Mechanism of Impaired Diabetic Oral Wound Healing

描述(申请人提供)：糖尿病已成为一个日益重要的健康问题。糖尿病的一个严重并发症是口腔伤口愈合受损，这对创伤和手术的反应产生了负面影响。尽管被美国国立卫生研究院确定为一个重要的健康问题，但令人惊讶的是，很少有研究研究糖尿病损害口腔结缔组织愈合的机制。这一点很重要，因为口腔伤口的愈合受到与口腔相关的独特压力的影响，包括沐浴在粘膜表面的细菌和咀嚼的力量等。初步数据表明，口腔创伤损害了愈合，它们表现出FOXO1 DNA结合活性增加和成纤维细胞核移位。根据初步数据和已发表的报道，糖尿病创面中FOXO1的增加可能通过增加成纤维细胞的凋亡、抑制成纤维细胞的增殖和增强促炎介质的表达来干预创面的愈合过程。FOXO1在口腔糖尿病创面愈合中的潜在作用将在目标1中进行研究，转基因小鼠携带FLOXO1，成年糖尿病和正常血糖小鼠的FoxO1将通过他莫昔芬诱导的Cre重组酶缺失。此外，Cre重组酶将被限制在成纤维细胞系的细胞中，这将避免FOXO1全局缺失的陷阱。通过这种方法，我们将确定糖尿病增强的成纤维细胞中FOXO1的上调是否有助于糖尿病的改变愈合。通过研究FOXO1缺失调节的基因，我们还将研究FOXO1可能影响愈合过程的特定下游靶点。目的2将研究糖尿病增强FOXO1的三个具体机制。这些因素包括肿瘤坏死因子-α的增加，晚期糖基化终末产物通过RAGE的更多信号，以及口腔细菌的影响。这将通过使用特定的肿瘤坏死因子、RAGE和抗生素抑制剂来实现，以减少细菌负担。这些机制与初步数据一致，表明肿瘤坏死因子和RAGE信号可在体外刺激成纤维细胞中FOXO1的激活，并表明通过抗生素治疗可以促进免疫缺陷动物的口腔伤口愈合。综上所述，我们建议验证糖尿病增强的FOXO1干扰口腔伤口愈合的假说，并研究糖尿病导致成纤维细胞中FOXO1增加的机制。这些研究将探索糖尿病损害口腔伤口愈合的一种新机制，以及一种以前从未被研究过的任何形式的伤口愈合。我们期望他们能更好地了解导致糖尿病并发症的步骤。 公共卫生相关性：拟议研究的目标是调查糖尿病通过增加FOXO1干扰口腔伤口愈合的机制。这是基于初步数据和已发表的报告，FOXO1刺激成纤维细胞凋亡，减少增殖，增加炎症介质的表达。这是引人注目的，因为糖尿病伤口愈合的特点是更多的成纤维细胞凋亡和减少的增殖以及长期的炎症。将研究糖尿病增强FOXO1的三种可能机制，包括更高水平的肿瘤坏死因子-a，通过RAGE传递晚期糖基化终末产物的信号，以及口腔细菌的影响。

DESCRIPTION (provided by applicant): Diabetes has become an increasingly important health issue. A serious complication of diabetes is impaired oral wound healing, which negatively affects the response to trauma and surgery. Despite being identified by NIH as an important health issue there have been surprisingly few studies that have examined mechanisms by which oral connective tissue healing is impaired by diabetes. This is significant since oral wound healing is subject to unique stresses associated with the oral cavity including bacteria which bathe the mucosal surfaces and forces of mastication, amongst others. Preliminary data demonstrate that oral wounds have impaired healing and that they exhibit increased FOXO1 DNA binding activity and nuclear translocation in fibroblastic cells. Based upon Preliminary Data and published reports increased FOXO1 in diabetic wounds could interfere with the wound healing process by increasing fibroblast apoptosis, decreasing fibroblast proliferation and enhancing the expression of pro- inflammatory mediators. The potential role of FOXO1 in oral diabetic wound healing will be studied in Aim 1 in transgenic mice with floxed FOXO1 that will be deleted in adult diabetic and normoglycemic mice by tamoxifen induced Cre recombinase. Moreover, Cre recombinase will be restricted to cells of the fibroblast lineage, which will avoid the pitfalls of global FOXO1 deletion. By this approach we will determine whether diabetes-enhanced FOXO1 upregulation in fibroblasts contributes to altered diabetic healing. By investigating genes that are modulated by FOXO1 deletion we will also examine specific downstream targets through which FOXO1 may affect the healing process. Aim 2 will investigate three specific mechanisms for diabetes enhanced FOXO1. These include increased TNF-a, greater signaling of advanced glycation end products through RAGE and the impact of oral bacteria. This will be accomplished by the use of specific inhibitors of TNF and of RAGE and antibiotics to decrease the bacterial burden. These mechanisms are consistent with Preliminary Data which indicate that TNF and RAGE signaling stimulate FOXO1 activation in fibroblasts in vitro and that oral wound healing can be improved in immunodeficient animals through antibiotic treatment. In summary, we propose to test the hypothesis that diabetes enhanced FOXO1 interferes with oral wound healing and to investigate mechanisms through which diabetes leads to greater FOXO1 in fibroblastic cells. These studies will investigate a novel mechanism for diabetes impaired oral wound healing and one that has not been investigated previously in any form of wound healing. We expect that they will provide a better understanding of the steps that lead to this diabetic complication. PUBLIC HEALTH RELEVANCE: The goal of the proposed studies is to investigate mechanisms by which diabetes interferes with the oral wound healing through increased FOXO1. This is based on Preliminary Data and published reports that FOXO1 stimulates fibroblast apoptosis, reduces proliferation and increases the expression of inflammatory mediators. This is striking since diabetic wound healing is characterized by greater fibroblast apoptosis and reduced proliferation as well as prolonged inflammation. Three potential mechanisms for diabetes enhanced FOXO1 will be examined including greater levels of TNF-a, signaling of advanced glycation end products through RAGE and the effect of oral bacteria.