AKR1C3介导炎性内皮微粒靶向调节血管间质干细胞分化在血管重构中的作用及机制

The main pathological characteristic of vascular remodeling is the abnormal accumulation of proliferation vascular smooth muscle cells (pSMC) in the intima, which is caused by inflammation. In recent reports, pSMC is derived from vascular mesenchymal stem cell (vMSC), but the mechanism is still unknown. Our hypothesis is “inflammation inhibits the differentiation of pSMC”. It means under the stimulation of inflammatory factors, endothelial cells (ECs) will release AKR1CR to vSMC via endothelial microparticles (EMPs). The activity of retinene reductase of AKR1C3 leads retinal insufficient supply, retinoic acid (RA) synthesis reduction and intracellular RA reduction. Furthermore, it will block the differentiation process, “vMSC→pSMC→cSMC”, in pSMC, which will lead to a massive accumulation of pSMC and causes vascular remodeling. Our research will use LPS to treat AKR1C3.overexpression ECs or AKR1C3 knockout ECs. Collecting EMPs and incubating it with vMSC and detecting the influence of AKR1C3 in vMSC differentiation; Co-culture of EC and vMSC by Transwell system to observe the role of EMPs in the transmission of AKR1C3; investigating the influence of AKR1C3 in vascular remodeling through endothelial specific AKR1C3 knockout mice. Our objective is to look for a new target for the prevention and treatment of vascular remodeling.

增殖型血管平滑肌细胞(pSMC)是血管重构的病理基础。近年报道pSMC主要由血管间质干细胞(vMSC)分化而来，但其调节机制不清楚。我们提出“炎症导致pSMC分化阻滞”的思路：炎症刺激下，内皮细胞(EC)通过释放内皮微粒(EMPs)将AKR1C3传递给vMSC, AKR1C3通过其视黄醛还原酶活性导致细胞内RA水平降低，使“vMSC→pSMC→cSMC”分化过程阻滞在pSMC节点，导致pSMC大量增加并引起血管重构。本研究拟采用脂多糖（LPS）处理AKR1C3过表达或敲除的EC，收集EMPs与vMSC共孵育，观察AKR1C3对vMSC分化的影响；利用Transwell培养体系进行EC/vMSC共培养，观察EMPs在传递AKR1C3中的作用；利用内皮特异性AKR1C3基因敲除小鼠，观察AKR1C3对血管重构的影响。为阐明炎症诱导血管重构的发生机制和寻找新的药物靶点提供理论指导及实验依据。

血管平滑肌细胞(VSMC)是血管重构的病理基础。近年报道增殖型VSMC主要由血管间质干细胞分化而来，但其调节机制不清楚。我们提出“炎症导致增殖型VSMC异常分化”的思路：即炎症刺激下，VSMC过度增殖或由收缩型向分泌增殖型分化，从而导致血管过度增生或重构。这一过程与AKR1C3、Wnt-5a、microRNA、内皮微粒(如外泌体)等信号分子或载体有关。药物可通过干预上述分子或载体调节VSMC分化增殖，进而抑制血管过度增生或重构。.本项目在进行充分文献调研的基础上，分别采用大鼠骨髓间充质干细胞(BMSCs)-VSMCs分化模型、小鼠股动脉内膜拉伤诱导血管增生和血管重构模型及Apo E-/-动脉粥样硬化模型，应用基因沉默等现代分子生物学技术，研究了炎性应激对VSMC分化、表型转变、增殖与血管增生、重构的影响及调节机制。发现：1)炎症可加速BMSCs-VSMCs，使其收缩特性发生改变，Caveolin-1/notch-1/Myocardin通路参与了VSMC收缩特性改变的调节过程，姜黄素（AKR1蛋白家族的抑制剂）通过上调Caveolin-1可解除由Notch-1介导的Myocardin的表达抑制进而增强细胞收缩；2)LncRNA-SRA高表达导致VSMC增殖、迁移增强，该过程与MEK、ERK和CREB磷酸化增强有关;3）Wnt5a通过促进NF-kB入核和炎症因子/细胞因子释放介导了VSMC增殖、脂质蓄积和血管重构，Wnt5a敲除可逆转这一过程；4）炎症/氧化应激可通过干扰自噬诱导VSMC蓄脂及增殖，雷公藤红素则通过激活自噬促进p62与c-MYC结合，加速c-MYC的泛素化降解，进而有效抑制VSMC增殖和内膜增生；5）外泌体在血管内皮细胞和血管平滑肌细胞之间发挥重要的信号传递及相互调节作用，正常内皮细胞释放的外泌体可抑制VSMC增殖，而由LPS和ox-LDL等炎性因子诱导的内皮细胞外泌体则诱导VSMC增殖，姜黄素及其衍生物烟酸姜黄素酯通过促进正常内皮外泌体释放、抑制炎性或氧化应激因子诱导的外泌体释放，参与血管内皮细胞对VSMC的调节。.上述研究共完成论文16篇，已发表12篇，申报专利1项，对阐明炎性因素在干预VSMC分化、增殖、表型转变及血管增生、重构中的作用与调控机制，指导研发干预药物具有一定指导意义。