAP),CD90 and alpha-smooth muscle actin(α-SMA)was observed in the keloid tissues and normal skin tissues by dual immunofluorescence staining.Furthermore,fibroblasts were isolated from 3 normal skin and 3 keloid tissue samples,and subjected to primary culture.Subsequently,the fibroblasts were treated with 10 ng/ml transforming growth factor-β1(TGF-β1)for 48 hours in vitro,during which,changes in fibroblast phenotypes were observed in the 2 groups.Fluorescence-based quantitative RT-PCR and Western blot analysis were performed to determine the mRNA and protein expression of FAP,CD90 andα-SMA.Measurement data were compared between 2 groups by using t test.Results Immunofluorescence staining of the normal skin tissues revealed that FAP+/CD90-fibroblasts were predominantly distributed in the superficial dermis,FAP-/CD90+fibroblasts in the deep dermis,and CD90+cells hardly expressedα-SMA;however,a large number of FAP+fibroblasts and CD90+fibroblasts were observed in the deep keloid tissues,and many CD90+fibroblasts also expressedα-SMA.Dual immunofluorescence staining showed that normal tissue-derived fibroblasts hardly expressedα-SMA,and keloid-derived fibroblasts expressedα-SMA.The fluorescence intensity ofα-SMA+cells significantly increased in the normal tissue-and keloid-derived fibroblasts after 24-hour treatment with TGF-β1(21.058±0.709,27.112±0.097,respectively)compared with that in the corresponding untreated fibroblasts(11.312±0.636,21.306±0.464,t=22.430,13.370,resp
机构地区 武汉大学人民医院皮肤科
出处 《中华皮肤科杂志》 CAS CSCD 北大核心 2021年第6期504-509,共6页 Chinese Journal of Dermatology
基金 国家自然科学基金(81972919)。
关键词 瘢痕疙瘩 成纤维细胞 肌成纤维细胞 乳头层成纤维细胞 网状层成纤维细胞 CD90 Α平滑肌肌动蛋白 Keloid Fibroblasts Myofibroblasts Papillary fibroblast Reticular fibroblast CD90 α-SMA
分类号 R73 [医药卫生—肿瘤]
