LOX-1在代谢综合征小鼠脑缺血再灌注损伤加重中的关键作用

Stroke is the number 2 cause of death worldwide. It is reported that stroke is more prevalence in people with metabolic syndrome, and for whom a poorer prognosis is predicted. However, no effective treatment has been developed yet. Our preliminary experiments showed that mice fed on a high-fat diet induced obese, hyperglycemia and hyperlipidemia. They also resulted in aggravated damage after cerebral ischemia reperfusion injury comparing to mice fed with normal diet, the mechanism of which is still unknown. Inflammation after ischemia is one of the most important steps toward cell injury. And microglia activation is a key step of inflammatory injury after brain ischemia. Recent studies showed an over-expression of oxidized low concentration lipoprotein (ox-LDL) in high fat fed animals, while its major receptor - leptin like oxLDL receptor (LOX-1) was found widely expressed in many type of cells in the mouse brain including endothelia cells, neurons and microglia cells in our preliminary study. And it was up-regulated in the microglia in the penumbra region after ischemia, thus might induce NLRP3 inflammasome activation since NLRP3 was showed to increase after oxidative stress in macrophages. We also found that LOX-1 inhibitor reduced cultured microglia damage caused by OGD and reduced the elevation level of NLRP3 in microglia after OGD injury. However, as the macrophages in CNS, the role of microglia and its activation in the aggravation effect of ischemic brain injury in metabolic syndrome mice was not determined yet. This project is designed to explore the role of LOX-1/NLRP3 inflammasomes in the aggravation of brain ischemic damage in mice with metabolic syndrome, preferably the role of LOX-1/NLRP3 pathway in ox-LDL induced aggravation of microglia oxidative stress and OGD injury. Furthermore, we are to explore the possibility of reducing ischemic brain damage in metabolic syndrome mice through the interference with LOX-1 or NLRP3 inflammasomes activation and expression. The sequel will provide important information and novel target for the treatment of ischemic brain injury in patients with metabolic syndrome.

脑卒中是全球第二大致死性疾病，研究显示以肥胖高脂血症为代表的代谢综合征人群脑卒中风险增高。我们前期研究发现：高脂饲养所致的代谢综合征小鼠脑缺血损伤明显加重，但机制尚不清楚。脑缺血后的炎症反应是导致神经细胞损伤的主要原因之一，而小胶质细胞的激活是缺血后炎症反应的关键步骤。我们前期研究显示高脂饮食造成体内oxLDL上调，其受体LOX-1在小鼠脑内多种细胞中表达，且在缺血半影区小胶质细胞中表达增强，可能引起巨噬细胞内NLRP3炎症小体的激活介导细胞损伤；LOX-1抑制剂可减轻小胶质细胞氧糖剥夺损伤并抑制NLRP3炎症小体的表达。因此我们推测：代谢综合征动物可能由于LOX-1/NLRP3炎症小体的过度激活而加重脑缺血再灌注损伤。本项目拟以LOX-1/NLRP3通路在小胶质细胞缺血缺氧损伤中的作用为切入点，探讨其在代谢异常小鼠脑缺血再灌注损伤加重中的作用，为代谢综合征患者脑卒中损伤提供新的治疗靶点。

本课题以高脂饮食致肥胖与高脂血症为基础，从体内、体外两方面探索了该条件下脑缺血损伤是否加重，以及其中的重要机制。发现高脂饮食可诱导小鼠脑血管重塑，脑缺血损伤加重、水肿、渗出增加，出血性转化发生率及严重程度均增高，而MMP9的活性，而非表达含量在其中起着重要作用。MMP9-/-后高脂饮食，不在能诱发小鼠脑缺血损伤的加重。另一方面，高脂饮食引起脑缺血后半影区LOX-1前体蛋白（50kDa）表达显著增高，但Nlrp3则在正常饮食动物缺血前后变化更加明显。高脂高糖培养均能在体外诱发小胶质细胞LOX-1高表达与Nlrp3表达的增加，并且在缺血或氧糖剥夺后不同时间均高于正常培养对照，可能是加重氧糖剥夺后炎症反应的关键因素。抑制Lox-1的配体受体结合可逆转正常与高脂高糖培养下OGD损伤加重的现象，并在体内也可引起神经保护作用。此外研究发现高脂饮食还可引起MCAO后14d的脑损伤面积增加并遗留环境相关恐惧记忆缺损，Nlrp3的抑制剂治疗可减少正常饮食动物的脑损伤但对高脂饮食动物无显著疗效。而主动脉内膜的LOX-1染色以及巨噬细胞浸润也在高脂饮食后明显增加，脑缺血可加重这一现象，说明高脂饮食可引起全身血管炎症，更容易产生斑块，而脑组织损伤可加重这种全身的血管炎症，甚至影响板块的稳定性，引起斑块脱落，造成二次损伤。