Melanocyte-Keratinocyte Cross-Talk In Relation To Barrier Function

描述（由申请人提供）：我们已经证明，与种族或民族无关的深色色素沉着赋予表皮增强的屏障功能和抗菌防御。因此，浅色皮肤本来就更容易发生炎症性皮肤疾病，如特应性皮炎和成人湿疹，以及所有类型的皮肤感染。但色素沉着与表皮功能之间联系的基础仍不清楚。使用与黑素细胞一起生长的器官型人角质细胞培养物，以及选定的小鼠模型，我们将首先评估黑素细胞密度和黑素细胞色素沉着在调节表皮功能中的作用。功能终点将包括渗透性屏障稳态、角质层（SC）完整性/内聚性的变化，以及某些有助于皮肤抗菌防御的关键抗菌肽的表达。由于我们最近的研究表明，深色黑色素细胞通过酸化外表皮来影响表皮功能，我们接下来将评估黑色素细胞酸化表皮的细胞基础；然后在突变小鼠模型中，黑素体形成，酸化或分泌的步骤调节了这一过程。如果酸化本身不能解释黑素细胞对表皮功能的影响，我们计划寻找其他黑素细胞发起的以旁分泌方式影响表皮功能的调节信号。在目标3中，我们将研究相反的问题；也就是说，表皮屏障需求的改变、表面pH值的升高或环境湿度（UV-B）的降低如何调节滤泡间（表皮）色素沉着，从而以前馈式方式改善屏障功能。我们之前的研究已经表明，屏障破坏会增加IL-1a和NGF的产生，而破坏屏障的UV-B剂量会上调p53的表达。已知这些信号通过增加表皮POMC表达和aMSH的蛋白水解加工来增加表皮色素沉着，和/或黑素细胞功能的MC1R信号。我们已经确定了另外两种角化细胞调节信号也可以将表皮屏障功能的变化与色素沉着联系起来；即1)表皮干细胞因子，诱导黑色素细胞从毛囊迁移到毛囊间表皮；ii)转录因子foxn1，它将黑素细胞定位到基底层。我们将评估屏障需求的改变是否会调节这5种信号机制中的一种或多种，从而导致黑色素生成的关键下游酶的表达或活性增加；即酪氨酸酶、酪氨酸酶羟化酶和/或黑素小体酸化。最后，我们将评估TRPV4（表皮传感器或外部湿度）是否调节色素对环境湿度降低的反应。总之，这些研究将为色素沉着在调节表皮功能中的作用提供新的见解，可能解释不同色素组中疾病易感性的差异，并可能导致浅色素表皮的疾病预防。相反，他们应该查明表皮来源的信号，这些信号调节毛囊间色素沉着的发展，以应对外源应激。

DESCRIPTION (provided by applicant): We have shown that dark pigmentation independent of race or ethnicity, endows the epidermis with enhanced barrier function and antimicrobial defense. As a result, lightly-pigmented skin should be inherently more at risk for development of inflammatory skin disorders, such as atopic dermatitis and adult eczemas, well as all types of skin infections. But the basis for the link between pigmentation and epidermal function remains unknown. Using organotypic human keratinocyte cultures, grown with melanocytes, as well as selected mouse models, we will assess first, the role of melanocyte density and the extent of melanocyte pigmentation in regulating epidermal function. The functional end-points will comprise changes in permeability barrier homeostasis, stratum corneum (SC) integrity/cohesion, and expression of certain key antimicrobial peptides that contribute to cutaneous antimicrobial defense. Since our recent studies suggest that darkly-pigmented melanocytes influence epidermal function by acidifying the outer epidermis, we next will assess the cellular basis for epidermal acidification by melanocytes; and then in mutant mouse models, which step in melanosome formation, acidification, or secretion regulates this process. If acidification alone fails to account for the impact of melanocytes on epidermal function, we plan to search for other melanocyte-initiated regulatory signals that influence epidermal function in a paracrine fashion. In Aim 3 we will examine the converse issue; i.e., how alterations in epidermal barrier requirements, elevations in surface pH, or a reduced environmental humidity ( UV-B) could regulate interfollicular (epidermal ) pigmentation, which then would improve barrier function in a feed forward fashion. Our prior studies have already shown that barrier disruption increases both IL-1a and NGF production, and that dose of UV-B that damage the barrier upregulate p53 expression. These signals are known to either increase epidermal pigmentation by increasing epidermal POMC expression and proteolytic processing to aMSH, and/or MC1R signalling of melanocyte function. We have identified two other keratinocyte regulatory signals could also link changes in epidermal barrier function to pigmentation; i.e., i) epidermal stem cell factor, which induces melanocytes to migrate from follicles to interfollicular epidermis; and ii) the transcription factor, foxn1, which targets melanocytes to the basal layer. We will assess whether altered barrier requirements regulate one or more of these 5 signaling mechanisms, leading to increased expression or activity of the key downstream enzymes of melanin production; i.e., tyrosinase, tyrosinase hydroxylase, and/or melanosome acidification. Finally, we will assess whether TRPV4, the epidermal sensor or external humidity, regulates pigmentary responses to decreased environmental humidity. Together, these studies will provide new insights into the role of pigmentation in regulating epidermal function, potentially explaining differences in disease susceptibility in different pigment groups, and perhaps leading to disease prevention in lightly-pigmented epidermis. Conversely, they should pinpoint epidermal-derived signals that regulate the development of interfollicular pigmentation in response to exogenous stress. PUBLIC HEALTH RELEVANCE: Decreased pigmentation reduces veterans' threshold for the development of inflammatory skin conditions and infections; while conversely, more pigmentation provides protection. These studies will reveal mechanisms whereby melanocytes regulate epidermal function, and vice-versa; i.e., how epidermal functional requirements in turn regulate pigmentation. Our research could point to straight-forward methods that could improve the health status of individuals at risk due to fair pigmentation, including veterans with atopic dermatitis and psoriasis.