Differential Macro- and Micro-vascular Remodeling in Type 2 Diabetes and Metaboli

项目总结 2型糖尿病患者的心脏病与冠状动脉病变的严重程度直接相关 心脏病(CAD)，导致冠状动脉血流受损，增加心肌梗死(MI)的风险。这 在美国和全世界造成了非常严重的健康和经济负担，因为心血管疾病 2型糖尿病的主要死因或死亡率，占糖尿病患者死亡总数的65%。此外， 糖尿病患者发生心肌梗死的可能性是非糖尿病患者的2-4倍。而动脉粥样硬化 内皮功能障碍是MI的已知因素，MI是冠状动脉阻力的相对贡献 T2 DM心脏微血管(CRM)重构与大血管(导管动脉)重构的比较 疾病在很大程度上仍是未知的。 我们以前发现，从T2 DM db/db小鼠或临床前Ossabaw小鼠分离出的CRM Met的猪模型经历了与僵硬降低相关的向内肥厚重塑(新颖和 反教条主义)面对大血管僵硬(教条主义)。伴随着CRM重塑 冠脉血流量减少，部分依赖血管平滑肌细胞(VSMC) 增殖，弹性细胞外基质(ECM)表达增加，局部表达降低 黏附/细胞骨架蛋白。相反，局灶性黏附/细胞骨架和细胞外基质蛋白表达 T2 DM db/db小鼠大血管数量增加。这些发现表明， 大血管和冠脉微血管的细胞硬度和细胞外基质决定了整体血管硬度。 T2 DM。我们还观察到，CRMVSMC表现出与增强相关的增殖表型 晚期糖基化终产物(AGE)/RAGE信号通路与半胱氨酸表达降低 富蛋白2(CRP2)是VSMC增殖的负性调节因子。目前还没有研究表明 直接研究了刚性对T2 DM VSMC表型的影响。根据我的初步数据， 不那么僵硬的CRM壁促进体内增殖的VSMC表型，而较硬的主动脉(大血管) WALL支持合成的VSMC表型。这些数据表明，客户关系管理背后的机制 重塑与那些调节大血管重塑的重塑有根本的不同，可能代表着 决定VSMC表型的病理机制导致糖尿病心肌梗死。因此，我的总体假设是 大血管和冠状动脉微血管重塑和僵硬的改变反映了不同的 控制VSMC表型的细胞和细胞外基质硬度的贡献可能部分由其决定 按年龄/愤怒和/或CRP2。 这项提案中概述的研究将利用一种新的 主动脉和CRM的脱细胞技术，原代培养的VSMC，分离的冠脉阻力 微血管，糖尿病小鼠血管重塑和血流的体内分析。在接受指导的过程中 阶段，我们将确定ECM和VSMCs对整体血管硬度的相对贡献 进一步评估僵硬是否可以调节大血管和CRM的经典VSMC表型。 指导研究将主要集中在确定细胞和生物力学机制上。 在原代VSMC培养中，这些变化是潜在的。在独立阶段，我将确定是否 僵硬、年龄/年龄和CRP2的相互作用调节从两者分离的VSMC的表型 糖尿病小鼠和临床前服用蛋氨酸的猪。最后，我将评估一种新的RAGE的治疗潜力 拮抗剂FPS-ZM1减少T2 DM小鼠不良血管重构。 此过渡奖项将使我能够获得未来成功所需的额外指导 有竞争力且成功的独立调查员。该提案包括一项全面的培训计划。 这将由一个指导委员会的成员监督，该委员会拥有涵盖血管的专业知识 信号和生物力学、细胞外基质与糖尿病和代谢的冠状动脉并发症 综合症。一旦完成，我将在假设生成、实验方面获得更多的专业知识 与体外细胞信号、血管力学、细胞外基质和冠状动脉相关的设计和数据解释 这将加强我在体外和体内生理学方面已经很好的专业知识。它将进一步提供 为今后的研究奠定坚实的基础，我想在其中研究(1)内皮细胞在 T2 DM的不良血管重塑，以及(2)冠脉循环和 糖尿病心脏，因为冠脉循环位于一个复杂的组织内，显示出舒张期的迹象 代谢功能障碍，如糖尿病和代谢综合征。我致力于维护 来自国家卫生研究院等来源的外部资金，以维持独立的实验室和培训 环境，同时继续在代谢性疾病的心血管并发症方面取得进展。我的 过去的表现证明了我出色的技术技能、解决问题的能力、职业道德和干劲 这将使我为成功做好准备。

PROJECT SUMMARY Heart disease in type 2 diabetes mellitus (T2DM) is directly related to the severity of coronary artery disease (CAD), which results in impaired coronary flow and increased risk of myocardial infarction (MI). This poses a very significant health and economic burden in the U.S. and worldwide since cardiovascular disease is the leading cause or mortality in T2DM, accounting for >65% of all deaths in diabetic patients. Moreover, diabetic patients are 2-4 times more likely to experience MI than non-diabetic patients. While atherosclerosis and endothelial dysfunction are known contributors to MI, the relative contribution of coronary resistance microvessel (CRM) remodeling versus macrovessel (conduit artery) remodeling to T2DM-induced cardiac disease remains largely unknown. We previously showed that CRMs isolated from either T2DM db/db mice or the pre-clinical Ossabaw porcine model of MetS undergo inward hypertrophic remodeling associated with reduced stiffness (novel and anti-dogmatic) in the face of increased macrovascular stiffness (dogmatic). CRM remodeling was accompanied by reduced coronary blood flow and was partially dependent upon vascular smooth muscle cell (VSMC) proliferation, increased expression of elastic extracellular matrix (ECM), and decreased expression of focal adhesion/cytoskeletal proteins. In contrast, focal adhesion/cytoskeletal and ECM protein expression were increased in macrovessels of T2DM db/db mice. These findings point to an intimate interaction between cellular stiffness and the ECM in macro- and coronary micro-vessels that dictates overall vascular stiffness in T2DM. We also observed that CRM VSMCs exhibit a proliferative phenotype that is associated with enhanced signaling by the advanced glycation end products (AGE)/RAGE pathway and decreased expression of cysteine rich-protein 2 (CRP2), which is a negative regulator of VSMC proliferation. There are currently no studies that have directly investigated the effect of stiffness on T2DM VSMC phenotype. Based on my preliminary data, a less stiff CRM wall promotes an in vivo proliferative VSMC phenotype, while the stiffer aortic (macrovascular) wall favors a synthetic VSMC phenotype. These data demonstrate that mechanisms underlying CRM remodeling are fundamentally different from those that regulate macrovascular remodeling and may represent pathologic mechanisms dictating VSMC phenotype, contributing to diabetic MI. Thus, my overall hypothesis is that alterations in macro- and coronary micro-vascular remodeling and stiffness reflect differential contributions of cellular and ECM stiffness that controls VSMC phenotype and may be dictated in part by AGE/RAGE and/or CRP2. The studies outlined in this proposal will investigate these mechanisms comprehensively utilizing a novel decellularization technique in aorta and CRMs, primary VSMC cultures, isolated coronary resistance microvessels, and in vivo analysis of vascular remodeling and blood flow in diabetic mice. During the mentored phase, we will determine the relative contribution of both the ECM and VSMCs to overall vascular stiffness further assess whether stiffness can modulate classical VSMC phenotypes in both macro-vessels and CRMs. The mentored studies will focus heavily on determining the cellular and biomechanical mechanisms underlying these alterations in primary VSMC cultures. During the independent phase, I will determine whether the interplay of stiffness, AGE/RAGE, and CRP2 modulates the phenotype of VSMCs isolated from both diabetic mice and pre-clinical pigs with MetS. Finally, I will evaluate the therapeutic potential of a novel RAGE antagonist, FPS-ZM1, in reducing adverse vascular remodeling in T2DM mice. This transition award will allow me access to additional mentoring necessary for future success as a competitive and successful independent investigator. This proposal includes a comprehensive training plan that will be overseen by members of a mentoring committee with expertise that encompasses vascular signaling and biomechanics, extracellular matrix, and coronary complications of diabetes and metabolic syndrome. Once complete, I will have gained additional expertise in hypothesis generation, experimental design, and data interpretation related to in vitro cellular signaling, vascular mechanics, ECM, and the coronary circulation that will enhance my already sound expertise in ex vivo and in vivo physiology. It will further provide a solid foundation for future studies in which I would like to investigate (1) the role of endothelial cells in adverse vascular remodeling in T2DM, and (2) the dynamic cross-talk between the coronary circulation and the diabetic heart, since the coronary circulation lies within a complex tissue that exhibits signs of diastolic dysfunction in metabolic conditions such as diabetes and metabolic syndrome. I am committed to maintaining extramural funding from sources such as the NIH to maintain an independent laboratory and training environment, while continuing to make inroads in the cardiovascular complications of metabolic diseases. My past performance is a testament to my excellent technical skills, problem-solving skills, work ethic, and drive that will collectively make me well poised for success.