Intestinal phosphate absorption occurs through both a paracellular mechanism involving tight junctions and an active transcellular mechanism involving the type II sodium-dependent phosphate cotransporter NPT2b (SLC34a2). To define the contribution of NPT2b to total intestinal phosphate absorption, we generated an inducible conditional knockout mouse, Npt2b Inorganic phosphate is an essential mineral critical for cellular processes and bone mineralization. Severe disruptions in serum phosphate have pathologic consequences. 1,2 Hypophosphatemic disorders are associated with rickets, osteomalacia, and a host of secondary dysfunctions. 3 In contrast, hyperphosphatemia associated with chronic kidney disease (CKD) is linked tightly to increased risk of cardiovascular morbidity and mortality. 4 -6 Recent studies show that elevated phosphate concentrations within the high normal range in individuals with functional kidneys also are correlated with increased cardiovascular risk and mortality. 7,8 Thus, an elevated serum phosphate level is an emerging health risk. Despite the importance of maintaining a relatively narrow serum phosphate range, nearly 70% of dietary phosphate is absorbed, resulting in transient postprandial increases in serum phosphate concentrations. 9 Normalization of serum phosphate appears to be managed primarily within the renal proximal tubule by the type II sodium-dependent phosphate cotransporters NPT2a (SLC34a1) and NPT2c (SLC34a3). Genetic knockout mouse models demonstrate that 80% and 20% of total urinary phosphorus are managed by the Npt2a and Npt2c transporters, respectively. 10,11 Chronic and acute regulation of these renal transporters is modulated by changes in dietary and serum phosphate

Chronic kidney disease–mineral bone disorder (CKD‐MBD) is defined by abnormalities in mineral and hormone metabolism, bone histomorphometric changes, and/or the presence of soft‐tissue calcification. Emerging evidence suggests that features of CKD‐MBD may occur early in disease progression and are associated with changes in osteocyte function. To identify early changes in bone, we utilized the jck mouse, a genetic model of polycystic kidney disease that exhibits progressive renal disease. At 6 weeks of age, jck mice have normal renal function and no evidence of bone disease but exhibit continual decline in renal function and death by 20 weeks of age, when approximately 40% to 60% of them have vascular calcification. Temporal changes in serum parameters were identified in jck relative to wild‐type mice from 6 through 18 weeks of age and were subsequently shown to largely mirror serum changes commonly associated with clinical CKD‐MBD. Bone histomorphometry revealed progressive changes associated with increased osteoclast activity and elevated bone formation relative to wild‐type mice. To capture the early molecular and cellular events in the progression of CKD‐MBD we examined cell‐specific pathways associated with bone remodeling at the protein and/or gene expression level. Importantly, a steady increase in the number of cells expressing phosphor‐Ser33/37‐β‐catenin was observed both in mouse and human bones. Overall repression of Wnt/β‐catenin signaling within osteocytes occurred in conjunction with increased expression of Wnt antagonists (SOST and sFRP4) and genes associated with osteoclast activity, including receptor activator of NF‐κB ligand (RANKL). The resulting increase in the RANKL/osteoprotegerin (OPG) ratio correlated with increased osteoclast activity. In late‐stage disease, an apparent repression of genes associated with osteoblast function was observed. These data confirm that jck mice develop progressive biochemical changes in CKD‐MBD and suggest that repression of the Wnt/β‐catenin pathway is involved in the pathogenesis of renal osteodystrophy. © 2012 American Society for Bone and Mineral Research.

The incidence of cardiovascular events and mortality strongly correlates with serum phosphate in individuals with CKD. The Npt2b transporter contributes to maintaining phosphate homeostasis in the setting of normal renal function, but its role in CKD-associated hyperphosphatemia is not well understood. Here, we used adenine to induce uremia in both Npt2b-deficient and wild-type mice. Compared with wild-type uremic mice, Npt2b-deficient uremic mice had significantly lower levels of serum phosphate and attenuation of FGF23. Treating Npt2b-deficient mice with the phosphate binder sevelamer carbonate further reduced serum phosphate levels. Uremic mice exhibited high turnover renal osteodystrophy; treatment with sevelamer significantly decreased the number of osteoclasts and the rate of mineral apposition in Npt2b-deficient mice, but sevelamer did not affect bone formation and rate of mineral apposition in wild-type mice. Taken together, these data suggest that targeting Npt2b in addition to using dietary phosphorus binders may be a therapeutic approach to modulate serum phosphate in CKD.

Altered bone turnover is a key pathologic feature of chronic kidney disease-mineral and bone disorder (CKD-MBD). Expression of TGF-β1, a known regulator of bone turnover, is increased in bone biopsies from individuals with CKD. Similarly, TGF-β1 mRNA and downstream signaling is increased in bones from jck mice, a model of high-turnover renal osteodystropy. A neutralizing anti-TGF-β antibody (1D11) was used to explore TGF-βs role in renal osteodystrophy. 1D11 administration to jck significantly attenuated elevated serum osteocalcin and type I collagen C-telopeptides. Histomorphometric analysis indicated that 1D11 administration increased bone volume and suppressed the elevated bone turnover in a dose-dependent manner. These effects were associated with reductions in osteoblast and osteoclast surface areas. μCT confirmed the observed increase in trabecular bone volume and demonstrated improvements in trabecular architecture and increased cortical thickness. 1D11 administration was associated with significant reductions in expression of osteoblast marker genes (Runx2, alkaline phosphatase, osteocalcin) and the osteoclast marker gene, Trap5. Importantly, in this model, 1D11 did not improve kidney function or reduce serum PTH levels indicating that 1D11 effects on bone are independent of changes in renal or parathyroid function. 1D11 also significantly attenuated high turnover bone disease in the adenine-induced uremic rat model. Antibody administration was associated with a reduction in pSMAD2/SMAD2 in bone but not bone marrow as assessed by quantitative immunoblot analysis. Immunostaining revealed pSMAD staining in osteoblasts and osteocytes but not osteoclasts, suggesting 1D11 effects on osteoclasts may be indirect. Immunoblot and whole genome mRNA expression analysis confirmed our previous observation that repression of Wnt/β catenin expression in bone is correlated with increased osteoclast activity in jck mice and bone biopsies from CKD patients. Furthermore, our data suggests that elevated TGF-β may contribute to the pathogenesis of high turnover disease partially through inhibition of β-catenin signaling.

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