E1A激活基因阻遏子通过调控溶酶体稳态对高血压血管重塑的保护作用研究

From the perspective of vascular homeostasis regulation, exploring novel protective molecules repressing hypertensive vascular remodeling has become an active hot spot in the research field of prevention and treatment of hypertension. Cellular repressor of E1A-stimulated gene (CREG), a novel lysosomal protein, is widely expressed in mature tissues. Serial studies from our laboratory have demonstrated that CREG plays essential role in maintaining vascular homeostasis, and antagonizes proliferative vascular diseases including restenosis after percutaneous coronary intervention (PCI) and atherosclerosis. Our recent studies suggested that CREG expression in the blood vessels is inversely related to hypertensive vascular remodeling. In addition, CREG deficiency aggravates Ang II-induced vascular remodeling in hypertension, indicating that CREG may be a key molecule in regulating hypertensive vascular remodeling. To further clarify the role of CREG in the regulation of hypertensive vascular remodeling, we firstly intend to establish animal models of secondary hypertension based on wild-type mice, CREG-deficient heterozygous mice and smooth muscle cells specific CREG transgenic mice, respectively. Then adenovirus-based CREG gene delivery will be applied to identify the role of CREG in the treatment of hypertension and vascular remodeling. Finally, we sought to uncover the key molecules mediating the CREG’s role against vascular injury from the view of protection of lysosomal function. The findings will highlight that CREG might serve as a novel therapeutic target for the prevention and treatment of hypertensive vascular remodeling.

从血管自身稳态调控角度，寻找保护分子，遏制高血压血管重塑发生，已成为高血压防治研究新的热点。E1A激活基因阻遏子基因（CREG）是成熟细胞溶酶体中广泛表达的小分子糖蛋白。本室系列研究证实，CREG能维持血管分化稳态，对抗PCI术后再狭窄和动脉粥样硬化等增生性血管病发生。近期研究发现：高血压血管重塑发生过程中伴随CREG表达下调，CREG表达减少加重血管紧张素II诱导的高血压和血管重塑，提示CREG可能是高血压血管重塑的重要调控分子。本项目拟建立CREG正常表达、CREG表达减少和平滑肌过表达CREG的小鼠继发性高血压模型，明确CREG在高血压血管重塑发生及演进中的调控作用；通过腺病毒载体介导的CREG基因外源性干预，阐明CREG对小鼠高血压和血管重塑的治疗作用；并尝试从CREG对溶酶体功能保护角度，揭示CREG对抗高血压血管重塑的关键分子，为高血压防治提供新的靶点。

从血管自身稳态调控角度，寻找保护分子，遏制高血压血管重塑发生，已成为高血压防治研究新的热点。因此，本项目旨在明确内源性分子E1A激活基因阻遏子（CREG）在高血压血管重塑发生及演进中的调控作用，并探讨其溶酶体稳态调控的分子机制。研究内容主要包括3各方面：（1）CREG在高血压血管重塑发生及演进中的调控作用；（2）CREG保护溶酶体稳态调控高血压血管重塑的机制；（3）CREG对高血压血管重塑的治疗作用及机制。我们通过尸检收集高血压患者和正常人颈外动脉血管标本，通过高盐饮食喂养Dahl盐敏感大鼠、小鼠皮下埋植微渗透泵在体输注血管紧张素II（Ang II）或去甲肾上腺素成功构建高血压血管重塑模型，发现高血压血管重塑过程中伴随管壁中CREG基因表达下调，且下调时间早于血管重塑发生时间，提示CREG基因参与高血压血管重塑过程。应用CREG表达减少的CREG杂合子（CREG+/-）小鼠建立高血压血管重塑模型，发现CREG基因表达减少加重Ang II诱导的高血压血管重塑发生，提示CREG基因是高血压血管重塑过程中的关键调控基因。进一步观察CREG基因表达对血管壁及原代血管平滑肌细胞（VSMCs）中溶酶体数量及功能的影响，发现与CREG+/+小鼠相比，Ang II灌注的CREG+/-小鼠主动脉中溶酶体标志物LAMP1和LAMP2表达增多，但溶酶体酶Cathepsin D表达不变，而外源性添加重组人CREG（rhCREG）蛋白可抑制Ang II诱导的VSMCs中I型胶原的表达，同时增加溶酶体数量及溶酶体酶的表达。外源性添加巴弗洛霉素A1、氯喹和NH4Cl抑制溶酶体功能，rhCREG蛋白抑制Ang II诱导的VSMCs表达I型胶原的作用消失。外源性给予CREG+/+小鼠rhCREG蛋白或携带CREG基因的腺病毒可剂量依赖性地抑制Ang II诱导高血压血管重塑，并上调管壁中溶酶体酶的数量及功能，外源性给予氯喹抑制溶酶体功能后，rhCREG蛋白抑制高血压血管重塑的作用消失，提示CREG基因通过调控溶酶体酶数量级功能抑制高血压血管重塑。本项目首次阐明CREG介导的溶酶体稳态调控机制在高血压血管重塑中的作用，为临床高血压及血管稳态失衡引起血管性疾病的防治研究提供重要思路和治疗靶点。