COORDINATE REGULATION OF SMOOTH MUSCLE BY PDGF & MATRIX

在动脉粥样硬化病变的发展过程中，包括血小板衍生生长因子(PDGF)在内的炎性产物会导致平滑肌细胞(SMC)积聚。在体内，SMC通常被包括1型胶原在内的细胞外基质(ECM)包围，而在动脉粥样硬化形成过程中，ECM被降解，新的ECM成分被合成和组装。我们已经证明，人SMC在纤维胶原上的培养模拟了正常培养液中SMC的表型状态，在体外，纤维胶原使SMC滞留在细胞周期的G1期，而不依赖于PDGF的存在，而单体胶原支持SMC的增殖。我们进一步证明，不允许和允许SMC增殖的环境似乎针对一个共同的途径--CDK2抑制剂p27/Kip1的基质整合素调节。单独的研究已经证实，降解的胶原蛋白转导了导致局灶性粘连溶解的不同信号，包括快速裂解局灶性粘附性激酶和巴西林。因此，来自各种I型胶原的整合素介导的信号导致整合素信号复合体的特异性和快速调节以及SMC对PDGF的反应性。这导致了一种假设，即正常培养液中的细胞外基质可能不允许SMC迁移和增殖，而降解的基质可能会将SMC从这种不允许的状态释放出来。拟议的研究将在体内验证这一假说和PDGF的作用，具体目的如下：i.评估在允许和不允许的条件下调控CDK抑制物p27/Kip1水平的信号通路；2.检查导致局部黏附复合体I型胶原降解的分子通路；以及3.通过评估嵌合体来测试PDGF在动脉粥样硬化病变形成中的作用。在嵌合体中，Apo E-/-小鼠的循环细胞(形成病变的主要来源是巨噬细胞和血小板)被来自PDGF-/-胚胎的胎肝细胞取代。

Smooth muscle cells (SMC) accumulate in developing lesions of atherosclerosis in response to inflammatory products, including platelet- derived growth factor (PDGF). SMC in vivo are normally surrounded by extracellular matrix (ECM), including type 1 collagen, while in atherogenesis, the ECM is degraded and new ECM components are synthesized and assembled. We have demonstrated that culture of human SMC on fibrillar collagen mimics the phenotypic state of SMC in vivo of the normal media, and in vitro fibrillar collagen arrests SMC in the G1 phase of the cell cycle, independent of the presence of PDGF, while monomer collagen supports SMC proliferation. We have further demonstrated that non-permissive and permissive environments for SMC proliferation appear to target a common pathway-matrix-integrin regulation of the cdk2 inhibitor, p27/Kip1. Separate studies have established that degraded collagen transduced distinct signals that lead to dissolution of focal adhesions, including rapid cleavage of focal adhesion kinase and paxillin. Thus, integrin-mediated signals from various forms of type I collagen lead to specific and rapid modulation of the integrin signaling complex and the responsiveness of SMC to PDGF. This has led to the hypothesis that the extracellular matrix within the normal media may be non-permissive for SMC migration and proliferation, and that degraded matrix may release SMC from this non-permissive state. The proposed studies will test this hypothesis and the role of PDGF in vivo with the following specific aims: I. evaluate the signaling pathways responsible for modulating levels of the cdk inhibitor p27/Kip1 under permissive and non-permissive conditions for SMC; 2. examine the molecular pathways responsible for degraded type I collagen dissolution of the focal adhesion complex; and 3. test the role of PDGF in the formation of lesions of atherosclerosis by evaluation chimeras in which circulating cells (major sources of PDGF in developing lesions are macrophages and platelets) of Apo E-/- mice are replaced with fetal liver cells from PDGF-/- embryos.