The role of smooth muscle PPAR gamma in neointima formation

描述（由申请人提供）：项目总结本项目的目的是确定过氧化物酶体增殖物激活受体（PPAR）3激活在新生内膜形成中的作用。血管成形术后经常发生新生内膜形成，并导致显著的发病率;血管平滑肌细胞（SMC）是新生内膜形成过程中的关键细胞。我们将研究两种临床可用的药物对SMC生物学和新生内膜形成的影响。PPAR 3是一种配体激活的核受体，已显示对血管疾病具有有益作用。我们将比较两种药物的作用：吡格列酮（仅激活PPAR 3）和贝沙罗汀（一种激活PPAR 3、PPAR 1、PPAR 4、LXR和FXR的RXR激动剂）。我们的假设是，特异性地在平滑肌细胞（SMC）中激活PPAR 3将通过减少驻留SMC迁移和增殖以及SMC衍生的趋化因子产生和随后的骨髓衍生细胞的募集来减少新生内膜形成。我们认为吡格列酮和贝沙罗汀都是有效的，但贝沙罗汀可能更有效，因为它激活了其他核受体。在目的一中，我们将比较吡格列酮和贝沙罗汀对SMCs的作用。我们将测量增殖、细胞因子产生和平滑肌基因表达的变化。在目标二中，我们将确定药物是否影响对维持SMC表型至关重要的microRNA水平，如miR- 143，miR-145和miR-221。在目标三中，我们将研究在股动脉钢丝损伤过程中药物的体内作用。为了追踪骨髓来源的细胞向动脉损伤部位的募集，所有小鼠将接受来自GFP阳性供体的骨髓移植。导丝损伤后，将在多个时间点对小鼠进行分析。沿着新生内膜大小，我们将测量趋化因子（IL-6、MCP-1、SDF-11和KC）的产生、骨髓源性细胞和巨噬细胞的募集以及细胞增殖。我们还计划研究PPAR 3活化在新生内膜形成过程中在平滑肌细胞中的作用。使用可诱导的组织特异性敲除模型，我们将在小鼠接受来自GFP阳性供体的骨髓移植后耗尽平滑肌细胞中的PPAR 3。诱导型PPAR 3敲除小鼠和对照小鼠将接受吡格列酮、贝沙罗汀或对照的治疗，并经受股动脉线损伤。在多个时间点，将再次测量新生内膜尺寸。在Aim 3B中使用的所有小鼠将具有R26 R报告基因等位基因，其中平滑肌细胞用2-半乳糖苷酶标记。由于我们可以测量骨髓来源的和驻留的SMC，我们将确定每种细胞类型对新生内膜的相对贡献。我们还将能够确定是否特异性地在平滑肌细胞中的PPAR 3介导贝沙罗汀或吡格列酮的作用。 公共卫生相关性：项目叙述贝沙罗汀和吡格列酮是两种临床使用的激活PPAR 3的药物。在目标一中，我们将测试这两种药物是否能同等地影响平滑肌细胞生物学，在目标二中，我们将确定这两种药物是否能改变microRNA水平。在目标三中，我们将确定哪种药物可以最大限度地减少导丝损伤后的新生内膜尺寸。

DESCRIPTION (provided by applicant): PROJECT SUMMARY The goal of this project is to define the role of peroxisome proliferator-activated receptor (PPAR)3 activation in neointima formation. Neointima formation occurs frequently after angioplasty and causes significant morbidity; vascular smooth muscle cells (SMCs) are key cells during neointima formation. We will study the effects of two clinically available agents on SMC biology and neointima formation. PPAR3 is a ligand-activated nuclear receptor that has been shown to have beneficial effects on vascular disorders. We will compare the effects of two agents: pioglitazone (activates PPAR3 only) and bexarotene (an RXR agonist which activates PPAR3, PPAR1, PPAR4, LXR, and FXR). Our hypothesis is that PPAR3 activation specifically in smooth muscle cells (SMC) will reduce neointima formation by decreasing resident SMC migration and proliferation as well as SMC-derived chemokine production and subsequent recruitment of bone marrow-derived cells. We believe both pioglitazone and bexarotene will be effective but bexarotene may be more effective due to activation of other nuclear receptors. In Aim One, we will compare the effects of pioglitazone to bexarotene on SMCs. We will measure changes in proliferation, cytokine production, and smooth muscle gene expression. In Aim Two, we will determine if the agents affect levels of microRNAs crucial to maintaining SMC phenotype, such as miR- 143, miR-145, and miR-221. In Aim Three, we will examine the effects of the agents in vivo during femoral artery wire injury. To track recruitment of bone-marrow derived cells to the site of arterial injury, all mice will receive bone marrow transplants from a GFP positive donor. After wire injury, mice will be analyzed at multiple time points. Along with neointima size, we will measure production of chemokines (IL-6, MCP-1, SDF-11, and KC), recruitment of bone marrow-derived cells and macrophages, and cellular proliferation. We also plan to study the role of PPAR3 activation specifically in smooth muscle cells during neointima formation. Using an inducible tissue-specific knockout model, we will deplete PPAR3 in smooth muscle cells after mice have received bone marrow transplants from GFP positive donors. Inducible PPAR3 knockout mice and control mice will receive therapy with either pioglitazone, bexarotene or control and be subjected to femoral artery wire injury. At multiple time points, neointima size will again be measured. All mice used in Aim 3B will have the R26R reporter allele in which smooth muscle cells are labeled with 2-galactosidase. Since we can measure both bone marrow derived and resident SMCs, we will determine the relative contribution each cell type makes to the neointima. We will also be able to determine if PPAR3 specifically in smooth muscle cells mediates the effects of bexarotene or pioglitazone. PUBLIC HEALTH RELEVANCE: PROJECT NARRATIVE Bexarotene and pioglitazone are two clinically used drugs that activate PPAR3. In Aim One, we will test whether both agents can affect smooth muscle cell biology equally and, in Aim Two, we will determine whether the agents can change microRNA levels. In Aim Three, we will determine which agent can reduce neointima size the greatest after wire injury.