TRPM7通道介导的血管平滑肌特异性Ca2+/Zn2+失稳态促腹主动脉瘤发生的机理研究

Abdominal aortic aneurysm (AAA) is a severe vascular disease with vascular rupture being the major cause of death. Disruption of medial elastic lamina is the hallmark of AAA but the precise mechanisms remains largely undefined. This study aims to investigate the role of ion channels in the pathogenesis of AAA. To this end, we screened a total of 28 TRP (Transient Receptor Potential) channels and found for the first time that only TRPM7 (subtype melastatin 7) channel was upregulated specifically in vascular smooth muscle cells (VSMC) in AAA. Treatment of AAA mice with a newly characterized TRPM7 blocker - waixenicin A - significantly reduced AAA incidence. Genetic silencing of TRPM7 suppressed the enzymatic activity of VSMC-derived matrix metalloproteinase (MMP2 & MT1-MMP). Activation of the 2 matrix-degrading enzymes required the presence of intracellular Zn2+, and expression of MMP2 was regulated by intracellular Ca2+. Given that TRPM7 is a unique cation channel that permeates Ca2+ & Zn2+, we proposed that perturbation of vascular smooth muscle cell intracellular Ca2+/Zn2+ levels by TRPM7 upregulation underlies the occurrence of AAA. This study will employ a novel TRPM7 blocker - waixenicin A and VSMC-specific TRPM7 knockout mice, use a mouse model of aneurysm (infusion of Apoe knockout mice with angiotensin II) and AAA patient samples and conduct in vivo and in vitro experiments, in order to fully elucidate the mechanisms by which TRPM7 promotes AAA. This study will evaluate if TRPM7 represents a novel therapeutic target for AAA treatment as well as if waixenicin A could be utilized to treat AAA.

腹主动脉瘤（abdominal aortic aneurysm，AAA）是一种严重血管病变，血管破裂是主要死因。中膜弹力层降解是特征病理改变，机理未完全阐明。本研究以离子通道异常为新切入点首次发现AAA时TRPM7在血管平滑肌细胞特异性上调；新型TRPM7阻断药waixenicin A显著降低小鼠AAA发病率； TRPM7基因沉默抑制血管平滑肌基质金属蛋白酶活性；MT1-MMP与MMP2酶激活依赖胞内锌离子，MMP2表达受钙离子调节。TRPM7为一独特Ca2+/Zn2+通道，因此假设：TRPM7上调致血管平滑肌胞内Ca2+/Zn2+失稳态而活化MMP进而促AAA发病。本研究拟使用waix及血管平滑肌特异性TRPM7敲除小鼠，结合AAA动物模型与病人标本，进行体内外实验以阐明TRPM7介导钙/锌失稳态而促AAA的机制。本研究将评估TRPM7是否为防治AAA新靶点及waix治疗AAA的应用前景

腹主动脉瘤（Abdominal aortic aneurysm, AAA）以中膜弹力层破坏为主要病理特征，危害大但发病机制尚未阐明，药物治疗手段缺乏。本研究首次从“胞内离子失稳态”角度，聚焦Trpm7离子通道探讨该病发病机制。构建内皮（Trpm7fl/fl;Tie2-Cre）、血管平滑肌（Trpm7fl/fl;Myh11-CreERT2）及巨噬细胞（Trpm7fl/fl;Cx3cr1-CreERT2, Trpm7fl/fl;Lyz2-Cre）特异性Trpm7敲除小鼠品系后复制CaCl2诱导的AAA模型，发现仅平滑肌敲除Trpm7降低AAA致病关键酶pro-MMP2表达与活化，减轻AAA发生发展。进一步构建Apoe-/-背景内皮、平滑肌及巨噬细胞敲除小鼠，复制AngII诱导的AAA模型，亦发现仅血管平滑肌特异性敲除Trpm7（Apoe-/-;Trpm7fl/fl;Myh11-CreERT2）显著抑制pro-MMP2表达与活化，减轻AAA发生发展。动物及细胞实验均证实，Trpm7通道开放后介导Ca2+内流激活Calcineurin/TORC2/CREB，介导Zn2+内流激活锌敏感转录因子MTF1，两条信号通路协同促进pro-MMP2基因转录。因此，本研究揭示了Trpm7通道介导的平滑肌特异性Ca2+/Zn2+失稳态促进AAA发生。本研究首次从离子失稳态角度，多细胞类型多疾病模型相互比较，共同确证了Trpm7是防治AAA的新靶点，为开发高效特异通道阻断药物奠定了实验基础。此外，该研究亦发现前体mRNA可变剪接（alternative splicing-coupled nonsense mediated decay, AS-NMD）是调控Trpm7表达的重要新机制，是该通道于疾病状态下表达改变的可能原因。该项目已标注发表论文6篇，共培养10名研究生，3名已完成学位论文。