Urinary CD80 and Serum suPAR as Biomarkers of Glomerular Disease among Adults in Brazil.

INTRODUCTION: Urinary CD80 has been shown to have good specificity for minimal change disease (MCD) in children. However, the investigation of circulating factors such as soluble urokinase plasminogen activator receptor (suPAR) as biomarkers of focal segmental glomerulosclerosis (FSGS) is quite controversial. The objective of this study was to determine whether urinary CD80 and serum suPAR can be used for the diagnosis of MCD and FSGS, respectively, in the adult population of Brazil. We also attempted to determine whether those biomarkers assess the response to immunosuppressive treatment. METHODS: This was a prospective study in which urine and blood samples were collected for analysis of CD80 and suPAR, respectively, only in the moment of renal biopsy, from patients undergoing to diagnostic renal biopsy. At and six months after biopsy, we analyzed serum creatinine, serum albumin, and proteinuria in order to evaluate the use of the CD80 and suPAR collected in diagnosis as markers of response to immunosuppressive treatment. In healthy controls were collected urinary CD80 and proteinuria, serum suPAR, and creatinine. RESULTS: The results of 70 renal biopsies were grouped, by diagnosis, as follows: FSGS (n = 18); membranous nephropathy (n = 14); MCD (n = 5); and other glomerulopathies (n = 33). There was no significant difference among the groups in terms of the urinary CD80 levels, and serum suPAR was not significantly higher in the FSGS group, as would have been expected. Urinary CD80 correlated positively with nephrotic syndrome, regardless of the type of glomerular disease. Neither biomarker correlated with proteinuria at six months after biopsy. CONCLUSION: In adults, urinary CD80 can serve as a marker of nephrotic syndrome but is not specific for MCD, whereas serum suPAR does not appear to be useful as a diagnostic or treatment response marker.

The Protein-Independent Role of Phosphate in the Progression of Chronic Kidney Disease.

Several factors contribute to renal-function decline in CKD patients, and the role of phosphate content in the diet is still a matter of debate. This study aims to analyze the mechanism by which phosphate, independent of protein, is associated with the progression of CKD. Adult Munich-Wistar rats were submitted to 5/6 nephrectomy (Nx), fed with a low-protein diet, and divided into two groups. Only phosphate content (low phosphate, LoP, 0.2%; high phosphate, HiP, 0.95%) differentiated diets. After sixty days, biochemical parameters and kidney histology were analyzed. The HiP group presented worse renal function, with higher levels of PTH, FGF-23, and fractional excretion of phosphate. In the histological analysis of the kidney tissue, they also showed a higher percentage of interstitial fibrosis, expression of α-actin, PCNA, and renal infiltration by macrophages. The LoP group presented higher expression of beclin-1 in renal tubule cells, a marker of autophagic flux, when compared to the HiP group. Our findings highlight the action of phosphate in the induction of kidney interstitial inflammation and fibrosis, contributing to the progression of renal disease. A possible effect of phosphate on the dysregulation of the renal cell autophagy mechanism needs further investigation with clinical studies.

Effect of parathyroidectomy on bone tissue biomarkers and body composition in patients with chronic kidney disease and secondary hyperparathyroidism.

BACKGROUND/OBJECTIVE: Loss of renal function may induce secondary hyperparathyroidism (s-HPT), which triggers several complications leading to an extreme decline in quality of life and increased mortality in affected patients. We evaluated whether parathyroidectomy (PTx), as surgical treatment for s-HPT, modifies body composition, and hormones involved in the protein-energy metabolism of affected patients. SUBJECTS/METHODS: Overall, 30 s-HPT patients were evaluated at two times, before PTx (pre PTx) and 6 months after PTx (post PTx). Patients were evaluated by biochemistry analysis, anthropometry, electrical bioimpedance (BIA), food intake diary, handgrip strength, and modified global subjective nutritional assessment (SGA). RESULTS: After PTx, patients showed decreased serum levels of total and ionic calcium, as well as decreased alkaline phosphatase and PTH, and increased 25 (OH) vitamin D. These results demonstrate that PTx was efficient to correct part of the mineral disorder. We also observed an increase in caloric intake, body weight, body mass index (BMI), phase angle, handgrip strength, SGA score, and a decreasing in the percentage of weight loss. The osteocalcin concentration of both carboxylated (cOC) and undercarboxylated form was diminished post PTx. The cOC correlated with bone metabolism markers and SGA score. CONCLUSIONS: PTx modified body composition improving nutritional status and preventing the progression of weight loss with increased of energy intake, BMI, handgrip strength, phase angle of BIA, and SGA score. The present study also suggests an association of cOC with bone markers and SGA score. Further studies are needed to better clarify these associations with larger sample size.

A Randomized Trial of Zoledronic Acid to Prevent Bone Loss in the First Year after Kidney Transplantation.

BACKGROUND: Bone and mineral disorders commonly affect kidney transplant (KTx) recipients and have been associated with a high risk of fracture. Bisphosphonates may prevent or treat bone loss in such patients, but there is concern that these drugs might induce adynamic bone disease (ABD). METHODS: In an open label, randomized trial to assess the safety and efficacy of zoledronate for preventing bone loss in the first year after kidney transplant, we randomized 34 patients before transplant to receive zoledronate or no treatment. We used dual-energy x-ray absorptiometry (DXA), high-resolution peripheral quantitative computed tomography (HR-pQCT), and bone biopsies to evaluate changes in bone in the 32 evaluable participants between the time of KTx and 12 months post-transplant. RESULTS: Both groups of patients experienced decreased bone turnover after KTx, but zoledronate itself did not affect this outcome. Unlike previous studies, DXA showed no post-transplant bone loss in either group; we instead observed an increase of bone mineral density in both lumbar spine and total hip sites, with a significant positive effect of zoledronate. However, bone biopsies showed post-transplant impairment of trabecular connectivity (and no benefit from zoledronate); HR-pQCT detected trabecular bone loss at the peripheral skeleton, which zoledronate partially attenuated. CONCLUSIONS: Current immunosuppressive regimens do not contribute to post-transplant central skeleton trabecular bone loss, and zoledronate does not induce ABD. Because fractures in transplant recipients are most commonly peripheral fractures, clinicians should consider bisphosphonate use in patients at high fracture risk who have evidence of significantly low bone mass at these sites at the time of KTx.

A prospective study of the influence of the skeleton on calcium mass transfer during hemodialysis.

BACKGROUND: Calcium gradient, the difference between serum calcium and dialysate calcium d[Ca], is the main contributor factor influencing calcium transfer during hemodialysis. The impact, however, of bone turnover, on calcium mass transfer during hemodialysis is still uncertain. METHODS: This prospective cross-sectional study included 10 patients on hemodialysis for a 57.6±16.8 months, with severe hyperparathyroidism. Patients were submitted to 3 hemodialysis sessions using d[Ca] of 1.25, 1.5 and 1.75 mmol/l in three situations: pre-parathyroidectomy (pre-PTX), during hungry bone (early post-PTX), and after stabilization of clinical status (late post-PTX). Biochemical analysis and calcium mass transfer were evaluated and serum bone-related proteins were quantified. RESULTS: Calcium mass transfer varied widely among patients in each study phase with a median of -89.5, -76.8 and -3 mmol using d[Ca] 1.25 mmol/L, -106, -26.8 and 29.7 mmol using d[Ca] 1.50 mmol/L, and 12.8, -14.5 and 38 mmol using d[Ca] 1.75 mmol/L during pre-PTX, early post-PTX and late post-PTX, respectively, which was significantly different among d[Ca] (p = 0.0001) and among phases (p = 0.040). Ca gradient and delta of Ca also differed among d[Ca] and phases (p<0.05 for all comparisons), whether ultrafiltration was similar. Serum Osteocalcin decreased significantly in late post-PTX, whereas Sclerostin increased earlier, in early post-PTX. CONCLUSIONS: The skeleton plays a key role in Ca mass transfer during dialysis, either by determining pre-dialysis serum Ca or by controlling the exchangeable Ca pool. Knowing that could help us to decide which d[Ca] should be chosen in a given patient.

Effect of variations in dietary Pi intake on intestinal Pi transporters (NaPi-IIb, PiT-1, and PiT-2) and phosphate-regulating factors (PTH, FGF-23, and MEPE).

Hyperphosphatemia is a common condition in patients with chronic kidney disease (CKD) and can lead to bone disease, vascular calcification, and increased risks of cardiovascular disease and mortality. Inorganic phosphate (Pi) is absorbed in the intestine, an important step in the maintenance of homeostasis. In CKD, it is not clear to what extent Pi absorption is modulated by dietary Pi. Thus, we investigated 5/6 nephrectomized (Nx) Wistar rats to test whether acute variations in dietary Pi concentration over 2 days would alter hormones involved in Pi metabolism, expression of sodium-phosphate cotransporters, apoptosis, and the expression of matrix extracellular phosphoglycoprotein (MEPE) in different segments of the small intestine. The animals were divided into groups receiving different levels of dietary phosphate: low (Nx/LPi), normal (Nx/NPi), and high (Nx/HPi). Serum phosphate, fractional excretion of phosphate, intact serum fibroblast growth factor 23 (FGF-23), and parathyroid hormone (PTH) were significantly higher and ionized calcium was significantly lower in the Nx/HPi group than in the Nx/LPi group. The expression levels of NaPi-IIb and PiT-1/2 were increased in the total jejunum mucosa of the Nx/LPi group compared with the Nx/HPi group. Modification of Pi concentration in the diet affected the apoptosis of enterocytes, particularly with Pi overload. MEPE expression was higher in the Nx/HPi group than in the Nx/NPi. These data reveal the importance of early control of Pi in uremia to prevent an increase in serum PTH and FGF-23. Uremia may be a determining factor that explains the expressional modulation of the cotransporters in the small intestine segments.

Parathyroidectomy in patients with chronic kidney disease: Impacts of different techniques on the biochemical and clinical evolution of secondary hyperparathyroidism.

BACKGROUND: Parathyroidectomy (PTx) decreases the mortality rate of refractory secondary hyperparathyroidism (rSHP) due to chronic kidney disease. A consensus regarding which techniques of PTx are associated with better outcomes is not available. The aims of this study are to evaluate the clinical and laboratory evolution of 49 hemodialysis patients with rSHP who underwent PTx using different techniques. METHODS: Patients underwent subtotal PTx (sub-PTx) or total PTx with autotransplantation (AT) of 45 (PTx-AT45) or 90 parathyroid fragments (PTx-AT90) and were followed for 12 months. We analyzed the expression of proliferating cell nuclear antigen (PCNA), calcium-sensing receptor (CasR), vitamin D receptor (VDR), fibroblast growth factor receptor-1 (FGFR1), sodium-dependent phosphate cotransporter-1 (PIT1), and Klotho in parathyroid glands. RESULTS: Baseline median serum intact parathyroid hormone (iPTH) levels were 1,466 (1,087-2,125) pg/mL; vascular calcification scores correlated with serum iPTH (r = 0.529; P = .002) and serum phosphate levels (r = 0.389; P = .028); and Klotho expression was negatively correlated with serum phosphate levels (r = -0.4; P = .01). After 12 months, serum iPTH and alkaline phosphatase levels were significantly controlled in all groups, as was bone pain. The proportions of patients with serum iPTH levels within the ranges recommended by Kidney Disease: Improving Global Outcomes were similar among the treatment groups. During the hungry bone disease (HBS), patients received 3,786 g (1,412-7,580) of elemental calcium, and a trend toward a positive correlation between the cumulative calcium load at the end of follow up and VC score post-PTx was noted (r = 0.390; P = .06). Two cases evolved to clinically uncontrolled hyperparathyroidism in the sub-PTx group. The expression patterns of PCNA, VDR, CasR, PIT1, FGFR1, and Klotho in parathyroid glands did not correlate with serum systemic iPTH levels or the duration of HBS. CONCLUSIONS: All 3 operative techniques were effective at controlling rSHP, both in clinical and laboratory terms. Neither the quantity nor quality of parathyroid fragments influenced serum systemic iPTH and AT-iPTH levels. The cumulative calcium load appeared to correlate with the VC score and may have affected its progression. The effects of phosphate restriction on Klotho expression in human parathyroid glands and the subsequent decrease in FGF23 resistance must be addressed in further studies.

Renal osteodystrophy in the obesity era: Is metabolic syndrome relevant?

BACKGROUND: Observational studies have shown a beneficial effect of obesity on bone health; however, most of those studies were not based on bone biopsies. Metabolic syndrome (MetS) could have an effect on bone remodeling. However, there are no data on the effects of MetS in the presence of renal osteodystrophy. OBJECTIVE: The aim of this study was to investigate associations between MetS and renal osteodystrophy using the bone histomorphometric turnover-mineralization-volume (TMV) classification. DESIGN, SETTING, PARTICIPANTS & MEASUREMENTS: This observational cross-sectional study included 55 hemodialysis patients (28 women/27 men) who were evaluated for MetS and bone histomorphometry. Biochemical parameters included calcium, phosphorus, alkaline phosphatase, intact parathyroid hormone (iPTH), 25-hydroxyvitamin D, free serum leptin, fibroblast growth factor 23 (FGF23), intact osteocalcin, sclerostin (Scl), glucose, insulin, and thyroid hormones. Robust models of multivariate linear regressions were used for the statistical analyses. RESULTS: Females had higher iPTH levels (1,143 vs. 358, p = 0.02). Patients with normal bone volume (BV/TV) had a higher prevalence of MetS (73.6% vs. 41.7%, p = 0.02) and higher serum phosphorus, C-terminal FGF23 and insulin levels. The multivariate regression analysis showed that low-density lipoprotein cholesterol (LDL) was positively correlated with bone formation rate (BFR/BS) and negatively associated with mineralization lag time. Bone volume was negatively associated with age but positively associated with MetS. Body mass index (BMI) was not correlated with any of the bone histomorphometric parameters. CONCLUSION: Our results suggest that MetS is not a risk factor for low bone volume in hemodialysis patients. Furthermore, BMI alone was not related to bone volume in this population.

Histomorphometric analysis of the femoral neck in patients with and without femoral neck fracture.

OBJECTIVE: To determine, through bone histomorphometry in femoral neck, whether there are differences in the cancellous bone of the proximal femur from female patients over 60 years old who had femoral neck fracture and similar patients who did not have such fracture. METHODS: We analyzed the trabecular part of the femur of 13 female patients, aged over 60 years old, by the bone histomorphometry method. Seven of these patients had femoral neck fracture. All of them were subjected to hip arthroplasty. RESULTS: Bone densitometry showed no significant difference. There was no significant difference on the average thickness of the trabecular bone (124.38µm versus 147.09µm). The number of bone trabeculae was lower (1.52, versus 1.88) and the separation between them was larger (541,19µm versus 391,14µm) in the fracture group. CONCLUSION: A difference in histomorphometric parameters of cancellous bone of the femur neck was observed among patients who had fractures as compared to patients who had not.

Histomorphometric bone assessment in patients with fracture of the proximal end of the femur.

OBJECTIVE: To determine whether there is a difference on the bone architecture in patients with femoral neck fracture compared to patients with intertrochanteric fractures and assess the importance of aging on bone microarchitecture in patients with proximal femoral fracture. METHODS: Biopsy of the iliac crest was made in seventeen patients between 55 and 90 years old who were admitted to the emergency room with fractures of the proximal end of the femur. After a small fragment was removed, we made a histomorphometric analysis of it. RESULTS: There was no significant difference between patients with femoral neck fracture and trochanteric fracture in structural parameters, formation and resorption. Comparing age groups we also did not find any significant change between the groups in the parameters volume and trabecular separation. CONCLUSION: There are no difference in the morphometric parameters analyzed between the different types of fracture and age is not a significant factor in the alteration of these parameters.

Bone disease in newly diagnosed lupus nephritis patients.

INTRODUCTION: Bone loss in Lupus Nephritis (LN) patients is common and multifactorial. The aim of this study was to evaluate the bone status of newly diagnosed LN patients and their correlation with inflammatory factors involved in LN physiopathology. METHODS: We studied 15 pre-menopausal patients with ≤2 months of diagnosed SLE and LN. Patients with prior kidney or bone disease were excluded. In addition to biochemical evaluation (including 25-hydroxyvitamin D3 [25(OH)D] and Monocyte Chemotactic Protein (MCP1) dosage), we performed bone biopsies followed by osteoblast culture, histomorphometric and immunohistochemistry analysis. RESULTS: LN patients presented a mean age of 29.5±10 years, a proteinuria of 4.7±2.9 g/day and an estimated glomerular filtration rate (GFR) of 37(31-87) ml/min/1,73 m2. They were on glucocorticoid therapy for 34±12 days. All patients presented vitamin D insufficiency (9.9±4.4 ng/ml, range 4-20). Urinary MCP1 correlated negatively with 25(OH)D (r = -0.53, p = 0.003) and positively with serum deoxypyridinoline (r = 0.53, p = 0.004). Osteoblasts isolated from LN bone biopsies presented a significantly higher expression of MCP-1 when compared to controls (32.0.±9.1 vs. 22.9±5.3 mean fluorescence intensities, p = 0.01). LN patients presented a significantly reduced osteoid volume, osteoid thickness, osteoid surface, mineralization surface and bone formation rate, associated with an increased eroded surface and osteoclast surface. Patient's bone specimens demonstrated a reduced immunostaining for osteoprotegerin (0.61±0.82 vs. 1.08±0.50%, p = 0.003), and an increased expression of Receptor Activator of NF-κB ligand (RANKL) (1.76±0.92 vs. 0.41±0.28%, p<0.001) when compared to controls. DISCUSSION: Newly diagnosed LN patients presented a significant disturbance in bone metabolism, characterized by an impaired bone formation and mineralization, associated with an increase in resorption parameters. Glucocorticoid use, vitamin D insufficiency and inflammation might be involved in the physiopathology of bone metabolism disturbance.

Prefabricated bone flap: an experimental study comparing deep-frozen and lyophilized-demineralized allogenic bones and tissue expression of transforming growth factor ß.

BACKGROUND: Extensive bone defects are still a challenge for reconstructive surgery. Allogenic bones can be an alternative with no donor area morbidity and unlimited amount of tissue. Better results can be achieved after allogenic bone preparation and adding a vascular supply, which can be done along with flap prefabrication. The purpose of this study was to evaluate demineralized/lyophilized and deep-frozen allogenic bones used for flap prefabrication and the tissue expression of transforming growth factor ß (TGF-ß) in these bone fragments. METHODS: Fifty-six Wistar rat bone diaphyses were prepared and distributed in 4 groups: demineralized/lyophilized (experimental group 1 and control group 2) and deep freezing (experimental group 3 and control group 4). Two bone segments (one of each group) were implanted in rats to prefabricate flaps using superficial epigastric vessels (experimental groups) or only transferred as grafts (control groups). These fragments remained in their respective inguinal regions until the death that occurred at 2, 4, and 6 weeks after the operation. Semiquantitative histologic (tetracycline marking, cortical resorption, number of giant cells, and vascularization) and histomorphometrical quantitative (osteoid thickness, cortical thickness, and fibrosis thickness) analyses were performed. Transforming growth factor ß immunohistochemistry staining was also performed. RESULTS: Group 1 fragments presented an osteoid matrix on their external surface in all periods. Cartilage formation and mineralization areas were also noticed. These findings were not observed in group 3 fragments. Group 1 had more mineralization and double tetracycline marks, which were almost not seen in group 3. Cortical resorption and the number of giant cells were greater in group 3 in all periods. Vascularization and fibrosis thickness were similar in both experimental groups. Group 1 had more intense TGF-ß staining within 2 weeks of study. Nevertheless, from 4 weeks onward, group 3 presented statistically significant stronger staining. CONCLUSIONS: Although there are some differences between the preparation methods of allogenic bone, it is possible to prefabricate flaps with demineralized/lyophilized and deep-frozen bones.

Phosphorus is associated with coronary artery disease in patients with preserved renal function.

High serum phosphorus levels have been associated with mortality and cardiovascular events in patients with chronic kidney disease and in the general population. In addition, high phosphorus levels have been shown to induce vascular calcification and endothelial dysfunction in vitro. The aim of this study was to evaluate the relation of phosphorus and coronary calcification and atherosclerosis in the setting of normal renal function. This was a cross-sectional study involving 290 patients with suspected coronary artery disease and undergoing elective coronary angiography, with a creatinine clearance >60 ml/min/1.73 m(2). Coronary artery obstruction was assessed by the Friesinger score and coronary artery calcification by multislice computed tomography. Serum phosphorus was higher in patients with an Agatston score >10 than in those with an Agatston score ≤ 10 (3.63 ± 0.55 versus 3.49 ± 0.52 mg/dl; p = 0.02). In the patients with Friesinger scores >4, serum phosphorus was higher (3.6 ± 0.5 versus 3.5 ± 0.6 mg/dl, p = 0.04) and median intact fibroblast growth factor 23 was lower (40.3 pg/ml versus 45.7 pg/ml, p = 0.01). Each 0.1-mg/dl higher serum phosphate was associated with a 7.4% higher odds of having a Friesinger score >4 (p = 0.03) and a 6.1% greater risk of having an Agatston score >10 (p = 0.01). Fibroblast growth factor 23 was a negative predictor of Friesinger score (p = 0.002). In conclusion, phosphorus is positively associated with coronary artery calcification and obstruction in patients with suspected coronary artery disease and preserved renal function.

Repression of osteocyte Wnt/ß-catenin signaling is an early event in the progression of renal osteodystrophy.

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-ß-catenin was observed both in mouse and human bones. Overall repression of Wnt/ß-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/ß-catenin pathway is involved in the pathogenesis of renal osteodystrophy.

Chronic kidney disease bone and mineral disorder (CKD-MBD) in apolipoprotein E-deficient mice with chronic renal failure.

BACKGROUND: Chronic kidney disease (CKD) is associated with disorders of mineral and bone metabolism (MBD) which include renal osteodystrophy and vascular calcifications. This is of clinical concern because the high risk of cardiovascular (CVD) complications observed in uremic patients may be linked with bone disease. In this context, our aim was to characterize the bone lesions in CKD-apolipoprotein E-deficient mice (apoE(-/-)) and analyze their relationships with the vascular calcifications which these animals develop rapidly in this model. With ApoE being also involved in bone metabolism, we compared the effects of CRF on the bone of apoE(-/-) mice to those observed in wild type mice (WT) of the same genetic background, C57/BL6. METHODS: After CRF creation or sham surgery, 10 week-old female apoE(-/-) and WT mice were randomized to 4 groups (n=10-14/group) and fed with standard diet. Eight weeks later, animals were euthanized. Serum, aorta and femur were sampled. Femurs were imaged with 3-dimensional microtomography (microCT) and processed for bone histomorphometry (BHM). Additional quantitative histology was performed on atherosclerotic and calcified lesions in the aortas of apoE(-/-) mice. RESULTS: First, apoE(-/-) mice exhibited higher cortical (10%) and trabecular (31%) bone mass than WT. CRF led to a further increase in trabecular BV/TV in WT and in apoE(-/-) mice (10.2% and 77.2%, respectively). We observed a similar increase in osteoid surface and osteoblastic parameters in CRF mice of both genotypes while resorption parameters were less augmented by CRF in apoE(-/-) mice. Finally, based on either BHM or microCT we found positive correlations between the extent of atherosclerotic lesions and bone volume parameters, and between the size of plaque calcification and osteoclast parameters in apoE(-/-) mice. CONCLUSION: ApoE deficiency is associated with an increase in bone mass and volumetric mineral density in 20 week-old female mice. Bone mass is further increased, whereas bone mineral density is decreased, in response to CRF in association with histological features of osteitis fibrosa. Finally, our findings of correlations between changes in bone and aortic lesions in apoE(-/-) mice, are compatible with the hypothesis of a link between bone and vascular disease and require further study.

RANKL is a mediator of bone resorption in idiopathic hypercalciuria.

BACKGROUND AND OBJECTIVES: This study aimed to determine the expression of osteoprotegerin, receptor activator of nuclear factor kappaB ligand, interleukin-1alpha, transforming growth factor-beta, and basic fibroblast growth factor in stone-forming patients with idiopathic hypercalciuria. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Immunohistochemical analysis was performed in undecalcified bone samples previously obtained from 36 transiliac bone biopsies of patients who had idiopathic hypercalciuria and whose histomorphometry had shown lower bone volume, increased bone resorption, and prolonged mineralization lag time. RESULTS: Bone expression of receptor activator of nuclear factor kappaB ligand and osteoprotegerin was significantly higher in patients with idiopathic hypercalciuria versus control subjects. Transforming growth factor-beta immunostaining was lower in patients with idiopathic hypercalciuria than in control subjects and correlated directly with mineralization surface. Interleukin-1alpha and basic fibroblast growth factor staining did not differ between groups. Receptor activator of nuclear factor kappaB ligand bone expression was significantly higher in patients who had idiopathic hypercalciuria and exhibited higher versus normal bone resorption. CONCLUSION: A higher expression of receptor activator of nuclear factor kappaB ligand in bone tissue suggests that increased bone resorption in patients with idiopathic hypercalciuria is mediated by receptor activator of nuclear factor kappaB ligand. Osteoprotegerin bone expression might have been secondarily increased in an attempt to counteract the actions of receptor activator of nuclear factor kappaB ligand. The low bone expression of transforming growth factor-beta could contribute to the delayed mineralization found in such patients.

Usefulness of a quick decalcification of bone sections embedded in methyl methacrylate[corrected]: an improved method for immunohistochemistry.

Immunohistochemistry of undecalcified bone sections embedded in methyl methacrylate (MMA) is not commonly employed because of potential destruction of tissue antigenicity by highly exothermic polymerization. The aim of the present study was to describe a new technique in which a quick decalcification of bone sections embedded in MMA improves the results for immunohistochemistry. The quality of interleukin 1alpha (IL-1alpha) immunostaining according to the present method was better than the conventional one. Immunostaining for osteoprotegerin (OPG) and the receptor activator of NF-kappaB ligand (RANKL) in bone sections of chronic kidney disease patients with mineral bone disorders (CKD-MBD) was stronger than in controls (postmortem healthy subjects). The present study suggested that this method is easy, fast, and effective to perform both histomorphometry and immunohistochemistry in the same bone fragment, yielding new insights into pathophysiological aspects and therapeutic approaches in bone disease.

Effects of calcitriol on parathyroid function and on bone remodelling in secondary hyperparathyroidism.

BACKGROUND: Secondary hyperparathyroidism (2HPT) develops in chronic renal failure due to disturbances of calcium, phosphorus and vitamin D metabolism. It is characterized by high turnover bone disease and an altered calcium-parathyroid hormone (PTH) relationship. Calcitriol has been widely used for the treatment of 2HPT. However, it remains controversial whether calcitriol is capable of inducing changes of the calcium-PTH curve. The aim of the present study was to examine this issue and to determine the effect of calcitriol on bone remodelling in patients with severe 2HPT. METHODS: We evaluated 16 chronic haemodialysis patients with severe 2HPT (PTH 899+/-342 pg/ml). Each patient underwent a dynamic parathyroid function test (by infusion of calcium gluconate and sodium citrate) and a bone biopsy before and after a 6 month period of i.v. calcitriol therapy (CTx). RESULTS: After treatment, eight patients were identified as calcitriol responders and the other eight as non-responders, based on plasma PTH level (<300 pg/ml for responders and >300 pg/ml for non-responders). The first group had higher plasma 25OHD(3) levels (39+/-8 vs 24+/-7 ng/ml, P<0.005). As to the calcium-PTH curve, we found differences in slope (-12.7+/-5.2 vs -21.7+/-11.4, P=0.05), basal/maximum PTH ratio (48.8+/-14.9 vs 71.05+/-20.1%, P=0.01) and time to achieve hypocalcaemia (79.0+/-13.5 vs 94.3+/-13.7 min, P<0.001). Initial histomorphometric parameters did not allow identification of the different groups. After the 6-month CTx, alterations in the calcium-PTH curve were clearly seen in responders [a drop in maximum PTH (from 1651+/-616 to 938+/-744 pg/ml, P<0.05) and minimum PTH (from 163+/-75.4 to 102.2+/-56.7 pg/ml, P<0.005)], associated with an increase in minimum/basal PTH ratio (from 23.3+/-11.6 to 34.5+/-20.4%, P<0.05) and maximum calcium (from 0.99+/-0.07 to 1.1+/-0.09 mmol/l, P<0.05). Set point and slope were not altered after calcitriol treatment, in responders (set point=1.17+/-0.08 vs 1.15+/-0.1 mmol/l, ns; slope=-12.7+/-5.2 vs -12.9+/-9.3, ns) or non-responders (set point=1.21+/-0.05 vs 1.21+/-0.2 mmol/l, ns; slope=-21.7+/-11.4 vs -17.3+/-8.4, ns). Bone formation parameters were reduced in all patients [osteoid surface (OS/BS)=from 57.1+/-21.6 to 41.6+/-26%, P<0.05 for responders, and from 76.7+/-12 to 47.1+/-15%, P<0.001 in non-responders], but non-responders had increased bone resorption [eroded surface (ES/BS)=7.1+/-3.4 vs 16.6+/-4.9, P<0.05]. CONCLUSION: Calcitriol had non-uniform effects on parathyroid function and bone remodelling in uraemic patients. Non-responders exhibited a decoupled remodelling process that could further influence mineral balance or possibly also bone structure. To avoid such undesirable effects, early identification of non-responder patients is crucial when using calcitriol for the treatment of 2HPT.