B cell anergy, modified LDL uptake, and atherosclerosis

项目摘要 心血管疾病是世界范围内主要的死亡原因。它是一种多方面的慢性炎症性疾病 一种疾病，其特征是修饰的脂蛋白和免疫细胞在主动脉、血管内积聚 功能障碍和低度慢性炎症。尽管在过去的十年中取得了长足的进步， 关于疾病进展背后的机制，仍有许多未知之处。改良剂的吸收 巨噬细胞的低密度脂蛋白(MLDL)已被很好地描述为 动脉粥样硬化的发展。我们的实验室表明B细胞可以摄取mLDL，这种摄取 诱导一些显著的B细胞表型变化和上调LXR依赖的靶点。而当 MS摄取低密度脂蛋白的机制和作用已被很好地描述，B细胞摄取低密度脂蛋白的途径 目前尚不清楚，目前尚不清楚mLDL摄取对B细胞功能的影响。我们的预赛 数据表明，B细胞摄取acLDL诱导了一种独特的基因图谱，这支持抑制途径 包括GRAIL。GRAIL是一种众所周知的T细胞耐受和T细胞激活的调节因子。我们的数据 证明GRAIL在B细胞中也有表达，并在B细胞活化和B细胞分化中发挥重要作用 无能。重要的是，B细胞特异性缺陷会导致动脉粥样硬化的增加。在本申请的目标1中， 我们建议测试GRAIL表达和mLDL摄取控制B细胞的分子机制 动脉粥样硬化中的激活。首先，我们将使用SR-B1，Marco， CXCL16和CD163缺陷小鼠或阻断Abbbcr转基因和FcRIIB缺陷小鼠。我们将测试 以LXR-mTOR-Otub1-GRAIL为中心探讨mLDL摄取如何诱导B细胞抑制途径 轴心。接下来，我们将确定GRAIL塑造B细胞激活的途径，研究GRAIL依赖 BCR信号、B细胞存活和IL-4依赖的B细胞信号转导，并检测B细胞特异性的作用 动脉粥样硬化的圣杯。在目标2中，我们将确定动脉粥样硬化如何影响B细胞无能，并测试 GRAIL和mLDL摄取对无能B细胞稳定性和功能的影响我们将比较无能的表型。 来自年轻的、老年的APOE-/-小鼠的B细胞，以及年龄匹配的自然产生的无能的BL/6 B细胞。我们还将 在ARS/A1过继转移实验中检测动脉粥样硬化对无能B细胞稳定性的影响 转基因小鼠，B细胞无能模型。接下来，我们将研究GRAIL如何影响B细胞无能和 在B细胞特异性启动子下接受AAV-shRNA-Rnf128的ARS/A1小鼠的动脉粥样硬化形成 AAV-PSCK9。这项工作的意义是重大的，因为结果将为 MLDL摄取在B细胞生物学中的作用--通过GRAIL依赖的B细胞无能和B细胞激活调节 动脉硬化。

Project Summary Cardiovascular disease is the leading cause of death worldwide. It is a multifaceted chronic inflammatory disease characterized by the accumulation of modified lipoproteins and immune cells in the aorta, vascular dysfunction, and low-grade chronic inflammation. Despite the strides that have been made in the past decade, there is still much that is unknown about the mechanisms behind disease progression. The uptake of modified low density lipoproteins (mLDL) by macrophages has been well characterized as being a major mechanism in the development of atherosclerosis. Our laboratory show that B cells can uptake mLDL and this uptake induces some significant phenotypical changes in B cells and upregulation of LXR-dependent targets. While mechanisms and role of mLDL uptake by Ms are well-characterized, the pathways of mLDL uptake in B cells are unclear and nothing is yet known about the impact of mLDL uptake on B cell functions. Our preliminary data suggest that acLDL uptake by B cells induces a unique gene profile, which supports inhibitory pathways including GRAIL. GRAIL is a well-known regulator for T-cell tolerance and T cell activation. Our data demonstrate that GRAIL is also expressed in B cells and play an important role in B cell activation and B cell anergy. Importantly, B cell-specific deficiency results in increased atherosclerosis. In Aim 1 of this application, we propose to test molecular mechanisms whereby GRAIL expression and mLDL uptake control B cell activation in atherosclerosis. First, we will test mechanisms of mLDL uptake by B cells using SR-B1, MARCO, CXCL16, and CD163 deficient mice or blocking Abs BCR-transgenic and FcRIIB- deficient mice. We will test how mLDL uptake induces some inhibitory pathways in B cells with the focus on LXR-mTOR-Otub1-GRAIL axis. Next, we will determine pathways by which GRAIL shapes B cell activation, investigate GRAIL-dependent BCR signaling, B cell survival and IL-4-dependent signaling in B cells, and examine the role of B cell specific GRAIL in atherosclerosis. In Aim 2, we will determine how atherosclerosis affects B cell anergy and test a role of GRAIL and mLDL uptake in stability and functions of anergic B cells. We will compare phenotypes of anergic B cells from young, aged Apoe-/- mice, and age-matched naturally occurring anergic BL/6 B cells. We will also test how atherosclerosis affects stability of anergic B cells in adoptive transfer experiments using Ars/A1 transgenic mice, a model of B cell anergy. Next, we will investigate how GRAIL affects B cell anergy and atherogenesis using Ars/A1 mice that would receive AAV–shRNA-Rnf128 under B cell specific promoter and AAV-PSCK9. The implications from this work are significant since the results will provide the first evidence for a role of mLDL uptake in B cell biology via GRAIL-dependent regulation of B cell anergy and B cell activation in atherosclerosis.