Skin Barrier Adaptation

摘要 皮肤表皮为环境保护提供了关键的第一道防线。然而，调解机制 表皮角质形成细胞是如何适应和响应外部环境的，人们对此知之甚少。Involucrin (IVL)是角质形成细胞周围角化包膜的主要支架蛋白。我们的 以前的工作支持总环蛋白在表皮对环境的适应性反应中的功能作用， 急性微生物暴露和炎症。(1)我们最近发现了对IVL单倍型的选择性扫描 北欧(CEU)人口。IVL CEU单倍型与IVL表达增加相关 日晒皮肤与非日晒皮肤的比较[根据基因-组织表达 项目(GTEx)数据]。IVL CEU的正选择与日益增长的北纬直接相关， 这表明在北部的克隆中增加IVL的表达对健康有好处。(2)IVL是唯一大的 表皮分化复合体(EDC)编码的皮肤屏障蛋白表现出显著更高的水平 在常规饲养(CR)实验室小鼠的表皮中与无菌(GF)和重组小鼠的表皮进行比较 皮肤微生物组(RSM)小鼠(急性暴露于CR共生微生物群的GF小鼠)，表明 角质形成细胞对微生物相互作用的反应。(3)IVL-/-小鼠皮肤具有保护作用。 MC903诱导炎症并降低维生素D受体的表达，提示IVL的调节作用 在表皮中传递维生素D信号。我们假设IVL水平是专门校准的，以产生 一种适应性皮肤屏障，可根据环境进行微调。我们将通过以下方式验证这一假设 实现三个目标。目的1.确定增加IVL基因表达的致病基因变异 级别。IVL CEU包括增强子923，如下面所证明的那样，该增强子是表达总状蛋白靶基因所必需的 923-/-小鼠IVL表达缺失。角质形成细胞的细胞和分子分析将被用来鉴定 增强子变异(S)调控靶基因表达的分子机制。目标2.确定 总蛋白在皮肤对共生微生物区系适应性反应中的作用。我们会检查皮肤和 再生IVL-/-CV小鼠的炎症反应作为皮肤适应的功能测量。目标3. 确定包皮蛋白调节维生素D受体表达的分子机制 在表皮中发出信号。MC903处理的IVL/耳皮肤没有表现出严重的炎症，并表现出 VDR表达降低。我们假设Involucrin的作用是积极调节VDR的表达和 通过磷酸化的p38δ发出信号，这是一种在其他类型的人类细胞中已知的vdr激活剂。我们的假设是 使用成分活性p38δ和体外基因敲除研究进行了严格测试，预期结果 VDR表达和信号转导分别增加和减少。这项研究将确定分子 参与人类皮肤屏障进化和适应的机制，可发展为创新 治疗皮肤屏障受损的炎症性皮肤病的治疗方法。

ABSTRACT The skin epidermis provides a critical first line of environmental defense. However, the mechanisms mediating how epidermal keratinocytes adapt and respond to the external environment are poorly understood. Involucrin (IVL) is the major scaffold protein of the cornified envelope surrounding the differentiated keratinocyte. Our previous work supports a functional role for involucrin in epidermal adaptive responses to the environment, acute microbial exposure, and inflammation. (1) We discovered recent selective sweep for an IVL haplotype in the northern European (CEU) population. This IVL CEU haplotype is associated with increased IVL expression in sun-exposed skin compared to that in non-sun-exposed skin [according to Genotype-Tissue Expression Project (GTEx) data]. Positive selection of IVL CEU is directly correlated with increasingly northern latitudes, suggesting a fitness benefit of increased IVL expression in northern clines. (2) IVL was the only large Epidermal Differentiation Complex (EDC)-encoded skin barrier protein that exhibited significantly higher levels in the epidermis of conventionally raised (CR) lab mice compared to those in germ-free (GF) and reconstituted skin microbiome (RSM) mice (GF mice acutely exposed to CR commensal microbiota), suggesting an adaptive keratinocyte response to microbial interactions. (3) Ivl-/- mouse skin exhibited a protective effect against MC903-induced inflammation with reduced vitamin D receptor expression, suggesting a regulatory role for IVL in vitamin D signaling in the epidermis. We hypothesize that IVL levels are specifically calibrated to generate an adaptive skin barrier that is finely tuned in response to the environment. We will test this hypothesis by performing three aims. Aim 1. Determine the causative genetic variants that increase IVL gene expression levels. IVL CEU includes enhancer 923 that is required for involucrin target gene expression as evidenced by loss of Ivl expression in 923-/- mice. Cellular and molecular assays in keratinocytes will be performed to identify the molecular mechanisms by which enhancer variation(s) modulate target gene expression. Aim 2. Determine the role of involucrin in the skin adaptive response to commensal microbiota. We will examine the skin and inflammatory responses in rederived Ivl-/- CV mice as a functional measurement of skin adaptation. Aim 3. Determine the molecular mechanism by which involucrin regulates Vitamin D receptor expression and signaling in the epidermis. MC903-treated Ivl-/- ear skins did not exhibit severe inflammation and exhibited reduced Vdr expression. We hypothesize a role for Involucrin to positively regulate Vdr expression and signaling via phosphorylated p38δ, a known VDR activator in other human cell types. Our hypothesis will be rigorously tested using constitutively active p38δ and knockdown in vitro studies with anticipated results for increased and decreased Vdr expression and signaling, respectively. This study will identify the molecular mechanisms involved in human skin barrier evolution and adaptation, which can be developed into innovative therapies for treating skin-barrier impaired inflammatory skin diseases.