VIRAL CHEMOKINE RECEPTORS: TRANSPLANT VASCULAR SCLEROSIS

该项目的长期目标是确定与人类巨细胞病毒（HCMV）加速移植血管硬化有关的机制。所有带血管的器官移植的移植物损失的主要原因是由于血管病变相关的慢性排斥反应。这种形式的血管病变被称为TVS，其特征是同心内膜平滑肌细胞增殖，导致血管闭塞并最终导致移植物衰竭。迄今为止，治疗严重TVS的唯一方法是再移植。移植受者的临床研究表明HCMV与TVS加速之间存在直接联系。基于这些发现，我们开发了一种心脏和小肠移植大鼠模型，其中巨细胞病毒感染加速了TVS的时间和严重程度。虽然CMV介导这一过程的确切机制尚不清楚，但我们小组最近的工作表明，CMV可能通过表达病毒编码的趋化因子受体（US28），诱导平滑肌细胞（SMC）向趋化因子产生位点迁移，从而促进TVS的发生。我们还观察到，大鼠和小鼠CMV也通过各自的病毒趋化因子受体R33和M33诱导SMC迁移，这两个受体是HCMV US28的功能同源物。我们假设cmv加速TVS的机制涉及病毒编码趋化因子受体的表达导致内膜SMC迁移，从而导致TVS的特征性血管病变。因此，我们将利用我们的体外和体内模型来扩展我们的观察，并确定病毒编码趋化因子受体在TVS发展中的作用。首先，使用慢性排斥的大鼠心脏移植模型，我们将确定病毒对TVS疾病进展动力学的影响，病毒在组织中的表达程度以及参与该过程的细胞类型。我们还将确定宿主因子，如趋化因子和细胞因子，以及R33在rcmv诱导TVS发展的各个阶段所参与的贡献。其次，我们将描述参与诱导SMC迁移的信号传导的R33结构域和作为激动剂或拮抗剂诱导细胞运动的配体。在最后一篇文章中，我们将生成重组RCMV，该RCMV包含与信号相关的结构域突变，以了解它们在大鼠心脏移植模型中对TVS的贡献。最后，R33诱导SMC迁移的拮抗剂将在体内大鼠模型中测试其阻断TVS的能力。这些研究将为理解巨细胞病毒在加速TVS中的作用提供有价值的动物模型。此外，这些研究的完成将为新的和合理的治疗策略设计奠定基础，以提高HCMV感染移植受者的长期移植存活率。

The long term goal of this project is to identify mechanisms involved with human cytomegalovirus (HCMV) acceleration of transplant vascular sclerosis. The primary cause of graft loss of all vascularized organ transplants is due to a vascular lesion associated with chronic rejection. This form of vasculopathy referred to as TVS is characterized by concentric neointimal smooth muscle cell proliferation that results in vessel occlusion and ultimately graft failure. To date the only therapy available to treat severe TVS is retransplantation. Clinical studies in transplant recipients have demonstrated a direct link between HCMV and the acceleration of TVS. Based on these findings, we have developed a rat model of heart and small bowel transplantation in which CMV infection accelerates the time and severity of TVS. While the exact mechanism through which CMV mediates this process is unknown, recent work from our group suggests that CMV may contribute to TVS through induction of smooth muscle cell (SMC) migration towards sites of chemokine production through expression of a virally encoded chemokine receptor (US28). We have also observed that Rat and Murine CMV also induce SMC migration through their respective viral chemokine receptors R33 and M33, which are functional homologues of HCMV US28. We hypothesize that the mechanism of CMV-accelerated TVS involves the expression of virally encoded chemokine receptors leading to intimal SMC migration, which results in the characteristic vascular lesions of TVS. Therefore, we will utilize our in vitro and in vivo models to extend our observations and determine the role of virally encoded chemokine receptors in the development of TVS in three specific aims. First, using a rat cardiac transplant model of chronic rejection, we will determine the effects of virus on the kinetics of disease progression of TVS and the extent of viral expression in tissues as well as the cell types involved in the process. We will also determine the host factors such as chemokines and cytokines, and the contribution of R33 involved at various stages of the development of RCMV-induced TVS. Secondly, we will characterize the R33 domains involved in signaling which induce SMC migration and the ligands, which serve as agonists or antagonists to induced cellular movement. In the last specific, we will generate recombinant RCMV which contain mutations in domains involved in signaling to understand their contribution to TVS in the rat cardiac transplant model. Lastly, antagonists of R33 induced SMC migration in vitro will be tested for their ability to block TVS in the in vivo rat model. These studies will provide a valuable animal model to understand the role of CMV in the acceleration of TVS. In addition completion of these studies will form the basis for novel and rational design of therapeutic strategies to enhance long-term graft survival in HCMV infected transplant recipients.