管周脂肪PEDF通过抑制血管外膜滋养管生成抗动脉粥样硬化的机制研究

Perivascular adipose tissue (PVAT) is demonstrated to contribute to atherosclerosis (AS). Adventitia vasa vasorum(VV) is the major pathway to drain pro-inflammatory adipocytokines, which result in subsequent vascular injury. Inhibition of neovascularization of VV is a potential therapeutic target of AS. The previous research has confirmed that PEDF has protective effect on increasing AS plaque stability. As a target gene, PEDF involves the effects of miR-33a on angiogenesis. PEDF is mainly derived from adipose tissue. In this study, we propose the hypothesis that upregulation of PEDF in PVAT is potent to inhibit VV angiogenesis, resulting to ameliorate AS. We will build adipose tissue allotransplant murine model by transplanting adipose tissue with PEDF knockout(PEDFfl/fl;adipoq-cre) to carotid artery, and treat with PEDF agonist Ginsenoside Rb1 (Rb1). Concentration of PVAT and circulating PEDF , density of VV, formation and stability of AS plaque were determined via immunohistochemistry and immunofluorescence, molecular pathology, ELISA and western blotting technology to investigate the relationship between PEDF in PVAT and VV formation in AS, and explore whether PEDF is the key compound of Rb1 in anti-atherosclerosis or not. Endothelial cells and adipocytes will be co-cultured to elucidate the effect of adipose-derived PEDF on human umbilical vein endothelial cells (HUVECs) and confirm the pathway of Rb1→PPARγ→miR-33a→PEDF→ATGL. The results of the study will provide fundamental evidences to illuminate the internal relationship between PVAT and vascular injury, will assist to develop new therapeutic targets in prevention and treatment of AS.

管周脂肪组织（PVAT）是动脉粥样硬化（AS）的重要危险因素，血管外膜滋养管（VV）通过管周脂肪输送炎症因子造成血管损伤，抑制VV生成能够延缓动脉粥样硬化的发生发展。前期研究已证实色素上皮衍生因子（PEDF）稳定AS斑块，且PEDF作为靶基因介导miR-33a的血管新生作用，而PEDF的主要来源是脂肪组织，因此，我们提出上调管周脂肪PEDF抑制VV生成和减缓AS的假说。我们拟通过小鼠颈动脉管周PEDF条件性敲除脂肪移植模型，并同时给予可上调PEDF的人参皂苷Rb1，探究管周脂肪PEDF与VV形成和AS的关系，明确管周脂肪PEDF是否为Rb1发挥抗AS的关键活性产物；并通过脂肪细胞-内皮细胞共培养模型验证脂肪来源PEDF对内皮功能的影响及Rb1→PPARγ→miR-33a→PEDF→ATGL的作用通路是否成立，为阐明管周脂肪与血管损伤的内在联系提供新的依据。

动脉粥样硬化（AS）斑块易损、破裂和血栓形成是导致急性心脑血管事件的主要病理机制。在AS发生发展过程中，起源于外膜的血管外膜滋养管（VV）增生进而造成斑块内出血，增加斑块易损性。色素上皮衍生因子（PEDF）具备很强的抑制血管新生作用及抗炎、抗氧化、抑制血栓形成等作用，因此在稳定斑块的作用中可能具有更多的优势。但PEDF在VV增生中的作用尚未明确。经典中药人参的主要活性物质人参皂苷Rb1可抑制AS的发生发展。课题组前期实验证实，人参皂苷Rb1通过促进PEDF表达抑制人脐静脉内皮细胞（HUVEC）血管新生作用。本项目通过收集非冠心病及冠心病患者心包脂肪，检测心包脂肪中PEDF在动脉粥样硬化中的表达变化；在PEDF敲减的动脉粥样硬化小鼠模型的基础上构建颈动脉管周脂肪移植模型，干预移植脂肪PEDF的表达，验证PEDF在VV增生及动脉粥样硬化中的作用；在PEDF敲除的ApoE-/-小鼠进一步验证PEDF的抗AS作用及分子机制；ApoE-/-小鼠过表达miR-33，验证Rb1抗动脉粥样硬化的分子机制；在肥胖小鼠及脂肪细胞中验证Rb1的抗肥胖作用。结果证实：冠心病患者心包脂肪及血清中PEDF含量较非冠心病患者降低；管周脂肪组织来源的PEDF，抑制血管外膜滋养管新生，降低局部血管的斑块内炎症水平，增加斑块稳定性，抑制动脉粥样硬化的发展；PEDF通过作用于层粘连蛋白受体（LR）和活化激酶C受体1（RACK1），促进内皮细胞自噬，进而抑制内皮细胞的血管新生；PEDF作为mi-R33的靶基因，受到Rb1的调控，是Rb1发挥抗AS的关键活性产物；Rb1通过PPARγ-AQP7途径促进脂质排除、抑制肥胖。本研究为探讨脂肪组织与动脉粥样硬化的内在联系及内在机制提供了新的依据。项目资助发表SCI论文6篇，待发表1篇。培养博士生1名，已取得博士学位。硕士生2名，其中2名已经取得硕士学位。