Stefania Sella

Persistent secondary hyperparathyroidism and vertebral fractures in kidney transplantation: role of calcium-sensing receptor polymorphisms and vitamin D deficiency.

Bone morbidity remains a major problem even after successful renal transplantation. We investigated the role of calcium‐sensing receptor (CaSR) polymorphisms and 25‐hydroxyvitamin D levels on the persistence of secondary hyperparathyroidism (SHPT) and their relationships with vertebral fractures (VFx) in 125 renal allograft recipients transplanted 44 ± 23 months before. All patients underwent evaluation of the main biochemical parameters of calcium metabolism as well as vertebral and femoral bone density. In 87 patients, CaSR polymorphisms (A986S, R990G, and Q1011E) also were assessed. X‐ray images of the lateral spine were obtained in 102 subjects to perform vertebral morphometry. High parathyroid hormone (PTH) and 25‐hydroxyvitamin D lower than 80 nmol/L were found in 54% and 97% of patients, respectively, with 40% of these showing vitamin D levels lower than 30 nmol/L. VFx were detected in 57% of the subjects. After multiple adjustments, 25‐hydroxyvitamin D, age, and hemodialysis duration, but not CaSR polymorphisms, were found to be significant predictors of high PTH, whereas age and time since transplant were positively related with lower 25‐hydroxyvitamin D values. PTH and time since transplant were significantly associated with VFx. Patients with two or more VFx showed serum PTH levels 50% higher than patients without fractures. We therefore conclude that persistent SHPT is a very common feature after renal transplantation and that, unlike CaSR polymorphisms, low 25‐hydroxyvitamin D is involved in its pathogenesis. High PTH levels, in turn, are associated with an increased VFx risk, which confirms the need for strategies aimed at lowering serum PTH in this setting as well.

BONE DISEASE IN PRIMARY HYPERCALCIURIA

Primary hypercalciuria (PH) is very often accompanied by some degree of bone demineralization. The most frequent clinical condition in which this association has been observed is calcium nephrolithiasis. In patients affected by this disorder, bone density is very frequently low, and increased susceptibility to fragility fractures is reported. The very poor definition of this bone disease from a histomorphometric point of view is a crucial aspect. At present, the most common finding seems to be a low bone turnover condition. Many factors are involved in the complex relationships between bone loss and PH. Since bone loss was mainly reported in patients with fasting hypercalciuria, a primary alteration in bone metabolism was proposed as a cause of both hypercalciuria and bone demineralization. This hypothesis was strengthened by the observation that some bone resorbing-cytokines, such as interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor nechrosis factor-α (TNF-α), are high in hypercalciuric patients. An excessive response to the acid load induced by dietary protein intake seems to be an additional factor explaining a primitive alteration of bone. The intestine plays a major role in the clinical course of bone disease in PH. Patients with absorptive hypercalciuria less frequently show bone disease, and a reduction in dietary calcium greatly increases the probability of bone loss in PH subjects. It has recently been reported that greater bone loss is associated with a larger increase in intestinal calcium absorption in PH patients. Considering the absence of parathyroid hormone (PTH) alterations, it was proposed that this is not a compensatory phenomenon, but probably the marker of disturbed cell calcium transport, involving both intestinal and bone tissues. While renal hypercalciuria is rather uncommon, the kidney still seems to play a role in the pathogenesis of bone loss in PH patients, possibly via the effect of mild-to-moderate urinary phosphate loss with secondary hypophosphatemia. In conclusion, bone loss is very common in PH patients. Even if most of the factors involved in this process have been identified, many aspects of this intriguing clinical condition remain to be elucidated.

Densitometric Threshold and Vertebral Fractures in Heart Transplant Patients

Background. Bone disease is one of the major complications of solid organ transplantation, causes considerable morbidity, and most patients are treated with immunosuppressant drugs after graft. The majority of studies reported rapid bone loss and an increased incidence of fractures after transplantation. The aim of our study was to evaluate osteoporosis and fracture prevalence, bone metabolism, and the effect of immunosuppressant agents on bone after heart transplantation. Methods. We planned a cross-sectional study in 180 heart transplant patients recruited from 3 different centers with a less than 10 years from graft. Each patient underwent a densitometric scan, and in 157 of them, an x-ray of the spine was performed to evaluate fractures. Biochemical assessment of bone metabolism was made at the time of the visit. Physical activity, diet, and calcium intake were evaluated using a specific questionnaire. Results. Vertebral fractures were diagnosed in 40% of subjects, but densitometric osteoporosis was observed only in 13% of spine and in 25% of hip scans. Interestingly, increasing T-score threshold up to −1.5 standard deviation, the prevalence of fractured patient improved significantly, reaching 60% in both genders. Bone content was inversely correlated with glucocorticoids, while a positive correlation was found with cyclosporine A. Almost all subjects had vitamin D deficiency. Conclusions. Standard densitometric criteria are unreliable to identify bone fragility after transplantation, and a different threshold (−1.5 standard deviation) should be considered. Transplanted patients should be adequately supplemented with vitamin D, and the effects of immunosuppressant agents on bone need further investigation.

Vitamin K plasma levels determination in human health.

Abstract Vitamin K (phylloquinone or vitamin K1 and menaquinones or vitamin K2) plays an important role as a cofactor in the synthesis of hepatic blood coagulation proteins, but recently has also aroused an increasing interest for its action in extra-hepatic tissues, in particular in the regulation of bone and vascular metabolism. The accurate measurement of vitamin K status in humans is still a critical issue. Along with indirect assays, such as the undercarboxylated fractions of vitamin K-dependent proteins [prothrombin, osteocalcin (OC), and matrix gla protein], the direct analysis of blood levels of phylloquinone and menaquinones forms might be considered a more informative and direct method for assessing vitamin K status. Different methods for direct quantification of vitamin K serum levels are available. High-performance liquid chromatography (HPLC) methods coupled with post-column reduction procedures and fluorimetric or electrochemical detection are commonly used for food and blood analysis of phylloquinone, but they show some limitations when applied to the analysis of serum menaquinones because of interferences from triglycerides. Recent advancements include liquid chromatography tandem mass spectrometry (LCMS/MS) detection, which assures higher specificity. The optimization and standardization of these methods requires specialized laboratories. The variability of results observed in the available studies suggests the need for further investigations to obtain more accurate analytical results.

Low vitamin K1 intake in haemodialysis patients

BACKGROUND & AIMS Vitamin K acts as a coenzyme in the γ-carboxylation of vitamin K-dependent proteins, including coagulation factors, osteocalcin, matrix Gla protein (MGP), and the growth arrest-specific 6 (GAS6) protein. Osteocalcin is a key factor for bone matrix formation. MGP is a local inhibitor of soft tissue calcification. GAS6 activity prevents the apoptosis of vascular smooth muscle cells. Few data on vitamin K intake in chronic kidney disease patients and no data in patients on a Mediterranean diet are available. In the present study, we evaluate the dietary intake of vitamin K1 in a cohort of patients undergoing haemodialysis. METHODS In this multi-centre controlled observational study, data were collected from 91 patients aged >18 years on dialysis treatment for at least 12 months and from 85 age-matched control subjects with normal renal function. Participants completed a food journal of seven consecutive days for the estimation of dietary intakes of macro- and micro-nutrients (minerals and vitamins). RESULTS Compared to controls, dialysis patients had a significant lower total energy intake, along with a lower dietary intake of proteins, fats, carbohydrates, fibres, and of all the examined minerals (Ca, P, Fe, Na, K, Zn, Cu, and Mg). With the exception of vitamin B12, vitamins intake followed a similar pattern, with a lower intake in vitamin A, B1, B2, C, D, E, folates, K1 and PP. These finding were confirmed also when normalized for total energy intake or for body weight. In respect to the adequate intakes recommended in the literature, the prevalence of a deficient vitamin K intake was very high (70-90%) and roughly double than in controls. Multivariate logistic model identified vitamin A and iron intake as predictors of vitamin K deficiency. CONCLUSIONS Haemodialysis patients had a significantly low intake in vitamin K1, which could contribute to increase the risk of bone fractures and vascular calcifications. Since the deficiency of vitamin K intake seems to be remarkable, dietary counselling to HD patients should also address the adequacy of vitamin K dietary intake and bioavailability. Whether diets with higher amounts of vitamin K1 or vitamin K supplementation can improve clinical outcomes in dialysis patients remains to be demonstrated.

Risedronate prevents the loss of microarchitecture in glucocorticoid-induced osteoporosis in rats

Osteoporosis is a severe complication of glucocorticoid treatment. Bisphosphonates are a powerful therapeutic option to prevent osteoporotic fractures. The aims of this study were: a) to determine bone alterations induced by therapy with glucocorticoids (GC); b) to establish the efficacy of risedronate (Ris) in the prevention of these effects. We studied 40 female Sprague-Dawley rats randomly divided into 4 groups of treatment, administered 3 times a week sc: 1. Control: vehicle of methylprednisolone (GC) + vehicle of Ris; 2. Ris: Ris 5 μg/kg body weight vehicle of GC; 3. GC: GC 7 mg/kg + vehicle of Ris; 4. GC+Ris: GC 7 mg/kg, Ris 5 μg/kg. Animals were treated for 30 days and then were sacrificed. Densitometry was performed at baseline and at the end of the treatment. Right tibiae were removed for histomorphometric analyses. The GC group showed a 7% decrease in bone density vs controls (p<0.05), while the GC+Ris group was associated with a 3.5% increase in bone density vs controls (p<0.05). In the GC group, histomorphometric evaluations showed reduced bone volume (BV/TV) and thinning of trabeculae (Tb.Th) vs controls (BV/TV: 31±1 vs 35±1%, p<0.05; Tb.Th: 43±2 vs 50±3 μm, p<0.01; Ac.f: 1.8±0.2 vs 1.6±0.3 N/yr). The GC+Ris group had increased BV/TV and Tb.Th, and reduced Ac.f vs the GC group. Ris also maintained trabecular microarchitecture. At the histological level, glucocorticoid-induced osteoporosis was characterized by decreased bone volume, reduced osteoblastic activity, and deterioration of microarchitecture. Ris counteracted these effects both by prolonging osteoblast activity, and by maintaining bone microarchitecture.

Declining trends in the incidence of hip fractures in people aged 65 years or over in years 2000–2011

BACKGROUND The aim of this study was to explore hip fracture (HFx) incidence in the Veneto Region of Italy, looking at potential differences with the national data. METHODS We analyzed HFx incidence for people aged 65years or over, in years 2000-2011, using data from the Regional Hospitalization Database. Patients were stratified by sex, calendar year and 5-year age class. Data for the single provinces of the Region were also obtained. Absolute number of HFx, crude incidence for 10,000 inhabitants and age-standardized fracture rates were calculated. RESULTS During the study period, there were 53,917 hospitalizations for HFx (77.7% in females). In the whole 11year period of observation, the absolute HFx number increased by 17.7% in males and 10.6% females, respectively. However, age-standardized incidence rates declined by 18% in the same period (IRR 0.82, 95% CI 0.78-0.87). This decreasing trend was almost identical through all the age-cohorts up to 84years. In the whole study period, HFx incidence was lower for Padova (IRR 0.63, 95% CI 0.60-0.66) and Verona (IRR 0.66, 95% CI 0.63-0.70) provinces as compared to the others. This regional profile was quite different with respect to the data published, for the same calendar years, for Italy as a whole, in spite of an almost identical demography of the population. CONCLUSIONS HFx incidence is declining in the Veneto Region of Italy. Further studies, aimed to investigate factors involved in this figure are needed.

The relationship between the Spine Deformity Index, biochemical parameters of bone metabolism and vascular calcifications: results from the Epidemiological VERtebral FRACtures iTalian Study (EVERFRACT) in dialysis patients.

Abstract Background: The Spine Deformity Index (SDI) is a measure of vertebral fractures (VFs), providing information on both their number and severity. Methods: We evaluated the relationships between SDI and clinical, biochemical and arterial calcification parameters in 387 hemodialysis (HD) patients. VFs, assessed by quantitative vertebral morphometry, and vascular calcifications were identified in the same lateral spinal X-ray. To improve the detection of fracture severity, we created a corrected SDI (c-SDI), by dividing SDI for the number of VFs. We assessed routine biochemistry, bone-Gla-protein (BGP), undercaboxylated BGP (ucBGP), and matrix-Gla-protein (MGP). Results: VFs prevalence was 55.3%. HD patients with a SDI >1 were more frequently males (p<0.05), and had lower BGP (p<0.01). Patients with a c-SDI >1 had higher LDL-cholesterol (p<0.05) and lower ucBGP (p<0.05) and MGP (p<0.05). Calcifications of the abdominal aorta (AAoC) were more frequent in patients with SDI >1 (p<0.05) and with c-SDI >1 (p<0.05). Multivariate logistic regression showed that male sex (OR 1.86, CI 1.20–2.91), age (OR 1.03, CI 1.01–1.05) and albumin ≥3.5 g/dL (OR 0.54, CI 0.31–0.93) were predictors of a SDI >1. Age (OR 1.05, CI 1.03–1.07), LDL-cholesterol (OR 1.74, CI 1.04–2.92) and ucBGP (OR 0.35, CI 0.18–0.70) were associated with c-SDI >1. Conclusions: We conclude that the severity of VFs was associated with age, atherogenic factors and bone metabolism markers.

The effect of risedronate on osteogenic lineage is mediated by cyclooxygenase-2 gene upregulation.

IntroductionThe purpose of this study was to evaluate the effects of risedronate (Ris) in the modulation of bone formation in rats with glucocorticoid (GC)-induced osteoporosis by histomorphometric, immunohistochemical and gene expression analyses.MethodsWe analyzed structure, turnover and microarchitecture, cyclooxygenase 2 (COX-2) levels and osteocyte apoptosis in 40 female rats divided as follows: 1) vehicle of methylprednisolone (vGC) + vehicle of risedronate (vRis); 2) Ris 5 μg/Kg + vGC; 3) methylprednisolone (GC) 7 mg/Kg + vRis; 4) GC 7 mg/Kg +Ris 5 μg/Kg. In addition, we evaluated cell proliferation and expression of COX-2 and bone alkaline phosphatase (b-ALP) genes in bone marrow cells and MLO-y4 osteocytes treated with Ris alone or in co-treatment with the selective COX-2 inhibitor NS-398 or with dexametasone.ResultsRis reduced apoptosis induced by GC of osteocytes (41% vs 86%, P < 0.0001) and increased COX-2 expression with respect to controls (Immuno-Hystochemical Score (IHS): 8.75 vs 1.00, P < 0.0001). These positive effects of Ris in bone formation were confirmed by in vitro data as the viability and expression of b-ALP gene in bone marrow cells resulted increased in a dose dependent manner.ConclusionsThese findings suggest a positive effect of Ris in bone formation and support the hypothesis that the up-regulation of COX-2 could be an additional mechanism of anabolic effect of Ris.

Osteoporosi nel paziente sottoposto a trapianto d’organo

RiassuntoNegli ultimi anni il trapianto è diventato un concreto ed efficace approccio terapeutico per molte patologie terminali d’organo, quali quelle di rene, cuore, fegato, polmone e pancreas, nonché per molte patologie ematologiche. Tuttavia, è ben noto che il trapianto d’organo può essere gravato da diverse complicanze cliniche, tra le quali l’osteoporosi è una delle più frequenti e rilevanti. Gradi diversi di fragilità ossea sono spesso presenti già prima del trapianto, in relazione alle diverse alterazioni del metabolismo scheletrico che caratterizzano insufficienza renale, epatica, cardiaca, polmonare e pancreatica. Si stima che oltre il 10% dei pazienti possa presentare fratture prima del trapianto. Tuttavia, la percentuale di pazienti con osteoporosi e fratture cresce in modo drastico dopo il trapianto potendo raggiungere oltre il 50% e fino al 35% dei pazienti, rispettivamente. Il trapianto di cuore e il trapianto di polmone sembrano essere le tipologie maggiormente gravate dal rischio di fratture osteoporotiche. In epoca post-trapianto, aldilà delle alterazioni metaboliche ancora presenti in relazione alla malattia che ha condotto all’insufficienza terminale d’organo (come, ad esempio, l’iperparatiroidismo secondario persistente nel trapianto di rene), il principale fattore di rischio per osteoporosi e fratture è certamente rappresentato dall’assunzione della terapia immunosoppressiva. Se non è ancora del tutto chiaro l’effetto della ciclosporina A e del tacrolimus sul metabolismo osseo, l’impiego di glucocorticoidi costituisce senza dubbio il fattore causale di maggiore rilievo. Nel soggetto trapiantato, infatti, i glucocorticoidi sono adoperati, almeno in fase iniziale, a dosi molto elevate, nella maggioranza dei casi vengono mantenuti in terapia per un tempo indefinito e spesso usati ad alto dosaggio per il trattamento degli episodi di rigetto. Nuove prospettive terapeutiche sembrano affacciarsi oggi per il trattamento di questa condizione. Seppure in studi di dimensioni contenute, infatti, diversi bisfosfonati si sono dimostrati efficaci nel prevenire la perdita di massa ossea e, in qualche caso, nel ridurre il rischio di frattura. In conclusione, l’osteoporosi post-trapianto d’organo rap-presenta una delle più gravi forme di malattia fragilizzante dello scheletro. Tuttavia, la crescente attenzione nei confronti di questa condizione potrà consentire un sempre più valido approccio diagnostico e terapeutico alla malattia.