PATHOGENESIS OF HUMAN AND MURINE HYPERCALCIURIA

这项拟议研究的总体目标是了解人类特发性高钙尿(IH)高钙尿的分子基础。遗传性高钙尿性结石(GHS)大鼠血清CA正常，肠道钙吸收增加，钙负平衡和骨丢失加重，是研究人类高钙尿症和肾结石的独特动物模型。与一些IH患者一样，GHS大鼠的血清1，25(OH)2D3水平正常。肠、肾、骨和脾单核细胞中维生素D受体(VDR)水平升高可能通过夸大1，25(OH)2D3的生物学作用而介导GHS大鼠的高钙尿。IH患者外周血单核细胞(PBM)过度分泌具有破骨细胞刺激活性的细胞因子(IL-1α和β，IL-6)、肿瘤坏死因子α和GM-CSF。VDR的病理性增加和/或细胞因子的过度产生可能导致IH表型，这两种假说都将在IH患者和GHS大鼠中进行验证。这些假说将在三个特定目标上进行检验：1.血清1，25(OH)2D3正常的IH患者有过度的VDR和夸大的维生素D靶组织对1，25(OH)2D3的反应，以及外周血巨噬细胞过度产生细胞因子。对IH患者的研究将测量：a)真皮成纤维细胞和PBM VDR水平、24-羟基酶(24-OHase)活性和PBM细胞因子产生；b)十二指肠VDR水平和钙吸收；以及c)1，25(OH)2D3依赖和独立的骨吸收。2.在调节正常大鼠VDR水平的条件下，GHS大鼠VDR呈结构性增加：a)衰老、高钙饮食、甲状旁腺激素降低VDR AS；b)低钙饮食、低磷饮食增加VDR AS；c)肾单位共定位VDR、钙结合蛋白28kd和钙敏感受体。3.GHS大鼠细胞因子的产生：a)体外内毒素和1，25(OH)2D3刺激；b)低钙饮食；c)体内抑制IL-1β、肿瘤坏死因子α对尿钙、肠钙转运和VDR水平的影响；d)体外细胞因子对单核细胞VDR和24-OHase活性的影响。建议的研究将导致治疗和预防高钙尿症和钙性肾结石的新方法。

The overall objective of the proposed research is to understand the molecular basis for the hypercalciuria of human idiopathic hypercalciuria (IH). Features of IH including normal serum CA, increased intestinal Ca absorption, and negative Ca balance and bone loss exaggerated by low Ca diet are also found in the genetic hypercalciuric stone-forming (GHS) rat, which is a unique animal model to study human hypercalciuria and nephrolithiasis. Like some patients with IH, GHS rats have normal serum 1,25 (OH)2D3 levels. Elevated vitamin D receptor (VDR) levels in intestine, kidney, bone, and splenic monocytes may mediate the hypercalciuria in GHS rats by exaggerating 1,25 (OH)2D3 biologic actions. Peripheral blood monocytes (PBMs) from IH patients overproduce cytokines (IL-1alpha and beta, IL-6), TNFalpha, and GM- CSF) that have osteoclast-stimulating activity. Pathologic increases in VDR and/or over-production of cytokines may result in the IH phenotype and both hypotheses will be tested in IH patients and GHS rats. The hypotheses will be tested in three specific aims: 1. That IH patients with normal serum 1,25 (OH)2D3 have excessive VDR and exaggerated vitamin D target tissue response to 1,25(OH)2D3 and that PBMs overproduce cytokines. Studies in IH patients will measure: a) dermal fibroblast and PBM VDR levels, 24-hydroxylase(24- OHase) activity, and PBM cytokine production; b) duodenal VDR levels and Ca absorption; and c) 1,25 (OH)2D3-dependent and independent bone resorption. 2. That VDR is constitutively increased in GHS rats by measuring VDR under conditions that modulate VDR levels in normal rats: a) decreased VDR as during aging, high Ca diet, parathyroid hormone; b) increased VDR as during low Ca diet, and low phosphate diet; and c) nephron co- localization of VDR, calbindin 28 kd, and Ca-sensing receptor. 3. Cytokine production in GHS rats: a) in vitro LPS- and 1,25 (OH)2D3- stimulated; b) during low Ca diet; c) effect of in vivo inhibition of IL- 1beta, TNFalpha action on urine Ca, intestinal Ca transport, and VDR levels; and d) effect of in vitro cytokines on monocyte VDR, and 24 - OHase activity. The proposed studies should lead to novel approaches to the treatment and prevention of hypercalciuria and Ca nephrolithiasis.