MSX2-Wnt Signaling in Cardiovascular Calcification

描述（由申请人提供）：心血管钙化是衰老、糖尿病、高胆固醇血症、高血压、异常瓣膜力学和炎症以及慢性肾功能不全的常见后果。曾经被认为是良性的，钙化性血管病变的有害临床后果现在变得越来越清楚;它是导致CVA、MI和PVD的病理生理学的关键组成部分--钙化性血管病变的解剖结构和程度预示着截肢和心血管死亡。治愈或基本上逆转大血管钙化（MVC）的能力代表了未满足的临床需求。需要更好地理解MVC的启动和进展。成骨和炎症基因调控程序在MVC中被激活-包括外膜，中膜，内膜和瓣膜组织-通过与控制骨生理学的机制重叠的机制。我们已经证明，糖尿病和血脂异常激活了主动脉肌成纤维细胞中的促成骨程序--一种“前馈”BMP 2-Msx 2-Wnt信号级联反应。相比之下，PTH/PTHrP受体激动剂（例如，teriparabine）抑制血管肌成纤维细胞钙化和主动脉Msx 2-Wnt基因表达。我们现在研究肌成纤维细胞Msx 2-Wnt信号在MVC中的作用和调节。在目标1中，我们使用LDLR-/-.TOPGAL+报告小鼠（Wnt响应性TCF/LEF元件控制下的LacZ转基因）将主动脉Wnt信号传导的空间激活与饮食诱导的Msx 2-Wnt表达和从外膜、中层和瓣膜肌成纤维细胞募集骨祖细胞联系起来。我们还测试了我们在CMV-Msx 2转基因小鼠中观察到的增强的主动脉钙化和Wnt表达是否激活主动脉LacZ表达。原代主动脉肌成纤维细胞的共培养研究将测试Msx 2激活的旁分泌Writ信号传导是否被PTH抑制（1-34）。在目标2中，使用TOPGAL小鼠，我们测试PTH（1-34）是否在体内抑制经典Wnt信号传导。在目标3中，我们绘制了Msx 2启动子蛋白-DNA相互作用，该相互作用在肌成纤维细胞中传递对PTH（1-34）的转录抑制，强调了介导BMP 2和Wnt活化的元件。

DESCRIPTION (provided by applicant): Cardiovascular calcification is a common consequence of aging, diabetes, hypercholesterolemia, hypertension, abnormal valve mechanics & inflammation, & chronic renal insufficiency. Once thought benign, the deleterious clinical consequences of calcific vasculopathy are now becoming clear; it is a key component of pathophysiology leading to CVA, Ml, and PVD -- with amputation and cardiovascular mortality portended by the anatomy and extent of calcific vasculopathy. The ability to cure or substantially reverse macrovascular calcification (MVC) represents an unmet clinical need. A better understanding of MVC initiation and progression is required. Osteogenic & inflammatory gene regulatory programs are activated in MVC-- variably involving adventitial, medial, intimal, and valvular tissues -- via mechanisms overlapping those that control bone physiology. We've shown that a pro-osteogenic program -- a "feed-forward" BMP2-Msx2-Wnt signaling cascade -- is activated in aortic myofibroblasts by diabetes and dyslipidemia. By contrast, PTH/PTHrP receptor agonists (e.g., teriparatide) suppress vascular myofibroblast calcification and aortic Msx2-Wnt gene expression. We now study the role and regulation of myofibroblast Msx2-Wnt signaling in MVC. In Aim 1, we use LDLR-/-.TOPGAL+ reporter mice (LacZ transgene under control of Wnt-responsive TCF/LEF element) to relate spatial activation of aortic Wnt signaling to diet-induced Msx2-Wnt expression and recruitment of osteoprogenitors from adventitial, medial, & valvular myofibroblasts. We also test if the enhanced aortic calcification and Wnt expression we observe in CMV-Msx2 transgenic mice activates aortic LacZ expression. Co-culture studies of primary aortic myofibroblasts will test if Msx2--activated paracrine Writ signaling is inhibited by PTH (1-34). In Aim 2, using TOPGAL mice, we test if PTH(1-34) inhibits canonical Wnt signaling in vivo. In Aim 3, we map Msx2 promoter protein-DNA interactions that convey transcriptional suppression to PTH(1-34) in myofibroblasts, emphasizing elements that mediate BMP2 and Wnt activation.