CAJ | 학술논문

目的探讨组织蛋白酶B在光老化皮肤中的表达意义.方法 6例成人曝光和非曝光皮肤标本,采用免疫组化法定位及对比组织蛋白酶B的表达.体外培养原代人皮肤成纤维细胞,甲氧沙林+UVA法体外诱导培养细胞光老化.衰老相关-β-半乳糖苷酶染色证明老化诱导成功.Western印迹技术及RT-PCR对比检测光老化成纤维细胞及正常成纤维细胞组织蛋白酶B蛋白及基因表达.结果 6例成人曝光和非曝光活体皮肤均见组织蛋白酶B阳性染色,和非曝光部位相比,曝光部位皮肤阳性染色A值降低.Western印迹结果示,成纤维细胞光老化诱导组蛋白表达较UVA诱导组、甲氧沙林孵育组及空白组明显下调.光老化成纤维细胞诱导后l周,组织蛋白酶B与内参的灰度比由28.099±0.054下降为25.103±0.102,诱导后3周灰度值进一步下降为17.693±0.099.实时定量RT-PCR结果示,光老化细胞组织蛋白酶B mRNA表达下调为正常组的64%(P<0.05).结论组织蛋白酶B在光老化皮肤及老化成纤维细胞中表达降低,且有时间依赖性,与光老化皮肤自我修复能力下降有关.
목적 탐토조직단백매B재광노화피부중적표체의의.방법 6례성인폭광화비폭광피부표본,채용면역조화법정위급대비조직단백매B적표체.체외배양원대인피부성섬유세포,갑양사림+UVA법체외유도배양세포광노화.쇠로상관-β-반유당감매염색증명노화유도성공.Western인적기술급RT-PCR대비검측광노화성섬유세포급정상성섬유세포조직단백매B단백급기인표체.결과 6례성인폭광화비폭광활체피부균견조직단백매B양성염색,화비폭광부위상비,폭광부위피부양성염색A치강저.Western인적결과시,성섬유세포광노화유도조단백표체교UVA유도조、갑양사림부육조급공백조명현하조.광노화성섬유세포유도후l주,조직단백매B여내삼적회도비유28.099±0.054하강위25.103±0.102,유도후3주회도치진일보하강위17.693±0.099.실시정량RT-PCR결과시,광노화세포조직단백매B mRNA표체하조위정상조적64%(P<0.05).결론 조직단백매B재광노화피부급노화성섬유세포중표체강저,차유시간의뢰성,여광노화피부자아수복능력하강유관.
Objective To investigate the expression of cathepsin B in photoaging skin and its signifi-cance.Methods Skin specimens were obtained from the right forearm(sun-exposed sites)and buttock (unexposed sites)of 6 healthy volunteers with informed consent and subjected to immunohistochemistry for the detection of cathepsin B expression.Primary human fibroblasts derived from the prepuce of children aged 3 to 6 years were cultured in vitro;after 10 to 15 passages,cells were divided into four groups to be treated with methoxsalen of 50 mg/L for 24 hours followed by ultraviolet A(UVA)exposure(premature senescence group),phosphate buffered saline(PBS)only(control group),UVA exposure only(UVA group),methoxsalen only(methoxsalen group).Then,the protein and mRNA expressions of cathepsin B were detected by Western blot and RT-PCR,respectively,in these fibroblasts 1,2,3 weeks after the treatment.Results Cathepsin B was observed in both exposed and unexposed sites of all volunteers,and the average absorbence of cathepsin B was significantly lower in exposed sites than in unexposed sites(0.2130±0.7997 vs 0.4520±0.5921,t=5.37,P<0.05).Decreased protein expression of cathepsin B was also noted in the premature senescence group compared with the other three groups.Moreover,the gray ratio between cathepsin B protein and glyceraldehyde phosphate dehydrogenase(GAPDH)in premature senescence group reduced from 28.099±O.054 before treatment to 25.1 03±0.102 in week 1 and 17.693±0.099 in week 3 after UVA exposure.RT-PCR revealed that the mRNA expression level of cathepsin B in fibroblasts of premature senescence group decreased by 36 percent compared with that in the control group.Conclusions The expression of cathepsin B decreases in photoaged skin as well as in UVA-exposed fibroblasts in a time-dependent manner,which may be associated with the self-repair of photoaged skin.