HSP22在高糖致内皮损伤中的作用及罗格列酮的干预研究

High glouse is one of initiating factors of endothelial damage.Rosiglitazone (RGS) could protect the vascular endothelial function against antioxidant stress except for controlling blood glucose. Heat shock protein (HSP) was a kind of stress-protective proteins. Studies showed HSP27 expression was up-regulated in endothelial cells(ECs) in high glouse environment and played a protective effect.RGS could upregulate the level of HSP27 in ECs. HSP22 was a novel member of small HSP. In our preliminary study showed that HSP22 could protect ECs from hypoxia and reoxygenation injury. We observed the expression of HSP22 was also significantly increased in high glucose (33.3mmol/l) environment.Therefore we hypothesize that HSP22 may be play a protective role in endothelial injury induced by high glouse, and the protective effect on endothelial may be via increasing expression of HSP22.In this study ,we will observe the levelof HSP22 expression in diabetes mice and the impact of RGS intervention. In the 2nd part of study,we will establish HSP22 knock-out mice diabetes model, then the relationship of HSP22 and ECs injury will be discussed. The 3rd part is in vitro experiment: we will observe the HSP22 expression in high glucose medium and intervention by RGS at the same time, the HSP22 recombinant plasmids and siRNA plasmid will be used to increase and inhibition of HSP22 expression, then the role of HSP22 in the injury of endothelial induced by high glucose and its mechanism will be clarified.

高糖是血管内皮损伤的始动因素之一，罗格列酮(RGS)对高糖致内皮损伤有保护作用。热休克蛋白（HSP）是一种应激保护蛋白，研究表明HSP27在高糖环境内皮表达升高并具保护作用,RGS可上调内皮细胞HSP27表达。HSP22属新发现小分子HSP,我们前期研究显示HSP22对缺氧复氧损伤HUVEC具显著保护作用,并发现高糖环境HUVEC中HSP22表达亦明显升高。据此提出假设：HSP22在高糖致内皮损伤中起保护作用,而RGS可能通过HSP22发挥保护作用。本研究首先观察糖尿病鼠主动脉内皮HSP22表达水平及RGS干预影响；然后建立HSP22基因敲除鼠糖尿病动物模型，观察其与内皮损伤的关系；离体实验部分，将采用高糖离体培养HUVEC,同时行RGS干预，并利用HSP22重组及RNAi质粒分别增加及抑制HSP22表达,以进一步探讨其机制,从而阐明HSP22在高糖所致内皮损伤中的作用和RGS干预影响。

高糖诱导产生的线粒体氧化应激是血管内皮细胞损伤的关键因素。热休克蛋白22（Heat shock protein 22， HSP22）是一种应激保护蛋白。我们前期研究显示HSP22对缺氧复氧损伤人脐静脉内皮细胞（Human umbilical vein endothelial cells，HUVEC）具显著保护作用,并发现高糖环境HUVEC中HSP22表达明显升高。然而，HSP22能否对抗高糖诱导的血管内皮细胞损伤还未明确。据此提出假设：HSP22通过减少线粒体活性氧（Mitochondrial reactive oxygen species，mtROS）保护高糖损伤的内皮细胞。本项目采用高脂联合链脲佐菌素构建小鼠2型糖尿病模型和高糖干预HUVEC模拟体外实验，同时采用过表达和沉默HSP22基因技术，探讨HSP22在高糖诱导内皮损伤中的作用及可能机制。本项目研究结果提示，HSP22通过抑制内皮细胞粘附作用和抑制细胞因子释放明显降低内皮细胞活化和损伤。体外激光共集聚和流式细胞术检测提示HSP22可减少高糖刺激HUVEC中mtROS产生和线粒体损伤。机制上，采用线粒体特异性抗氧化抑制剂 MitoTEMPO干预内皮细胞，发现HSP22通过减少高糖介导产生的mtROS抑制内皮细胞活化和损伤。此外，我们研究还发现HSP22通过抑制mtROS维持内皮细胞线粒体融合和分裂动态平衡。本研究为HSP22可能作用2型糖尿病血管损伤的治疗靶点提供了依据。