富氢液通过自噬途径下调中波紫外线诱导的HaCaT细胞炎症因子的表达  

作　　者：张秉新[1] 邢卫斌[2] 付国俊 陈红光[4] 

机构地区：[1]天津中医药大学第一附属医院皮肤科,300193 [2]天津市第五中心医院皮肤科 [3]河北省沧州市人民医院皮肤科 [4]天津医科大学总医院麻醉科

出　　处：《中华皮肤科杂志》2016年第2期123-127,共5页Chinese Journal of Dermatology

基　　金：国家自然科学基金（81101409、81471842）

摘　　要：目的探讨氢气是否能通过自噬途径调节中波紫外线（UVB）诱导的HaCaT细胞炎症因子的表达。方法将培养的HaCaT细胞分为空白对照组（不做任何处理），氢气组（仅用富氢培养液培养），1、10、50mJ／cm^2UVB组，1、10、50mJ／cm^2UVB＋氢气组，50mJ／cm^2UVB＋3甲基腺嘌呤（3MA）组，50mJ／cm^2UVB＋雷帕霉素组，50mJ／cm^2UVB＋3MA＋氢气组，50mJ／cm^2UVB＋雷帕霉素＋氢气组。细胞经过不同处理培养12h后，噻唑蓝（M1TI＇）法检测细胞增殖，LDH试剂盒检N-％酸脱氢酶（LDH），细胞提取蛋白检测自噬蛋白LC3和Beclin1的表达，上清液用ELISA检测炎症因子肿瘤坏死因子（TNF）ot、白细胞介素（IL）-1β、HMGB1和IL-6的水平。结果与空白对照组相比，UYB诱导的HaCaT细胞LDH释放增多，细胞活力下降，自噬蛋白LC3和Beclinl表达增加，炎症因子TNF—d、IL-1β、HMGB1和IL-6释放增加（均P〈0．05）。与UVB组相比，氢气可减少LDH释放，提高细胞活力，自噬蛋白LC3和Beclin1表达进一步增加，炎症因子TNF—α、IL-1β、HMGB1和IL-6表达减少（均P〈0．05）。与50mJ／cm^2UVB组相比，50mJ／cm^2UVB＋3MA组炎症因子TNF-α、IL-1β、HMGB1和IL-6表达进一步增加（P〈0．05），而50mJ／cm^2UVB＋雷帕霉素组炎症因子TNF-α、IL-1β、HMGB1和IL-6表达减少（P〈0．05）。与50mJ／cm^2UVB＋氢气组相比，50mJ／cm^2UVB＋氢气＋3MA组炎症因子TNF-α、IL-1β、HMGB1和IL-6表达增加（P〈0．05）。结论UVB照射可以诱发自噬蛋白表达增加；富氢液可以下调UVB诱导的HaCaT细胞炎症因子的表达，而这一过程是通过激活自噬途径实现的。Objective To investigate whether hydrogen can regulate the expressions of inflammatory factors by ultraviolet B (UVB)-induced human HaCaT keratinocytes through the autophagy pathway.Methods Cultured HaCaT keratinocytes were divided into several groups:blank control group receiving no treatment,hydrogen group cultured in hydrogen-rich medium,three UVB groups irradiated with UVB at 1,10,50 mJ/cm2 respectively,three UVB + hydrogen groups irradiated with UVB at 1,10,50 mJ/cm2 respectively followed by culture in hydrogen-rich medium,UVB + 3MA group pretreated with the autophagy inhibitor 3MA for 1 hour followed by UVB radiation at 50 mJ/cm2,UVB + rapamycin group pretreated with the autophagy activator rapamycin for 1 hour followed by UVB radiation at 50 mJ/cm2,UVB + 3MA +hydrogen group pretreated with 3MA for 1 hour followed by UVB radiation at 50 mJ/cm2 and culture in hydrogen-rich medium,UVB + rapamycin + hydrogen group pretreated with rapamycin for 1 hour followed by UVB radiation at 50 mJ/cm2 and culture in hydrogen-rich medium.After additional culture with or without hydrogen for 12 hours,methyl thiazolyl tetrazolium (MTT) assay was performed to evaluate cellular proliferative activity,Western-blot analysis to measure the expressions of autophagy-associated protein 1 light chain 3 (LC3) and Beclin 1,and enzyme-linked immunosorbent assay (ELISA) to measure the supernatant levels of inflammatory factors including tumor necrosis factor (TNF)-α,interleukin (IL)-1β,IL-6 and high mobility group protein B1 (HMGB1),and a test kit was used to determine the level of lactate dehydrogenase (LDH).Results Compared with the blank control group,the 10-and 50-mJ/cm2 UVB groups showed significantly increased release of LDH,expressions of LC3 and Beclin1 and supernatant levels of TNF-α,IL-1 β,IL-6 and HMGB 1,but decreased cellular proliferative activity (all P ＜ 0.05).Hydrogen significantly attenuated the release of LDH,down-regulated the supernatant levels of TNF-α,IL-1β,IL-6 and HMGB1,but up-regulated cellular proliferati

关 键 词：紫外线 自噬 炎症趋化因子类 氢气 HACAT细胞 

分 类 号：R751[医药卫生—皮肤病学与性病学]