Chromatin accessibility and epigenetic deoxyribose nucleic acid (DNA) modifications in chronic kidney disease (CKD) osteoblasts: a study of bone and osteoblasts from pediatric patients with CKD.

Maturation defects are intrinsic features of osteoblast lineage cells in CKD patients. These defects persist ex vivo, suggesting that CKD induces epigenetic changes in bone cells. To gain insights into which signaling pathways contribute to CKD-mediated, epigenetically driven, impairments in osteoblast maturation, we characterized RNA expression and DNA methylation patterns by RNA-Seq and MethylationEpic in primary osteoblasts from nine adolescent and young adult dialysis patients with end-stage kidney disease and three healthy references. ATAC-Seq was also performed on a subset of osteoblasts. Bone matrix protein expression was extracted from the iliac crest and evaluated by proteomics. Gene set enrichment analysis was used to establish signaling pathways consistently altered in chromatin accessibility, DNA methylation, and RNA expression patterns. Single genes were suppressed in primary osteoblasts using shRNA and mineralization characterized in vitro. The effect of nuclear factor of activated T cells (NFAT) signaling suppression was also assessed using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) incorporation. We found that signaling pathways critical for osteoblast differentiation were strongly downregulated in CKD osteoblasts. Gene set enrichment analysis identified highly significant methylation changes, differential chromatin accessibility, and altered RNA expression in NFAT signaling targets. NFAT inhibition reduced osteoblast proliferation. Combined analysis of osteoblast RNA expression and whole bone matrix composition identified 13 potential ligand-receptor pairs. In summary, epigenetic changes in CKD osteoblasts associate with altered expression of multiple osteoblast genes and signaling pathways. An increase in NFAT signaling may play a role in impaired CKD osteoblast maturation. Epigenetic changes also associate with an altered bone matrix, which may contribute to bone fragility. Further studies are necessary to elucidate the pathways affected by these genetic alterations since elucidating these pathways will be vital to correcting the underlying biology of bone disease in the CKD population.

Utility of Blood Biomarkers to Predict Marrow Iron Stores in Children.

BACKGROUND: Iron deficiency is common in children with kidney failure, but current guidelines are based on biomarkers of iron stores that may be influenced by inflammation. This is the first study that examined which serum iron indices were associated with stainable marrow iron stores (the gold standard) in this population with kidney failure who underwent bone biopsies. METHODS: This cross-sectional study enrolled 71 clinically stable children and young adults receiving dialysis who underwent bone biopsy for chronic kidney disease-mineral bone disorder between 2007 through 2011. Bone biopsies were stained with Perls' Prussian blue and independently interpreted by a pathologist blinded to participants' iron parameters and clinical status. Marrow staining was scored absent vs. present to facilitate receiver operator curve (ROC) analysis. In ROC analysis, the ability of serum ferritin to detect stainable marrow iron stores was compared with that of transferrin saturation (TSAT), serum hepcidin, and clinical guideline-based iron deficiency cut-offs for serum iron, TSAT, and their combinations. RESULTS: Mean age was 17.2 ± 4.4 years (range 2-28), and 30% of patients were female. Median dialysis vintage was 1.2 (IQR 0.7, 2.0) years, and 56% were supported by peritoneal dialysis. Mean hemoglobin was 12.4 ± 1.7 g/dl, and 35% were receiving iron supplementation at the time of biopsy. Based on the gold standard of depleted marrow iron stores, 46.5% of patients were iron-deficient. As an indicator of marrow iron staining, serum ferritin provided a higher area under the ROC curve than serum hepcidin, TSAT, or clinical guidelines-based evaluation of TSAT + ferritin. CONCLUSIONS: In this cohort of children and young adults with kidney failure, serum ferritin provided the best indication of stainable marrow iron stores, followed by transferrin saturation.

Sclerostin, Osteocytes, and Wnt Signaling in Pediatric Renal Osteodystrophy.

The pathophysiology of chronic kidney disease-mineral and bone disorder (CKD-MBD) is not well understood. Specific factors secreted by osteocytes are elevated in the serum of adults and pediatric patients with CKD-MBD, including FGF-23 and sclerostin, a known inhibitor of the Wnt signaling pathway. The molecular mechanisms that promote bone disease during the progression of CKD are incompletely understood. In this study, we performed a cross-sectional analysis of 87 pediatric patients with pre-dialysis CKD and post-dialysis (CKD 5D). We assessed the associations between serum and bone sclerostin levels and biomarkers of bone turnover and bone histomorphometry. We report that serum sclerostin levels were elevated in both early and late CKD. Higher circulating and bone sclerostin levels were associated with histomorphometric parameters of bone turnover and mineralization. Immunofluorescence analyses of bone biopsies evaluated osteocyte staining of antibodies towards the canonical Wnt target, ß-catenin, in the phosphorylated (inhibited) or unphosphorylated (active) forms. Bone sclerostin was found to be colocalized with phosphorylated ß-catenin, which suggests that Wnt signaling was inhibited. In patients with low serum sclerostin levels, increased unphosphorylated "active" ß-catenin staining was observed in osteocytes. These data provide new mechanistic insight into the pathogenesis of CKD-MBD and suggest that sclerostin may offer a potential biomarker or therapeutic target in pediatric renal osteodystrophy.

All the might of the osteocyte: emerging roles in chronic kidney disease.

Osteocytes are the most abundant type of bone cell and play crucial roles in bone health. Osteocytes sense mechanical stress and orchestrate osteoblasts and osteoclasts to maintain bone density and strength. Beyond this, osteocytes have also emerged as key regulators of organ crosstalk, and they function as endocrine organs via their roles in secreting factors that mediate signaling within their neighboring bone cells and in distant tissues. As such, osteocyte dysfunction has been associated with the bone abnormalities seen across a spectrum of chronic kidney disease. Specifically, dysregulated osteocyte morphology and signaling have been observed in the earliest stages of chronic kidney disease and have been suggested to contribute to kidney disease progression. More important, US Food and Drug Administration-approved inhibitors of osteocytic secreted proteins, such as fibroblast growth factor 23 and sclerostin, have been used to treat bone diseases. The present mini review highlights new research that links dysfunctional osteocytes to the pathogenesis of chronic kidney disease mineral and bone disorder.

Effects of Primary Kidney Disease Etiology on Renal Osteodystrophy in Pediatric Dialysis Patients.

Congenital diseases of the kidney and urinary tract (CAKUT) and glomerulonephritis are the main causes of chronic kidney disease (CKD) in children. Although renal osteodystrophy (ROD) and indices of mineral metabolism have been characterized in dialyzed children, the impact of primary kidney disease on ROD is unknown. We performed a cross-sectional study of bone biopsies performed in 189 pediatric dialysis patients aged 12.6 ± 5.4 years. Patients were classified into three groups according to primary kidney disease: CAKUT (n = 82), hereditary (n = 22), or glomerular disease (n = 85). Serum concentrations of calcium, phosphate, alkaline phosphatase (ALP), parathyroid hormone (PTH), and 25(OH) vitamin D were measured at the time of biopsy. Fibroblast growth factor 23 (FGF23) levels were measured in a subset of 59 patients. Levels of calcium, phosphate, PTH, and 25(OH) vitamin D were similar across groups. CAKUT patients had higher serum ALP and lower C-terminal FGF23 levels. Bone turnover and bone volume parameters did not differ across groups. However, osteoid volume (OV/BV), osteoid surface (OS/BS), and osteoid maturation time (OMT) were highest in the CAKUT group and lowest in the hereditary group. Multiple regression analysis revealed that calcium, phosphate, ALP, and PTH were independently associated with OV/BV and osteoid thickness (O.Th). PTH was an independent factor affecting bone formation rate. The relationship between CKD etiology and bone histomorphometric variables was abrogated after adjustment for biochemical parameters in the multivariable models. Overall, bone histology differed according to CKD etiology in the unadjusted analysis; however, this association could not be confirmed independently of biochemical parameters. Although CAKUT patients had a greater mineralization defect with elevated serum ALP levels, longitudinal studies will be needed to elucidate mediation pathways that might be involved in the complex interplay of CKD-mineral bone disease (MBD). © 2022 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Erythroid overproduction of erythroferrone causes iron overload and developmental abnormalities in mice.

The hormone erythroferrone (ERFE) is produced by erythroid cells in response to hemorrhage, hypoxia, or other erythropoietic stimuli, and it suppresses the hepatic production of the iron-regulatory hormone hepcidin, thereby mobilizing iron for erythropoiesis. Suppression of hepcidin by ERFE is believed to be mediated by interference with paracrine bone morphogenetic protein (BMP) signaling that regulates hepcidin transcription in hepatocytes. In anemias with ineffective erythropoiesis, ERFE is pathologically overproduced, but its contribution to the clinical manifestations of these anemias is not well understood. We generated 3 lines of transgenic mice with graded erythroid overexpression of ERFE and found that they developed dose-dependent iron overload, impaired hepatic BMP signaling, and relative hepcidin deficiency. These findings add to the evidence that ERFE is a mediator of iron overload in conditions in which ERFE is overproduced, including anemias with ineffective erythropoiesis. At the highest levels of ERFE overexpression, the mice manifested decreased perinatal survival, impaired growth, small hypofunctional kidneys, decreased gonadal fat depots, and neurobehavioral abnormalities, all consistent with impaired organ-specific BMP signaling during development. Neutralizing excessive ERFE in congenital anemias with ineffective erythropoiesis may not only prevent iron overload but may have additional benefits for growth and development.

Bone marrow adiposity inversely correlates with bone turnover in pediatric renal osteodystrophy.

Bone marrow adiposity is associated with bone disease in the general population. Although chronic kidney disease (CKD) is associated with increased bone fragility, the correlation between marrow adiposity and bone health in CKD is unknown. We evaluated the relationship between bone marrow adipocytes and bone histomorphometry in 32 pediatric patients. We also evaluated the effects of growth hormone and calcitriol (1,25(OH)2D3)-two therapies commonly prescribed for pediatric bone disease-on marrow adiposity and bone histomorphometry. Finally, the adipogenic potential of primary human osteoblasts from CKD patients was assessed in vitro, both alone and in the presence of 1,25(OH)2D3. In cross-sectional analysis, marrow adipocyte number per tissue area (Adi.N/T.Ar) correlated with bone formation rate/bone surface (BFR/BS) in patients with high bone turnover (r = -0.55, p = 0.01) but not in those with low/normal bone turnover. Changes in bone formation rate correlated with changes Adi.N/T.Ar on repeat bone biopsy(r = -0.48, p = 0.02). In vitro, CKD and control osteoblasts had a similar propensity to transition into an adipocyte-like phenotype; 1,25(OH)2D3 had very little effect on this propensity. In conclusion, marrow adiposity correlates inversely with bone turnover in pediatric patients with high turnover renal osteodystrophy. The range of adiposity observed in pediatric patients with low/normal bone turnover is not explained by intrinsic changes to precursor cells or by therapies but may reflect the effects of circulating factors on bone cell health in this population.

An ARHGAP25 variant links aberrant Rac1 function to early-onset skeletal fragility.

Ras homologous guanosine triphosphatases (RhoGTPases) control several cellular functions, including cytoskeletal actin remodeling and cell migration. Their activities are downregulated by GTPase-activating proteins (GAPs). Although RhoGTPases are implicated in bone remodeling and osteoclast and osteoblast function, their significance in human bone health and disease remains elusive. Here, we report defective RhoGTPase regulation as a cause of severe, early-onset, autosomal-dominant skeletal fragility in a three-generation Finnish family. Affected individuals (n = 13) presented with multiple low-energy peripheral and vertebral fractures despite normal bone mineral density (BMD). Bone histomorphometry suggested reduced bone volume, low surface area covered by osteoblasts and osteoclasts, and low bone turnover. Exome sequencing identified a novel heterozygous missense variant c.652G>A (p.G218R) in ARHGAP25, encoding a GAP for Rho-family GTPase Rac1. Variants in the ARHGAP25 5' untranslated region (UTR) also associated with BMD and fracture risk in the general population, across multiple genomewide association study (GWAS) meta-analyses (lead variant rs10048745). ARHGAP25 messenger RNA (mRNA) was expressed in macrophage colony-stimulating factor (M-CSF)-stimulated human monocytes and mouse osteoblasts, indicating a possible role for ARHGAP25 in osteoclast and osteoblast differentiation and activity. Studies on subject-derived osteoclasts from peripheral blood mononuclear cells did not reveal robust defects in mature osteoclast formation or resorptive activity. However, analysis of osteosarcoma cells overexpressing the ARHGAP25 G218R-mutant, combined with structural modeling, confirmed that the mutant protein had decreased GAP-activity against Rac1, resulting in elevated Rac1 activity, increased cell spreading, and membrane ruffling. Our findings indicate that mutated ARHGAP25 causes aberrant Rac1 function and consequently abnormal bone metabolism, highlighting the importance of RhoGAP signaling in bone metabolism in familial forms of skeletal fragility and in the general population, and expanding our understanding of the molecular pathways underlying skeletal fragility. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Characterization of Primary Cilia in Osteoblasts Isolated From Patients With ADPKD and CKD.

Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited cause of chronic kidney disease (CKD) and leads to a specific type of bone disease. The primary cilium is a major cellular organelle implicated in the pathophysiology of ADPKD caused by mutations in polycystin-1 (PKD1) and polycystin-2 (PKD2). In this study, for the first time, cilia were characterized in primary preosteoblasts isolated from patients with ADPKD. All patients with ADPKD had low bone turnover and primary osteoblasts were also obtained from patients with non-ADPKD CKD with low bone turnover. Image-based immunofluorescence assays analyzed cilia using standard markers, pericentrin, and acetylated-α-tubulin, where cilia induction and elongation were chosen as relevant endpoints for these initial investigations. Osteoblastic activity was examined by measuring alkaline phosphatase levels and mineralized matrix deposition rates. It was found that primary cilia can be visualized in patient-derived osteoblasts and respond to elongation treatments. Compared with control cells, ADPKD osteoblasts displayed abnormal cilia elongation that was significantly more responsive in cells with PKD2 nontruncating mutations and PKD1 mutations. In contrast, non-ADPKD CKD osteoblasts were unresponsive and had shorter cilia. Finally, ADPKD osteoblasts showed increased rates of mineralized matrix deposition compared with non-ADPKD CKD. This work represents the first study of cilia in primary human-derived osteoblasts from patients with CKD and patients with ADPKD who have normal kidney function, offering new insights as bone disease phenotypes are not well recapitulated in animal models. These data support a model whereby altered cilia occurs in PKD-mutated osteoblasts, and that ADPKD-related defects in bone cell activity and mineralization are distinct from adynamic bone disease from patients with non-ADPKD CKD. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC. on behalf of American Society for Bone and Mineral Research.

Bone material properties and response to teriparatide in osteoporosis due to WNT1 and PLS3 mutations.

CONTEXT: Patients with osteoporosis-associated WNT1 or PLS3 mutations have unique bone histomorphometric features and osteocyte-specific hormone expression patterns. OBJECTIVE: To investigate the effects of WNT1 and PLS3 mutations on bone material properties. DESIGN: Transiliac bone biopsies were evaluated by quantitative backscattered electron imaging, immunohistochemistry, and bone histomorphometry. SETTING: Ambulatory patients. PATIENTS: Three pediatric and eight adult patients with WNT1 or PLS3 mutations. INTERVENTION: Bone mineralization density distribution and osteocyte protein expression was evaluated in 11 patients and repeated in six patients who underwent repeat biopsy after 24 months of teriparatide treatment. MAIN OUTCOME MEASURE: Bone mineralization density distribution and protein expression. RESULTS: Children with WNT1 or PLS3 mutations had heterogeneous bone matrix mineralization, consistent with bone modeling during growth. Bone matrix mineralization was homogenous in adults and increased throughout the age spectrum. Teriparatide had very little effect on matrix mineralization or bone formation in patients with WNT1 or PLS3 mutations. However, teriparatide decreased trabecular osteocyte lacunae size and increased trabecular bone FGF23 expression. CONCLUSION: The contrast between preserved bone formation with heterogeneous mineralization in children and low bone turnover with homogenous bone mineral content in adults suggests that WNT1 and PLS3 have differential effects on bone modeling and remodeling. The lack of change in matrix mineralization in response to teriparatide, despite clear changes in osteocyte lacunae size and protein expression, suggests that altered WNT1 and PLS3 expression may interfere with coupling of osteocyte, osteoblast, and osteoclast function. Further studies are warranted to determine the mechanism of these changes.

Vitamin C overload may contribute to systemic oxalosis in children receiving dialysis.

BACKGROUND: Malnutrition and anorexia are common in children with chronic kidney disease (CKD) and gastrostomy tubes (GT) as well as nasogastric tubes (NGT) have been recommended to maximize nutritional support. The optimal requirement of vitamin C in children with CKD remains to be defined but oxalate is a breakdown product of vitamin C. Elevated vitamin C intake and bone oxalate were identified in two formula-fed dialyzed children with negative genetic testing for primary hyperoxaluria. METHODS: We evaluated the impact of nutritional support on serum ascorbic acid and plasma oxalate levels in 13 dialyzed infants and young children. RESULTS: All patients were fed by GT or NGT since the first months of life; overall patients were receiving between 145 and 847% of the age-specific DRI for vitamin C. Mean serum ascorbic acid and plasma oxalate levels were elevated (244.7 ± 139.7 µM/L and 44.3 ± 23.1 µM/L, respectively), and values did not differ according to the degree of residual kidney function. Ascorbic acid levels did not correlate with oxalate levels (r = 0.44, p = 0.13). CONCLUSIONS: Excessive vitamin C intake may contribute to oxalate accumulation in dialyzed children.

Regional variation in bone turnover at the iliac crest versus the greater trochanter.

BACKGROUND: Iliac crest bone biopsy with histomorphometry is the gold standard for diagnosis of abnormalities in bone turnover, yet fractures more frequently occur at the greater trochanter of the hip. Whether bone turnover is similar at these two anatomic sites within individuals is uncertain. METHODS: We collected bone biopsy samples from the ipsilateral iliac crest and greater trochanter in 9 deceased individuals undergoing autopsies at an academic medical center between March-August 2018. We measured 14 static bone histomorphometry parameters including osteoclast number (N.Oc/T.A), eroded surface (ES/BS), trabecular separation (Tb.Sp), osteoclast surface (Oc.S/BS) and osteoid volume (OV/BV) as markers of bone turnover, mineralization, and volume (TMV), and evaluated the correlation of these markers between the iliac crest and greater trochanter. RESULTS: Average age at time of death was 58 ± 15 years, 2 were women, and average time from death to autopsy was 2.9 ± 1.8 days. Overall, correlations of the markers of bone turnover across the two sites were poor, ranging from as low as 0 for Tb.Sp (p = 1.0) to as high as 0.583 for Oc.S/BS (p = 0.102). CONCLUSIONS: Static histomorphometric measures of bone turnover at the iliac crest may not provide reliable information about turnover at other anatomic sites.

Inhibition of miR-29-3p isoforms via tough decoy suppresses osteoblast function in homeostasis but promotes intermittent parathyroid hormone-induced bone anabolism.

miRNAs play a vital role in post-transcriptional regulation of gene expression in osteoblasts and osteoclasts, and the miR-29 family is expressed in both lineages. Using mice globally expressing a miR-29-3p tough decoy, we demonstrated a modest 30-60% decrease all three miR-29-3p isoforms: miR-29a, miR-29b, and miR-29c. While the miR-29-3p decoy did not impact osteoclast number or function, the tough decoy decreased bone formation in growing mice, which led to decreased trabecular bone volume in mature animals. These data support previous in vitro studies suggesting that miR-29-3p is a positive regulator of osteoblast differentiation. In contrast, when mice were treated with intermittent parathyroid hormone (PTH1-34), inhibition of miR-29-3p augmented the effect of PTH on cortical bone anabolism, increased bone formation rate and osteoblast surface, and increased levels of Ctnnb1/ßcatenin mRNA, which is a miR-29 target. These findings highlight differences in the mechanisms controlling basal level bone formation and bone formation induced by intermittent PTH. Overall, the global miR-29-3p tough decoy model represents a modest loss-of-function, which could be a relevant tool for assessing the possible impact of systemically administered miR-29-3p inhibitors. Our studies provide a potential rationale for co-administration of PTH1-34 and miR-29-3p inhibitors, to boost bone formation in severely affected osteoporosis patients, particularly in the cortical compartment.

Unraveling the osteocyte in CKD-MBD post-renal transplantation.

Changes in indices of mineral metabolism, bone protein expression, and bone turnover were assessed between pre- and post-renal transplant bone biopsies obtained 12 months apart. Circulating sclerostin and fibroblast growth factor 23 (FGF-23) levels decreased, and a low bone turnover state was highly prevalent on follow-up. In contrast, bone sclerostin expression increased, whereas FGF-23 bone expression was unchanged/decreased. These findings underscore the limitations of circulating biomarkers and the critical role of bone biopsy to understand osteocyte biology in chronic kidney disease-mineral bone disorder.

Vitamin D sterols increase FGF23 expression by stimulating osteoblast and osteocyte maturation in CKD bone.

Impaired osteoblast and osteocyte maturation contribute to mineralization defects and excess FGF23 expression in CKD bone. Vitamin D sterols decrease osteoid accumulation and increase FGF23 expression; these agents also increase osteoblast maturation in vitro but a link between changes in bone cell maturation, bone mineralization, and FGF23 expression in response to vitamin D sterols has not been established. We evaluated unmineralized osteoid accumulation, osteocyte maturity markers (FGF23: early osteocytes; sclerostin: late osteocytes), and osteocyte apoptosis in iliac crest of 11 pediatric dialysis patients before and after 8 months of doxercalciferol therapy. We then evaluated the effect of 1,25(OH)2vitamin D on in vitro maturation and mineralization of primary osteoblasts from dialysis patients. Unmineralized osteoid accumulation decreased while numbers of early (FGF23-expressing) increased in response to doxercalciferol. Osteocyte apoptosis was low but increased with doxercalciferol. Bone FGF23 expression correlated with numbers of early, FGF23-expressing, osteocytes (r = 0.83, p < 0.001). In vitro, 1,25(OH)2vitamin D increased expression of the mature osteoblast marker osteocalcin (BGLAP) but only very high (100 nM) concentrations affected in vitro osteoblast mineralization. High doses (10 and 100 nM) of 1,25(OH)2vitamin D also increased the ratio of RANKL/OPG expression in CKD osteoblasts. Vitamin D sterols directly stimulate osteoblast maturation. They also increase osteocyte turnover and increase osteoblast expression of osteoclast differentiation factors, thus likely modulating osteoblast/osteoclast/osteocyte coupling. By increasing numbers of early osteocytes, vitamin D sterols increase FGF23 expression in CKD bone.

Racial differences in bone histomorphometry in children and young adults treated with dialysis.

BACKGROUND: Healthy African-Americans are known to have greater bone mineral density and decreased risk of fracture when compared to Caucasians. In fact, comparisons of bone histomorphometry in healthy South African children and adults reveal greater cortical thickness in Black subjects as compared to White. How these differences are reflected in the bone of American children and young adults on dialysis is unknown. METHODS: Using tetracycline-labeled, iliac crest bone biopsies obtained in prior research protocols in pediatric and young adult dialysis patients, we compared trabecular and cortical parameters between non-Hispanic African-American subjects and non-Hispanic Caucasian subjects matched by age and gender. A linear regression model controlled for trabecular turnover and mineralization was used to further investigate the association of race with cortical thickness. RESULTS: The matched cohort consisted of 52 subjects-26 African-American and 26 Caucasian. Turnover, mineralization and volume parameters in trabecular bone did not show significant differences between racial groups. Characterizing subjects by renal osteodystrophy type did not show a statistically significant difference although Caucasian patients had double the prevalence of mineralization defects. Consistent with this was a trend toward better mineralization parameters in African-Americans including shorter osteoid maturation time and lower osteoid volume. A sub-cohort of patients with cortical measures demonstrated greater median (IQR) cortical thickness in African-Americans (541 µm [354, 694]) than in Caucasians (371 µm [336, 446], p = 0.08). In a linear regression model controlling for trabecular turnover and mineralization, African-American subjects had 36.2% (95% CI 0.28 to 85.1%, p = 0.048) greater cortical thickness as compared to White subjects. There was no significant difference in cortical porosity. CONCLUSIONS: Although likely limited by sample size, our findings suggest that, similar to findings in populations of normal children, African-American race in pediatric and young adults on dialysis is associated with greater cortical thickness. Additionally, there was a trend toward greater mineralization defects in Caucasian children. Both findings require further exploration with larger patient samples in order to thoroughly explore these racial differences and the implications on CKD-MBD treatment.

Impaired osteocyte maturation in the pathogenesis of renal osteodystrophy.

Pediatric renal osteodystrophy is characterized by skeletal mineralization defects, but the role of osteoblast and osteocyte maturation in the pathogenesis of these defects is unknown. We evaluated markers of osteocyte maturation and programmed cell death in iliac crest biopsy samples from pediatric dialysis patients and healthy controls. We evaluated the relationship between numbers of fibroblast growth factor 23 (FGF23)-expressing osteocytes and histomorphometric parameters of skeletal mineralization. We confirmed that chronic kidney disease (CKD) causes intrinsic changes in bone cell maturation using an in vitro model of primary osteoblasts from patients with CKD and healthy controls. FGF23 co-localized with the early osteocyte marker E11/gp38, suggesting that FGF23 is a marker of early osteocyte maturation. Increased numbers of early osteocytes and decreased osteocyte apoptosis characterized CKD bone. Numbers of FGF23-expressing osteocytes were highest in patients with preserved skeletal mineralization indices, and packets of matrix surrounding FGF23-expressing osteocytes appeared to have entered secondary mineralization. Primary osteoblasts from patients with CKD retained impaired maturation and mineralization characteristics in vitro. Addition of FGF23 did not affect primary osteoblast mineralization. Thus, CKD is associated with intrinsic changes in osteoblast and osteocyte maturation, and FGF23 appears to mark a relatively early stage in osteocyte maturation. Improved control of renal osteodystrophy and FGF23 excess will require further investigation into the pathogenesis of CKD-mediated osteoblast and osteocyte maturation failure.

Skeletal Consequences of Nephropathic Cystinosis.

Nephropathic cystinosis is a rare lysosomal storage disorder. Patients present in the first year of life with renal Fanconi syndrome that evolves to progressive chronic kidney disease (CKD). Despite the multiple risk factors for bone disease, the frequency and severity of skeletal disorders in nephropathic cystinosis have not been described. We performed systematic bone and mineral evaluations of subjects with cystinosis seen at the NIH (n = 30), including history and physical examination, serum and urine biochemistries, DXA, vertebral fracture assessment, skeletal radiographs, and renal ultrasound. Additionally, histomorphometric analyses are reported on six subjects seen at the UCLA Bone and Mineral Metabolism Clinic. In NIH subjects, mean age was 20 years (range, 5 to 44 years), 60% were CKD stages G1 to G4, and 40% had a renal transplant. Mean bone mineral density (BMD) Z-scores were decreased in the femoral neck, total hip, and 1/3 radius (p < 0.05). Low bone mass at one or more sites was present in 46% of subjects. Twenty-seven percent of subjects reported one or more long bone fractures. Thirty-two percent of subjects had incidental vertebral fractures, which were unrelated to transplant status. Long-bone deformity/bowing was present in 64%; 50% had scoliosis. Diffuse osteosclerosis was present in 21% of evaluated subjects. Risk factors included CKD, phosphate wasting, hypercalciuria, secondary hyperparathyroidism, hypovitaminosis D, male hypogonadism, metabolic acidosis, and glucocorticoid/immunosuppressive therapy. Sixty-one percent of the non-transplanted subjects had ultrasonographic evidence of nephrocalcinosis or nephrolithiasis. Histomorphometric analyses showed impaired mineralization in four of six studied subjects. We conclude that skeletal deformities, decreased bone mass, and vertebral fractures are common and relevant complications of nephropathic cystinosis, even before renal transplantation. Efforts to minimize risk factors for skeletal disease include optimizing mineral metabolism and hormonal status, combined with monitoring for nephrocalcinosis/nephrolithiasis. © 2018 This article is a U.S. Government work and is in the public domain in the USA.