PTH通过ATF3/V-ATPase信号通路调节终末期肾脏病血磷的机制研究

The disorder of phosphorus is a common complication and cause of death in end stage renal disease (ESRD). Our and other researcher’s data have shown that the release of phosphorus from bone is very important for regulating blood phosphorus in patients with ESRD. However, the exact mechanism of the bone releases phosphorus is unclear. Our preliminary experiments revealed that there was parathyroid hormone receptor, and parathyroid hormone (PTH) increased the expression of activating transcription factor 3 (ATF3) on osteoclast. ATF3 may be the transcription factor for the V-ATPase A3 gene. We speculate that PTH increase blood phosphorus by increasing ATF3 participation in V-atpase a3 transcription regulation, and release H+ from cytoplasm to the outside of a cell. In order to elucidate the mechanism, we intends to use ChIP experiment to obtain the reliable evidences that ATF3 directly participates in V-ATPase a3 gene regulation, and to observe its effect on osteoclast activity and phosphorus release by using gene transfection and silencing technique, and by using ATF3 transgenic and knockout mice to construct ESRD model. This study will be the first to elucidate the new pathway of PTH in regulating blood phosphorus, which provides a new theory and target for the treatment of patients with higher phosphorus in ESRD.

磷代谢紊乱是终末期肾脏病(ESRD)常见并发症及死亡原因。最新研究显示：ESRD患者骨骼释放磷对调节血磷非常重要，甲状旁腺激素(PTH)参与骨骼磷的释放，但机制不清。我们预实验发现：PTH通过其受体上调破骨细胞激活转录因子3(ATF3)，促进空泡型氢离子三磷酸腺苷转运酶a3亚基（V-ATPase a3）基因表达，提示PTH通过ATF3/V-ATPase途径增加骨溶解，从而上调血磷。为了验证上述假设，本课题拟采用ChIP实验，获得ATF3直接参与V-ATPase a3基因转录调控的可靠证据；用基因转染、沉默技术干预ATF3，观察其对破骨细胞活性及磷释放的影响；用ATF3转基因及敲除基因小鼠构建ESRD模型，观察干扰ATF3对血磷的影响。本研究首次阐明PTH调节血磷的新途径，为ESRD高磷血症的治疗提供新靶点。

慢性肾脏病(CKD)是常见病及多发病，据估计终末期肾脏病（ESRD）的患病率为2~3人/万，全国需要透析的人数达到200~300万人。血透患者继发性甲状旁腺功能亢进(SHPT)发病率高达50~70%，甲状旁腺切除(PTX) 术是治疗严重SHPT重要措施。PTX术改善临床症状及降低甲状旁腺素（PTH），但是导致低钙、低磷血症及低血压。既往认为PTX术后低磷血症与“骨饥饿”有关，目前研究认为可能与骨骼释放磷减少有关，但骼释放磷的确切机制尚未明了。PTX术后低血压患者即使充分补钙，仍然不能纠正低血压，而PTH有扩血管降压作用，提示PTX术后低血压可能有其他机制存在。　.维持性血液透析患者严重继发性甲状旁腺功能亢进患者，给予手术切除甲状旁腺术后，患者PTH明显下降，同时血磷也明显降低。同时，患者的软组织钙化也减轻。实验发现PTH作用破骨细胞PTH受体，上调活化转录因子3(ATF3)，活化空泡型氢离子三磷酸腺苷转运酶（V-ATPase 3）及泌H+，促进骨骼释放磷。采用基因转染方法制备了制备永生化的甲状旁腺细胞株及其构建方法，申请发明专利，探讨GCM2对PTH分泌的影响。.实验发现高血压相关钙调节蛋白 (HCaRG/COMMD5)在PTX术后血中增多，而HCaRG/COMMD5刺激胸主动脉，对肾上腺素收缩有抵抗作用，提示HCaRG/COMMD5有舒张血管功能。对SHPT大鼠行甲状旁腺切除术，比较各器官HCaRG/COMMD5表达水平。并探讨HCaRG/COMMD5作用心房钠尿肽（ANP）及一氧化氮合酶(NOS)分子途径，以明确PTX术后导致低血压的机制。.本研究结果填补了PTH调节血磷的机制，为临床ESRD患者的高磷血症治疗提供了新的治疗靶点。并且明确了继发性甲状旁腺功能亢进患者甲状旁腺切除术后低血压的机制。

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