MGP 调控 BMP2 信号通路影响肾草酸钙结石形成的机制

Randall’s Plaque (RP) theory, one of the classical theories explaining the pathogenesis of nephrolithiasis, suggests that the renal interstitial ectopic calcification may serve as the initial nidi for renal stones. Our previous research found that the transdifferentiation of tubular epithelial cells to osteoblast-like cells which was probably mediated by the abnormal activation of BMP2 signaling pathway was involved in the formation of RP. Matrix Gla protein (MGP) which has strong relationships with both BMP2 signaling pathway and ectopic calcification was found to play an important role in renal stone formation recently. We also have found that MGP was highly expressed in the renal tissue of genetic hypercalciuria stone-forming rats which are susceptible to nephrolithiasis. However, the expression of active carboxylated MGP was significantly reduced. And base on the above we can make this speculation reasonably: the dysfunction of MGP which can be caused by many different reasons results in abnormal BMP2 activation and tubular cells transdifferentiation, and subsequently leads to the RP and stones formation. This research program tries to clarify the roles of MGP in tubular cells transdifferentiation, RP and renal stone formation and their mechanisms by investigating the gene polymorphism and posttranslational modification of MGP, the effect of MGP on tubular cells biological functions related to stone formation, the regulation of BMP2 by MGP, and whether calcification and nephrolithiasis can be prevented by regulating the function of MGP in three aspects including cell, animal and human. We hope this research could provide a new theoretical and experimental basis for the prevention and control of nephrolithiasis.

Randall钙斑理论是阐释肾结石形成的经典理论之一，该理论认为肾间质异位钙化斑是肾结石形成的起始病变。我们前期研究发现，BMP2通路介导的肾小管上皮细胞向成（软）骨样细胞转分化参与钙斑形成。最新研究提示，与BMP2通路和异位钙化关系密切的基质Gla蛋白(MGP)与肾结石密切相关。我们在易自发形成结石的遗传性高钙尿大鼠肾组织中发现其总 MGP 表达升高，但其中具有功能活性的羧化cMGP不升反降，藉此推测：多种因素可导致的MGP功能障碍使其无法充分发挥对BMP2通路及肾小管上皮细胞表型转化的抑制作用，最终导致钙斑和结石形成。本课题拟在细胞、动物和人体层面，从MGP基因多态性和翻译后羧化修饰、MGP对细胞相关生物学功能影响、MGP对BMP2的调控、调节MGP功能对钙斑和结石的预防等方面，探究MGP在肾小管上皮细胞表型转化、钙斑和肾结石形成中的作用及其机制，以期为肾结石防治提供新的理论基础。

MGP是维生素K依赖性蛋白家族的成员，是目前发现最强大的组织钙化抑制剂，其活性形式是cMGP，无活性形式为ucMGP。有文献报道MGP参与了肾结石的形成，但其发挥的作用并不清楚。本研究构建了新型高钙尿结石形成大鼠模型，发现该大鼠肾组织MGP的表达较正常大鼠明显减少。通过分析肾结石患者的肾乳头芯片数据，我们也证实MGP在结石患者肾乳头中表达减少。我们发现高钙可以抑制大鼠肾小管上皮细胞NRK-52E中MGP的表达，MGP在钙盐沉积中发挥了抑制作用。通过测定肾结石患者血ucMGP水平，我们发现结石复发患者的血ucMGP水平明显高于结石初发者，血ucMGP≥2400ng/mL是肾结石复发的危险因素。我们还发现，正常情况下调控GGCX并不影响MGP的表达水平，但是抑制GGCX的表达可以使ucMGP水平升高。在高钙环境下，过表达GGCX可以使ucMGP水平下降并抑制BMP2和OPN的表达，而干扰GGCX则促进BMP2和OPN的表达。补充VitK1、Vitk2和华法令对NRK-52E细胞中MGP的表达并无影响，但是补充VitK1和Vitk2可以降低细胞上清中ucMGP的水平，而补充华法令使得ucMGP的水平升高。为高钙尿结石形成大鼠补充VitK2和氢氯噻嗪后，大鼠肾组织中的钙盐沉积、凋亡发生和OPN的表达较1,25(OH)2D3组明显减少。补充VitK2的结石大鼠血ucMGP水平以及肾组织中BMP2的表达明显低于结石大鼠，而补充氢氯噻嗪组的结石大鼠血ucMGP水平和肾组织中BMP2的表达与结石大鼠相比无明显差异。补充华法令后，结石形成大鼠肾组织中钙盐沉积明显增多，且该组大鼠血ucMGP水平、肾组织中BMP2和OPN的表达水平也明显升高。我们的研究表明MGP可以通过抑制BMP2信号通路抑制结石形成，补充维生素K有望为结石的防治提供一种简单、有效的方法。