BMP-7 induced Macrophage Polarization in Atherosclerosis

BMP-7 诱导动脉粥样硬化中的巨噬细胞极化

• 批准号: 8599047
• 负责人: Sampath Parthasarathy
• 金额: $ 34.36万
• 依托单位: UNIVERSITY OF CENTRAL FLORIDA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-19 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8599047
• 关键词: Address; Affect; Animals; Anti-Inflammatory Agents; Anti-inflammatory; Apolipoprotein E; Area; Arterial Fatty Streak; Arteries; Atherosclerosis; BMP7 gene; Backcrossings; Biological Models; Blood; Blood Vessels; Bone Morphogenetic Proteins; Cardiovascular Diseases; Carotid Arteries; Cell Culture Techniques; Cells; Cessation of life; Clinical; Colony-Stimulating Factor Gene; Coronary artery; Data; Development; Disease; Disease Progression; Gene Deletion; Gene Expression; Generations; Genes; Growth Factor; Human; In Vitro; Infiltration; Inflammation; Inflammatory; Interleukin-1; Interleukin-10; Knock-out; Knockout Mice; Left; Lesion; Ligation; MAP2K1 gene; MAPK14 gene; MAPK8 gene; Macrophage Colony-Stimulating Factor; Measures; Mediating; Methodology; Mitogen-Activated Protein Kinases; Modeling; Molecular; Mononuclear; Mus; Myocardial Infarction; Operative Surgical Procedures; Osteoblasts; Osteoporosis; Pathway interactions; Phenotype; Play; Proto-Oncogene Proteins c-akt; Research; Role; Small Interfering RNA; Smooth Muscle Myocytes; Staging; Techniques; Testing; Thromboplastin; Time; Up-Regulation; arginase; base; bone morphogenetic protein 7; calcification; cytokine; experience; follow-up; improved; in vivo; inhibitor/antagonist; intravenous injection; loss of function; macrophage; monocyte; novel; prevent; public health relevance; receptor; receptor function; scavenger receptor; shear stress; success; transdifferentiation

DESCRIPTION (provided by applicant): A novel concept has recently emerged demonstrating that monocytes, following infiltration into arteries, can differentiate into two major classical (M) or alternative (M2) macrophages during the development of atherosclerosis (ATH). In vitro preliminary data indicates that BMP-7 stimulated monocytes (THP-1 cells) differentiate into M2 polarized macrophages, which was confirmed with increased M2-specific marker CD206 and up regulation of released IL-10. In vivo preliminary data suggests that intravenous injection of BMP-7 in ApoE KO mice experiencing a partial ligated coronary artery (PLCA) have increased M2 polarized macrophages, increased BMP-7R on monocytes and M2 polarized macrophages, increased M2 macrophage- specific released IL-10 and IL-1 receptor antagonist, and increased blood velocity compared with sham controls at two weeks follow up. Accordingly, we propose that monocytes differentiate into M2 macrophages, an anti-inflammatory phenotype, under the influence of BMP-7 at different stages of ATH development. Additionally, we propose to understand whether M-CSF action is also required along with BMP-7 on monocyte differentiation to M2 polarized macrophages and to correlate the presence and influence of BMP-7 on M1 and M2 macrophages in ATH using heterozygous mice obtained from backcrossing op mice (M-CSF deficient) and ApoE deficient mice. We also propose to establish the mechanisms by which BMP-7 would affect macrophage polarization. Specific aims are as follows: Aim1a: To examine the presence of BMP-7 in the lesions during the development of ATH in ApoE KO animals and to correlate its presence with M1 and M2 types of macrophages. We will determine this using immunohistochemical and real-time PCR techniques and measuring pro-and anti- inflammatory cytokine levels, specific expression of inflammatory genes, and specific genes for M1 and M2 phenotype macrophages such as arginase and scavenger receptors. Furthermore, the presence and preponderance of specific macrophage phenotypes will be identified. Aim 1b: We will use loss of function of BMP-7R using siRNA methodology and inhibitors for BMP-7 to determine macrophage polarization. Aim 2: To determine if BMP-7 induced monocytes to M2 polarization is mediated via BMP-7 alone or if influence is provided by released M-CSF from the monocytes. We will establish whether a) BMP-7 induces differentiation of monocytes into M2 macrophages, or whether b) BMP-7 and M-CSF both induce monocytes into M2 polarized macrophages. Genetically altered op mice specific for M-CSF gene deletion will be backcrossed with the ApoE KO mice, heterozygous mice will be subjected to PLCA surgery, and all parameters will be established as proposed in Aim 1. Aim 3: We will determine SMAD-dependent and p38, JNK, and ERK pathways and their correlation with the release/expression of specific phenotype-associated gene expressions. Results of these studies will define the impact of BMP-7 on M2 polarized macrophages, anti-inflammatory cytokines, their effects on gene expression, and molecular mechanisms in the progression of ATH.

描述（由申请人提供）：最近出现了一种新的概念，证明单核细胞在浸润到动脉中后，在动脉粥样硬化（ATH）的发展过程中可以分化为两种主要的经典（M）或替代（M2）巨噬细胞。体外初步数据表明，BMP-7刺激的单核细胞（THP-1细胞）分化成M2极化的巨噬细胞，这通过增加的M2特异性标志物CD 206和释放的IL-10的上调得到证实。体内初步数据表明，在两周随访时，与假手术对照相比，在经历部分结扎冠状动脉（PLCA）的ApoE KO小鼠中静脉内注射BMP-7增加了M2极化的巨噬细胞，增加了单核细胞和M2极化的巨噬细胞上的BMP-7 R，增加了M2巨噬细胞特异性释放的IL-10和IL-1受体拮抗剂，并且增加了血流速度。因此，我们提出，单核细胞分化成M2巨噬细胞，一种抗炎表型，在ATH发展的不同阶段的BMP-7的影响下。此外，我们建议了解M-CSF作用是否也需要沿着BMP-7对单核细胞分化为M2极化巨噬细胞的作用，并使用从回交op小鼠（M-CSF缺陷型）和ApoE缺陷型小鼠获得的杂合子小鼠，将BMP-7对ATH中M1和M2巨噬细胞的存在和影响关联起来。我们还建议建立BMP-7影响巨噬细胞极化的机制。具体目标如下：目标1a：在ApoE KO动物中检查ATH发展过程中病变中BMP-7的存在，并将其存在与M1和M2型巨噬细胞相关联。我们将使用免疫组织化学和实时PCR技术，并测量促炎和抗炎细胞因子水平，炎症基因的特异性表达，以及M1和M2表型巨噬细胞的特异性基因，如胰蛋白酶和清道夫受体来确定这一点。此外，将鉴定特定巨噬细胞表型的存在和优势。目的1b：我们将使用siRNA方法和BMP-7的抑制剂，使用BMP-7 R的功能丧失来确定巨噬细胞极化。目标二：确定BMP-7诱导的单核细胞向M2极化是否仅通过BMP-7介导，或者是否由单核细胞释放的M-CSF提供影响。我们将确定a）BMP-7是否诱导单核细胞分化为M2巨噬细胞，或者B）BMP-7和M-CSF是否都诱导单核细胞分化为M2极化巨噬细胞。M-CSF基因缺失特异性遗传改变的op小鼠将与ApoE KO小鼠回交，杂合子小鼠将接受PLCA手术，所有参数将按照目标1中的拟定确定。目标三：我们将确定SMAD依赖性和p38，JNK和ERK途径及其与特定表型相关基因表达的释放/表达的相关性。这些研究的结果将确定BMP-7对M2极化巨噬细胞、抗炎细胞因子的影响，它们对基因表达的影响，以及ATH进展中的分子机制。