Correcting the Cellular Ecology of Diabetic Complications

项目摘要/摘要 糖尿病影响了近10%的成年人口(3000万)，预计到2019年，这一数字将翻一番 2050年。糖尿病的病理生理学严重损害了所有组织修复过程，导致 受影响患者的慢性不可愈合伤口。糖尿病足溃疡影响15%-30%的糖尿病人 在美国，这是导致截肢的主要原因。传统型 治疗糖尿病的方法，如胰岛素或口服降糖药，可以控制疾病，但不能。 预防糖尿病并发症，即使在几十年后仍有持续进行性器官功能障碍 医疗优化。这凸显了对新的治疗方法的明显需求。在过去的15年里 在美国国立卫生研究院的资助下，我们的实验室为我们理解关键分子和 正常和糖尿病组织修复中的细胞通路。我们已经确定了与高血糖相关的损害 局部微环境和祖细胞归巢和细胞因子的产生 糖尿病并发症的发病机制。我们已经证明，糖尿病会导致关键细胞耗尽。 亚群，导致新生血管减少和组织愈合受损。要了解 糖尿病对细胞种群动力学的影响更精确，我们开发了新的单细胞 -组学技术，在单细胞水平上识别细胞亚群中的关键扰动。这是我们的 糖尿病改变异质细胞群“细胞生态”的基本假设 组织修复和这些细胞亚群正常化可以治疗或逆转糖尿病并发症。 在这个提案中，我们将整合新兴的多模式组学技术，以明确地描述 细胞亚群在糖尿病并发症中的行为，包括伤口愈合。我们将继续开展这项工作。 通过使用基于细胞的方法正常化这些缺陷来治疗和预防糖尿病 并发症。首先，我们将使用一种新的多路方法进行高通量的单细胞测序 明确描述人类糖尿病患者驻留组织和祖细胞亚群的行为 和非糖尿病创面(具体目标1)。然后我们将在动物模型中证实这些人类观察到的结果 通过将单元格测序与下一代空间技术相结合，以空间分辨率定义这些变化 转录组和蛋白质组学技术并精确地描绘出三维伤口的位置 环境存在这些差异(具体目标2)。最后，我们将利用这些信息来优化系统 提供以细胞为基础的治疗药物，以防止或逆转糖尿病引起的相关细胞缺陷 减少糖尿病患者的风险，从而纠正糖尿病并发症(具体目标3)。综上所述，这一新颖的方法 识别、空间表征和纠正糖尿病并发症中的亚群缺陷将提供 对糖尿病病理生理学的新见解，并为预防和治疗这些并发症提供新的策略。

PROJECT SUMMARY / ABSTRACT Diabetes affects nearly 10% of the adult population (30 million), and these numbers are expected to double by the year 2050. The pathophysiology of diabetes profoundly impairs all tissue reparative processes, leading to chronic non-healing wounds in affected patients. Diabetic foot ulcers affect between 15-30% of all diabetic individuals and represent the leading cause of lower limb amputations in the United States. Conventional methods to treat diabetes, such as with insulin or oral hypoglycemic agents, can control the disease but do not prevent diabetic complications, as demonstrated by continued progressive organ dysfunction even decades after medical optimization. This highlights a clear need for new therapeutic approaches. Over the past 15 years of NIH funding, our laboratory has made important contributions to our understanding of the critical molecular and cellular pathways in normal and diabetic tissue repair. We have identified hyperglycemia-related impairments in both the local microenvironment and progenitor cell homing and cytokine production that contribute to the pathogenesis of diabetic complications. We have demonstrated that diabetes results in depletion of critical cell subpopulations, resulting in decreased neovascularization and impaired tissue healing. To understand the effects of diabetes on cell population dynamics with greater precision, we have developed novel single cell “-omics” techniques to identify critical perturbations in cell subpopulations at the single cell level. It is our fundamental hypothesis that diabetes alters the “cellular ecology” of heterogeneous cell populations involved in tissue repair and that normalization of those cell subpopulations can treat or reverse diabetic complications. In this proposal, we will integrate emerging multimodal -omics technologies to definitively characterize the behavior of cell subpopulations in diabetic complications, including wound healing. We will extend this work therapeutically by using cell-based approaches to normalize these defects to treat and prevent diabetic complications. To begin, we will employ a novel multiplex approach for high-throughput single cell sequencing to definitively characterize the behavior of resident tissue and progenitor cell subpopulations in human diabetic and non-diabetic wounds (Specific Aim 1). We will then confirm these human observations in animal models and define these changes with spatial resolution by integrating single cell sequencing with next-generation spatial transcriptomic and proteomic technologies and precisely delineate where in the three-dimensional wound environment these differences exist (Specific Aim 2). Finally, we will use this information to optimize the systemic delivery of cell-based therapeutics in order to prevent or reverse diabetes-induced defects in relevant cell populations and thereby correct diabetic complications (Specific Aim 3). Taken together, this novel approach for identifying, spatially characterizing, and correcting subpopulation deficits in diabetic complications will provide new insights into diabetic pathophysiology and inform novel strategies to prevent and treat these complications.