Mast cell Stat5-regulatory pathway in atopic dermatitis

描述(申请人提供)：特应性皮炎(AD)是一种慢性瘙痒性炎症性皮肤病，发病率不断上升。虽然阿尔茨海默病的病因还不完全清楚，但大量研究表明皮肤屏障功能受损和免疫失调是该病的基础。我们开发了模拟人类AD的新型过敏原诱导和自发的小鼠模型：由过敏原表面治疗引起的AD样皮肤损害需要肥大细胞和胸腺基质淋巴生成素受体(TSLP)。我们发现，磷脂酶C(PLC)基因缺失的小鼠自发地发生AD样皮肤损害，并以肥大细胞依赖的方式增加TSLP和Periostin的表达；PLC-3-/-小鼠的过敏原诱导的皮炎比野生型小鼠更严重。皮损皮肤的基因表达谱在过敏原诱导的皮炎、PLC？3-/-小鼠的自发性皮炎和人类AD之间有很高的相似性。我们最近报道，PLC-β3通过形成含有SHP-1(蛋白磷酸酶)、PLC-β3和Stat5(转录因子)的多分子SPS复合体，抑制包括肥大细胞在内的造血细胞的增殖；PLC-β3与SHP-1和Stat5相互作用，增强SHP-1对Stat5的去磷酸化活性，并抑制Stat5的活性。Stat5和JAK2(一种Stat5激活蛋白)对肥大细胞的发育和增殖是必不可少的。我们的初步数据表明，肥大细胞Stat5和SHP-1参与了AD的发病机制，表现为肥大细胞特异性缺失的Stat5和SHP-1分别降低或增强了变应原诱导的皮炎的严重程度，以及Jak-Stat5抑制剂对预防变应原诱导的皮炎的强烈效果。此外，人类AD皮肤中具有高激活水平的STAT5的肥大细胞数量增加，编码SPS成分的基因与人类AD的风险和/或其严重性有关。这些和其他初步数据导致了以下假设：(1)肥大细胞中的Stat5调节通路在AD中起着关键的致病作用。(2)SPS成分调节由Th2细胞因子(肥大细胞)、Periostin(成纤维细胞)和TSLP(角质形成细胞)组成的皮炎放大/持续恶性循环中的细胞成分的活动。(3)SPS组分不仅对小鼠有调节作用，对人肥大细胞的生物学也有调节作用。因此，(目的1)利用SPS组分基因条件性敲除小鼠以及药理学方法，我们将进一步评估肥大细胞Stat5调节通路在PLC-3-/-小鼠皮炎发生和持续中的作用。(目的2)利用体外培养和体内模型，我们将研究SPS成分如何调节肥大细胞产生Th2细胞因子、成纤维细胞产生Periostin以及角质形成细胞产生TSLP。(目的3)最后，我们将研究SPS组分在人肥大细胞生长、存活和高亲和力IgE受体(Fc？RI)介导的激活中的作用，并比较人AD和健康人的肥大细胞。总体而言，该项目可能会建立AD发病机制的新范例，并提供一种新的治疗策略。

DESCRIPTION (provided by applicant): Atopic dermatitis (AD) is a chronic pruritic inflammatory skin disease with an increasing prevalence. Although the etiology of AD is not completely understood, numerous studies suggest that impaired skin barrier function and immune dysregulation underlie the disease. We have developed novel allergen-inducible and spontaneous mouse models that mimic human AD: AD-like skin lesions induced by epicutaneous treatment with allergen require mast cells and the receptor for thymic stromal lymphopoietin (TSLP). We found that phospholipase C (PLC)-¿3-deficient mice spontaneously develop AD-like skin lesions with increased expression of TSLP and periostin in a mast cell-dependent manner; allergen-induced dermatitis was more severe in PLC-¿3-/- than in wild-type mice. Gene expression profiles in lesional skin exhibited high similarity between allergen-induced dermatitis, spontaneous dermatitis in PLC-¿3-/- mice, and human AD. We recently reported that PLC-¿3 inhibits the proliferation of hematopoietic cells, including mast cells, through formation of the multi-molecular SPS complex containing SHP-1 (protein phosphatase), PLC-¿3, and Stat5 (transcription factor); PLC-¿3 interacts with SHP-1 and Stat5, augments the dephosphorylating activity of SHP-1 toward Stat5, and inhibits Stat5 activity. Stat5 and Jak2 (a Stat5-activating kinase) are essential for the development and proliferation of mast cells. Our preliminary data indicated the involvement of mast-cell Stat5 and SHP-1 in AD pathogenesis, as revealed by reduced or enhanced severity of allergen-induced dermatitis in mice with mast cell-specific deletion of Stat5 or SHP-1, respectively, as well as strong efficacy of a Jak-Stat5 inhibitor to prevent allergen-induced dermatitis. Moreover, human AD skin had increased numbers of mast cells with high activation levels of STAT5 and the genes encoding the SPS components were linked to the risk of human AD and/or its severity. These and other preliminary data have led to the following hypotheses: (1) The Stat5- regulatory pathway in mast cells plays a critical pathogenic role in AD. (2) SPS components regulate the activity of the cellular elements of the dermatitis-amplifying/perpetuating vicious cycle consisting of Th2 cytokines (mast cells), periostin (fibroblasts), and TSLP (keratinocytes). (3) The SPS components regulate not only mouse, but also human mast cell biology. Therefore, (Aim 1) using conditional knockout mice of SPS component genes as well as pharmacological approaches, we will further evaluate the role of the mast cell Stat5-regulatory pathway in development and persistence of dermatitis in PLC-¿3-/- mice. (Aim 2) Using in vitro cultures and in vivo models, we will study how SPS components regulate Th2 cytokine production from mast cells, periostin production from fibroblasts, and TSLP production from keratinocytes. (Aim 3) Finally, we will study the role of SPS components in the growth, survival, and Fc?RI (high-affinity IgE receptor)-mediated activation of human mast cells and compare mast cells between human AD and healthy individuals. Overall, this project will likely establish a novel paradigm of AD pathogenesis and provide a novel therapeutic strategy.