HMGB1在氧化应激诱导白癜风黑素细胞免疫损伤中的作用及机制研究

Vitiligo is an autoimmune skin disease. Under oxidative stress, the level of chemokine in skin is elevated, thus inducing the colonization of CD8+ T cells and in turn destroy the cutaneous melanocytes, which has been demonstrated to be the key event in the pathogenesis of vitiigo. In our previous research, we have found that oxidative stress could induce the keratinocytes to produce low level of chemokine, which, however, could be significantly elevated after the involvement of melanocytes. Resorting to screening analysis, we found that oxidative stress could facilitate the melanocytes to release abundant HMGB1 that has been proved to have a role in the production and secretion of chemokine by activating the pattern recognition receptors. Based on this, we hypothesize that oxidative stress can promote the release of HMGB1 from the melanocytes and activate its receptors on keratinocytes, thus facilitating the secretion of chemokine and in turn inducing CD8+ T cells to undermine melanocytes, which ultimately results in the development of vitiligo. To test our hypothesis, this project will investigate the role of oxidative stress in the release of HMGB1 from melanocytes as well as the involved mechanism, and elucidate the effect of HMGB1 on the secretion of chemokine from keratinocytes as well as the related signaling pathways. Moreover, we will construct the 3-dimensional model of epidermis culture to testify the phenomenon and mechanism as described previously. The results of this project can help to complement the immune pathogenesis of vitiligo, and provide new target molecules for its treatment.

白癜风是一种自身免疫性皮肤病，在氧化应激环境下，皮损局部趋化因子水平升高，诱导CD8+T细胞杀伤黑素细胞（MC）是其发病的关键环节，但其中趋化因子的产生机制尚不清楚。我们在前期研究中发现氧化应激仅可诱导角质形成细胞（KC）产生少量趋化因子，但在MC参与后其趋化因子分泌水平显著升高；同时通过分子筛选发现氧化应激诱导MC释放大量HMGB1，其已被证实可通过活化模式识别受体参与趋化因子的生成与分泌。据此我们提出研究假说：氧化应激诱导MC释放HMGB1，活化其在KC上的受体，促进KC分泌趋化因子，诱导CD8+T细胞杀伤MC，导致白癜风的发生。为了验证这一假说，本项目拟在细胞水平验证氧化应激诱导MC释放HMGB1的功能及其机制，同时明确HMGB1对KC分泌趋化因子的作用及相关信号通路，并建立三维表皮培养模型对上述现象和机制加以验证，预期结果将完善白癜风的免疫发病机制，并为其治疗提供新靶点。

白癜风是因黑素细胞（MC）被破坏，局部或全身形成白斑的一种多发、难治性皮肤病，严重影响患者的容貌和心理健康。氧化应激环境下，皮损局部趋化因子水平升高，诱导CD8+ T细胞杀伤MC是其发病的关键环节，但其中趋化因子的产生机制尚不清楚。本课题在前期研究中发现氧化应激仅可诱导角质形成细胞（KC）产生少量趋化因子，但在MC参与后其趋化因子分泌水平显著升高；同时通过分子筛选发现氧化应激诱导MC释放的大量HMGB1，其已被证实可通过活化模式识别受体参与趋化因子的生成与分泌。据此提出以下科研假说：氧化应激诱导MC释放HMGB1，活化其在KC上的受体，促进KC分泌趋化因子，诱导CD8+ T细胞杀伤MC，导致白癜风的发生。为了验证这一假说，本项目主要的研究内容为拟在细胞水平验证氧化应激诱导MC释放HMGB1的功能及其机制，同时明确HMGB1对KC分泌趋化因子的作用及相关信号通路。目前已获得如下重要结果和关键数据：①本课题首次在白癜风患者皮损中观察发现黑素细胞中的HMGB1由细胞核转位至细胞浆，同时在过氧化氢体外构建的黑素细胞氧化应激模型上清液中可检测到HMGB1的分泌增加；②我们进一步研究证实HMGB1通过与角质形成细胞表面的RAGE受体结合，启动下游NF-κB及MAPK-erk信号通路，调控趋化因子IL-8及CXCL16的分泌；③其中受HMGB1调控的趋化因子CXCL16对患者外周血中CD8+ T细胞的招募起关键作用；④同时观察发现HMGB1可以促进白癜风患者外周血树突状细胞细胞表面成熟标志物CD80、CD86及HLA-DR的表达。通过上述实验结果，我们证实了白癜风患者氧化应激状态下，可以引起MC释放HMGB1，与KC表面受体RAGE结合后，激活下游NF-κB及MAPK-erk信号通路，诱导KC产生大量趋化因子CXCL16及IL-8，继而募集特异性CD8+ T细胞杀伤黑素细胞，此过程中树突状细胞可能发挥了抗原呈递的作用。以上结果完善了白癜风的免疫发病机制，一定程度上为白癜风氧化应激启动自身免疫提供了有力的理论依据，在受体及相关通路机制的研究结论为白癜风的治疗提供了新的靶点。