基于REV-ERBα/RORγt调控Th17细胞分化探讨昼夜节律对银屑病的影响及凉血解毒方干预机制研究

Circadian rhythm is the organism self-feedback regulating mechanism. Rhythm disorders can induce or aggravate psoriasis. During the study on pathological mechanism of psoriasis, we used sequencing technique to find circadian clock core genes (Per2 and REV-ERBα) in two animal models of psoriasis, and initially confirmed the relevance of psoriasis and circadian clock. Th17 cells differentiation, which plays a critical role in the pathogenesis of psoriasis, is regulated by REV-ERBα. LiangXueJieDu Fang (LXJDF) is an effective TCM compound for treatment of psoriasis blood heat syndrome, which could significantly inhibit nuclear factor RORγt in Th17 cell and related inflammatory cytokines such as IL-17A expression by clinical observation and basic experiments. In this study, we will observe the effects of LXJDF on Th17 cell differentiation and related inflammatory cytokines and the expressions of circadian clock genes through the pathological features of psoriasis mice models (imiquimod induced circadian rhythm disorder mice and mPer2m/m mice), and Th17 cell and REV-ERBα transfected Th17 cell, try to investigate the effect of circadian rhythm on psoriasis and the molecular mechanism of LXJDF intervene via REV-ERBα/RORγt regulating the Th17 cell differentiation in time medicine.

昼夜节律是生命体内在的自我反馈调控机制，节律紊乱可诱发或加重银屑病。我们在研究银屑病病理机制过程中，应用测序技术在两种银屑病动物模型中发现了昼夜节律钟核心基因（Per2和REV-ERBα），初步证实银屑病与昼夜节律钟存在关联性。在银屑病发病机制中占主导作用的Th17细胞分化受REV-ERBα的调控。凉血解毒方是治疗银屑病血热证的有效中药复方，临床观察和基础实验均证实该方能明显抑制Th17细胞核转录因子RORγt和炎症因子IL-17A等表达。本研究通过分别构建咪喹莫特诱导昼夜节律紊乱小鼠和mPer2m/m小鼠银屑病模型，体外Th17细胞和REV-ERBα转染后Th17细胞实验，观察凉血解毒方对Th17细胞分化及相关细胞因子和昼夜节律钟基因表达的影响，尝试从时间医学角度通过REV-ERBα/RORγt调控Th17细胞分化途径探讨昼夜节律对银屑病的影响和凉血解毒方干预的分子机制。

昼夜节律是生命体内在的自我反馈调控机制，节律紊乱影响银屑病的发生发展过程。本研究采用网络药理学研究凉血解毒方对银屑病和昼夜节律相关机制的干预作用；通过咪喹莫特外涂照明紊乱和Per2敲除小鼠皮肤构建昼夜节律紊乱的银屑病样皮损动物模型，观察凉血解毒方对小鼠皮损、褪黑素水平、Th17细胞及炎症因子和昼夜节律钟相关基因的表达；通过体外Th17细胞模型，观察凉血解毒方有效成分（新穿心莲内酯）对Th17细胞分化及分泌炎症因子、昼夜节律钟基因的影响，探讨昼夜节律对银屑病的影响及凉血解毒方干预的分子机制。研究结果表明：网络药理学分析HIF-1通路和Th17细胞分化可能是凉血解毒方干预银屑病和昼夜节律相关机制的主要途径；凉血解毒方通过调控昼夜节律钟相关基因（CLOCK/BMAL1）、Th17细胞分化（REV-ERBα/RORγt）和HIF-1通路改善咪喹莫特诱导照明紊乱小鼠银屑病样皮损，降低PASI评分，减轻表皮厚度，抑制表皮增殖和角化不全，降低脾指数，抑制血清和皮损中炎症因子IL-17A和IL-17F；凉血解毒方通过调控昼夜节律基因（BMAL1）和Th17细胞分化（REV-ERBα/RORγt和NFIL3）改善咪喹莫特诱导Per2敲除小鼠银屑病样皮损，降低PASI评分，减轻表皮厚度，抑制表皮增殖和角化不全，调节褪黑素水平，降低脾指数和抑制脾脏Th17细胞数，抑制血清和皮损中炎症因子IL-17A和IL-17F；凉血解毒方中有效成分（新穿心莲内酯）能够激活REV-ERBα抑制Th17细胞分化及其分泌炎症因子IL-17A和IL-22。因此，凉血解毒方可通过昼夜节律相关基因调控Th17细胞分化改善银屑病，从而为银屑病或相关疾病的时间治疗实践提供一些初步的启示。