肾性骨病方调节BMP/Smad信号通路防治肾性骨病的研究

Early prevention and treatment of renal osteodystrophy can decrease the incidence rates of complications such as fracture and improve life quality of patients. But there is still no safe and effective clinical medicine. Recent studies had revealed that decreased bone formation, increased FGF23 expression, abnormal serum phosphorus and calcitriol production and vascular calcification induced by chronic renal failure could be reversed by treatment with BMP7 at the early stage, which indicates that BMP7 can be the target molecule for medicine development. Kidney-reinforcing formula is used clinically for primary osteoporosis, and has been proved to be effective. Basic research reveled that BMP7 expression and the activity of BMP/Smad signaling pathway could be upregulated, and bone formation could be promoted furthermore by the effective componants of kidney-reinforcing formula. Therefore, we presume that kidney-reinforcing formula can also be used for early prevention and treatment in renal osteodystrophy, and the mechamism may be related to upregulation of BMP7 expression and the activity of BMP/Smad signaling pathway. At first, this study is to observe bone histomorphometry and metabolism in mice when Bmp7 is specifically knockout in kidney, and to observe whether overexpression BMP7 specifically in kidney can reverse bone mass loss and vascular calcification induced by chronic renal failure. Part 2 of this study is to observe whether kidney-reinforcing formula can pomote bone formation and alleviate bone mass loss and vascular calcification in mice with chronic renal failure through regulation of BMP/Smad signaling pathway.

早期防治肾性骨病，可以降低由此带来的骨折等并发症，提高患者生活质量，但临床上缺乏安全有效的药物。研究表明在小鼠肾功能不全早期给予BMP7治疗，可以逆转骨形成降低，进而抑制FGF23的高表达，调节钙磷代谢紊乱和PTH表达，抑制血管钙化等一系列并发症，提示可以以BMP7为调控靶点，研究早期防治的药物。我们在临床上运用补肾方治疗原发性骨质疏松症，疗效显著；基础研究发现其有效组分可以上调骨组织细胞中BMP7表达或上调BMP/Smad信号通路活性，促进成骨。据此我们推测，补肾方也可早期防治肾性骨病，机制与调节BMP7表达和BMP/Smad信号通路活性，促进骨形成有关。本研究将首先观察特异性肾脏敲除BMP7对骨代谢的影响，以及肾脏特异性过表达BMP7是否可以逆转肾功能不全小鼠的骨丢失和血管钙化；并观察补肾方药是否能通过调节BMP7/Smad信号通路，促进骨形成，预防或减轻肾性骨病和血管钙化等。

在肾功能不全早期针对肾性骨病进行有效防治，可以延迟患者肾性骨病的发生。本研究拟从BMP7角度观察肾对骨的调节机制，并以此为治疗药物开发切入点。①完成繁殖不同月龄肾小管上皮细胞条件性基因敲除BMP7小鼠（Ksp-cre，BMP7fx/fx），通过microCT检测，未发现敲除肾小管上皮细胞中的BMP7小鼠骨代谢存在显著异常。②重新繁殖获得肾小管上皮细胞条件性基因敲除Axin1小鼠（Ksp-cre，Axin1fx/fx），使Wnt/β-catenin信号通路持续激活，研究发现6月龄雌性转基因小鼠雌激素水平明显降低，骨量明显降低。通过micro array芯片筛选，发现6月龄雌性转基因小鼠相关P450酶等表达出现明显变化，可能与雌激素水平变化相关，本研究切入点变更为Wnt/β-catenin信号通路，该方案调整已于2016年底进展汇报中上报国家自然基金委。③上述研究提示，肾脏中的Wnt/β-catenin信号通路的激活并不能增加骨量；课题组前期研究表明，直接激活骨组织细胞中的Wnt/β-catenin信号通路能有效增加骨量。因此，从Wnt/β-catenin信号通路角度对肾性骨病早期治疗药物进行筛选，仍应针对性筛选能够激活骨组织细胞中Wnt/β-catenin信号通路的中药复方及其有效单体。建立了Wnt/β-catenin信号通路报告基因筛查系统，筛选可以激活Wnt/β-catenin信号通路的有效组分补肾中药单体。④通过体内药效学研究，证明肾性骨病方及其有效组分蛇床子素、特女贞苷等可以部分缓解肾功能不全小鼠骨丢失，且早期干预效果优于晚期干预。并从体内外研究发现蛇床子素可能通过激活成骨细胞中Wnt/β-catenin信号通路，上调成骨细胞OPG/RANKL表达水平，抑制破骨细胞形成，从而有效逆转肾功能不全小鼠骨丢失。