Dermatan sulfate and its role in tissue repair

描述(由申请人提供)：GAG已成为一系列细胞信号事件中的关键信号分子。尽管硫酸皮肤素(DS)是皮肤和伤口液中含量最丰富的GAG，但这代表了伤口愈合的一个区域，仍需通过评估修复所需的细胞信号事件来更广泛地评估。DS通过成纤维细胞生长因子-2和成纤维细胞生长因子-7介导的信号通路刺激细胞增殖，比研究更广泛的GAG-硫酸乙酰肝素(HS)更能刺激细胞增殖。这项研究的目的是进一步表征DS在体外对成纤维细胞生长因子信号的影响，并将这些发现外推到人类生理环境中。总体假设是，与HS相比，成纤维细胞生长因子-10和成纤维细胞生长因子-22更倾向于DS，并且伤口环境中DS结合蛋白的数量可以显著影响组织修复。在目标I中，我们将使用体外生化方法确定在FGF10和成纤维细胞生长因子-22介导的信号转导中，DS是否优于HS。本课程将评估依赖DS的成纤维细胞生长因子/成纤维细胞生长因子受体相互作用的结合能力、增殖能力和结构特异性。目的II旨在评估人伤口液，以确定通过体内分子方法更强地结合DS和HS的FGFs。此外，还将评估DS对成纤维细胞生长因子定位和伤口闭合的影响。这些实验将阐明DS在最佳组织修复所需的成纤维细胞生长因子信号事件中的作用，并表征DS结合蛋白的表达模式，这些蛋白最终可用作病理疾病的诊断标记物。临床上，GAG在皮肤病的发病机制中起一定作用。通过必需的FGFR2-IIIB配体发现DS的依赖性将预测DS合成或降解的紊乱将扰乱正常的伤口修复。具体地说，我们怀疑异常创面中DS的平衡与正常修复时观察到的不同，这一点得到了一些临床研究的支持。因此，分析正常人或疾病患者伤口液中DS的含量和/或硫酸盐化程度是这项研究的一个潜在的临床方向，这可能为伤口异常愈合提供一种诊断工具，并为促进正常伤口修复提供新的治疗方法。总体而言，所提出的技术允许接触到新的分子和生化方法，用来分析蛋白质和碳水化合物的功能以及皮肤伤口愈合过程中的先天免疫。

DESCRIPTION (provided by applicant): GAGs have emerged as critical signaling molecules in an array of cell signaling events. Although dermatan sulfate (DS) is the most abundant GAG in the skin and in wound fluid, this represents an area of wound healing which remains to be more extensively evaluated through the assessment of cellular signaling events required for repair. DS has been shown to stimulate cellular proliferation via Fibroblast Growth Factor-2 (FGF-2) and FGF-7 mediated signaling more so than the more extensively studied GAG, heparan sulfate (HS). The purpose of this study is to further characterize the effect of DS on Fibroblast Growth Factor signaling in vitro, and extrapolate these findings to a human physiologic setting. The overall hypothesis is that FGF-10 and FGF-22 prefer DS over HS, and that the population of DS-binding proteins in the wound milieu can significantly influence tissue repair. In Aim I, we will determine if DS is preferred over HS in FGF10 and FGF-22 mediated signaling using an in vitro biochemical approach. The binding capability, proliferative capacity, and structural specificity of DS-dependent FGF/FGF receptor interactions will be assessed. Aim II is designed to evaluate human wound fluid to identify the FGFs that more strongly bind DS over HS through an in vivo molecular approach. In addition, the effect of DS on FGF localization and wound closure will be assessed. The proposed experiments will elucidate the role of DS in FGF-cell signaling events required for optimal tissue repair and characterize the expression patterns of DS-binding proteins, which can ultimately be used as a diagnostic marker in pathologic disease. Clinically, it has been suggested that GAGs play a role in the pathogenesis of skin disease. Finding a dependence for DS by the essential FGFR2-IIIB ligands would predict that a disorder of DS synthesis or degradation would disrupt normal wound repair. Specifically, we suspect that the balance of DS present in abnormal wounds will be different than that observed during normal repair, which is supported by a few clinical studies. Hence, the analysis of the levels and/or sulfation of DS in human wound fluid from normal or diseased individuals is a prospective clinical direction for this research proposal, which may yield a diagnostic tool for abnormal wound healing and novel therapeutic approaches to promote normal wound repair. Overall, the techniques proposed allow for exposure to new molecular and biochemical methods with which to analyze protein and carbohydrate function and innate immunity in dermal wound healing.