Prolonged vitamin D intoxication: presentation, pathogenesis and progress.

Vitamin D toxicity from unactivated vitamin D (calciferol) therapy is currently a rare cause of hypercalcaemia. However, the frequency of this event may increase as high-dose unactivated vitamin D preparations become available. Prolonged vitamin D toxicity can cause reversible hypercalcaemia and partially reversible renal impairment. Parathyroid hormone may not be suppressed with unactivated vitamin D toxicity, especially if renal disease coexists.

Association between 25-OH vitamin D and insulin is independent of lipoatrophy in HIV.

OBJECTIVE: Vitamin D deficiency (VDD) is prevalent in HIV, and following antiretroviral therapy (ART), increased rates of lipoatrophy and metabolic abnormalities are described. We investigated the relationships between 25-hydroxyvitamin D [25(OH)D] and other metabolic parameters in a group of HIV patients with and without lipoatrophy to examine whether lipoatrophy could explain the high prevalence of VDD and metabolic abnormalities. BACKGROUND: Vitamin D receptors are expressed in adipose tissue implicating vitamin D, through paracrine/autocrine mechanism, in exerting effects on fat metabolism. HIV patients frequently suffer from VDD, and those treated with thymidine analogues frequently suffer from lipoatrophy so we investigated whether lipoatrophy could explain these associations. DESIGN AND PATIENTS: Cross-sectional study of HIV-infected male patients (n = 107; 39 with lipoatrophy) from the West Australian cohort with measurements of 25(OH)D, adiponectin, insulin, lipids and leg fat as a percentage of mass. RESULTS: Reduced 25(OH)D levels were common and significantly associated with higher serum insulin in the entire cohort (P = 0·006), but there was no difference in 25(OH)D between untreated and antiretroviral-treated patients with or without lipoatrophy. Treated patients with lipoatrophy were more likely to take thymidine analogue therapy, were older and on therapy longer than treated patients without lipoatrophy. Adiponectin levels did not correlate with 25(OH)D, but lipoatrophic-treated patients had lower levels of adiponectin compared with nonlipoatrophic-treated patients. CONCLUSIONS: Lower 25(OH)D is associated with higher serum insulin but not lipoatrophy or hypoadiponectinemia in HIV-infected patients. The association between VDD and insulin resistance is likely to be mediated by independent mechanisms.

Efficacy, side effects and route of administration are more important than frequency of dosing of anti-osteoporosis treatments in determining patient adherence: a critical review of published articles from 1970 to 2009.

The purposes of the study were to review available published literature on magnitude of non-adherence with osteoporosis regimens and to determine the association between frequency and modality of medication administration with patient preference and adherence. We searched peer-reviewed journal databases--MEDLINE, EMBASE, Biosis and Derwent Drug File for publications (January 1979 to January 2009) including MeSH terms--"patient preference", "adherence" and "compliance" based on "dosing frequency" and "modality". Since adherence was difficult to accurately quantify, preference, compliance and persistence were evaluated. Patients' preference and adherence at 12 months were higher with weekly over daily bisphosphonates (≥ 84% preference for weekly, medication possession ratios (MPR) 60-76% vs 46-64%; persistence 43.6-69.7% vs 31.7-55.7%). MPR reported for oral bisphosphonates were 68-71% at 12 months. At 2 years, only 43% of patients had MPR ≥ 80% for daily and weekly bisphosphonates. Observational studies (6-12 months) reported discontinuation rates of 18-22% for daily and 7% for weekly bisphosphonates. Data on monthly bisphosphonates are conflicting and confounded by cost differences, patient support programmes and definition of persistence. Studies suggest patient preference for annual zoledronic acid infusions over weekly bisphosphonates (66.4-78.8% vs 9.0-19.7%, respectively), but no data on compliance or persistence are available. Drug effectiveness, side effects and route of administration were more important than frequency. Although less frequent dosing is preferred, other factors such as perceived efficacy, side effects, medication cost, availability of patient support programmes and route of delivery are equally important. Adherence is complex and difficult to quantify and may not be exclusively influenced by frequency of medication administration.

Association between 25-hydroxyvitamin D, somatic muscle weakness and falls risk in end-stage renal failure.

OBJECTIVE: Suboptimal levels of 25-hydroxyvitamin D (25OHD) are common in haemodialysis patients (Chronic Kidney disease-5D: CKD-5D) and may be associated with reduced muscle strength and increased falls risk. We tested the hypothesis that 25OHD levels may be independently associated with falls risk in CKD-5D. BACKGROUND: Supplementation with calcium and cholecalciferol reduces hip and other nonvertebral fractures in elderly individuals, and this effect may in part be attributable to reduction in falls frequency. The relationship between 25OHD and falls risk has not been investigated in CKD-5D. DESIGN AND PATIENTS: This is a cross-sectional study of 25 CKD-5D patients with predialysis 25OHD, 1,25-dihydroxyvitamin D (1,25(OH)(2)D) and intact parathyroid hormone (iPTH) measurement. Falls risk was assessed by quadriceps muscle strength, FallsScreen((c)) test (FST), Berg Balance Scale (BBS), timed 'up and go' (TUG) test, Modified Barthel Index (MBI) and Falls Efficacy Scale (FES). RESULTS: Mean age was 69.8 +/- 12.1 years, and median time on dialysis was 3.1 years. Median 25OHD level was 55.3 nmol/l (range 20.8-125.8 nmol/l). Muscle strength was significantly positively correlated with 25OHD (P = 0.024) but not with 1,25(OH)(2)D (P = 0.477) or PTH (P = 0.461). Statistically significant correlation between 25OHD levels and FST (P = 0.028) plus MBI (P = 0.0046) was noted. No significant correlation was detected between falls risk and 1,25(OH)(2)D or PTH. CONCLUSIONS: Suboptimal levels of 25OHD in CKD-5D are associated with reduced quadriceps muscle strength and increased falls risk. 25OHD may be more important than the active renal metabolite 1,25(OH)(2)D for muscle strength with implications for vitamin D choice and goals of supplementation. Further investigation is required to examine effectiveness of calciferol supplementation on the incidence of falls in CKD-5D.

Developments in multicavity hot runner tooling.

Technological developments in specific areas of multicavity hot runner systems are improving quality control and repeatability and reducing cycle times. Advances such as closed loop filling control of multicavity moulds within the mould itself, servo-actuated valve gates and synchronised valve stems are discussed.

Current assays overestimate 25-hydroxyvitamin D3 and underestimate 25-hydroxyvitamin D2 compared with HPLC: need for assay-specific decision limits and metabolite-specific assays.

BACKGROUND: Clinical demand for quick, cheap, precise and accurate 25-hydroxyvitamin D (25(OH)D) results has led to the development of a variety of assay methods. Lack of standardization of these methods has resulted in inter-method disagreement and challenged whether current assays recognize 25(OH)D2 and 25(OH)D3 equally. METHODS: We studied 172 patient samples from hip fracture cases using DiaSorin (DS) and IDS radioimmunoassays and the Nichols Advantage-automated protein binding assay (NA-CLPBA) in comparison to high-performance liquid chromatography (HPLC). 52 patient samples were analysed before and after three months treatment with 1000 IU of daily ergocalciferol (vitamin D2). RESULTS: Linear regression analysis in pre-treatment samples demonstrated a positive Y-intercept for each immunoassay compared with HPLC, and a slope that varied from 0.64 (IDS) to 0.97 (DS, NA-CLPBA). Bland Altman analysis demonstrated that all the three assays had a proportional positive bias relative to HPLC at values from 20 to 50 nmol/L. Regression analysis of post-treatment samples demonstrated a slope that was not significantly different from zero for the IDS and NA-CLPBA and 0.2 for the DS method, with a positive intercept for all assays of between 8 and 22, indicating less than 50% of 25(OH)D2 measured by HPLC was detected. CONCLUSIONS: These results demonstrate the need for assay-specific decision limits for 25(OH)D3 in order to define appropriate thresholds for treatment institution. Treatment with vitamin D2 may not be accurately monitored with any of the three commercial assays studied. Clinicians and biochemists who continue to use 25(OH)D assays need to be urgently informed of these issues.

Diagnosis of primary hyperparathyroidism: controversies, practical issues and the need for Australian guidelines.

Primary hyperparathyroidism (PHPT) is one of the most common endocrine disease processes, however the clinical presentation in 2003 is typically characterized by minimal signs or symptoms of hypercalcaemia or para-thyroid hormone (PTH) excess. Recent developments in imaging and management of PHPT have been published, however the area of biochemical investigation has been relatively neglected. A group of experts convened in April 2002 to consider whether changes were needed to the 1990 consensus guidelines which defined criteria for the diagnosis and management of asymptomatic PHPT. It is appropriate to review the revised recommendations, which have been disseminated by the panel and were recently published. Each of the laboratory -analytes used to establish the diagnosis of PHPT and exclude alternative diagnoses or complications will be considered in succession in this review: (i) calcium, (ii) intact PTH, (iii) urinary calcium and (iv) 25 hydroxy-vitamin D. Furthermore, critical appraisal of the new diagnostic criteria and their applicability to Australian laboratories will be addressed. Finally, limitations and problems associated with the measurement of each analyte will be reviewed.

Regulation of the 1b isoform of the plasma membrane calcium pump by 1,25-dihydroxyvitamin D3 in rat osteoblast-like cells.

The first isogene of the plasma membrane calcium pump (PMCA1) is expressed on the apical plasma membrane of osteoblasts, but its regulation by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] has not been studied in this cell type. We studied 1,25(OH)2D3 effects on PMCA1 function, protein, messenger RNA (mRNA), and isoform expression in osteoblasts. Of seven rat and human immortalized osteoblast-like cell lines studied, PMCA1 mRNA expression was confirmed in all. Only ROS 17/2.8 cells expressed measurable PMCA1 protein by Western analysis. Immunocytochemistry indicated that PMCA1 was expressed predominantly on the plasma membrane of ROS 17/2.8 cells. The 1,25(OH)2D3 but not 24,25-dihydroxyvitamin D3 [24,25(OH)2D3] treatment of confluent ROS 17/2.8 cells resulted in an approximate 3- to 5-fold dose-dependent increase in PMCA1 expression of message and protein as assessed by Western and Northern analysis and vesicular 45Ca uptake of membrane vesicles. 1,25(OH)2D3 had no effect on PMCA1 posttranscriptional splicing. The 1b isoform of PMCA was expressed under all experimental conditions. 1,25(OH)2D3 favored increased expression of the 5.5 kilobases (kb) over the 7.5-kb PMCA1b transcript, with a 2-fold proportional increase in the smaller transcript relative to the larger transcript evident at the highest dose of 1,25(OH)2D3 studied. The resultant proportional increase in the smaller 5.5-kb transcript may increase mRNA stability and account for the increase in PMCA1b protein and function with 1,25(OH)2D3. These data provide evidence for the role of 1,25(OH)2D3 and PMCA1b in the regulation of calcium transport in bone cells.

The promoter region of the human PMCA1 gene mediates transcriptional downregulation by 1,25-dihydroxyvitamin D(3).

The gene for plasma membrane calcium pump isoform1 (PMCA1) is expressed in calcium-transporting epithelia and bone mesenchymal cells and is upregulated to 1,25-(OH)(2)D(3) in those tissues. A candidate sequence for a vitamin D response element (VDRE) is present within a 1.7-kb promoter region of the human PMCA1 (hPMCA1) gene. We studied hPMCA1 promoter activity in MDBK and ROS 17/2.8 cell lines as PMCA1 mRNA expression is upregulated by 1,25-(OH)(2)D(3) in both. Structural analysis of the putative hPMCA1 VDRE sequence was performed using mobility shift analysis (EMSA) and nuclear extracts from COS-1 cells expressing human VDR (hVDR) and RXRalpha (hRXRalpha). 1,25-(OH)(2)D(3) induced transrepression of the entire 1.7-kb hPMCA1 promoter and of one promoter deletion construct in ROS 17/2.8 cells but not MDBK cells when assayed by luciferase reporter gene assays. Three additional hPMCA1 promoter deletion constructs were unaffected by 1,25-(OH)(2)D(3) in either cell line. While hVDR and hRXRalpha were capable of complexing with a rat osteocalcin DR3 VDRE, EMSA analysis of the potential VDRE from the hPMCA1 gene did not show interaction of either nuclear receptor. Our results indicate tissue-specific sensitivity of the promoter region of the hPMCA1 gene to direct transcriptional downregulation by 1,25-(OH)(2)D(3) and suggest that any positive regulatory VDRE must lie outside of the 1.7-kb core promoter.

Calcitriol upregulates expression and activity of the 1b isoform of the plasma membrane calcium pump in immortalized distal kidney tubular cells.

The plasma membrane calcium pump (PMCA) is ubiquitously expressed in calcium transporting epithelia. PMCA is encoded by four distinct genes (PMCA1-4) which can be further posttranscriptionally modified. PMCA1b is the only isoform of PMCA1 expressed in kidney and intestine. Calcitriol upregulates PMCA protein expression and activity and PMCA1 mRNA expression in the intestine. Calcitriol has a similar effect on kidney distal tubule PMCA activity in vivo but the cellular basis for this effect has not been studied. PMCA expression in Madin-Darby bovine kidney (MDBK) cells, a distal kidney tubule cell line, was compared with a proximal tubule (LLC-PK1) and embryonic (HEK 293) kidney cell line. Only MDBK cells express PMCA1b mRNA and PMCA protein. In MDBK cells, calcitriol increased steady state expression of PMCA1b mRNA and protein and upregulated the functional activity of PMCA on calcium transport to a similar degree. Furthermore, calcitriol enhanced PMCA1b mRNA stability. These data are consistent with in vivo localization studies demonstrating the distal kidney tubule localization of PMCA protein. Furthermore, they indicate that calcitriol is an important regulator of PMCA activity in the kidney distal tubule by a pathway that includes translation and posttranscriptional modification of PMCA1b.

High prevalence of osteoporosis in cardiac transplant recipients and discordance between biochemical turnover markers and bone histomorphometry.

OBJECTIVE: All patients attending the cardiac transplantation clinic at the Royal Perth Hospital were investigated to determine the prevalence of osteoporosis and to assess changes in bone metabolism and histomorphometry in a cohort of cardiac transplant recipients. DESIGN: Retrospective cross-sectional study. PATIENTS: Thirty-two patients (27 male; 5 female) who had received a cardiac transplant during the past 10 years and who were receiving immunosuppressive therapy with cyclosporin, azathioprine and prednisolone were studied. MEASUREMENTS: All patients had bone densitometry by DEXA of the lumbar spine and femoral neck and X-rays of the thoracolumbar spine. Fasting serum ionized calcium, intact PTH, creatinine, 25 hydroxy-vitamin D, alkaline phosphatase, osteocalcin, testosterone and free thyroxine and urine calcium, creatinine, hydroxyproline and deoxypyridinoline were measured. Six osteoporotic patients consented to transiliac bone biopsy following double tetracycline labelling. RESULTS: Osteoporosis was present at the lumbar spine in eight patients, femoral neck in seven patients and was present at one or more sites in 13 patients (41%). Seven patients (22%) had vertebral fractures which were asymptomatic in five patients. Secondary hyperparathyroidism was present in 16 patients (53%) but significant renal failure (creatinine clearance < 70 ml/min) was only found in 8 (50%). Levels of biochemical markers of bone turnover were increased in 23 patients (72%). Serum osteocalcin (P = 0.02) and alkaline phosphatase (P = 0.04) were significantly higher in osteoporotic patients than in nonosteoporotic patients. Histomorphometric findings varied markedly between patients. Microscopic features of hyperparathyroidism were not observed. CONCLUSIONS: Osteoporosis and asymptomatic vertebral fractures are common following cardiac transplantation. Biochemical markers of bone turnover were increased in the majority of patients. Many had biochemical evidence of secondary hyperparathyroidism but this could be attributable to significant renal failure in only 50% of cases. Osteocalcin and alkaline phosphatase correlated inversely with bone density. Histomorphometric findings did not correlate with these biochemical changes in most cases. These results suggest that multiple factors are responsible for osteoporosis in cardiac transplant recipients. Osteocalcin and alkaline phosphatase may be useful biochemical markers, predicting patients at highest risk of fracture.

High prevalence of normal total calcium and intact PTH in 60 patients with proven primary hyperparathyroidism: a challenge to current diagnostic criteria.

BACKGROUND: Others have reported a clear distinction between patients with primary hyperparathyroidism (PHPT) and normal subjects using the intact PTH (iPTH) assay. AIM: We reviewed our last 60 surgically proven cases of PHPT, who had adequate preoperative biochemical assessment, to determine the usefulness of the iPTH assay, ionised calcium and other biochemical criteria in differentiating between normal subjects and patients with PHPT. METHODS: We conducted a retrospective cross-sectional study of all patients with surgically proven PHPT who had been referred to Sir Charles Gairdner Hospital, Perth, Western Australia for preoperative biochemical assessment. All cases had fasting preoperative blood and urine samples collected for ionised calcium, plasma total calcium, albumin, urine calcium excretion, renal phosphate threshold and iPTH. RESULTS: Fifty cases had a single or double adenoma and ten had hyperplasia. All except one had ionised hypercalcaemia but only 47 (78%) had an elevated corrected total calcium (cCa). Therefore 13 cases (22%) had a normal cCa and five of those patients (8%) had both an iPTH and cCa within the reference range. Forty-nine (82%) had an elevated ionised calcium (iCa) and iPTH; the remaining 11 (18%) had an iPTH within the reference range. Of this latter 18%, ten (91%) had a low renal phosphate threshold and five (45%) had significant renal calcium conservation: all 11 cases had at least one abnormality in the renal handling of calcium or phosphate and all normalised their plasma calcium postoperatively (ionised and corrected total calcium). CONCLUSIONS: One in five patients with proven PHPT have a non-elevated cCa and/or intact PTH. Ionised calcium should be measured in all suspected cases. Additional studies of renal calcium and phosphate handling are helpful to establish a diagnosis where any uncertainty exists.

Surgically proven primary hyperparathyroidism with a suppressed intact parathyroid hormone.

A patient with hypercalcaemia had persistently suppressed serum intact parathyroid hormone levels (measured by immunochemiluminometric assay). However, other biochemical tests and open-neck exploration confirmed a diagnosis of primary hyperparathyroidism.