Rapamycin induced Inhibition of aortic elastolysis in a murine model of marfan syndrome

马凡氏综合征是一种复杂的遗传性疾病，其症状多效性是由马凡氏蛋白-1基因突变引起的。主要症状是血管疾病、透镜脱位和骨骼变形。对于危及生命的并发症，如主动脉瘤和夹层，仍然没有系统的治疗方法。这种疾病的血管成分的病理生理学可以用主动脉中膜微纤维的不稳定来解释。这种不稳定性是由于基质金属蛋白酶（MMP）在主动脉平滑肌细胞中的异常高表达和活性所致。这组酶降解弹性蛋白纤维，并通过微纤维的断裂导致更高的TGF-β生物利用度。所有这些发现最终导致主动脉壁的不稳定。MMP表达较高的原因在于TGF-β驱动的信号传导途径的较高活性。相关的信号级联之一是mTOR（雷帕霉素的机制靶点）途径。mTOR是一种高度保守的蛋白质，调节细胞分化、细胞增殖和凋亡等中枢细胞机制。此外，文献综述表明，动物模型中诱导性主动脉瘤的扩张与雷帕霉素对mTOR的抑制之间存在相关性。我能够在初步实验中表明，在小鼠马凡氏小鼠的主动脉壁冷冻切片中也存在较高的mTOR活性。因此，本项目的工作假设是，马凡氏综合征等遗传性主动脉疾病是mTOR依赖性的。为了更深入地了解潜在的机制，需要进行一些实验。首先，将在来自Marfan小鼠的鼠主动脉平滑肌细胞（mAoSMC）中进行雷帕霉素诱导的mTOR体外抑制。这将提供mTOR抑制是否能够降低细胞培养物中MMP的表达和活性的见解。在此概念验证之后，将进行体内实验。这里的计划是抑制马凡氏小鼠的弹性蛋白溶解，并检测主动脉壁结构的机械变化。此外，我的目标是减少或完全抑制主动脉瘤的扩张，以提高生存率。到目前为止，对马凡氏患者唯一可用的治疗方法是手术矫正扩张的主动脉瘤。到目前为止，还没有一种系统性疗法能够实现其承诺。相比之下，雷帕霉素和其他mTOR抑制剂已经被很好地确立为移植医学中的免疫调节剂。随着雷帕霉素依赖性抑制主动脉瘤形成的成功，可以优化全身治疗。

Marfan syndrome is a complex genetic disorder with a pleiotropy of symptoms caused by a mutation in the fibrillin-1 gene. The leading symptoms are vascular disorders, lens luxations and skeletal deformations. There is still no systemic therapy for the life threatening complications like aortic aneurysms and dissections. The pathophysiology of this disease’s vascular components can be explained by instability of the microfibrils in the aortic media. The instability is due to an abnormally high expression and activity of matrix metallo proteinases (MMP) from the aortic smooth muscle cells in the media. This group of enzymes degrades the elastin fibers and through a fragmentation of the microfibrils leads to a higher TGF-β bioavailability. All of these findings ultimately cause a destabilisation of the aortic wall. The reason for a higher MMP expression is rooted in a higher activity of TGF-β driven signalling pathways. One of the associated signalling cascades is the mTOR (mechanistic target of rapamycin) pathway. mTOR is a highly conserved protein which regulates central cell mechanisms as cell differentiation, cell proliferation and apoptosis. In addition, literature reviews suggest that there is a correlation between the expansion of inducible aortic aneurysms in animal models and the inhibition of mTOR via rapamycin.I was able to show in preliminary experiments that there is also a higher activity of mTOR in cryosections of the aortic wall of murine Marfan mice.Therefore, the working hypothesis of this project is that hereditary aortic diseases like Marfan syndrome are mTOR dependent. For a closer understanding of the underlying mechanisms several experiments are needed. First a rapamycin induced inhibition of mTOR in vitro in murine aortic smooth muscle cells (mAoSMC) from Marfan mice will be performed. This should provide the insight of whether the mTOR inhibition is capable of reducing the expression and activity of MMPs in the cell culture. After this proof of concept in vivo experiments will be carried out. The plan here is to inhibit elastolysis in the Marfan mouse and detect mechanistic changes in the aortic wall structure. In addition I aim to reduce or totally inhibit the expansion of aortic aneurysms for a survival benefit.Up till now the only therapy available for Marfan patients is the surgical correction of the expanding aortic aneurysms. No systemic therapy has so far lived up to its promises. In contrast rapamycin and other mTOR inhibitors have been already well established as imunmodulators in transplantation medicine. With a successful rapamycin-dependent inhibition of aortic aneurysm formation a systemic therapy could be optimized.