Exosomes in the Pathogenesis of Diabetic Atherosclerosis & its Treatment Opportunities

这项提案的目的是了解为什么糖尿病患者会出现严重的富含脂肪的斑块， 动脉粥样硬化加速的动脉粥样硬化与几种类型的心血管疾病有关。 导致糖尿病患者残疾和过早死亡的疾病。不幸的是，如此严重的 医疗问题是非常频繁的退伍军人登记在VHA医疗保健系统，因为他们几乎 患糖尿病的可能性是普通人群的三倍。一个原因是 在糖尿病个体中被鉴定为增强动脉粥样硬化的是高血糖，也称为“高血糖症”。 但高血糖症究竟如何增强动脉粥样硬化尚不清楚。 我们最近的研究重点集中在探索细胞释放的微泡的作用，称为 在动脉粥样硬化中，“外来体”作为炎症的来源。我们在修订后的赠款中提出的初步结果 建议显示诊断为晚期外周动脉粥样硬化性心血管疾病人类糖尿病受试者 疾病在其血流中积累促炎性外泌体。我们的发现还表明，糖尿病小鼠 在血液中积累促炎性外来体。此外，我们的研究结果表明，这种糖尿病 血浆外泌体可增加动脉中积聚的白色血细胞的数量 植入非糖尿病小鼠体内。有趣的是，我们的数据表明，促炎性外泌体可以通过 在模拟糖尿病高血糖的富含葡萄糖的培养基中培养的巨噬细胞。值得注意的是，我们的数据还 表明巨噬细胞在含有以下物质的培养基中培养时可以产生抗炎外泌体： 保护性细胞因子如白细胞介素-4。 基于我们广泛的新发现，我们建议探索糖尿病高血糖是否会增强 动脉粥样硬化通过外泌体的进展，外泌体在免疫系统中传递促炎信号 系统和血管壁。我们的目标还在于产生外泌体， 糖尿病性动脉粥样硬化的进展。 在我们的第一个目标中，我们将定义高血糖导致促炎性细胞因子产生的程度。 患有晚期动脉粥样硬化疾病的退伍军人的血液中的外泌体。我们也将这样做 通过研究糖尿病小鼠模型。接下来，我们将测试糖尿病血浆外泌体增强 血管炎症和动脉粥样硬化。我们还将测试 在含有高水平葡萄糖或称为oxLDL的坏胆固醇的培养基中培养的巨噬细胞， 当注入非糖尿病小鼠时，产生可以诱导动脉粥样硬化的外泌体。最后，我们将探索 如果外泌体携带的一类称为microRNA的信号分子负责它们的炎症反应， 发信号。 在我们的第二个目标中，我们将寻求生产治疗性外泌体。我们的策略是 来自巨噬细胞的外泌体，所述巨噬细胞在含有包括白细胞介素-4的保护因子的培养基中培养。我们 第二种策略将是从培养的巨噬细胞中生产生物工程外泌体， 并测试这些外泌体是否可以发挥抗炎特性， 减少糖尿病小鼠动脉粥样硬化的进展。

The goal of this proposal is to understand why people with diabetes develop severe fatty-rich plaques called atherosclerosis in arteries. Accelerated atherosclerosis has been linked to several types of cardiovascular diseases that cause disabilities and premature death amongst diabetic individuals. Unfortunately, such serious medical problems are very frequent among veterans enrolled in the VHA Health Care System as they are almost three times more likely to develop diabetes than people in the general population. One reason that has been identified to enhance atherosclerosis among diabetic individuals is high blood sugar, also called “hyperglycemia”. But exactly how hyperglycemia enhances atherosclerosis is not known. Our recent research focus has centered on exploring the role of microvesicles released by cells, called “exosomes”, as a source of inflammation in atherosclerosis. Preliminary findings presented in our revised grant proposal show that human diabetic subjects diagnosed with advance peripheral atherosclerotic cardiovascular disease accumulate pro-inflammatory exosomes in their bloodstream. Our findings also show that diabetic mice accumulate pro-inflammatory exosomes in their bloodstream. Furthermore, our findings show that such diabetic plasma exosomes can increase the number of white blood cells that accumulate in arteries when they are infused into non-diabetic mice. Interestingly, our data demonstrate that pro-inflammatory exosomes can be produced by macrophages cultured in glucose-rich medium that simulates diabetic hyperglycemia. Remarkably, our data also show that macrophages can produce anti-inflammatory exosomes when they cultured in medium that contains protective cytokines such as interleukin-4. Based on our extensive new findings, we propose to explore whether diabetic hyperglycemia enhances the progression of atherosclerosis through exosomes that communicate pro-inflammatory signaling in the immune system and the vessel wall. We also aim to produce exosomes that can serve to overcome the effects of hyperglycemia to suppress the progression of diabetic atherosclerosis. In our First Aim we will define the extent to which hyperglycemia causes the production of proinflammatory exosomes in the bloodstream of veterans suffering from advanced atherosclerotic disease. We will do the same by studying mouse models of diabetes. Next, we will test the ability of diabetic plasma exosomes to enhance vascular inflammation and atherosclerosis when infused into hyperlipidemic mice. We will also test whether macrophages cultured in medium that contains elevated levels of glucose or bad cholesterol called oxLDL, will produce exosomes that can induce atherosclerosis when infused into non-diabetic mice. Lastly, we will explore if a class of signaling molecule called microRNA carried by exosomes are responsible for their inflammatory signaling. In our Second Aim we will seek to produce therapeutic exosomes. Our strategy will consist of isolating exosomes from macrophages cultured in medium that contains protective factors including interleukin-4. Our second strategy will be to produce bioengineered exosomes from cultured macrophage tailored to produce desired levels of protective miRNA and test whether such exosomes can exert anti-inflammatory properties to reduce the progression of atherosclerosis in diabetic mice.