自噬抑制剂通过氧化应激诱导人头皮毛乳头细胞早衰进程  

Autophagy Inhibitors Induce Premature Senescence of Human Scalp Dermal Papilla Cells by Oxidative Stress

作　　者：万梅 钟意 翁祖铨 吴亚光 宋潇 杨希川 WAN Mei;ZHONG Yi;WENG Zuquan;WU Yaguang;SONG Xiao;YANG Xichuan(Department of Dermatology,the First Affiliated Hospital of the Army Medical University,Chongqing 400038,China;College of Biological Science and Engineering,Fuzhou University,Fuzhou 350108,China)

机构地区：[1]陆军军医大学第一附属医院皮肤科,重庆400038 [2]福州大学生物科学与工程学院,福建福州350108

出　　处：《中国皮肤性病学杂志》2023年第7期748-754,共7页The Chinese Journal of Dermatovenereology

基　　金：国家自然科学基金项目(82073460)。

摘　　要：目的 探讨自噬失调对毛乳头细胞(dermal papilla cells,DPC)早衰的影响。方法 自噬激活剂雷帕霉素(rapamycin,RPM)处理DPC 24 h后再用自噬抑制剂U0126处理细胞24 h,采用透射电镜和Western blot分析自噬小体数和自噬相关蛋白LC3-Ⅱ/Ⅰ比值变化。U0126处理DPC 24 h后采用细胞迁移实验和SA-β-gal实验检测DPC迁移率和衰老阳性率;Western blot和qPCR分析衰老相关蛋白p16、生物活性因子C-myc、Survivin的表达变化;ELISA检测活性氧、DNA损伤程度及影响毛囊生长周期的因子IGF-1、TGF-β1、TGF-β2水平。结果 RPM显著提高了DPC自噬小体数和LC3-Ⅱ/Ⅰ比值,而U0126显著降低了DPC自噬小体数和LC3-Ⅱ/Ⅰ比值(P<0.05)。此外,U0126处理的DPC迁移能力显著降低,衰老阳性率显著增加以及生物活性因子C-myc、Survivin表达显著降低(P<0.05)。同时,ELISA的检测结果表明U0126显著提高DPC内活性氧水平、DNA损伤程度和分泌TGF-β的能力以及显著降低分泌IGF-1的能力(P<0.05)。结论 自噬被抑制后的DPC出现早衰表型和氧化应激损伤,同时生物活性因子的表达受到负向调控,因此调控DPC自噬有可能有助于雄激素性脱发治疗。Objective To investigate the effect of autophagy disorder on premature senescence of dermal papilla cells(DPC).Methods DPC were successively treated with autophagy activator rapamycin(RPM) and autophagy inhibitor U0126 for 24 h.Then the treated DPC were used for autophagy level analysis by detecting the number of autophagosomes and the ratio of autophagy-related protein LC3-Ⅱ/Ⅰ via transmission electron microscopy and Western blot.Cell migration assay and SA-β-gal assay were adopted to detect the migration rate and senescence positive rate of autophagy-inhibited DPC.Western blot and qPCR were used to analyze the expression of p16,C-myc and Survivin.ELISA assay was conducted to detect the degree of reactive oxygen species,DNA damage and the growth factors(IGF-1,TGF-β1 and TGF-β2) in the hair follicle growth cycle.Results RPM significantly increased the number of autophagosomes and LC3-Ⅱ/Ⅰ ratio of DPC,while the two metrics were significantly decreased by U0126(P<0.05).In addition,the migration ability and the expression of biological active factors were significantly decreased,while the positive rate of senescence was significantly increased inside the U0126-treated DPC(P<0.05).Moreover,results from ELISA assay on the U0126-treated DPC indicated that the reactive oxygen species level,DNA damage and the production of TGF-β were marked increased while the production of IGF-1 was remarkably declined(P<0.05).Conclusion Our experiments suggest that the premature senescence phenotype,oxidative stress damage and negative regulation of the expression of bioactive factors occur inside the autophagy-inhibited DPC.Therefore,the regulation of DPC autophagy may be a solution for the treatment of androgenetic alopecia induced by the premature senescence of DPC.

关 键 词：U0126 自噬 毛乳头细胞 早衰 

分 类 号：R758.71[医药卫生—皮肤病学与性病学]