成纤维生长因子23在磷代谢紊乱相关性疾病发病过程中的作用及其机制的研究

As the regulatory of inorganic phosphate mechanism has remained elusive until now, there is non-specific treatment for diseases related to disorders of phosphate metabolism. Fibroblast growth factor 23(FGF23), the central regulator of Pi metabolism, exerts regulatory function in type II sodium-dependent Pi cotransporter (Npt2), which mediate about 80% reabsorption process, located in renal proximal tubule(RPT).The applicant previous studies confirmed that severe phosphate disorders were caused by mutations of Npt2 gene, it is noteworthy that Npt2a played the key role. Furthermore, preliminary experiments showed that increased serum FGF23 level caused by high serum phosphate level could induce the significantly increased expression of FGFR4 and CaN in kidney proximal tubule epithelial cells (do not express Klotho); meanwhile, the expression and enzyme activity of CaN was also increased significantly. On the basis of previous studies we hypothesize that FGF23 can regulate Npt2a expression directly through activation of FGFR4-CaN-NFAT signaling pathway, furthermore, this effect was independent of Klotho. This project aims to further study this hypothesis using Klotho gene knockout mouse and OK cell (do not express Klotho) treated with different interventions and then further explore the changes of critical factors related to Pi metabolic, biochemical indicators and signaling pathway. This study will reveal the pathogenesis and development mechanism of diseases related to disorders of phosphate metabolism; furthermore, it will provide the theoretical basis for novel diagnosis and therapy of these diseases.

磷代谢调控机制尚不明确，因此磷代谢紊乱相关性疾病缺乏特异性治疗。成纤维生长因子23(FGF23)是肾近曲小管（RPT）Ⅱ型钠磷协同转运子Npt2（介导80%肾磷重吸收）表达调控的核心因子。申请人前期研究证实Npt2基因突变会导致严重磷代谢紊乱，其中Npt2a起主导作用。预实验发现血磷水平增高导致FGF23水平增高会引起RPT上皮细胞（不表达Klotho）上FGFR4、CaN、NFAT表达水平及CaN磷酸酶活性显著增高；据此提出假说：FGF23在RPT可不依赖Klotho通过FGFR4-CaN-NFAT通路调控Npt2a表达。本项目拟利用Klotho基因敲除鼠及OK细胞（不表达Klotho）模型，通过不同方法诱导磷及FGF23水平发生变化，进一步检测磷代谢、生化指标、通路中关键因子等相应的变化来验证此假说。本研究将揭示磷代谢紊乱相关性疾病发生发展机制，为此类疾病诊疗方案制定提供重要理论依据。

磷具有多种生理功能，其代谢平衡可以保证机体多项生理功能得以正常进行。磷在肾近曲小管刷状缘顶端细胞的跨膜转运过程是其代谢的限速步骤及调控的主要部位。此限速步骤过程是由Ⅱ型钠磷协同转运子（type II sodium-dependent Pi cotransporter, Npt2）辅助转运完成的。目前发现与肾脏磷转运相关的主要有Npt2a和Npt2c两种，因此Npt2是体内磷代谢平衡的关键因子。.磷代谢调控机制尚不明确，因此磷代谢紊乱相关性疾病缺乏特异性治疗。本研究证实成纤维生长因子23（fibroblast growth factor 23,FGF23）水平异常是磷代谢相关疾病发生、发展及预后的独立危险因素。本研究在前期研究的基础上，建立基因敲除鼠模型，通过特殊饮食诱导与体内血磷、血钙及尿磷、尿钙等水平发生变化，进一步检测与磷代谢及调控的生化指标尤其是FGF23、关键因子及肾脏矿化程度等相应的变化来进一步研究磷代谢及其调控的具体机制。.本研究主要包括两个部分：第一部分通过建立基因敲除鼠模型，并用不同的饮食进行干预，检测与磷代谢及调控相关的生化指标及关键因子水平尤其是FGF23水平对肾脏矿化程度的影响；并进一步研究由不同钙磷含量饮食所引起的不同血钙磷浓度及尿磷、尿钙排泄指数、及相关调节因子尤其是FGF23水平与磷代谢过程之间的作用。同时观察肾脏矿化程度的变化,研究HHRH患者肾脏矿化的预测因子。总之通过本研究可能够进一步明确磷代谢和调节的相关机制和关键因子，为临床上表现为磷代谢紊乱相关疾病寻找新的致病突变，优化现有治疗并寻求新的治疗方案。第二部分首先利用体内实验，通过不同成分进行对基因敲除鼠进行尾静脉注射进一步明确，FGF23调控磷代谢的信号通路；同时利用OK细胞模型，在体外通过不同的干预方式进一步证实并明确FGF23通过CaN-NFAT通路来调控Npt2A的表达进而调控磷代谢。