蛋白激酶SGK1调控巨噬细胞凋亡促进肺动脉高压的机制研究

Lung inflammation precedes the development pulmonary arterial hypertension (PAH). Macrophage plays a central role in inflammation-associated PAH, but the mechanism for regulation of the inflammation is still unclear. Our previous data demonstrated that the expression of Serum-glucocorticoid regulated kinase 1 (SGK1) in lungs of rats of monocrotaline (MCT)-induced PAH and mice of chronic hypoxia (CH)-induced PAH was significantly increased. SGK1 knockout (SGK1-/-) mice and rats with SGK1 inhibitor (EMD638683) treatment suppressed macrophage infiltration and PAH. As SGK1 exerts anti-apoptosis effects in a variety of cells, we hypothesize that SGK1 may participate in the regulation of inflammation-associated PAH by inhibiting macrophage apoptosis. We are going to confirm the important role of SGK1 in macrophage apoptosis, Perivascular inflammation and pulmonary vascular remodeling by using SGK1-/- mice in CH-PAH and SGK1 inhibitor in MCT-PAH rats with pathologic analysis, flow cytometry and western blot. In vitro, we plan to use hypoxic bone marrow-derived macrophage (BMDM) derived from WT and SGK1-/- mice to further investigate the potential molecular mechanisms of regulation macrophage apoptosis by SGK1 in PAH. And then we are going to test the SGK1 expression and apoptosis of Peripheral blood mononuclear cells from PAH patients after hypoxia. We aimed to determine the mechanism by which SGK1 regulates macrophage apoptosis, thus contributing inflammation-associated PAH and engage to find a novel target for PAH therapy.

炎症参与肺动脉高压（PAH）发生发展过程，新近研究发现巨噬细胞在PAH炎症过程中起关键作用。血清和糖皮质激素诱导蛋白激酶1（SGK1）可调控炎症及细胞凋亡。我们前期发现，PAH时SGK1可促进肺组织巨噬细胞浸润（已发表SCI论文）。但SGK1通过何种机制调控巨噬细胞浸润参与PAH仍不清楚。据此我们提出：“SGK1抑制巨噬细胞凋亡促进肺血管周围炎症”假说。我们拟应用SGK1抑制剂及SGK1敲除小鼠分别在大鼠及小鼠PAH模型中，通过病理分析、流式细胞术、Western blot等方法明确SGK1对巨噬细胞凋亡和肺血管周围炎症及肺血管重塑的影响；低氧刺激野生型及SGK1敲除小鼠的骨髓来源巨噬细胞，探讨SGK1抑制巨噬细胞凋亡的信号通路；培养PAH患者外周血单核细胞，低氧刺激并观察SGK1表达及其抗凋亡特征；试图阐明SGK1调控巨噬细胞凋亡参与炎症介导PAH的分子机制，为PAH治疗提供新靶点。

肺动脉高压是各种原因所致肺血管收缩及肺动脉重塑为特征的临床综合征，肺血管阻力进行性增加，最终致右心衰竭而死亡。临床及基础研究均证实肺血管周围炎症在肺动脉高压形成过程中起重要作用。巨噬细胞作为一种重要的炎症细胞，在肺动脉高压患者及动物模型中均可见巨噬细胞的浸润及活化。提示巨噬细胞功能在肺血管重塑及肺动脉高压形成过程中发挥关键作用。.血清和糖皮质激素诱导蛋白激酶1（SGK1）是一种快速诱导型蛋白激酶，参与多种心脏及血管周围炎症过程。本课题中我们发现其在低氧诱导肺动脉高压与肺血管重塑过程中发挥重要作用。我们在持续低氧诱导复制肺动脉高压小鼠模型中发现肺组织内SGK1表达显著上调。免疫荧光染色证实其定位于巨噬细胞。利用SGK1基因敲除小鼠，我们发现SGK1敲除可明显改善持续低氧刺激导致的肺动脉高压及肺血管重塑。免疫组化染色发现SGK1敲除减轻了低氧刺激后肺血管周围及肺组织内巨噬细胞浸润。进一步，我们体外应用LPS刺激野生型及SGK1敲除小鼠骨髓来源巨噬细胞，发现敲除SGK1可抑制炎症因子表达。.本课题证实SGK1促进了慢性低氧诱导肺动脉高压形成过程。SGK1在低氧诱导肺动脉高压的炎症过程中起重要作用，SGK1敲除减轻了炎症相关肺动脉高压。本课题为肺动脉高压的治疗提供了新的靶点。