UK national chronic hypoparathyroidism audit.

OBJECTIVES: Individuals with chronic hypoparathyroidism may experience suboptimal medical care with high frequency of unplanned hospitalisation and iatrogenic harm. In 2015 the European Society for Endocrinology published consensus guidelines on the management of chronic hypoparathyroidism. We set out to audit compliance with these guidelines. METHODS: Using these recommendations as audit standards we worked with the Society for Endocrinology and Parathyroid UK to conduct a national audit of management of chronic hypoparathyroidism in the United Kingdom. Endocrine leads in 117 endocrine departments were invited to participate in the survey by completing a data collection tool on up to 5 sequential cases of chronic hypoparathyroidism seen in their outpatient clinics in the preceding 12 months. Data were collected on 4 treatment standards and 9 monitoring standards. Data on hospitalisations and Quality of Life monitoring were also collected. RESULTS: Responses were received from 22 departments giving a response rate of 19%, concerning 80 individual cases. The mean age of subjects was 48.4 years. The main findings were that the commonest cause of hypoparathyroidism was post surgical (66.3%). Treatments taken by the group included activated vitamin D analogues (96.3%), oral calcium salts (66.3%), vitamin D supplements (17.5%), thiazide diuretics (5%) and rhPTH1-34 (1.3%). Compliance with the audit standards varied between 98.8% and 60% for the treatment standards and between 91.3% and 20% for the monitoring standards. Some of the areas of weakness revealed include low rates of 24 h urinary calcium excretion monitoring, serum magnesium monitoring and low rates of renal imaging where indicated. In addition and importantly, 16.3% of subjects had experienced at least one hospital admission in the preceding 12 months. CONCLUSION: We conclude that further improvements in the UK national standard of management of chronic hypoparathyroidism should be made and that this will benefit both quality of life, morbidity and potentially mortality in this group of patients.

Constitutive P2Y2 receptor activity regulates basal lipolysis in human adipocytes.

White adipocytes are key regulators of metabolic homeostasis, which release stored energy as free fatty acids via lipolysis. Adipocytes possess both basal and stimulated lipolytic capacity, but limited information exists regarding the molecular mechanisms that regulate basal lipolysis. Here, we describe a mechanism whereby autocrine purinergic signalling and constitutive P2Y2 receptor activation suppresses basal lipolysis in primary human in vitro-differentiated adipocytes. We found that human adipocytes possess cytoplasmic Ca2+ tone due to ATP secretion and constitutive P2Y2 receptor activation. Pharmacological antagonism or knockdown of P2Y2 receptors increases intracellular cAMP levels and enhances basal lipolysis. P2Y2 receptor antagonism works synergistically with phosphodiesterase inhibitors in elevating basal lipolysis, but is dependent upon adenylate cyclase activity. Mechanistically, we suggest that the increased Ca2+ tone exerts an anti-lipolytic effect by suppression of Ca2+-sensitive adenylate cyclase isoforms. We also observed that acute enhancement of basal lipolysis following P2Y2 receptor antagonism alters the profile of secreted adipokines leading to longer-term adaptive decreases in basal lipolysis. Our findings demonstrate that basal lipolysis and adipokine secretion are controlled by autocrine purinergic signalling in human adipocytes.

Identification of 70 calcium-sensing receptor mutations in hyper- and hypo-calcaemic patients: evidence for clustering of extracellular domain mutations at calcium-binding sites.

The calcium-sensing receptor (CaSR) is a G-protein-coupled receptor that has an extracellular bilobed venus flytrap domain (VFTD) predicted to contain five calcium (Ca(2+))-binding sites. To elucidate the structure-function relationships of the VFTD, we investigated 294 unrelated probands with familial hypocalciuric hypercalcaemia (FHH), neonatal severe primary hyperparathyroidism (NSHPT) or autosomal dominant hypocalcaemic hypercalciuria (ADHH) for CaSR mutations and performed in vitro functional expression studies and three-dimensional modelling of mutations involving the VFTD. A total of 70 different CaSR mutations were identified: 35 in FHH, 10 in NSHPT and 25 in ADHH patients. Furthermore, a CaSR variant (Glu250Lys) was identified in FHH and ADHH probands and demonstrated to represent a functionally neutral polymorphism. NSHPT was associated with a large proportion of truncating CaSR mutations that occurred in the homozygous or compound heterozygous state. Thirty-four VFTD missense mutations were identified, and 18 mutations were located within 10 Å of one or more of the predicted Ca(2+)-binding sites, particularly at the VFTD cleft, which is the principal site of Ca(2+) binding. Mutations of residues 173 and 221, which are located at the entrance to the VFTD cleft binding site, were associated with both receptor activation (Leu173Phe and Pro221Leu) and inactivation (Leu173Pro and Pro221Gln), thereby highlighting the importance of these residues for entry and binding of Ca(2+) by the CaSR. Thus, these studies of disease-associated CaSR mutations have further elucidated the role of the VFTD cleft region in Ca(2+) binding and the function of the CaSR.

Nonsteroidal anti-inflammatory drugs increase TNF production in rheumatoid synovial membrane cultures and whole blood.

Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase activity and hence PG production. However, the ability of NSAIDs to ameliorate pain and tenderness does not prevent disease progression in rheumatoid arthritis, a disease whose pathogenesis is linked to the presence of proinflammatory cytokines, such as TNF-alpha. To understand this observation, we have examined the effect of NSAIDs on the production of clinically validated proinflammatory cytokines. We show that a variety of NSAIDs superinduce production of TNF from human peripheral blood monocytes and rheumatoid synovial membrane cultures. A randomized, double-blinded, crossover, placebo-controlled trial in healthy human volunteers also revealed that the NSAID drug celecoxib increased LPS-induced TNF production in whole blood. NSAID-mediated increases in TNF are reversed by either the addition of exogenous PGE(2) or by a PGE(2) EP2 receptor agonist, revealing that PGE(2) signaling via its EP2 receptor provides a valuable mechanism for controlling excess TNF production. Thus, by reducing the level of PGE(2), NSAIDs can increase TNF production and may exacerbate the proinflammatory environment both within the rheumatoid arthritis joint and the systemic environment.

Diagnostic challenges due to phenocopies: lessons from Multiple Endocrine Neoplasia type1 (MEN1).

Phenocopies may confound the clinical diagnoses of hereditary disorders. We report phenocopies in Multiple Endocrine Neoplasia type 1 (MEN1), an autosomal dominant disorder, characterised by the combined occurrence of parathyroid, pituitary and pancreatic tumours. We studied 261 affected individuals from 74 families referred with a clinical diagnosis of MEN1 and sought inconsistencies between the mutational and clinical data. We identified four patients from unrelated families with phenocopies. Patients 1 and 2 from families with MEN1, developed prolactinomas as the sole endocrinopathy but they did not harbour the germline MEN1 mutation present in their affected relatives. Patient 3, had acromegaly and recurrent hypercalcaemia following parathyroidectomy, whilst patient 4 had parathyroid tumours and a microprolactinoma. Patients 3 and 4 and their relatives did not have MEN1 mutations, but instead had familial hypocalciuric hypercalcaemia (FHH) due to a calcium-sensing receptor mutation (p.Arg680Cys), and the hyperparathyroidism-jaw tumour (HPT-JT) syndrome due to a hyperparathyroidism type 2 deletional-frameshift mutation (c.1239delA), respectively. Phenocopies may mimic MEN1 either by occurrence of a single sporadic endocrine tumour in a patient with familial MEN1, or occurrence of two endocrine abnormalities associated with different aetiologies. Phenocopies arose in >5% of MEN1 families, and awareness of them is important in the clinical management of MEN1 and other hereditary disorders.

Hypercalcaemia - presentation and management .

Hypercalcaemia is a common disorder normally caused by primary hyperparathyroidism (PHPT) or malignancy. A proportion of cases present as an emergency, which carries a significant mortality. Emergency management of hypercalcaemia is based on intravenous rehydration with normal saline but when this is inadequate, bisphosphonate therapy is used; more recently the novel anti-resorbtive agent denosumab has been shown to have a useful role in treatment. It is estimated that up to 10% of all cases of PHPT presenting under the age of 45 years have an underlying genetic predisposition; nine potentially causative genes are now recognised and may be screened in routine clinical practice. Although parathyroidectomy is the only curative treatment for PHPT, this is indicated in a minority of cases. Many cases can be adequately managed conservatively and guidance from the 4th international workshop on the management of asymptomatic PHPT has recently been updated in a consensus statement.

Genetic mapping studies of familial juvenile hyperuricemic nephropathy on chromosome 16p11-p13.

Familial juvenile hyperuricemic nephropathy (FJHN), which is inherited as an autosomal dominant disorder, is characterized by hyperuricemia, a low fractional renal excretion of urate, and chronic renal failure that is associated with interstitial fibrosis. Studies in 4 families (3 European and 1 Japanese) have mapped the gene causing autosomal dominant FJHN to chromosome 16p11-p13. To refine this location we have pursued linkage studies in 7 European families with autosomal dominant FJHN and used 11 chromosome 16p11-p13 polymorphic loci whose order has been established as 16pter-D16S3069-D16S3060-D16S3041-D16S3036-D16S3046-[D16S403,D16S417]-D16S420-D16S3113-D16S401-D16S3133-16cen. Cosegregation between these polymorphic loci and FJHN was observed in 5 of the families, and linkage was established between FJHN and 6 loci (peak LOD score, 5.32 with D16S417, at 0% recombination), with the most likely location of FJHN being within a 22-centimorgan interval flanked centromerically by D16S401 and telomerically by D16S3069. Furthermore, FJHN in 2 families was found not to be linked to chromosome 16p11-p13, thereby demonstrating genetic heterogeneity. Thus, 5 additional families with FJHN showing linkage to chromosome 16p11-p13 loci have been identified, and genetic heterogeneity has been demonstrated in more than 25% of FJHN families. These results will facilitate the characterization of this gene regulating urate metabolism.

Frequent occurrence of an intron 4 mutation in multiple endocrine neoplasia type 1.

MEN1 is an autosomal dominant disorder characterized by parathyroid, pituitary, and pancreatic tumors. The MEN1 gene is located on chromosome 11q13 and encodes a 610-amino acid protein. MEN1 mutations are of diverse types and are scattered throughout the coding region, such that almost every MEN1 family will have its individual mutation. To further characterize such mutations we ascertained 34 unrelated MEN1 probands and undertook DNA sequence analysis. This identified 17 different mutations in 24 probands (2 nonsense, 2 missense, 2 in-frame deletions, 5 frameshift deletions, 1 frameshift deletional-insertion, 3 frameshift insertions, 1 donor splice site mutation, and a g-->a transition that resulted in a novel acceptor splice site in intron 4). The intron 4 mutation was found in 7 unrelated families, and the tumors in these families varied considerably, indicating a lack of genotype-phenotype correlation. However, this intron 4 mutation is the most frequently occurring germline MEN1 mutation ( approximately 10% of all mutations), and together with 5 others at codons 83-84, 118-119, 209-211, 418, and 516, accounts for 36.6% of all mutations, a finding that indicates an approach for identifying the widely diverse MEN1 mutations.

Comparison of human chromosome 19q13 and syntenic region on mouse chromosome 7 reveals absence, in man, of 11.6 Mb containing four mouse calcium-sensing receptor-related sequences: relevance to familial benign hypocalciuric hypercalcaemia type 3.

Familial benign hypocalciuric hypercalcaemia (FBHH) is a genetically heterogeneous disorder that consists of three designated types, FBHH1, FBHH2 and FBHH3, whose chromosomal locations are 3q21.1, 19p and 19q13, respectively. FBHH1 is caused by mutations of a calcium-sensing receptor (CaSR), but the abnormalities underlying FBHH2 and FBHH3 are unknown. FBHH3, also referred to as the Oklahoma variant (FBHH(Ok)), has been mapped to a 12cM interval, flanked by D19S908 and D19S866. To refine the location of FBHH3, we pursued linkage studies using 24 polymorphic loci. Our results establish a linkage between FBHH3 and 17 of these loci, and indicate that FBHH3 is located in a 4.1 Mb region flanked centromerically by D19S112 and telomerically by rs245111, which in the syntenic region on mouse chromosome 7 contains four Casr-related sequences (Gprc2a-rss). However, human homologues of these Gprc2a-rss were not found and a comparative analysis of the 22.0 Mb human and 39.3 Mb mouse syntenic regions showed evolutionary conservation of two segments that were inverted with loss from the human genome of 11.6 Mb that contained the four Gprc2a-rss. Thus, FBHH3 cannot be attributed to Gprc2a-rss abnormalities. DNA sequence analysis of 12 other genes from the interval that were expressed in the parathyroids and/or kidneys did not detect any abnormalities, thereby indicating that these genes are unlikely to be the cause of FBHH3. The results of this study have refined the map location of FBHH3, which will facilitate the identification of another CaSR or a mediator of calcium homeostasis.

Assessing glycemic control in maintenance hemodialysis patients with type 2 diabetes.

OBJECTIVE: Optimizing glycemic control in diabetic patients undergoing maintenance hemodialysis requires accurate assessment. We hypothesize that 1) 48-h continuous glucose monitoring (CGM) provides additional, clinically relevant, information to that provided by the A1C measurement and 2) glycemic profiles differ significantly between day on and day off dialysis. RESEARCH DESIGN AND METHODS: With the use of GlucoDay S, 48-h CGM was performed in 19 type 2 diabetic subjects undergoing hemodialysis to capture consecutive 24-h periods on and off dialysis. Energy intake was calculated using food diaries. A1C was assayed by a high-performance liquid chromatography method. RESULTS: CGM data were available for 17 subjects (13 male) with a mean (range) age of 61.5 years (42-79 years) and diabetes duration of 18.8 years (4-30 years). The 24-h CGM area under the glucose curve and 24-h mean glucose values were significantly higher during the day off dialysis than on dialysis (5,932.1 +/- 2,673.6 vs. 4,694 +/- 1,988.0 mmol x 3 min(-1) x l(-1), P = 0.022, and 12.6 +/- 5.6 vs. 9.8 +/- 3.8 mmol/l, P = 0.013, respectively), independent of energy intake. Asymptomatic hypoglycemia occurred in 4 subjects, 3 within 24 h of dialysis, and the glucose nadir in 14 subjects occurred within 24 h of dialysis. CONCLUSIONS: Glucose values are significantly lower on dialysis days than on nondialysis days despite similar energy intake. The risk of asymptomatic hypoglycemia was highest within 24 h of dialysis. Physicians caring for patients undergoing hemodialysis need to be aware of this phenomenon and consider enhanced glycemic monitoring after a hemodialysis session. CGM provides glycemic information in addition to A1C, which is potentially relevant to clinical management.

Functional characterization of calcium sensing receptor polymorphisms and absence of association with indices of calcium homeostasis and bone mineral density.

OBJECTIVES: Associations between calcium-sensing receptor (CaSR) polymorphisms and serum calcium, PTH and bone mineral density (BMD) have been reported by six studies. However, three other studies have failed to detect such associations. We therefore further investigated three CaSR coding region polymorphisms (Ala986Ser, Arg990Gly and Gln1011Glu) for associations with indices of calcium homeostasis and BMD and for alterations in receptor function. PATIENTS AND DESIGN: One hundred and ten adult, Caucasian, female, dizygotic twin pairs were investigated for associations between the three CaSR polymorphisms and serum calcium, albumin, PTH, 25-hydroxyvitamin D(3) (25OHD(3)), 1,25-dihydroxyvitamin D(3)[1,25(OH)(2)D(3)], urinary calcium excretion and BMD. Each polymorphic CaSR was also transfected into HEK293 cells and functionally evaluated. RESULTS: There was a lack of association between each of these three CaSR polymorphisms and serum calcium corrected for albumin, PTH, 25OHD(3), 1,25(OH)(2)D(3), urinary calcium excretion or BMD at the hip, forearm and lumbar spine. These findings were supported by a lack of functional differences in the dose-response curves of the CaSR variants, with the EC(50) values (mean +/- SEM) of the wild-type (Ala986/Arg990/Gln1011), Ser986, Gly990 and Glu1011 CaSR variants being 2.74 +/- 0.29 mm, 3.09 +/- 0.34 mm (P > 0.4), 2.99 +/- 0.23 mm (P > 0.4) and 2.96 +/- 0.30 mm (P > 0.5), respectively. CONCLUSIONS: Our study, which was sufficiently powered to detect effects that would explain up to 5%, but not less than 1%, of the variance has revealed that the three CaSR polymorphisms of the coding region have no major influence on indices of calcium homeostasis in this female population, and that they do not alter receptor function.