Calcium-sensing receptor mutations in familial benign hypercalcemia and neonatal hyperparathyroidism.

Familial benign hypercalcemia (FBH) and neonatal hyperparathyroidism (NHPT) are disorders of calcium homeostasis that are associated with missense mutations of the calcium-sensing receptor (CaR). We have undertaken studies to characterize such CaR mutations in FBH and NHPT and to explore methods for their more rapid detection. Nine unrelated kindreds (39 affected, 32 unaffected members) with FBH and three unrelated children with sporadic NHPT were investigated for mutations in the 3,234-bp coding region of the CaR gene by DNA sequencing. Six novel heterozygous (one nonsense and five missense) mutations were identified in six of the nine FBH kindreds, and two de novo heterozygous missense mutations and one homozygous frame-shift mutation were identified in the three children with NHPT. Our results expand the phenotypes associated with CaR mutations to include sporadic NHPT. Single-stranded conformational polymorphism analysis was found to be a sensitive and specific mutational screening method that detected > 85% of these CaR gene mutations. The single-stranded conformational polymorphism identification of CaR mutations may help in the distinction of FBH from mild primary hyperparathyroidism which can be clinically difficult. Thus, the results of our study will help to supplement the clinical evaluation of some hypercalcemic patients and to elucidate further the structure-function relationships of the CaR.

Prescribing a website.

AIM: To assess the value of directing the attention of patients to sources of medical information on the internet. DESIGN: Prospective qualitative study in an orthopaedic outpatient clinic. PARTICIPANTS: 253 patients agreed to complete electronic questionnaires before and after reviewing information relevant to their conditions on the internet. Patients were allocated randomly into two groups; one group was given indications of general sites and the other recommended specific non-commercial sites. Completed questionnaires were received from 44 patients. RESULTS: 95% of the patients found the internet information easy to understand and 84% said that it was helpful for coping. 86% of the patients were satisfied that their current treatment was appropriate in the light of what they had learned from the internet. Ten patients out of the 36 who expressed a view thought that the internet information contradicted that provided by the doctor. Despite these results most patients still said that the doctor represented the best source of patient education. CONCLUSIONS: Increasing numbers of patients are familiar with the internet. Most of our patients felt that the internet was, on balance, helpful in providing information. The main difficulties with the internet are the sheer volume of information, the potential for misleading and the danger of misunderstanding. We feel that there is a real place for the specific prescription of an internet site by a clinician who has personally reviewed it to a patient thought to be able to benefit from it.

Novel mutations in the 1alpha-hydroxylase (P450c1) gene in three families with pseudovitamin D-deficiency rickets resulting in loss of functional enzyme activity in blood-derived macrophages.

Pseudovitamin D-defiency rickets (PDDR) is an autosomal recessive disorder characterized by hypocalcemia, rickets (which are resistant to treatment with vitamin D), and low or undetectable serum levels of 1,25-dihydroxyvitamin D (1,25(OH)2D). The symptoms are corrected with 1,25(OH)2D treatment, and the disease is now believed to result from a defect in the cytochrome P450 component (P450c1; CYP27B1) of the renal 25-hydroxyvitamin D-1alpha-hydroxylase (1-OHase). We have studied genomic DNA from three families with PDDR and have identified the same homozygous mutation in the P450c1 gene in two of the index cases, causing a frameshift in exon 8, resulting in a premature stop codon in the heme-binding domain. The two cases in the third kindred were compound heterozygotes with missense mutations in exons 6 and 9. We have also identified a C/T polymorphism in intron 6 of the P450c1 genomic DNA. Interferon gamma-inducible 1-OHase activity in blood-derived macrophages was shown by 1,25(OH)2D synthesis in all control cells tested (37-184 fmol/h/106 cells) and those from the PDDR family parents (34-116 fmol/h/106 cells) but was totally absent from the patients' cells, indicating a defect in their macrophage 1-OHase, similar to the presumed renal defect. The assumption of similarity between the renal and macrophage P450c1 was supported by our ability to clone a 514 bp sequence, including the heme-binding region of the macrophage P450c1 cDNA from controls, which was identical to that published for both the renal and keratinocyte P450c1 cDNAs.

Vitamin D nutrition in relation to season and occupation.

Seasonal variations in vitamin D nutrition were assessed by measurements of serum 25-hydroxycholecalciferol levels in outdoor workers, indoor workers and long-term hospital inpatients. All three groups showed seasonal changes and the outdoor workers had, as might be expected, the highest levels at all seasons. However, the highest levels of 25-hydroxycholecalciferol were found in October in the indoor workers and in November for the outdoor workers whereas the peak in ultraviolet exposure was in July. The possible reasons for this long lag are discussed; the most likely explanation is that vitamin D continues to be formed and stored during the autumn especially in outdoor workers.

Osteogenesis imperfecta: the distinction from child abuse and the recognition of a variant form.

Unexplained fractures are characteristic of both osteogenesis imperfecta (OI) and non-accidental injury (NAI) but in most cases the diagnosis is straightforward. However, in a few OI patients an initial diagnosis of NAI is made. Factors contributing to such difficulties include failure to recognise that OI can occur without a family history, without blue sclerae, without osteopenia, without an excess of Wormian bones, or with metaphyseal fractures. In addition we report on 39 patients with an unusual history in that fractures only occurred in the first year of life. Rib fractures, metaphyseal abnormalities and periosteal reactions were common. The initial diagnosis was usually OI if the fractures occurred in hospital, but NAI if they appeared to have been sustained at home. Additional findings such as anaemia, vomiting, hepatomegaly, and apnoeic attacks were often found in these patients. The disorder has some similarities to the syndrome of infantile copper deficiency. Like the latter it is particularly common in preterm infants and in twins. Therefore, we are attempting to examine the incidence of significant hypocupraemia in unselected preterm infants. We suggest that the likely cause of this "temporary brittle bone disease" is a temporary deficiency of an enzyme, perhaps a metalloenzyme, involved in the post-translational processing of collagen.

Excess paternal age in apparently sporadic osteogenesis imperfecta.

The objective of this study was to examine whether parental age is associated with the occurrence of apparently sporadic osteogenesis imperfecta (OI). We compared parental age and the joint distribution of maternal and paternal age with expected distributions based on statutory birth records for each year and location of birth. The study included patients with OI based in the United Kingdom. The study was restricted to cases born in England, Wales, and Scotland between 1961 and 1998. Subgroup analysis was by clinical type [Sillence et al., 1979: J Med Genet 16:101-116] and apparent mode of inheritance based on pedigree analysis. Of 730 eligible cases, 357 were apparently sporadic. The mean age of fathers at birth of children with apparently sporadic OI was 0.87 years greater than expected (P = 0.010; 95% confidence interval = 0.21 to 1.54 years). The relative risk was 1.62 for fathers in the highest quintile of paternal age compared with fathers in the lowest quintile. The magnitude of the paternal age excess did not differ significantly between Sillence types (analysis of variance P = 0.534). In sporadic cases, paternal age was 0.51 years greater than expected, given maternal age, year, and location of birth (P = 0.033). In contrast, in familial cases, there was no significant paternal age excess, and paternal age was not significantly different from that expected given maternal age. Increased paternal age is a significant risk factor for sporadic OI. This effect is not accounted for by increasing maternal age. The magnitude of the paternal age excess is small in comparison with that in some other autosomal dominant disorders.

Causes of death in osteogenesis imperfecta.

AIMS: To determine the causes of death in patients with osteogenesis imperfecta, excluding infants with the perinatal lethal form (type II). METHODS: Seventy nine patients with known osteogenesis imperfecta were identified, 37 of whom had been seen clinically in life. Causes of death were identified from death certificates, postmortem reports, medical records, hospital consultants, relatives, and the Brittle Bone Society's records. RESULTS: Patients with the milder types of osteogenesis imperfecta, I and IV, often had a normal lifespan and died of unrelated illnesses such as myocardial infarction and malignancy. In some of these patients and in many patients with the more severe type III disease, it was clear that osteogenesis imperfecta contributed significantly to death, almost certainly to many of the respiratory deaths and to deaths from cardiac failure due to kyphoscoliosis. Osteogenesis imperfecta also caused six deaths, directly or indirectly, due to basilar invagination of the skull. Osteogenesis imperfecta may have contributed to deaths from intracranial bleeding. Apparently minor traumatic incidents may have disastrous consequences in patients with this disorder. CONCLUSIONS: Prompt care for respiratory infections and prevention of trauma in patients with osteogenesis imperfecta is essential.

Bone changes in polycythaemia vera and myelosclerosis.

To study the problem of new bone formation in myelosclerosis bone biopsies have been performed in cases of polycythaemia vera, which may be regarded as a pre-myelosclerotic condition, and in primary myelosclerosis; morphological studies of bone and reticulin were made and osteoid and bone matrix was quantitatively assessed. Relevant metabolic studies were also carried out, but were predominantly normal. No new bone formation was detected in polycythaemia vera and some bone was osteoporotic; no significant increase in osteoid was found. With the use of polarized light it was found that new bone formation in myelosclerosis resulted from the ossification of wavy argyrophilic fibres in marrow: the presence of a dense network of these wavy fibres was a pre-requisite of new bone formation and ossification of the non-refractile branched reticulin fibres was not observed.

The value of serum 25-hydroxyvitamin D measurements in hypoparathyroid and pseudohypoparathyroid patients treated with calciferol.

In 23 patients with hypoparathyroidism or pseudohypoparathyroidism treated with vitamin D, and in whom the dosage was adjusted downward or upward in response to hypercalcemia or hypocalcemia respectively, assays of serum 25-hydroxyvitamin D (25-OHD) were carried out in addition to the usual serum calcium assays. In 120 assays there was a significant correlation between serum 25-OHD levels and serum calcium levels (corrected for serum albumin). There was, however, no clear distinction between the 25-OHD levels of patients who were hypocalcemic, normocalcemic or hypercalcemic. The highest serum 25-OHD level found in a hypocalcemic patient was 1193 nmol/L and the lowest serum 25-OHD level found in a hypercalcemic patient was 605 nmol/L. It was not possible to predict subsequent episodes of hypocalcemia or hypercalcemia from the serum 25-OHD levels. The 25-OHD assay was found to be useful only in checking compliance. We conclude that the assay of serum 25-OHD is of no more value than serum calcium alone in the management of compliant patients.

Evaluation of urinary oestrogen assays after the menopause and their potential for screening.

Urinary oestrone-3-glucuronide, oestradiol-3-glucuronide, oestrone and oestradiol were measured by radioimmunoassay methods adapted for the very low levels found in postmenopausal women. Oestrogen concentrations related to creatinine in morning urine samples from ten postmenopausal women were found to correlate well with total oestrogen excreted in 24 h (r = 0.934, 0.867, 0.947, 0.909, respectively; p less than 0.002). Day to day variation in five individuals, measured over 5 weeks, showed random fluctuations, with no obvious cyclical variation. Reference ranges, based on two consecutive morning urine samples from 131 postmenopausal women, were 0.73-4.57 mumol/mol creatinine for oestrone-3-glucuronide, 0.66-3.00 mumol/mol creatinine for oestradiol-3-glucuronide, 4.8-30.9 nmol/mol creatinine for oestrone and 3.8-16.8 nmol/mol creatinine for oestradiol. We suggest that such assays may have a part to play in the screening for women at greatest risk of developing osteoporotic fractures.

Endogenous factors affecting bone mineral content in post-menopausal women.

Eighty-eight healthy post-menopausal women were divided into two groups, one of 35 subjects who had undergone menopause up to 9 years previously and the second of 53 subjects who were 10 or more years post-menopausal. In each individual we related the bone mineral content (BMC), measured by single photon absorptiometry in the distal forearm, to anthropometric variables and urinary oestrogen excretion. There was a positive association between BMC and both urinary oestrogen excretion and anthropometric variables, but this was statistically significant only in the older women. As expected, BMC in the distal forearm decreased with advancing age, the fall being greatest in the first 9 years after the menopause. We concluded that although a single measurement of urinary oestrogen and anthropometric variables does not provide enough information to predict an individual's BMC, the values obtained may prove of use, along with a single BMC determination, in helping to predict the rate of bone loss.

A technique for the tensile testing of demineralised bone.

The purpose of this study was to devise a method for the tensile testing of the organic component of demineralised bone. It was hoped that the method could be applied to determine whether an observed reduction in the strength of bone in copper-deficient rats could be attributed to a reduction in the cross-linking of collagen. Tensile testing of a fully demineralised specimen proved impossible to perform because of the difficulty in gripping the material, which tended to slide out of its holding clamps. The technique devised involved the demineralisation of only the diaphysis of the bone, leaving the bone ends intact. This enabled the bone to be gripped firmly at both ends in an Instron tensile testing machine. Elongation under loading of the demineralised diaphysis was measured by the Instron and a video recording was made of the elongation of the central portion. The strain was found to be uniform. Tensile strength was found to correlate with copper levels in the liver (r = 0.54, p < 0.05), although no significant differences were found between the strength, the stiffness, or the strain to failure for bone from the copper-deficient animals and bone from the normal controls. The test method itself should be useful in other studies in which the tensile properties of the organic phase of bone need to be measured.

Skin content of 7-dehydrocholesterol in patients with malabsorption.

The mechanism for the development of vitamin D deficiency in patients with malabsorption remains unclear. We wished to examine the hypothesis that one factor was a reduced skin content of 7-dehydrocholesterol, the precursor for the formation of vitamin D in the presence of ultraviolet radiation. We measured 7-dehydrocholesterol in skin samples from 9 patients who had previously had vitamin D deficiency due to malabsorption (6 with Crohn's disease, 2 with primary biliary cirrhosis, and 1 with idiopathic pseudo-obstruction). We found no evidence of reduced levels of 7-dehydrocholesterol in the skin in these patients. Lack of 7-dehydrocholesterol does not contribute to vitamin D deficiency in malabsorption.

Evaluation of peripheral dual energy X-ray absorptiometry: comparison with single photon absorptiometry of the forearm and dual energy X-ray absorptiometry of the spine or femur.

An evaluation of the Osteoscan peripheral dual energy X-ray absorptiometer (pDXA) was carried out to compare its performance with those of a single photon absorptiometer (SPA) (Molsgaard Medical ND1100A) and a dual energy X-ray absorptiometer (DXA) (Lunar DPX alpha) of the spine or femur. In 57 patients, correlation between bone mineral content (BMC) of the forearm at the ultradistal (UD) site by pDXA and by SPA was high (r = 0.94). Comparisons were also made with spine and femur bone mineral density (BMD) DXA measurements. The correlation of z-scores of UD BMD with z-scores for lumbar spine L2-L4 was r = 0.63 (n = 73 patients); and with z-scores for neck of femur was r = 0.72 (n = 33). With the Osteoscan the measurement error coefficient of variation in vivo was 2.6% for BMC, 1.8% for BMD at the ultradistal site; 2.1% for BMC and 1.9% for BMD at the mid-distal site. Repeat measurements were made of the European forearm phantom; precision for SPA was slightly better than either pDXA or Lunar DXA. The Osteoscan has the potential for a rapid throughput of patients and is not affected by calcification and degenerative changes that can corrupt DXA measurements on the anteroposterior spine in older women.

Osteogenesis imperfecta with dominant inheritance and normal sclerae.

Most patients with dominantly inherited osteogenesis imperfecta have blue sclerae and relatively mild symptoms. However, in a small group of families the patients have normal sclerae and this disorder has been classified as Type 4 osteogenesis imperfecta. This paper reports the clinical and radiographical features of 48 patients from 16 families with Type 4 osteogenesis imperfecta and compares the findings with those of the classical disorder with blue sclerae (Type 1 osteogenesis imperfecta). The two types are similar in usually causing a mild disease but with a wide range of severity, and in both types the rate of fracture declines in adolescence. There are, however, some significant differences apart from the colour of the sclerae. In Type 4 the first fracture more commonly occurs at birth, dentinogenesis imperfecta is more frequent than in Type 1 and bruising and nose-bleeds are less common. As in Type 1, the radiographic appearances of the bones may be normal. It is important that Type 4 osteogenesis imperfecta should be recognised because of the need for competent genetic counselling, because the management may be different from that appropriate for Type 1 and because it may be mistaken for idiopathic juvenile osteoporosis or child abuse.

Peripheral bone mineral density in patients with distal radial fractures.

Fractures of the distal forearm are widely regarded as the result of "fragility". We have examined the extent to which patients with Colles' fractures have osteopenia. We measured the bone mineral density (BMD) in the contralateral radius of 235 women presenting with Colles' fractures over a period of two years. While women of all ages had low values for ultra-distal BMD, the values, in age-matched terms, were particularly low among premenopausal women aged less than 45 years. This result was not due to the presence of women with an early menopause. This large survey confirms and extends the findings from earlier small studies. We consider that it is particularly important to investigate young patients with fractures of the distal forearm to identify those with osteoporosis, to seek an underlying cause and to consider treatment.

Bone mineral density of the radius in patients with Colles' fracture.

To ascertain whether patients with Colles' fracture should be investigated for osteoporosis and the risk of future fractures, we measured the bone mineral density of the distal radius of the other arm in 31 women patients and compared the results with those of a control group of 289 normal women. We divided the patients into two groups, those younger than 66 years and those older. In 25 patients we found values for bone mineral density which were lower than one standard deviation below the mean value for their age. Younger patients had a deficit greater than that expected for their ages. We believe that women with Colles' fracture should be evaluated routinely for osteoporosis, particularly if they are under 66 years of age.

Lindau's disease and familial hyperparathyroidism.

A case of medullary hemangioblastoma is described occurring in association with familial hyperparathyroidism. The possibility that this combination may represent a previously unreported variant of Lindau's disease is discussed.

Influence of injection site for low-dose heparin on wound complication rates after inguinal hernia repair.

A high incidence of complications related to bleeding was observed after open prosthetic inguinal hernia repair. The site of injection of low-dose heparin into the abdominal wall was thought to be a possible causal factor for these complications. The wound complication rate after repair of primary unilateral inguinal hernias was recorded for 51 patients who had been given abdominal wall injections of heparin. Subsequently the injection site was changed to the upper limb in a further 63 patients and the incidence of wound complications recorded. A significantly higher incidence of haematomas and seromas was found in the abdominal wall injection group (39.2% vs 17.5%, P = 0.01). The role of low-dose heparin prophylaxis in inguinal hernia repair is discussed. We conclude that in those patients receiving heparin prophylaxis the injections should be given at a site remote from the operative area.

How common is hearing impairment in osteogenesis imperfecta?

Hearing impairment has long been recognized as a common feature in osteogenesis imperfecta. The figures in some publications could be taken to imply that, with increasing age, the proportion of osteogenesis imperfecta patients with hearing impairment approaches 100 per cent. The incidence of hearing loss in a large survey of 1394 patients with osteogenesis imperfecta was examined. It was found that the most common age of onset was in the second, third and fourth decades of life. At the age of 50 approximately 50 per cent of the patients had symptoms of hearing impairment; over the next 20 years there was little further increase. Differences were shown between patients with different clinical types of osteogenesis imperfecta as delineated in the Sillence classification; hearing loss was significantly less common in the type IV disease than in the type I disorder. Among the 29 families with osteogenesis imperfecta type IA there were distinct differences in the likelihood of hearing loss. These findings provide insights which will be valuable in giving patients advice on the likelihood of developing hearing loss in the future.