Effects of altering TGF beta signaling on wound healing

描述（由申请人提供）：与各种疾病相关的慢性皮肤创伤（例如，糖尿病）和急性创伤的异常愈合（例如，增生性瘢痕）是主要的卫生保健负担。我们以前的研究表明，当转化生长因子21（TGF 21）过表达的水平与皮肤病中发现的水平相当时，它会诱导严重的皮肤炎症，延迟伤口愈合和过度的纤维化反应。相比之下，其拮抗剂Smad7具有有效的抗炎作用并加速皮肤伤口愈合，同时具有更快的上皮再形成、减少的炎症和减少的纤维化反应。因此，我们假设：1）向伤口床递送Smad7蛋白可以治疗受损的皮肤伤口愈合。2)Smad7通过其转录和蛋白质相互作用活性直接调节伤口愈合期间的角质形成细胞迁移和表皮再上皮化。3)Smad7调节表皮中参与肉芽组织形成和皮肤伤口愈合的基质重塑的分泌分子。为了验证这些假设，目标1将检查局部Smad7蛋白递送是否促进皮肤伤口的愈合和重塑。我们将使用带有达特细胞可渗透标签的人Smad 7重组蛋白来治疗两种受损伤口愈合模型中的切除皮肤伤口。首先，我们将利用K5.TGF21转基因伤口作为炎症相关的受损愈合模型，这是验证Smad7靶向作用的理想模型。第二，我们将治疗糖尿病小鼠的伤口。目的2分析Smad7调控角质形成细胞迁移的分子机制。将在目标1中产生的伤口样品中的病理条件下鉴定和验证调节表皮再上皮化的Smad7转录靶标和蛋白质伴侣。目的3将鉴定影响伤口基质的Smad7转录靶点。我们已经产生了诱导型，表皮特异性Smad7小鼠含有生物素标记的Smad7转基因（Smad7biotin）。在伤口愈合的每个特定阶段，来自这些小鼠的切除伤口将用于高通量分析，以鉴定影响伤口基质的时间Smad7靶标。目标1中产生的样品将用于检查这些靶点中的任何一个是否被Smad7处理改变，以及这些靶点的体内敲低是否影响愈合。这些研究有助于我们进一步了解受损皮肤伤口愈合的病理机制，通过局部递送Smad7或靶向Smad7下游蛋白来测试治疗方法。 公共卫生相关性：在美国，慢性皮肤伤口影响着650万患者。该应用程序执行实验疗法来治疗受损的皮肤伤口愈合，并分析所涉及的分子机制。

DESCRIPTION (provided by applicant): Chronic skin wounds associated with various diseases (e.g., diabetes) and aberrant healing from acute wounding (e.g., hypertrophic scarring) is a major health care burden. Our previous studies show that when transforming growth factor 21 (TGF21) is overexpressed at a level comparable to that found in skin diseases, it induces severe skin inflammation, delayed wound healing and excessive fibrotic response. In contrast, its antagonist, Smad7, has a potent anti-inflammatory effect and accelerated skin wound healing with faster re- epithelialization, reduced inflammation and reduced fibrotic response. Hence, we hypothesize: 1) Smad7 protein delivery to the wound bed can treat impaired skin wound healing. 2) Smad7 directly regulates keratinocyte migration and epidermal re-epithelialization during wound healing through its transcription- and protein-interaction activities. 3) Smad7 regulates secreting molecules in the epidermis involved in granulation tissue formation and stromal remodeling of skin wound healing. To test these hypotheses, Aim 1 will examine if local Smad7 protein delivery promotes healing and remodeling of skin wounds. We will use human Smad7 recombinant protein with a Tat cell permeable tag to treat excisional skin wounds in two impaired wound healing models. First, we will utilize K5.TGF21 transgenic wounds as an inflammation-related impaired healing model, which is an ideal model for validation of the targeted effect of Smad7. Second, we will treat wounds in diabetic mice. Aim 2 will analyze molecular mechanisms by which Smad7 regulates keratinocyte migration. Smad7 transcriptional targets and protein partners regulating epidermal re-epithelialization will be identified and validated under pathological conditions in wound samples generated in Aim 1. Aim 3 will identify Smad7 transcriptional targets affecting the wound stroma. We have generated inducible, epidermal-specific Smad7 mice containing a biotin-tagged Smad7 transgene (Smad7biotin). Excisional wounds from these mice at each specific stage of wound healing will be used for high throughput analyses to identify temporal Smad7 targets affecting the wound stroma. Samples generated in Aim 1 will be used to examine if any of these targets are altered by Smad7 treatment and if in vivo knock down of these targets affects healing. The proposed studies help us further understand pathological mechanisms of impaired skin wound healing, test therapeutic approaches by either local delivery of Smad7 or targeting proteins downstream from Smad7. PUBLIC HEALTH RELEVANCE: Chronic skin wounds affect 6.5 million patients in the United States. This application performs experimental therapeutics to treat impaired skin wound healing and analyzes the molecular mechanisms involved.