FABP4在动脉粥样硬化中调控的靶位及JAK/STAT信号通路的作用研究

Atherosclerosis(AS) is a chronic inflammatory disease which mainly is a disorder of lipid metabolism. The fatty acid binding protein 4(FABP4) is a key target gene which regulates lipid metabolism and inflammation. Our group has previously confirmed the role of FABP4 in AS. But it hasn't been elucidated about the mechanism and the downstream target of AS which is regulated by FABP4. On the basis of the preliminary work, this project plans to copy the AS animal and cell models, FABP4 acts as a target. On the overall level, screening the related target genes of lipid metabolism and inflammation by using gene and protein chip technology, determing the target significantly regulated by FABP4. In the foam cells level, constructing stable expression vector through the silence and overexpression of FABP4 with RNA interference and gene recombinant, and transfecting it to the cells in order to look for the regulatory sites of FABP4 which are consistent with the overall level. Taking blocking strategies from three levels of FABP4, JAK and STAT to clarify the main damage links and potential intervention targets of the downstream target regulated by FABP4, and the effects of JAK/STAT signaling pathway in the significant target regulated by FABP4. Transducting the mutated SOCS1,3 by lentiviral so as to explore the interaction of STAT3 and SOCS1,3, and provide a theoretical basis for finding a new therapy way for AS.

动脉粥样硬化(AS)是以脂代谢紊乱为主的慢性炎症性疾病，脂肪酸结合蛋白4(FABP4)是调控脂代谢和炎症反应的关键靶基因，课题组前期已证实了FABP4在AS中的作用，但FABP4调控AS的下游靶位及作用机制尚未阐明。本项目拟在前期工作基础上，复制AS动物和细胞模型，以FABP4为靶点，在整体水平，采用基因和蛋白芯片筛选脂代谢和炎症相关靶基因，确定FABP4显著调控的靶位；在泡沫细胞上，以RNA干扰和基因重组分别沉默和过表达FABP4，构建稳定表达载体并转染细胞，寻找与整体水平上一致的FABP4调控位点；从FABP4、JAK和STAT等三个层面采用阻断策略，明确FABP4调控下游靶位的主要受损环节和潜在干预靶点以及JAK/STAT信号通路在FABP4调控关键靶位中的作用；通过慢病毒转导突变的SOCS1、3，探讨STAT3和 SOCS1、3的相互作用，为寻找治疗AS的新途径提供理论依据。

背景：动脉粥样硬化(Atherosclerosis, AS)是一种发生于大、中动脉内膜以脂代谢紊乱、免疫炎症反应等因素导致的慢性炎症性疾病，是心脑血管疾病的重要基础性病变。目前，针对AS的主要治疗方案仍是降血脂，对于另一发生机制炎症的治疗方案尚处在探索中。FABP4是炎症和脂代谢的交叉靶基因，广泛参与炎症反应和脂质代谢，是促进AS形成的关键基因。.内容：复制ApoE-/-鼠AS动物模型，探讨FABP4及其下游转导通路的变化，关注JAK/STAT信号通路和效应靶点。培养泡沫细胞，油红O示泡沫细胞形成。将FABP4过表达腺病毒检测FABP4 mRNA及蛋白水平，检测MCP-1 mRNA及蛋白水平，油红O染色观察泡沫细胞脂质沉积；全基因组谱芯片分析过表达FABP4表达谱，检测JAK、STAT和SOCS1 mRNA及蛋白；检测JAK、STAT、SOCS1、MCP-1蛋白表达及脂质沉积变化。.结果：油红O证实泡沫细胞模型成功；Ad-FABP4后FABP4 mRNA和蛋白表达，差异有显著性（P<0.01），MCP-1 mRNA和蛋白表达平均明显升高（P<0.01）；基因表达谱示：JAK/STAT信号转导通路为较为明显通路之一，采用qRT-PCR和WB对此通路验证，结果与芯片结果一致；SOCS1 mRNA和蛋白的表达，与Control比较，Ad-FABP4组SOCS1 mRNA和蛋白表达水平均明显升高（P<0.01）；JAK特异性抑制剂AG490干预后，p-STAT1、p-STAT2、SOCS1、MCP-1蛋白表达较Ad-FABP4组明显降低（P<0.05）；三个不同的SOCS1 siRNA腺病毒转染细胞，结果以siRNA2最为明显；Ad-FABP4与SOCS1 siRNA2共转染，WB检测MCP-1蛋白表达，油红O染色看脂质沉积。.意义：明确FABP4调控下游靶位的主要受损环节和潜在干预靶点以及JAK/STAT信号通路在AS中的作用，为AS治疗新靶点的筛选及治疗新途径的建立提供理论依据。