慢性肾脏病致磷稳态失衡的机制研究

Phosphate homeostasis in the body is maintained by phosphate sensing and metabolism in the kidney and intestine, which is coordinately regulated by parathyroid hormone (PTH), fibroblast growth factor 23 (FGF23) and active vitamin D (1,25(OH)2D3). Chronic kidney disease (CKD) is the main reason for the disruption of phosphate homeostasis. However, the pathogenesis and systemic characteristic of this process remain unknown. Previous studies show that not only sodium-dependent phosphate cotransporters (NPTs) but also sodium hydrogen exchanger 3(NHE3) may participate in phosphate transport in both renal proximal tubular and intestinal epithelial cells. Our epidemiological data also indicated that serum phosphate levels were positively correlated with serum sodium levels in healthy Chinese population. Being the only humoral factor which acts on every organ responsible for phosphate metabolism (intestine, parathyroid gland, kidney and bone) and interacts with all the other phosphate regulatory hormones, 1,25(OH)2D3 has been evidenced recently to have potential gene regulatory effects on NPTs as well as NHE3. Therefore, we hypothesize that: (1) sodium ions and phosphate ions might synergistically regulate phosphate metabolism and sodium metabolism through the same cell sensors and their downstream pathway. (2) 1,25(OH)2D3 might be the key factor responsible for intracellular homeostasis in phosphate sensing cells. (3) Reduced 1,25(OH)2D3 synthesis due to nephron loss in CKD might be responsible for the abnormal phosphate sensing and dysregulation of phosphate metabolism in target organs and cells. We propose to validate the hypothesis through studies on proximal renal tubule, intestine, and parathyroid gland via in vitro intervention, conditional VDR gene knockout mice and CKD animals, to provide new theory and systemic evidence on the mechanisms of CKD-associated phosphate homeostasis disturbances.

机体磷稳态的维系依赖肾脏、肠道对磷的感应和代谢，以及PTH、FGF23、活性维生素D（1,25D）等激素的协调作用。慢性肾脏病（CKD）是导致磷稳态失衡的主因，目前学界对其发病机制的系统特性尚缺乏了解。以往研究及本课题组前期工作表明，正常人血磷与血钠正相关；近端肾小管及小肠上皮细胞钠磷转运子和钠氢交换子3均参与了磷转运；1,25D是唯一同时作用于磷代谢所有器官和调磷激素的体液因子，对上述磷转运蛋白具有潜在多向基因调控作用。故我们推测：钠离子和磷酸根离子通过共同细胞感应机制协同参与磷代谢和钠代谢；1,25D是磷感应细胞中调控磷转运的自稳机制；CKD肾单位减少致1,25D合成下降导致多部位磷感应及调控异常是磷稳态失衡的始动因素和进展机制。我们拟通过体外细胞干预和条件性VDR基因缺陷小鼠等方法，围绕近端肾小管、肠道和甲状旁腺验证上述假说，为阐明CKD致磷稳态失衡机制提供新理论和系统性研究依据。

慢性肾脏病（CKD）导致机体磷稳态失衡的确切机制不明，本项目围绕磷代谢主要器官肾脏、肠道、甲状旁腺，多角度验证“肾单位减少导致活性维生素D（1,25(OH)2D3，1,25D）合成及其受体（VDR）介导的生物学作用下降，引起多器官磷感应受损，共同参与磷稳态失衡”这一科学假说。主要研究成果如下，（一）临床研究：证明健康成人摄入不同磷饮食后体内维持磷稳态的调控网络出现明显差异，但肾脏合成分泌的循环激素1,25D始终是关键因素；高磷饮食可导致水钠潴溜和收缩压升高，低磷饮食则改善肠道菌群紊乱。（二）体外研究：（1）在人和大鼠甲状旁腺组织中发现高磷通过促进VDR泛素降解、刺激Pit-1/1,25D-VDR/NFκB信号通路等多种途径导致PTH合成分泌增加，且甲状旁腺嗜酸细胞增多是重要组织病理学改变；（2）在肾脏近端小管上皮细胞（HK2）中证实高磷通过Npt2a/NHE3介导引起胞内钙离子浓度升高，直接抑制细胞内1,25D生物合成，进而导致PTH1R及Klotho基因表达下降，参与调磷激素PTH和FGF23的肾脏作用抵抗，应用VDR激动剂可逆转上述异常；（3）自行构建小肠类器官，同样发现1,25D-VDR可调控肠上皮细胞中多种磷感应相关基因的表达，包括介导磷跨细胞主动转运的Npt2b及调节细胞旁被动转运的紧密连接蛋白，是影响肠道磷吸收的重要机制。（三）动物研究：成功构建不同VDR基因缺陷小鼠和CKD疾病模型，证实在全身VDR敲除小鼠和肾小管VDR敲除小鼠体内存在明显的负磷平衡，而肠道VDR敲除小鼠则呈现正磷平衡。因受疫情影响，有关饮食磷干预和VDR激动剂对这些动物磷稳态和器官磷感应的影响研究仍在进行中。本项目从系统生物学角度出发，率先在磷代谢主要器官中证明1,25D-VDR作用减弱是介导组织细胞磷感应异常的共性机制，为阐明CKD背景下磷稳态失衡的发病机制提供了新理论和干预新靶点。本项目已发表SCI收录论文13篇，获授权专利2项，软件著作权1项。培养跨学科青年学术骨干5人，晋升高级职称2人，并完成9名博士生和2名硕士生培养。