Augmentative effects of leukemia inhibitory factor reveal a critical role for TYK2 signaling in vascular calcification.

Medial vascular calcification in chronic kidney disease (CKD) involves pro-inflammatory pathways induced by hyperphosphatemia. Several interleukin 6 family members have been associated with pro-calcific effects in vascular smooth muscle cells (VSMCs) and are considered as therapeutic targets. Therefore, we investigated the role of leukemia inhibitory factor (LIF) during VSMC calcification. LIF expression was found to be increased following phosphate exposure of VSMCs. LIF supplementation aggravated, while silencing of endogenous LIF or LIF receptor (LIFR) ameliorated the pro-calcific effects of phosphate in VSMCs. The soluble LIFR mediated antagonistic effects towards LIF and reduced VSMC calcification. Mechanistically, LIF induced phosphorylation of the non-receptor tyrosine-protein kinase 2 (TYK2) and signal transducer and activator of transcription-3 (STAT3) in VSMCs. TYK2 inhibition by deucravacitinib, a selective, allosteric oral immunosuppressant used in psoriasis treatment, not only blunted the effects of LIF, but also interfered with the pro-calcific effects induced by phosphate. Conversely, TYK2 overexpression aggravated VSMC calcification. Ex vivo calcification of mouse aortic rings was ameliorated by Tyk2 pharmacological inhibition and genetic deficiency. Cholecalciferol-induced vascular calcification in mice was improved by Tyk2 inhibition and in the Tyk2-deficient mice. Similarly, calcification was ameliorated in Abcc6/Tyk2-deficient mice after adenine/high phosphorus-induced CKD. Thus, our observations indicate a role for LIF in CKD-associated vascular calcification. Hence, the effects of LIF identify a central pro-calcific role of TYK2 signaling, which may be a future target to reduce the burden of vascular calcification in CKD.

Secondary calciprotein particle size is associated with patient mortality in peripheral artery disease.

BACKGROUND AND AIMS: Secondary calciprotein particles (CPP-II) induce inflammation and contribute to vascular calcification. CPP-II size is associated with vascular calcification in patients with chronic kidney disease (CKD) and all-cause mortality in hemodialysis patients. Here, we investigate for the first time a possible role of CPP-II size in patients with peripheral artery disease (PAD) without severe CKD. METHODS: We measured the hydrodynamic radius (Rh) of CPP-II by using dynamic light scattering in a cohort of 281 PAD patients. Mortality was evaluated over a period of ten years by central death registry queries. 35% of patients died during the observation period (median of 8.8 (6.2-9.0) years). Cox-regression analyses were performed to estimate hazard ratios (HR) and 95% confidence intervals (CI) and to allow for multivariable adjustment. RESULTS: The mean CPP-II size was 188 (162-218) nm. Older patients, patients with reduced kidney function, and those with media sclerosis had larger CPP-II (p < 0.001, p = 0.008, and p = 0.043, retrospectively). There was no association between CPP-II size and overall atherosclerotic disease burden (p = 0.551). CPP-II size was independently significantly associated with all-cause (HR 1.33 (CI 1.01-1.74), p = 0.039) and cardiovascular mortality (HR 1.52 (CI 1.05-2.20), p = 0.026) in multivariable regression analyses. CONCLUSIONS: Large CPP-II size is associated with mortality in PAD patients and might be a new feasible biomarker for the presence of media sclerosis in this patient population.

Calcification Propensity in Serum and Cardiovascular Outcome in Peripheral Artery Disease.

Peripheral artery disease (PAD) has been shown to be linked to elevated cardiovascular risk. The novel T50 test quantifies calcification propensity of serum and has been associated with cardiovascular events and mortality in patients with chronic kidney disease (CKD) and in the general population. This study investigated the association of calcification propensity measured by the T50 test in 287 patients with PAD without severe CKD. Major cardiovascular events (MACEs) including nonfatal stroke and nonfatal myocardial infarction and all-cause death (MACE + ) were evaluated after a median follow-up of 4 years and long-term cardiovascular and all-cause mortality after a median follow-up of 8.7 years by Kaplan-Meier and Cox regression analyses. Mean T50 time was 268 ± 63 minutes in the study cohort (age 69 ± 10 years, 32% women, 47% diabetes). Low T50 values that signify high calcification propensity were significantly associated with the occurrence of MACE+ (hazard ratio [HR]: 0.72; 95% confidence interval [CI]: 0.55-0.94). This association sustained multivariate adjustment for cardiovascular risk factors (CVRFs), Fontaine PAD stage, and prevalent media sclerosis (HR: 0.65; CI: 0.47-0.91). Cardiovascular mortality was significantly associated with T50 after multivariate adjustment for CVRF (HR: 0.72; CI 0.53-0.99), but not all-cause mortality (HR: 0.80; CI: 0.64-1.01). In conclusion, calcification propensity associates with MACE+ and cardiovascular mortality in patients with PAD.

Comparison of Iron Dosing Strategies in Patients Undergoing Long-Term Hemodialysis: A Randomized Controlled Trial.

BACKGROUND AND OBJECTIVES: Whether iron supplementation in patients on hemodialysis could be delivered by less frequent but higher single doses compared with the currently more common higher-frequency schedules of lower single iron doses is unknown. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: We carried out an open-label, randomized, controlled noninferiority trial over 40 weeks in patients on prevalent hemodialysis (n=142). We administered in total 2 g iron as 100 mg iron sucrose biweekly in a continuous (20 × 100 mg) fashion or 500 mg ferric carboxymaltose every 10 weeks in a periodic (4 × 500 mg) fashion. The primary end point was the change in hemoglobin at week 40 from baseline with a noninferiority margin of -0.8 g/dl. Secondary end points were changes in ferritin, transferrin, transferrin saturation, and erythropoiesis-stimulating agent use. RESULTS: In total, 108 patients completed the study. At 40 weeks, hemoglobin changed by -0.27 g/dl (95% confidence interval, -0.64 to 0.09) in the iron sucrose arm and by -0.74 g/dl (95% confidence interval, -1.1 to -0.39) in the ferric carboxymaltose arm compared with baseline. Noninferiority was not established in the per-protocol population as hemoglobin changes compared with baseline differed by -0.47 g/dl (95% confidence interval, -0.95 to 0.01) in the ferric carboxymaltose arm compared with the iron sucrose arm. Proportional changes from baseline to week 40 differed by -31% (98.3% confidence interval, -52 to -0.1) for ferritin, by 1% (98.3% confidence interval, -7 to 10) for transferrin, and by -27% (98.3% confidence interval, -39 to -13) for transferrin saturation in the ferric carboxymaltose arm compared with the iron sucrose arm. Erythropoiesis-stimulating agent dosing did not differ between groups. The overall number of adverse events was similar; however, more infections were observed in the iron sucrose arm. CONCLUSIONS: An equal cumulative dose of ferric carboxymaltose administered less frequently did not meet noninferiority for maintaining hemoglobin levels compared with iron sucrose administered more frequently. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Comparison Study of Two Iron Compounds for Treatment of Anemia in Hemodialysis Patients (COPEFER), NCT02198495.

Propensity for Calcification in Serum Associates With 2-Year Cardiovascular Mortality in Ischemic Heart Failure With Reduced Ejection Fraction.

Background: The propensity of serum to calcify, as assessed by the T50-test, associates with mortality in patients with chronic kidney disease. In chronic heart failure, phosphate and fibroblast growth factor-23 (FGF-23), which are important components of the vascular calcification pathway, have been linked to patient survival. Here, we investigated whether T50 associates with overall and cardiovascular survival in patients with chronic heart failure with reduced ejection fraction (HFrEF). Methods: We measured T50, intact and c-terminal FGF-23 levels in a cohort of 306 HFrEF patients. Associations with overall and cardiovascular mortality were analyzed in survival analysis and Cox-regression models. Results: After a median follow-up time of 3.2 years (25th-75th percentile: 2.0-4.9 years), 114 patients (37.3%) died due to any cause and 76 patients (24.8%) died due to cardiovascular causes. 139 patients (45.4%) had ischemic and 167 patients (54.6%) had non-ischemic HFrEF. Patients with ischemic HFrEF in the lowest T50-tertile had significantly greater 2-year cardiovascular mortality compared to patients in higher tertiles (p = 0.011). In ischemic but not in non-ischemic HFrEF, T50 was significantly associated with cardiovascular mortality in univariate (p = 0.041) and fully adjusted (p = 0.046) Cox regression analysis. Significant associations of intact and c-terminal FGF-23 with all-cause and cardiovascular mortality in univariate Cox regression analysis did not remain significant after adjustment for confounding factors. Conclusion: T50 is associated with 2-year cardiovascular mortality in patients with ischemic HFrEF but not in non-ischemic HFrEF. More research on the role of T50 measurements in coronary artery disease is warranted.

The Role of Iron and Erythropoietin in the Association of Fibroblast Growth Factor 23 with Anemia in Chronic Kidney Disease in Humans.

Anemia in chronic kidney disease (CKD) is an almost universal complication of this condition. Fibroblast growth factor 23 (FGF23), a key-player in mineral metabolism, is reportedly associated with anemia and hemoglobin levels in non-dialysis CKD patients. Here, we sought to further characterize this association while taking into account the biologically active, intact fraction of FGF23, iron metabolism, and erythropoietin (EPO). Hemoglobin, EPO, iron, and mineral metabolism parameters, including both intact and c-terminal-FGF23 (iFGF23 and cFGF23, respectively) were measured cross-sectionally in 225 non-dialysis CKD patients (stage 1-5, median eGFR: 30 mL/min./1.73m2) not on erythropoiesis stimulating agents or intravenous iron therapy. Statistical analysis was performed by multiple linear regression. After adjustment for eGFR and other important confounders, only cFGF23 but not iFGF23 was significantly associated with hemoglobin levels and this association was largely accounted for by iron metabolism parameters. cFGF23 but not iFGF23 was also associated with mean corpuscular hemoglobin (MCH) and mean corpuscular volume (MCV), again in dependence on iron metabolism parameters. Similarly, EPO concentrations were associated with cFGF23 but not iFGF23, but their contribution to the association of cFGF23 with hemoglobin levels was marginal. In pre-dialysis CKD patients, the observed association of FGF23 with hemoglobin seems to be restricted to cFGF23 and largely explained by the iron status.

Diagnostic and Prognostic Value of Soluble Urokinase-type Plasminogen Activator Receptor (suPAR) in Focal Segmental Glomerulosclerosis and Impact of Detection Method.

The plasma soluble urokinase-type plasminogen activator receptor (suPAR) is a biomarker for focal segmental glomerulosclerosis (FSGS), but its value is under discussion because of ambiguous results arising from different ELISA methods in previous studies. The aim of this study was to compare diagnostic performance of two leading suPAR ELISA kits and examine four objectives in 146 subjects: (1) plasma suPAR levels according to glomerular disease (primary, secondary and recurrent FSGS after kidney transplantation, other glomerulonephritis) and in healthy controls; (2) suPAR levels based on glomerular filtration rate; (3) sensitivity and specificity of suPAR for FSGS diagnosis and determination of optimal cut-offs; (4) suPAR as prognostic tool. Patients with FSGS showed significant higher suPAR values than patients with other glomerulonephritis and healthy individuals. This applied to subjects with and without chronic kidney disease. Although both suPARnostic™ assay and Quantikine Human uPAR ELISA Kit exerted high sensitivity and specificity for FSGS diagnosis, their cut-off values of 4.644 ng/mL and 2.789 ng/mL were significantly different. Higher suPAR was furthermore predictive for progression to end-stage renal disease. In summary, suPAR values must be interpreted in the context of population and test methods used. Knowing test specific cut-offs makes suPAR a valuable biomarker for FSGS.

Taurolidine-based catheter lock regimen significantly reduces overall costs, infection, and dysfunction rates of tunneled hemodialysis catheters.

Catheter-related infections and dysfunction are the main catheter complications causing morbidity and mortality in hemodialysis patients. However, there are no consistent data for the choice of catheter lock solutions for tunneled hemodialysis lines. In this prospective, multicenter, randomized, controlled trial, two lock regimens using three commercial catheter lock solutions were compared in 106 hemodialysis patients with a newly inserted tunneled central catheter. In the taurolidine group, TauroLock™-Hep500 was used twice per week and TauroLock™-U25,000 once a week. In the citrate group, a four percent citrate solution was used after each dialysis. Both groups were compared regarding catheter-related infections, catheter dysfunction, and costs. Over a period of 15,690 catheter days, six catheter-related infections occurred in six of 52 patients in the taurolidine group, but 18 occurred in 13 of 54 patients in the citrate group, corresponding to 0.67 and 2.7 episodes of catheter-related infections per 1000 catheter days, respectively (Incidence Rate Ratio 0.25, 95% confidence interval, 0.09 to 0.63). Catheter dysfunction rates were significantly lower in the taurolidine group (18.7 vs. 44.3/1000 catheter days) and alteplase rescue significantly more frequent in the citrate group (9.8 vs. 3.8/1000 catheter days). These differences provided significant catheter-related cost savings of 43% in the taurolidine group vs. citrate group when overall expenses per patient and year were compared. Thus, use of taurolidine-based catheter lock solutions containing heparin and urokinase significantly reduced complications related to tunneled hemodialysis catheters when compared to four percent citrate solution and was overall more cost-efficient.

Calcification Propensity of Serum is Independent of Excretory Renal Function.

Vascular calcification is a component of cardiovascular disease, which is leading cause of death in patients with chronic kidney disease (CKD). A functional assay (T50-test) measuring the propensity of human serum to calcify associates with mortality and cardiovascular events in CKD patients. Calcification propensity is known to increase with CKD stage. We investigated whether the T50 readout is directly dependent on excretory kidney function (eGFR) or rather explained by deranged parameters of bone and mineral metabolism in the course of CKD. T50, along with markers implicated in calcification and mineral metabolism, were measured in a cross-sectional cohort of 118 patients with CKD stage 1-5. Associations of T50 with measured parameters were analysed and partial correlations performed to test to which extent the association of T50 with eGFR can be attributed to variation of these parameters. T50 correlates with eGFR, but serum levels of phosphate and calcium largely explain this association. Phosphate, magnesium, fetuin A, albumin, bicarbonate, and serum cross-laps but not Parathyroid Hormone or Fibroblast Growth Factor 23 are associated with T50 in multivariate adjusted models. These findings indicate that T50 values depend mainly on the concentration of promoters and inhibitors of calcification in serum, but not excretory kidney function.

Cortical porosity not superior to conventional densitometry in identifying hemodialysis patients with fragility fracture.

Hemodialysis (HD) patients face increased fracture risk, which is further associated with elevated risk of hospitalization and mortality. High-resolution peripheral computed tomography (HR-pQCT) has advanced our understanding of bone disease in chronic kidney disease by characterizing distinct changes in both the cortical and trabecular compartments. Increased cortical porosity (Ct.Po) has been shown to be associated with fracture in patients with osteopenia or in postmenopausal diabetic women. We tested whether the degree of Ct.Po identifies hemodialysis patients with prevalent fragility fractures in comparison to bone mineral density (BMD) assessed by dual X-ray absorptiometry (DXA). We performed a post-hoc analysis of a cross-sectional study in 76 prevalent hemodialysis patients. Markers of mineral metabolism, coronary calcification score, DXA-, and HR-pQCT-data were analyzed, and Ct.Po determined at radius and tibia. Ct.Po was significantly higher in patients with fracture but association was lost after adjusting for age and gender (tibia p = 0.228, radius p = 0.5). Instead, femoral (F) BMD neck area (p = 0.03), F T-score neck area (p = 0.03), radius (R) BMD (p = 0.03), R T-score (p = 0.03), and cortical HR-pQCT indices such as cortical area (Ct.Ar) (tibia: p = 0.01; radius: p = 0.02) and cortical thickness (Ct.Th) (tibia: p = 0.03; radius: p = 0.02) correctly classified patients with fragility fractures. Area under receiver operating characteristic curves (AUC) for Ct.Po (tibia AUC: 0.711; p = 0.01; radius AUC: 0.666; p = 0.04), Ct.Ar (tibia AUC: 0.832; p<0.001; radius AUC: 0.796; p<0.001), and F neck BMD (AUC: 0.758; p = 0.002) did not differ significantly among each other. In conclusion, measuring Ct.Po is not superior to BMD determined by DXA for identification of HD patients with fragility fracture.

Sclerostin declines during hemodialysis and appears in Dialysate.

BACKGROUND/AIMS: Sclerostin, a soluble inhibitor of Wnt-signaling, inhibits bone formation. We assessed the impact of dialysis on serum sclerostin and whether sclerostin is detectable in effluent dialysates. METHODS: In a prospective study of 54 prevalent hemodialysis patients, parameters of bone and mineral metabolism were assessed at the beginning and at the end of dialysis. In a subset of patients (n = 19) sclerostin was measured in serum at start, 1, 2, and 3 h of dialysis and in the effluent dialysate at several points in time. RESULTS: Sclerostin serum levels decreased from median 71.4 pmol/l to 43.5 pmol/l during dialysis (p < 0.0001). In patients with high pre-dialysis sclerostin serum levels, sclerostin was detected in the dialysate. Higher Kt/V correlated with greater relative decrease of sclerostin (r = 0.467; p = 0.001). CONCLUSION: Sclerostin is dialysable. Furthermore, sclerostin serum levels decrease during dialysis. Whether targeting lower sclerostin levels by means of improved dialysis adequacy has direct relevance to bone disease remains to be shown.

Correlations and time course of FGF23 and markers of bone metabolism in maintenance hemodialysis patients.

OBJECTIVE: Parathyroid hormone (iPTH) and fibroblast growth factor 23 (FGF23) are elevated in secondary hyperparathyroidism. In hemodialysis, higher dialysate calcium (1.5 mmol/L) induces intradialytic suppression of iPTH, whereas its impact on FGF23 and markers of bone metabolism is unknown. We assessed the time course of FGF23 and markers of bone metabolism in relationship to dialysate calcium. DESIGN AND METHODS: In this prospective cohort study of 19 patients on maintenance hemodialysis, we measured serum calcium (sCa), inorganic phosphate (iP), blood urea nitrogen (BUN), ß2-microglobulin (ßMG), iPTH, FGF23, aminoterminal propeptide type 1 procollagen (P1NP), C-telopeptide of type I collagen for bone degradation (CTX-I), osteocalcin (OC), bone specific alkaline phosphatase (BALP), and tartrate-resistant acid phosphatase (TRAP5b) during a single hemodialysis session at baseline, 1, 2, and 3h of dialysis. The time course of measured parameters was compared according to groups of prescribed dialysate calcium of 1.25 mmol/L and 1.5 mmol/L. RESULTS: iPTH declined in the 1.5 mmol/L dialysis group as serum calcium increased whereas it tended to increase in the 1.25 mmol/L group without significant changes in serum calcium. Patients on long-term dialysate calcium of 1.5 mmol/L had significantly lower CTX-I levels and tended to lower levels of iPTH, FGF23, OC, P1NP and TRAP5b at the start of dialysis compared to those on 1.25 mmol/L. CTX-I, FGF23 and OC but not BALP, P1NP and TRAP5b decreased during dialysis independent of dialysate calcium. CONCLUSIONS: In spite of immediate effects on iPTH, dialysate calcium does not acutely affect other parameters of bone and mineral metabolism. SHORT SUMMARY: Dialysate calcium concentration is known to have both immediate and longer-term impact on parathyroid hormone levels in hemodialysis patients. Little is known about the acute impact of dialysate calcium on bone metabolism. In this cross-sectional study of prevalent hemodialysis patients, we found no evidence of immediate short-term dialysate calcium-induced changes of fibroblast growth factor 23 or anabolic and catabolic markers of bone turnover during hemodialysis. However, differences in CTX-I and to a lesser extent other parameters between groups of higher and lower dialysate calcium suggest a longer-term effect that remains to be validated.

Clinical evaluation of two novel biointact PTH(1-84) assays in hemodialysis patients.

OBJECTIVES: In chronic kidney disease-mineral and bone disorder (CKD-MBD), most treatment decisions are guided by parathyroid hormone (PTH) levels. Here, we aimed at assessing the technical and clinical performance of two novel automated biointact PTH(1-84) assays, from Roche Diagnostics (Ro) and DiaSorin (DS), in hemodialysis patients. DESIGN AND METHODS: We recorded demographics, dialysis treatment characteristics, pharmacotherapy for CKD-MBD and laboratory work-up. Statistical methods included Passing-Bablok, and multiple linear regression. RESULTS: 121 patients, dialyzing on average for 3.5 years (range: 0.1-22.5), with serum phosphate 1.9±0.6 mmol/L (mean±SD), participated in the study. Median serum concentration for intact PTH was 223 ng/L (range: 5-2844), and for biointact PTH(1-84) was 136 ng/L (Ro; range: 1-1644), respectively 138 ng/L (DS; range: 4-1580). Both biointact assays were significantly correlated (r=0.98; Ro=0.87×DS+19.60). Bland-Altmann plots revealed an average bias ±2 SD of 10±27 ng/L below 200 ng/L, and -32±157 ng/L above 200 ng/L (Ro minus DS). The variably adjusted association between PTH and serum phosphate was very similar, regardless of the PTH assay, but this was not the case for PTH-derived measures (ratios biointact/intact; differences intact minus biointact). (Log)PTH concentrations as well as serum phosphate were significantly associated with serum creatinine, but only in patients with >0 mL urine per day. CONCLUSIONS: Results from Roche and DiaSorin biointact PTH(1-84) assays were well correlated, but showed increased deviations at higher concentrations. Biointact PTH(1-84) levels are roughly two third of intact PTH. The association between PTH and serum creatinine may depend on residual renal clearance of PTH and/or serum phosphate.

Inorganic phosphate and FGF-23 predict outcome in stable systolic heart failure.

BACKGROUND: Recent studies show associations between inorganic phosphate and risk of heart failure in the general population as well as between fibroblast growth factor 23 (FGF-23) and outcome in coronary heart disease. This study was carried out to assess whether circulating levels of inorganic phosphate and FGF-23, a new central hormone in mineral bone metabolism, predict outcome in systolic heart failure. MATERIALS AND METHODS: Ninety-nine consecutive outpatients with systolic heart failure were enrolled. Mean (SD) age was 61 years (11), mean left ventricular ejection fraction (LVEF) was 33% (10), 82 patients were men, median estimated creatinine clearance was 83 mL/min (Q(1) -Q(3) 58-106), median NTproBNP level was 803 pg/mL (Q(1) -Q(3) 404-2757), median inorganic phosphate was 1·12 mM (Q(1) -Q(3) 1·02-1·22), median FGF-23 was 39·02 pg/mL (Q(1) -Q(3) 32·45-55·86) and median follow-up was 35 months. Associations between inorganic phosphate, FGF-23 and endpoints were assessed using Cox regression analyses. RESULTS: Inorganic phosphate and FGF-23 levels were significantly higher (P < 0·001 and P = 0·009) in patients reaching the combined endpoint of cardiac hospitalization or death. FGF-23 (ln) predicted all-cause mortality (hazard ratio (HR) 5·042, P = 0·032) in a model adjusted for age, gender, estimated creatinine clearance, LVEF, New York Heart Association (NYHA) stage and NTproBNP level. Inorganic phosphate predicted heart failure hospitalization (HR 26·944, P = 0·021), cardiac hospitalization (HR 16·016, P = 0·017) and the combined endpoint (HR 13·294, P = 0·015) in models adjusted for the same co-variables. CONCLUSION: The results of this study demonstrate the independent prognostic value of inorganic phosphate and FGF-23 in heart failure even in the context of established risk markers.

Epithelial Notch signaling regulates interstitial fibrosis development in the kidneys of mice and humans.

Chronic kidney disease is a leading cause of death in the United States. Tubulointerstitial fibrosis (TIF) is considered the final common pathway leading to end-stage renal disease (ESRD). Here, we used pharmacologic, genetic, in vivo, and in vitro experiments to show that activation of the Notch pathway in tubular epithelial cells (TECs) in patients and in mouse models of TIF plays a role in TIF development. Expression of Notch in renal TECs was found to be both necessary and sufficient for TIF development. Genetic deletion of the Notch pathway in TECs reduced renal fibrosis. Consistent with this, TEC-specific expression of active Notch1 caused rapid development of TIF. Pharmacologic inhibition of Notch activation using a γ-secretase inhibitor ameliorated TIF. In summary, our experiments establish that epithelial injury and Notch signaling play key roles in fibrosis development and indicate that Notch blockade may be a therapeutic strategy to reduce fibrosis and ESRD development.

The Notch pathway in podocytes plays a role in the development of glomerular disease.

Albuminuria associated with sclerosis of the glomerulus leads to a progressive decline in renal function affecting millions of people. Here we report that activation of the Notch pathway, which is critical in glomerular patterning, contributes to the development of glomerular disease. Expression of the intracellular domain of Notch1 (ICN1) was increased in glomerular epithelial cells in diabetic nephropathy and in focal segmental glomerulosclerosis. Conditional re-expression of ICN1 in vivo exclusively in podocytes caused proteinuria and glomerulosclerosis. In vitro and in vivo studies showed that ICN1 induced apoptosis of podocytes through the activation of p53. Genetic deletion of a Notch transcriptional partner (Rbpj) specifically in podocytes or pharmacological inhibition of the Notch pathway (with a gamma-secretase inhibitor) protected rats with proteinuric kidney diseases. Collectively, our observations suggest that Notch activation in mature podocytes is a new mechanism in the pathogenesis of glomerular disease and thus could represent a new therapeutic target.

Unchanged expression of the sodium-dependent phosphate cotransporter NaPi-IIa despite diurnal changes in renal phosphate excretion.

Renal phosphate excretion is subjected to circadian rhythmicity. The bulk of filtered inorganic phosphate (P(i)) is reabsorbed by the sodium-dependent phosphate cotransporter NaPi-IIa. The regulation of proximal tubular phosphate reabsorptive capacity is largely attributed to the altered abundance of NaPi-IIa residing in the brush border membrane (BBM) of proximal tubular cells. Therefore, we examined if the diurnal rise in renal phosphate excretion is accompanied by a corresponding change in NaPi-IIa expression. Renal phosphate excretion, creatinine clearance, and serum phosphate were determined at consecutive time points in rats, starting from 8 a.m. until 5 p.m. During this period, renal phosphate excretion (fractional P(i) excretion) increased more than eightfold until 5 p.m. compared to the morning values at 8 a.m. In addition, serum phosphate and creatinine clearance as well as the calculated tubular phosphate threshold increased. Neither immunoblot analysis of BBMs nor immunohistochemical staining for NaPi-IIa yielded evidence for a lower abundance of NaPi-IIa in kidneys collected in the afternoon compared to those in the morning. However, kidneys sampled in the afternoon showed a small decrease (14%) in (32)P uptakes into BBM vesicles (BBMVs). Thus, the diurnal rise in renal phosphate excretion was associated with a mild reduction in the sodium-dependent phosphate transport rate in proximal tubular BBMs. There was no apparent downregulation of NaPi-IIa abundance and only a small reduction in Na(+)-dependent Pi-transport activity. Thus, the diurnal changes in urinary phosphate excretion appear to be mainly related to changes in serum phosphate and tubular threshold but not to NaPi-IIa expression.

Secreted frizzled-related protein-4 reduces sodium-phosphate co-transporter abundance and activity in proximal tubule cells.

The phosphatonin, secreted frizzled-related protein-4 (sFRP-4), induces phosphaturia and inhibits 25-hydroxyvitamin D 1alpha-hydroxylase activity normally induced in response to hypophosphatemia. To determine the mechanism by which sFRP-4 alters renal phosphate (P(i)) transport, we examined the effect of sFRP-4 on renal brush border membrane (BBMV) Na(+)-dependent P(i) uptake, and the abundance and localization of the major Na(+)-P(i)-IIa co-transporter in proximal tubules and opossum kidney (OK) cells. Infusion of sFRP-4 increased renal fractional excretion of P(i) and decreased renal beta-catenin concentrations. The increase in renal P(i) excretion with sFRP-4 infusion was associated with a 21.9 +/- 3.4% decrease in BBMV Na(+)-dependent P(i) uptake (P < 0.001) compared with a 39.5 +/- 2.1% inhibition of Na(+)-dependent P(i) transport in renal BBMV induced by PTH (P < 0.001). sFRP-4 infusion was associated with a 30.7 +/- 4.8% decrease in Na(+)-P(i)-IIa co-transporter protein abundance (P < 0.01) assessed by immunoblotting methods compared to a 45.4 +/- 8.8% decrease induced by PTH (P < 0.001). In OK cells, sFRP-4 reduced surface expression of a heterologous Na(+)-P(i)-IIa co-transporter. We conclude that sFRP-4 increases renal P(i) excretion by reducing Na(+)-P(i)-IIa transporter abundance in the brush border of the proximal tubule through enhanced internalization of the protein.

Renal phosphate loss in hereditary and acquired disorders of bone mineralization.

Three metabolic bone diseases display similar characteristics such as hypophosphatemia due to chronically elevated renal phosphate clearance, inappropriately low 1,25 (OH)2 vitamin D serum levels, and variable bone disease (rickets and osteomalacia). X-linked dominant hypophosphatemic rickets (XLH), also called vitamin D-resistant rickets and autosomal dominant hypophosphatemic rickets (ADHR) represent two inherited diseases, whereas oncogenic hypophosphatemia (OHO), also known as tumor induced osteomalacia (TIO), is an acquired paraneoplastic syndrome that, in certain aspects, has much in common with XLH and ADHR. Although the primary causes for these disorders are distinct and well established, their similar features suggest a unifying pathophysiological basis. This review summarizes what is known about the mechanisms that underlie these diseases and includes most up-to-date information about recently introduced factors that might be involved in the regulation of phosphate homeostasis and skeletal mineralization.