Renal regulation of phosphate homeostasis and its effect on bone

项目概要/摘要 持续升高的甲状旁腺激素（PTH）会导致高钙血症/低磷血症， 增强骨转换。这导致骨丢失，因为骨吸收增加更显着 比骨形成。相比之下，每日注射PTH仅通过短暂的矿物质促进骨形成， 离子变化。令人惊讶的是，在骨中表达组成型活性PTH/PTHrP受体（PTHR 1）的小鼠 （Col 1-H223 R或Dmp 1-H223 R）显示显著的骨量增加。同样，受影响的患者 假性甲状旁腺功能减退症（肾脏中的PTH抵抗，但骨中无），用钙和1，25（OH）2治疗 维生素D（1，25 D）可以显示骨形成的主要增加，尽管PTH水平升高。表现 因此，转基因小鼠和人类疾病表明，持续的PTHR 1激活可以增加骨密度， 如果防止过量尿磷酸盐排泄，则会出现肿块。大量证据表明， PTH的磷酸盐排泄不仅依赖于cAMP/PKA信号，还依赖于IP 3/PKC依赖性的磷酸盐代谢。 机制等事实上，表达磷脂酶C（PLC）缺陷型PTHR 1的D/D小鼠不能维持其表达。 当PTHR 1被持续激活时，磷酸盐排泄。此外，野生型动物接受 长期输注[Trp 1]PTH（1-34）（一种有偏见的PTH类似物，具有受损的IP 3信号传导）未能维持 磷酸盐排泄和1，25 D产生。除了肾脏的重要性，PTH刺激的 PLC/IP 3/PKC信号传导是骨形成所必需的。例如，对野生型小鼠进行了 持续的PTH依赖性PTHR 1激活显示成纤维细胞样基质骨（FSB）快速扩张 细胞;在PTH持续升高的D/D小鼠和 输注[Trp 1]PTH（1-34）的野生型小鼠，因此支持以下结论： PTHR 1是正常骨形成所必需的。我们现在将确定持续的IP 3/PKC信号传导是否 PTHR 1是长期调节磷酸盐稳态和1，25 D产生所必需的， 持续的PTHR 1激活是否可以增强骨形成，如果尿磷酸盐排泄过量， 是可以预防的。本研究的两个目的是：目的1将进一步探讨PTH依赖性IP 3/PKC是否与PTH相关。 持续的磷酸盐排泄、1，25-D的产生和细胞增殖都需要信号传导。 成骨细胞前体目的2将确定高骨量在Col 1-H223 R小鼠和动物中是否 骨细胞特异性SIK 2/3消融可通过促进肾磷酸盐排泄来逆转， PTH非依赖性干预，即通过新型NPT 2a抑制剂或通过重组FGF 23。 我们还将确定“趋骨”PTH类似物是否能诱导骨形成， 持续局部PTHR 1激活，但肾脏作用有限，即无磷酸盐尿和无1，25 D增加。 我们的研究将有助于进一步阐明PLC依赖的PTHR 1信号在肾脏和骨骼中的作用， 以及磷酸盐在两个基因改变的小鼠中维持高骨量的重要性。

PROJECT SUMMARY/ABSTRACT Continuously elevated parathyroid hormone (PTH) causes hypercalcemia/hypophosphatemia and enhances bone turnover. This results in bone loss because bone resorption increases more prominently than bone formation. In contrast, daily PTH injection enhances bone formation with only transient mineral ion changes. Surprisingly, mice expressing a constitutively active PTH/PTHrP receptor (PTHR1) in bone (Col1-H223R or Dmp1-H223R) show profound bone mass increases. Likewise, patients affected by pseudohypoparathyroidism (PTH-resistance in kidney, but not in bone), treated with calcium and 1,25(OH)2 vitamin D (1,25D), can show a major increase in bone formation, despite elevated PTH levels. Findings in transgenic mice and a human disease thus suggested that persistent PTHR1 activation can increase bone mass, if excess urinary phosphate excretion is prevented. Considerable evidence indicates that urinary phosphate excretion by PTH depends not only on cAMP/PKA signaling, but also on IP3/PKC-dependent mechanisms. In fact, D/D mice expressing a phospholipase C (PLC)-deficient PTHR1 do not sustain phosphate excretion when the PTHR1 is continuously activated. Furthermore, wild-type animals receiving long-term infusions of [Trp1]PTH(1-34), a biased PTH analog with impaired IP3 signaling, failed to sustain phosphate excretion and 1,25D production. In addition to its renal importance, PTH-stimulated PLC/IP3/PKC signaling is required for bone formation. For example, tibiae of wild-type mice undergoing prolonged PTH-dependent PTHR1 activation showed rapid expansion of fibroblast-like stromal bone (FSB) cells; such osteoblast precursors were not observed in D/D mice with continuous PTH elevations and in wild-type mice infused with [Trp1]PTH(1-34), thus supporting the conclusion that PLC-signaling at the PTHR1 is required for normal bone formation. We will now determine whether persistent IP3/PKC-signaling at the PTHR1 is required for long-term regulation of phosphate homeostasis and 1,25D production, and whether continuous PTHR1 activation can enhance bone formation, if excess urinary phosphate excretion can be prevented. Two aims will be pursued: Aim 1 will explore further whether PTH-dependent IP3/PKC signaling is required for sustained phosphate excretion, for 1,25D production, and for expansion of osteoblast precursors. Aim 2 will determine whether the high bone mass in Col1-H223R mice and animals with osteocyte-specific SIK2/3 ablation can be reversed by promoting renal phosphate excretion through PTH-independent interventions, namely through a novel NPT2a inhibitor or through recombinant FGF23. We will also determine whether “bone-seeking” PTH analogs can induce bone formation because of continuous local PTHR1 activation, yet limited renal actions, i.e. no phosphaturia and no 1,25D increase. Our investigations will help clarify further the role of PLC-dependent PTHR1 signaling in kidney and bone, and the importance of phosphate in maintaining high bone mass in two genetically altered mice.