Impact of Endotoxin on Fibroblast Growth Factor 23 Production by Osteocytes

说明(申请人提供)：成纤维细胞生长因子23(FGF23)是一种磷酸和维生素D调节激素，主要由骨骼中的骨细胞产生。这种激素的循环水平在肾脏损伤的最早阶段就开始升高，并随着肾脏疾病的进展而显著升高。FGF23不仅导致CKD患者的矿物质和骨骼异常，还可能直接导致该人群的心血管病变。尽管有大量证据表明FGF23与不良结局有关，但在慢性肾脏病中促进这种激素水平升高的机制仍不明确。最近的报道表明，CKD患者的血液中细菌内毒素水平较低，这可能是这些患者常见的炎症表型的原因之一。虽然这种内毒素血症的病因尚不清楚，但普遍的假设是，在肾脏损伤的情况下，肠道屏障功能受损，允许内毒素进入血液。有趣的是，CKD患者循环内毒素水平的升高模式与FGF23相似，在疾病的早期阶段有适度的增加，在晚期CKD阶段有更显著的上升。此外，最近的观察表明，CKD的全身炎症和FGF23水平之间存在独立的关联，这可能暗示着病理上的联系。众所周知，细菌内毒素对骨生理有多方面的影响，主要是内毒素。内毒素最成熟的骨骼作用之一是通过促进破骨细胞分化来刺激骨吸收 通过主要的内毒素共同受体CD14传递信号。尽管内毒素对成骨细胞和破骨细胞生物学的影响进行了大量的研究，但关于内毒素对骨细胞的影响的公开数据有限，骨细胞是产生FGF23的主要细胞来源。我们实验室的初步数据表明，小鼠一次注射脂多糖就能刺激骨骼中FGF23基因的表达。目前尚不清楚这是直接作用于骨骼中的骨细胞，还是继发于内毒素的其他全身作用。我们假设进行性肾功能不全患者中观察到的内毒素血症通过CD14依赖的机制导致骨中骨细胞产生FGF23增加。我们计划通过以下方式来检验这一假设：(I)进行体外和体内研究，以调查 (Ii)通过评估注射脂多糖后CD14-/-小鼠的FGF23的产生(目标2)，研究CD14信号在CD14-/-小鼠产生FGF23中的作用(目标2)；以及(Iii)通过评估腺嘌呤摄入诱导CKD后CD14-/-小鼠的骨产生FGF23，确定内毒素血症是否通过CD14依赖的机制导致CKD的FGF23的过度产生(目标3)。

DESCRIPTION (provided by applicant): Fibroblast growth factor 23 (FGF23) is a phosphaturic and vitamin D-regulatory hormone that is primarily produced by osteocytes in bone. Circulating levels of this hormone begin to rise at the earliest stages of kidney damage and become markedly elevated with progression of kidney disease. FGF23 not only contributes to mineral and bone abnormalities in patients with CKD, but may also directly induce cardiovascular pathology in this population. Despite substantial evidence linking FGF23 to adverse outcomes, the mechanisms that promote elevated levels of this hormone in CKD remain undefined. Recent reports indicate that patients with CKD exhibit low levels of bacterial endotoxin in their bloodstreams, which may contribute to the inflammatory phenotype that is commonly observed in these patients. While the etiology of this endotoxemia remains unclear, the prevailing hypothesis is that intestinal barrier function is compromised in the setting of kidney injury, allowing endotoxin entry into blood. Interestingly, the pattern of rise for circulating endotoxin levels in CKD mirrors that of FGF23, with modest increments occurring in the early stages of disease and a more dramatic rise in advanced CKD stages. Furthermore, recent observations suggest an independent association between systemic inflammation and FGF23 levels in CKD, possibly implying a pathologic link. It is well-documented that bacterial endotoxin, primarily lipopolysaccharide (LPS), has multiple effects on bone physiology. One of the most established skeletal actions of LPS is a stimulation of bone resorption by promoting osteoclast differentiation via signaling through CD14, a primary LPS co-receptor. Despite a substantial number of studies examining endotoxin effects on osteoblast and osteoclast biology, there is limited published data on LPS effects on osteocytes, the primary cell source for FGF23 production. Preliminary data from our lab suggests that a single injection of LPS in mice stimulates FGF23 gene expression in bone. It remains unclear if this is a direct action on osteocytes in bone or secondary to other systemic actions of LPS. We hypothesize that endotoxemia observed in patients with progressive renal dysfunction contributes to increased FGF23 production by osteocytes in bone in this setting through a CD14-dependent mechanism. We plan to test this hypothesis by: (i) performing both in vitro and in vivo studies to investigate the direct impact of LPS on osteocyte production of FGF23 (Aim 1); (ii) examining the role of CD14 signaling in LPS- mediated FGF23 production by bone by evaluating FGF23 production in CD14-/- mice following LPS injection (Aim 2); and (iii) determining if endotoxemia contributes to the overproduction of FGF23 in CKD through a CD14-dependent mechanism by evaluating FGF23 production by bone from CD14-/- mice following the induction of CKD by adenine ingestion (Aim 3).