Mechanisms of intermittent parathyroid hormone effects on calcific aortic disease

 描述(申请人提供)：钙化性主动脉血管和瓣膜疾病(CAVD)死亡率高，但目前尚无治疗方法。一个令人兴奋的可能性是，目前用于骨质疏松的间歇性甲状旁腺激素(IPTH)治疗可能为CAVD提供治疗潜力，因为在骨形成和CAVD之间存在年龄无关的反向关系。事实上，在一项动物研究中，iPTH被发现可以防止CAVD的发生。然而，iPTH在既往CAVD患者中的作用仍不清楚。由于CAVD广泛存在，尤其是在需要iPTH治疗的人群中，确定iPTH对既往CAVD患者是有益还是有害是至关重要的。值得注意的是，与持续高血清甲状旁腺素水平促进骨丢失和CAVD相反，每天注射甲状旁腺素产生的间歇性高水平促进骨骼形成。最近的研究表明，iPTH和氧化应激之间存在意想不到的联系，氧化应激是CAVD的已知贡献者。其他人在成骨细胞中的研究和我们在血管平滑肌细胞(VSMC)中的初步发现表明，iPTH减少了细胞氧化应激，这表明iPTH是通过直接受体介导的机制。我们最近的报告还提出了第二种机制，即在高脂血症小鼠中，iPTH通过诱导血清对氧磷酶-1(PON1)的活性来降低循环中促炎脂质氧化产物(氧合脂)的水平。基于这些发现和我们之前的工作表明氧脂促进CAVD，我们假设iPTH将减弱先前存在的CAVD，并且部分直接通过降低细胞氧化应激水平，部分间接通过PON1减少循环中的氧脂水平。我们提出了三个目标。在目标1中，将测试iPTH对高脂血症小鼠先前存在的CAVD的影响。由于钙化斑块含有两个潜在的iPTH作用靶点，即VSMC和破骨前巨噬细胞，iPTH可能通过抑制成骨软骨分化和/或通过诱导矿物质吸收来抑制CAVD。我们将评估：1)体内18F‘microPET-microCT成像的主动脉钙沉积，这允许对单个小鼠进行连续扫描；2)氧化应激以及通过调节和标记物水平进行的成骨和破骨细胞分化。在目标2中，将确定iPTH的作用部位。系统机制将通过PON1缺陷小鼠(PON1-/-ApoE-/-)进行评估，直接机制将通过血管甲状旁腺素受体1缺陷小鼠(Pth1rΔVSMCApoE-/-)进行评估。在目标3中，我们将使用一种新的移植技术，将携带突变的1°小鼠VSMC的扩散腔植入宿主小鼠皮下，并将测试：1)iPTH与内源性PTH相关肽(PTHrP)对钙化的贡献；2)iPTH在糖尿病低骨转换骨质疏松背景下的作用。这些发现将揭示iPTH对既往心血管疾病患者的CAVD是一种治疗还是一种风险，以及iPTH信号如何实现血管特异性效应。

 DESCRIPTION (provided by applicant): Calcific aortic vascular and valvular diseases (CAVD) carry a high mortality, yet no medical treatment is available. An exciting possibility is that intermittent parathyroid hormone (iPTH) treatment, currently used for osteoporosis, may offer therapeutic potential for CAVD, given that there is an age-independent, inverse relationship between bone formation and CAVD. Indeed, in an animal study, iPTH was found to prevent initiation of CAVD. However, the effect of iPTH in subjects with pre-existing CAVD is still not clear. Since CAVD is widespread and especially prevalent in the population requiring iPTH treatment, it is crucial to determine whether iPTH is still beneficial or even harmful in those wit pre-existing CAVD. Of note, in contrast to continuously high serum PTH levels, which promote bone loss and CAVD, intermittently high levels produced by daily injection of PTH, promote skeletal bone formation. Recent developments suggest an unexpected link between iPTH and oxidant stress, a known contributor of CAVD. Studies by others in osteoblasts and our preliminary findings in vascular smooth muscle cells (VSMC) show that iPTH reduced cellular oxidant stress, suggesting a direct receptor-mediated mechanism. A second mechanism is also suggested by our recent report showing that, in hyperlipidemic mice, iPTH reduces circulating levels of proinflammatory lipid oxidation products (oxylipids) by inducing serum paraoxonase-1 (PON1) activity. Based on these findings and our previous work showing that oxylipids promote CAVD, we hypothesize that, iPTH will attenuate pre-existing CAVD and will do so, in part, directly by reducing levels of cellular oxidant stress and, in part, indirectly by reducing circulaing levels of oxylipids via PON1. We propose 3 Aims. In Aim 1, iPTH effects will be tested on pre-existing CAVD in hyperlipidemic mice. Since calcific plaque contains two potential targets of iPTH action, VSMC and preosteoclastic macrophages, iPTH may attenuate CAVD by inhibiting osteochondrogenic differentiation and/or by inducing mineral resorption. We will assess: 1) aortic calcium deposition by in vivo 18F¯ microPET-microCT imaging, which allows serial scanning of individual mice and 2) oxidant stress and osteochondrogenic and osteoclastic differentiation by levels of regulators and markers. In Aim 2, the iPTH sites of action will be determined. The systemic mechanism will be assessed using PON1- deficient mice (Pon1-/-ApoE-/-), and the direct mechanism will be assessed using vascular deficient PTH receptor 1 mice (Pth1rΔVSMCApoE-/-). In Aim 3, we will employ a novel transplant technique, in which diffusion chambers carrying mutated 1°murine VSMC will be implanted subcutaneously into host mice, and we will test: 1) contributions of iPTH vs. endogenous PTH-related peptide (PTHrP) on calcification and 2) effects of iPTH in the context of low bone turnover osteoporosis of diabetes. The findings will reveal whether iPTH is a treatment or a risk for CAVD in patients with pre-existing cardiovascular disease, and how iPTH signaling achieves vascular-specific effects.