CAJ | 학술논문

目的研究长波紫外线(UVA)照射对皮肤成纤维细胞组织蛋白酶G(CatG)表达和分泌的影响.方法原代培养的皮肤成纤维细胞来自儿童包皮,10代以内的细胞行后续实验.①以10 J/cm2 UVA照射皮肤成纤维细胞,24、48、72 h后提取照射组和对照组细胞蛋白和mRNA,并收集细胞上清液;②分别以10、20、30 J/cm2UVA照射皮肤成纤维细胞,24 h后收集细胞和上清液.用RT-PCR和Western印迹分别检测各组细胞CatGmRNA及蛋白的表达,ELISA检测细胞上清液CatG的含量.结果 10 J/cm2 UVA照射后24、48、72 h,照射组细胞CatG mRNA表达分别为0.376±0.014、0.308±0.022和0.296±0.032,对照组分别为0.183±0.003、0.185±0.005、0.182±0.004;照射组细胞CatG蛋白灰度值分别为1.80±0.12、1.41±0.17和1.27±0.09,对照组分别为0.96±0.06、0.95±0.22、1.00±0.14;照射组细胞CatG mRNA、蛋白表达及细胞上清液CatG含量较相应的对照组升高(均P＜ 0.05),且以照射后24h表达最高.10、20、30 J/cm2 UVA照射后24 h,皮肤成纤维细胞CatG mRNA表达分别为对照组的1.90、2.51、3.04倍,细胞CatG蛋白表达分别为对照组的1.88、3.97、4.72倍,细胞上清液CatG含量分别为对照组的1.36、1.50、1.66倍,均随UVA剂量的升高而增加,其差异有统计学意义(均P＜ 0.01).结论急性UVA照射促进皮肤成纤维细胞表达和分泌CatG.
목적 연구장파자외선(UVA)조사대피부성섬유세포조직단백매G(CatG)표체화분비적영향.방법 원대배양적피부성섬유세포래자인동포피,10대이내적세포행후속실험.①이10 J/cm2 UVA조사피부성섬유세포,24、48、72 h후제취조사조화대조조세포단백화mRNA,병수집세포상청액;②분별이10、20、30 J/cm2UVA조사피부성섬유세포,24 h후수집세포화상청액.용RT-PCR화Western인적분별검측각조세포CatGmRNA급단백적표체,ELISA검측세포상청액CatG적함량.결과 10 J/cm2 UVA조사후24、48、72 h,조사조세포CatG mRNA표체분별위0.376±0.014、0.308±0.022화0.296±0.032,대조조분별위0.183±0.003、0.185±0.005、0.182±0.004;조사조세포CatG단백회도치분별위1.80±0.12、1.41±0.17화1.27±0.09,대조조분별위0.96±0.06、0.95±0.22、1.00±0.14;조사조세포CatG mRNA、단백표체급세포상청액CatG함량교상응적대조조승고(균P＜ 0.05),차이조사후24h표체최고.10、20、30 J/cm2 UVA조사후24 h,피부성섬유세포CatG mRNA표체분별위대조조적1.90、2.51、3.04배,세포CatG단백표체분별위대조조적1.88、3.97、4.72배,세포상청액CatG함량분별위대조조적1.36、1.50、1.66배,균수UVA제량적승고이증가,기차이유통계학의의(균P＜ 0.01).결론 급성UVA조사촉진피부성섬유세포표체화분비CatG.
Objective To investigate the effect of ultraviolet A (UVA) radiation on the expression and secretion of cathepsin G (CatG) by human dermal fibroblasts.Methods Dermal fibroblasts were isolated from the foreskins of boys,and subjected to primary culture and subculture.After 10 or less passages,the fibroblasts were collected and divided into several groups to be irradiated with 10 J/cm2 UVA followed by 24,48 and 72 hours of additional culture,or be irradiated with 10,20 and 30 J/cm2 UVA followed by 24 hours of additional culture,with those receiving no treatment serving as the control group.Subsequently,cells and culture supernatant were collected,real time PCR and Western blot were performed to detect the expressions of CatG mRNA and protein respectively in these cells,and enzyme-linked immunosorbent assay (ELISA) was conducted to measure the expression of CatG protein in the culture supernatant of these cells.Results Compared with the control group,the fibroblasts irradiated with 10 J/cm2 UVA showed a significant increase at 24,48 and 72 hours in the expressions of CatG mRNA (0.376 ± 0.014 vs.0.183 ± 0.003,0.308 ± 0.022 vs.0.185 ± 0.005,0.296 ± 0.032 vs.0.182 ± 0.004,respectively,all P＜ 0.05) and protein (1.80 ± 0.12 vs.0.96 ± 0.06,1.41 ± 0.17 vs.0.95 ± 0.22,1.27 ± 0.09 vs.1.00 ± 0.14,respectively,all P ＜ 0.05),as well as in the supernatant level of CatG protein ((161.35 ± 7.55) vs.(122.45 ± 6.46) ng/L,(141.76 ± 2.95) vs.(124.17 ± 6.15) ng/L,(139.63 ± 3.04) vs.(121.72 ± 3.17) ng/L respectively,all P ＜0.05),with the strongest increase observed at 24 hours.At 24 hours after 10,20 and 30 J/cm2 of UVA radiation,the expression of CatG mRNA in irradiated fibroblasts was 1.90,2.51 and 3.04 times respectively (all P ＜ 0.05),the expression of CatG protein was 1.88,3.97 and 4.72 times respectively (P ＜ 0.05),and the supernatant level of CatG protein was 1.36,1.50 and 1.66 times respectively (P ＜ 0.05),that in the control group,and there was an increasing trend in all the above three parameters with increasing dose of UVA.Conclusion Acute UVA radiation can promote the expression and secretion of CatG by human dermal fibroblasts.