Determining Host-microbiome guided oro-nasal fistula healing

摘要 摘要腭裂是儿童最常见的颅面畸形之一，其发病率为1：1000。 出生。患者一生中经历了多次手术，对许多人来说，第一次手术是修复 他们的腭裂。然而，60%的腭裂患者在手术后伤口愈合不良， 导致口鼻瘘(ONF)的发展，其特征是口鼻瘘之间持续沟通 口腔和鼻腔。修复ONF的主要方法是使用人类供体组织，它起作用 只是作为一种物理屏障，并带有与人类捐赠者组织相关的疾病风险，如普恩 或者是艾滋病毒感染。因此，迫切需要开发新的治疗策略来改善伤口。 腭裂修复术后愈合，减少ONF的病例。伤口愈合已被研究过 广泛地在皮肤上，特别是在肠道损伤之后。肠道创面愈合效率高 对环境因素敏感，特别是微生物组，在那里特定的微生物群落结构或 补充益生菌可增强伤口愈合过程。然而，人们对此知之甚少。 口腔微生物群如何影响ONF伤口愈合。我们直接解决了这一知识差距，并展示了 初步数据表明，在小鼠模型中创建ONF会导致微生物组的显著变化 某些微生物完全消失，其他细菌类群大量繁殖。基座 在这个前提下，在目标1中，我们将在无菌小鼠或常规饲养的小鼠身上制造ONF伤口 用抗生素治疗，评估治愈率和口腔微生物组组成的变化。我们预计 以显示破坏口腔微生物群对愈合过程和ONF形成的影响。在目标2中，我们 将使用水凝胶将手术后丢失的口腔共生微生物重新引入口腔伤口部位 并评估治愈率。我们期望证明口腔共生细菌对伤口部位的再补充可以 促进伤口愈合过程。此外，通过评估RNA-seq在ONFS中的转录丰度 无菌、抗生素治疗的小鼠，在口腔共生细菌被清除后，我们希望能识别出 在对口腔共生微生物群敏感的ONF内发挥愈合功能的基因网络。因此， 我们的总体假设是，口腔共生微生物群对口腔具有积极的调节作用。 伤口愈合，并可能限制神经营养因子的形成。在口腔或伤口内发现特定的细菌分类群 积极影响愈合的组织将是描述优生菌口腔的基本信息 用于伤口愈合的微生物群。我们的实验还将为临床医生提供有关 腭裂术后抗菌药物的谨慎使用及口腔微生物群耗竭的影响 治愈。我们还将证实使用水凝胶补充口腔共生菌的方法。 以科技为载体，促进口腔伤口愈合。总之，这些研究将产生关键数据 提示新的治疗方法，以降低腭裂手术后ONFS的患病率。

SUMMARY Cleft palate formation is one of the most common craniofacial anomalies in children and occurs in 1:1000 live births. Patients undergo multiple surgeries during their lifetime and, for many, the first surgery is the repair of their cleft palate. However, 60% of cleft palate patients suffer poor wound healing following surgery which leads to the development of an oro-nasal fistula (ONF) characterized by a continued communication between the oral and nasal cavities. The primary approach to repair an ONF is by using human donor tissue, which acts only as a physical barrier and carries the risks of disease associated with human donor tissue, such as prions or HIV infection. Therefore, there is an urgent need to develop new therapeutic strategies to improve wound healing after cleft palate surgical repair, and to reduce the cases of ONF. Wound healing has been studied extensively on the skin and particularly following intestinal injury. Intestinal wound healing efficiency is highly sensitive to environmental factors, especially the microbiome, where specific microbial community structures or supplementation with probiotic bacteria enhances the wound healing process. However, little is known about how the oral microbiome affects ONF wound healing. We directly address this gap in the knowledge and show preliminary data that creating an ONF in a murine model results in marked changes in the microbiome composition, with the complete disappearance of certain microbes, and blooms of other bacterial taxa. Based on this premise, in Aim 1, we will create ONF wounds in germ-free mice, or in conventionally raised mice treated with antibiotics, and assess healing rates and changes in the oral microbiome composition. We expect to show the influence of disrupting the oral microbiome on healing processes and ONF formation. In Aim 2, we will use a hydrogel to re-introduce commensal oral microbes lost following surgery to the site of the oral wound and assess healing rates. We expect to show that repletion of oral commensal bacteria to the wound site can promote wound healing processes. In addition, by assessing transcript enrichment by RNA-seq within ONFs in germ-free, antibiotic treated mice, and after the repletion of oral commensal bacteria, we expect to identify gene networks that function in healing within ONFs that are sensitive to the commensal oral microbiome. Thus, our overall hypothesis is that the oral commensal microbiome exerts positive modulatory influences on oral wound healing and may limit ONF formation. Discovering specific bacterial taxa within the oral cavity or wound tissue that positively influence healing will be essential information for the characterization of an eubiotic oral microbiome for wound healing. Our experiments will also generate critical information for clinicians about the prudent use of antibiotics after cleft palate surgery and the effects of depleting the oral microbiome during healing. We will also substantiate the approach of the repletion of the oral commensal bacteria using hydrogel technology as the vehicle to enhance oral wound healing. Together, these studies will yield critical data to inform new therapeutic modalities to lower the prevalence of ONFs following cleft palate surgery.