Gas6双重调控衰老心肌血管新生和成熟的作用及机制

Impairment of angiogenesis-new capillary blood vessel formation from pre-existing vessels, and immaturity of the newly formed vessels, are frequent in aging tissues and cells. So therapeutic angiogenesis which aims to promoting the development of new microvessels is not well performed in the elderly patients who suffer from ischemic heart diseases (IHD). Researches confirmed that growth arrest-specific protein 6 (Gas6) had significant role in the angiogenesis of neoplasia. Our previous study demonstrated that Gas6 participated in the mobilization of endothelial progenitor cells (EPCs) in acute coronary syndrome (ACS) patients, and identified Gas6 as a protector for IHD patients. As EPCs are widely acknowledged as the essential cells in angiogenesis, we hypothesize Gas6 would boost angiogenesis and vascular maturity in aging myocardium. To verify this hypothesis, we use the Gas6 gene knocked-down mice, the EPCs isolated from elderly people and aging rats, and heart microvascular endothelial cells isolated from aging rats. With immunofluorescence stain and confocal microscopy, we try to observe the functions of Gas6 in promoting angiogenesis. Electron microscope will be applied to examine the functions of Gas6 in vascular maturity in aging myocardial ischemia in vivo. With MTT assay, modified boyden chamber assay and tube formation assay, we assess the abilities of Gas6 in proliferation, migration and lumen formation of endothelial cell in vitro. Using the method of siRNA to silent or highly express Gas6 in the model of mice and cells, we investigate the mechanisms of Gas6 in both angiogenesis and vascular maturity from organic, histic and molecular levels. So as to provide academic basis for applying therapeutic angiogenesis in ischemic diseases for the eldly .

衰老心肌血管新生能力减弱和成熟障碍，极大地限制了治疗性血管新生在老年缺血性心脏病（IHD）人群中的应用。研究表明，生长停滞特异性蛋白6（Gas6）能促进肿瘤血管新生和成熟。我们前期研究发现，Gas6参与冠心病患者血管新生关键细胞内皮祖细胞（EPCs）的动员，并且是缺血性心脏病的保护因子，该保护作用是否与Gas6促进衰老心肌血管新生和成熟有关，尚未见报道。本项目拟在前期研究基础上，以缺血/缺氧为刺激因素，建立各年龄代Gas6基因敲除小鼠心梗模型、老年EPCs和老年大鼠微血管内皮细胞缺氧/复氧模型，运用免疫组化、影像学和细胞生物学等方法观察Gas6对血管新生和成熟的双重调控作用；用腺病毒混慢病毒siRNA 或过表达Gas6转染方法研究其可能的信号通路，以阐明Gas6促进衰老心肌血管新生和成熟的作用及机制，为老年缺血性疾病的防治提供新思路。

背景：生长抑制特异性蛋白6（Growth arrest-specific 6，Gas6）参与冠心病患者血管新生关键细胞内皮祖细胞（EPCs）的动员，并且是缺血性心脏病的保护因子，该保护作用是否与Gas6促进衰老心肌血管新生和成熟有关，尚未见报道。.研究内容：本项目在前期研究基础上，以缺血/缺氧为刺激因素，建立各年龄代小鼠心梗模型、老年EPCs和老年大鼠微血管内皮细胞缺氧/复氧模型，运用免疫组化、影像学和细胞生物学等方法观察Gas6对血管新生和成熟的双重调控作用；用腺病毒混慢病毒siRNA 或过表达Gas6转染方法研究其信号通路，以阐明Gas6促进衰老心肌血管新生和成熟的作用及机制，为老年缺血性疾病的防治提供新思路。.结果：通过细胞实验我们证明：过表达Gas6通过其受体Axl激活下游PI3K/Akt、mTOR/p70s6k及MEK/ERK通路，并上调HIF-1α的表达水平，可促进内皮细胞血管新生。.通过动物实验我们证明：外源性gas6干预组较对照组，小鼠心功能得到明显改善，射血分数（LVEF）和左心室缩短率（FS）显著提高；Masson染色表明缺血后心肌纤维化明显减轻；Tunnel实验显示心肌凋亡显著降低。在衰老缺血心肌中，外源性给予Gas6能通过促进心肌血管新生，减少心肌细胞凋亡，进而改善心脏功能，改善衰老缺血性心脏病动物模型的预后。.意义：本课题组阐明了Gas6促进衰老心肌血管新生和成熟的作用及机制，可为老年缺血性心脏病的防治提供新思路，有着较大的临床应用价值。