转录因子TFAP2B在健康人及白癜风患者表皮黑色素细胞中的表达模式

Expression patterns of transcription factor TFAP2B in epidermal melanocytes in healthy individuals and vitiligo patients

目的探究转录因子TFAP2B在健康人及白癜风患者表皮黑色素细胞中的表达模式.方法收集空军军医大学西京皮肤医院2020年1月至2022年12月明确诊断的5例进展期白癜风患者皮损组织,同时收集性别、年龄匹配的5例来源于整形外科手术剩余皮肤组织作为健康对照.培养永生化的健康人表皮黑色素细胞系PIG1、白癜风表皮黑色素细胞系PIG3V及人表皮原代黑色素细胞,其中原代黑色素细胞分离自西京医院泌尿外科手术切除的3例健康男性包皮组织.采用组织免疫荧光检测TFAP2B、多巴色素互变异构酶(DCT)在健康对照皮肤和白癜风皮损中的表达及定位,细胞免疫荧光和Western印迹法检测TFAP2B在人表皮黑色素细胞中的表达.两组间比较采用t检验,相关性分析采用Pearson相关分析法.结果组织免疫荧光结果显示,TFAP2B仅在人表皮黑色素细胞中表达,且定位于细胞核.Western印迹结果显示,TFAP2B在人表皮黑色素细胞系PIG1及原代黑色素细胞中强表达(相对表达量分别为0.45±0.05和0.36±0.04).组织免疫荧光结果显示,在10例人表皮组织(包括5例健康对照和5例白癜风)中,TFAP2B(623917.5±88784.0)与DCT(2232655.3±588810.4)的荧光强度呈显著正相关(r=0.91,P<0.001).此外,TFAP2B在白癜风皮损表皮黑色素细胞中相对荧光强度(0.12±0.05)显著低于健康对照表皮黑色素细胞(1,t=19.35,P<0.001).Western印迹结果显示,TFAP2B在白癜风表皮黑色素细胞系PIG3V中的相对表达值(0.62±0.09)亦显著低于PIG1细胞(1,t=5.92,P<0.027).结论TFAP2B在人表皮黑色素细胞中特异性高表达,其表达与黑色素细胞标记分子DCT的表达呈显著正相关,在白癜风患者表皮黑色素细胞中明显低表达.

Objective To explore expression patterns of transcription factor TFAP2B in epidermal melanocytes of healthy individuals and vitiligo patients.Methods Lesional tissues were collected from 5 patients confirmedly diagnosed with progressive vitiligo at the Department of Dermatology,Xijing Hospital,Air Force Medical University from January 2020 to December 2022.At the same time,some discarded normal skin tissues were obtained from 5 gender-and age-matched healthy individuals after plastic surgeries.The immortalized healthy human epidermal melanocyte cell line PIG1,the vitiligo epidermal melanocyte cell line PIG3V,and primary human epidermal melanocytes,which were isolated from the discarded foreskin tissues of 3 healthy males after urological surgeries in Xijing Hospital,were cultured in vitro.Tissue immunofluorescence assay was performed to determine the expression and localization of TFAP2B and dopachrome tautomerase(DCT)in healthy skin tissues and vitiligo lesions,and cell immunofluorescence assay and Western blot analysis were conducted to determine the TFAP2B expression in human epidermal melanocytes.Comparisons between two groups were performed using t test,and correlation analysis was performed using Pearson correlation coefficients.Results Tissue immunofluorescence assay showed that TFAP2B was specifically expressed in human epidermal melanocytes and localized in the nuclei.Western blot analysis showed that TFAP2B was strongly expressed in the human epidermal melanocyte cell line PIG1 and primary melanocytes,with the relative expression levels being 0.45±0.05 and 0.36±0.04,respectively.Tissue immunofluorescence analysis showed that the fluorescence intensity of TFAP2B(623917.5±88784.0)was significantly and positively correlated with that of DCT(2232655.3±588810.4;r=0.91,P<0.001)in human epidermal tissues from 5 healthy controls and 5 vitiligo patients.In addition,the relative fluorescence intensity of TFAP2B in epidermal melanocytes was significantly lower in the vitiligo lesions(0.12±0.05)than in the healthy skin tissues(1,t=19.35,P<0.001).Western blot analysis showed that the relative expression level of TFAP2B was also significantly lower in the PIG3V cells(0.62±0.09)than in the PIG1 cells(1,t=5.92,P<0.027).Conclusions TFAP2B was specifically and highly expressed in human epidermal melanocytes,and its expression level was significantly and positively correlated with that of the melanocyte marker DCT.Additionally,TFAP2B was obviously lowly expressed in the epidermal melanocytes of patients with vitiligo.