Increased upstream methylation has no influence on the overexpression of the parathyroid hormone-related protein gene in squamous cell carcinoma of the lung.

Humoral hypercalcaemia of malignancy (HHM) commonly results from the excessive production of a parathyroid hormone-related protein (PTHrP) by tumours. We have previously shown malignancy is associated with increased DNA methylation in the 5' region of the PTHrP gene. In a series of patients with lung carcinoma and relatively high serum calcium levels, 3 patients showed substantially increased PTHrP gene methylation while 5 patients showed no change in methylation status in this region. Patients showed marked tumour-specific expression of PTHrP through the P1 and P3 promoters with more general tumour and non-tumour expression through the P2 promoter. The lack of potential key regulatory CpG sites in the P1 promoter and the complete demethylation in the P2 and P3 promoters suggests methylation does not influence tumour-specific expression of PTHrP. Although demethylation may be a prerequisite for P2 and P3 expression, the overexpression of the PTHrP gene in cancer cells must be mediated through mechanisms other than DNA methylation.

Mobility and dementia: is physiotherapy treatment during respite care effective?

BACKGROUND: Mobility problems experienced by elderly people with a dementia are associated with falls, fractures and admission to long-term care. A hospital respite care admission is therefore often seen as an opportunity to provide physiotherapy treatment. AIM: To find whether elderly people with a dementia and a mobility problem show a greater improvement in mobility skills if given physiotherapy treatment than if given non-physical activities intervention during a hospital respite admission. METHOD: A controlled randomized multicentre trial with independent blinded assessment. The Southampton Mobility Assessment (mobility score) and Two Minute Walking Test (distance walked) were undertaken at the beginning and end of the study admission and beginning of the next respite admission. Following the first assessment, participants were randomized to either physiotherapy or activities. RESULTS: Eighty-one participants, from 12 clinical centres, with a mean age of 81.9 years and CAPE I/O score of 2. During the study admission there was a non-significant trend for a lower reduction in mobility score of the physiotherapy group (Mann-Whitney; p = 0.614) and a non-significant trend for greater decrease in distance walked in the activities group (t-test; p = 0.325). DISCUSSION: The results of this trial do not support the positive changes demonstrated elsewhere. However, changes in respite care during the early stages of this trial may have produced differences between the sample for this trial and that for the pilot study. This trial was therefore underpowered. CONCLUSION: This trial suggests that future research needs to change the focus from clinical settings to presentations.

CpG island methylation and promoter usage in the parathyroid hormone-related protein gene of cultured lung cells.

Excessive production of a parathyroid hormone-related protein (PTHrP) by tumours commonly results in the syndrome of humoral hypercalcaemia of malignancy. We have investigated whether epigenetic changes play a role in over-expression of the PTHrP gene, using cultures lung cells as a model system. Study of the methylation status of CpG dinucleotides in the 5' region of the gene showed that in normal cells the CpG island was completely unmethylated. In the lung squamous cell carcinoma cell line, BEN, two-thirds of the CpG island was substantially methylated. RT-PCR analysis showed that this heavy methylation did not prevent expression of any of the three PTHrP gene promoters. This is a surprising finding, since methylation is usually associated with inhibition of gene activity. Methylation of the 5' non-coding region of the PTHrP gene may not play a role in the regulation of adjacent promoters. Alternatively, maintenance of a demethylated state in the 170 bp at the 3' end of the CpG island may be fundamental for the use of PTHrP promoters.

Patterns of DNA methylation of the parathyroid hormone-related protein gene in human lung carcinoma.

Humoral hypercalcaemia of malignancy often results from production of parathyroid hormone-related protein (PTHrP) by the tumour. We have investigated whether malignancy is associated with epigenetic changes in the PTHrP gene in lung. In normal and tumour tissue, there was a general background of nonmethylation in the PTHrP gene. In the 5' region, there appeared to be increased methylation of sites upstream of the promoter, P2. The extent of methylation increased from germ line to normal tissue to tumour tissue to tumour cell line, indicating that new methylation events in this region mark neoplastic change in lung cells.

NMDA receptor status in elderly normal individuals and those with Alzheimer's disease.

The status of NMDA receptors in the brains of normal aged individuals and those with Alzheimer's disease was investigated. The binding of [3H]3-((+-)-2-carboxypiperazin-4-yl)propyl phosphonic acid ([3H]CPP) to NMDA antagonist-preferring sites on frontal and temporal cortical synaptic membranes was assessed. Binding could be resolved into two components, one of high and the other of low affinity. Pharmacologically, the two sites were qualitatively similar. Considerable intersubject variation in binding parameters was detected, but no significant differences were found between the mean values for the control and Alzheimer's disease groups. This study indicates that, when changes in receptor integrity occur in individual patients, these may be occluded because of the large variations between individuals.

Energy balance in healthy elderly women.

There is a paucity of data on which to base estimates of the energy requirements of the elderly. In general, ageing appears to be associated with a reduction in energy requirement arising from a reduction in physical activity and loss of fat-free mass. The aim of the present study was to measure the total energy expenditure (TEE), basal metabolic rate (BMR), and energy expended on physical activity (calculated as TEE-BMR) in a group of healthy elderly women living in the community in Southampton. Mean rates of TEE (9.21 (SD 1.48) MJ/d) and energy expended on physical activity (4.12 (SD 1.19) MJ/d) were higher than those observed in some studies of younger adults in the UK, and higher than the factors used to estimate the average energy requirement for the elderly. The results suggest that an age-related reduction in energy requirement is not inevitable and support the hypothesis that the effects of ageing on physical activity, body composition, and hence energy requirements, are variable.

L-aspartate-beta-hydroxamate exhibits mixed agonist/antagonist activity at the glutamate metabotropic receptor in rat neonatal cerebrocortial slices.

L-aspartate-beta-hydroxamate, a glutamate uptake inhibitor, was investigated for activity at a glutamate metabotropic receptor (mGluR) in neonatal rat cerebral cortical slices. Stimulation of phosphatidylinositol hydrolysis by 100 microM (1S,3R)-ACPD was inhibited only very weakly, to a maximal extent of 28%, L-aspartate-beta-hydroxamate did however exhibit agonist activity (EC50 = 760 microM) and, although much less potent than (1S,3R)-ACPD (EC50 = 20 microM), its efficacy was approximately 70% of the latter. These results indicate that, at least in this preparation, offspartate-beta-hydroxamate is of little value as an antagonist at the mGluR receptor.

Modulation of [3H]3-((+-)-2-carboxypiperazin-4-yl)propyl-1-phosphonic acid ([3H]CPP) binding by ligands acting at the glycine and the polyamine sites of the rat brain NMDA receptor complex.

The competitive N-methyl-D-aspartate (NMDA) receptor antagonist [3H]3-((+-)-2-carboxypiperazin-4-yl)propyl-1-phosphonic acid ([3H]CPP) interacts with two discrete binding sites, one of high- and the other of low-affinity, on rat forebrain synaptic plasma membranes. The high affinity site exhibited a Kd of 101.5 nM and a Bmax of 2.01 pmol/mg, while for the low affinity site the Kd was 4.11 microM with a Bmax of 19.7 pmol/mg. The glycine site antagonists 3-amino-1-hydroxy-2-pyrrolidone (HA-966), 1-aminocyclobutanecarboxylic acid (ACBC), the glycine site agonist 1-aminocyclopropanecarboxylic acid (ACC) and glycine itself (as well as the polyamines spermine and spermidine), enhanced [3H]CPP binding. When subjected to saturation analysis, this enhancement was found primarily to involve an increase in the affinity of the high affinity component of [3H]CPP binding. Neither of the parameters of the low affinity component of binding were affected. Although a similar enhancement was observed with the polyamines, the effects of these two classes of ligands were additive, consistent with their having actions at different recognition sites on the NMDA receptor complex.

Heterogeneity of NMDA receptors labelled with [3H]3-((+-)-2-carboxypiperazin-4-yl) propyl-1-phosphonic acid ([3H]CPP): receptor status in Alzheimer's disease brains.

The binding of [3H]3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid ([3H]CPP) was studied in rat and human brain synaptic membranes. Specific binding was saturable, reversible and inhibited by a range of compounds active at N-methyl-D-aspartate (NMDA) receptors such as 2-amino-5-phosphonopentanoate (AP5), 2-amino-7-phosphonoheptanoate (AP7), NMDA and cis-2,4-methanoglutamate. Binding was heterogeneous as evidenced by non-linear Scatchard plots and Hill coefficients for binding inhibitors significantly different from unity. LIGAND analysis of the binding data indicated the likely presence of two distinct binding components for CPP, one of high (Kd values approx. = 70 nM) and the other of low (Kd values approx. 5 microM) affinity. Possible alterations in the binding of [3H]CPP to either site were investigated in medial frontal and medial temporal cortex from Alzheimer's disease brains and compared with control tissues, carefully matched for age and postmortem delay. While there were considerable inter-individual variations in binding, no significant differences were detected either between brain regions in either Alzheimer or control subjects, or between Alzheimer's disease and control brains. These data suggest the presence of at least two components of [3H]CPP binding in both rat and human brain tissue. The integrity of neither of these components is altered in Alzheimer's disease, consistent with a lack of gross alterations of NMDA receptors in this disorder.

Raised blood urea in the elderly: a clinical and pathological study.

We have attempted to define a normal range for blood urea and creatinine for elderly inpatients and to determine the relative importance of pre-renal, renal and post-renal pathology in those with renal impairment. A total of 118 admissions to an acute geriatric unit and 67 separate post mortems in patients over 67 years of age were studied prospectively. Up to 123 items of data were coded and analysed including blood urea and creatinine, clinical or pathological changes associated with renal disease, clinical outcome and post mortem findings. We determined our own 'normal' hospital ranges for urea (1.4-13.2 mmol/l) and creatinine (48-141 mumol/l) from plasma values in 76 patients with no evidence of renal impairment, either on admission or in the past. Using these values 41% of post mortem cases and 25% of clinical admissions had a raised blood urea. Pre-renal conditions such as cardiac failure, dehydration and gastrointestinal haemorrhage, either alone or in combination, were present in 56% of these patients. Urea and creatinine values were substantially higher in patients who died in hospital as opposed to those who were discharged or transferred. Creatinine values were greater in those with intrinsic renal disease or post-renal obstruction as compared to patients with pre-renal causes of renal impairment. Patients with histological evidence of extensive glomerulosclerosis or nephrosclerosis had higher urea and creatinine levels than those with only minor ageing changes.

Orthopaedic geriatric care--is it effective? A prospective population-based comparison of outcome in fractured neck of femur.

Orthopaedic geriatric care has become widely accepted despite relatively little formal evaluation. In the East Dorset health authority all patients with fractured neck of femur initially share common orthopaedic facilities but only those from one geographical sector have subsequent access to an orthopaedic geriatric unit, patients from the other sector receiving standard care. We have carried out a prospective population-based comparison of the outcome of 155 consecutive incident cases of fractured neck of femur aged over 65 years. On admission to hospital the two populations were similar in respect of age, sex, fracture type, social status, medical and orthopaedic problems, mental status and dependency (Clifton assessment procedure for the elderly). Multiple regression analysis showed that the mean length of stay was 9.5 days shorter in patients from the sector with access to orthopaedic geriatric care (p less than 0.05, 95% confidence interval 0.6 to 18.4 days). This reduction was not due to any difference between the two groups as regards dependency or the level of support provided on discharge. There was no difference in outcome at 6 months post-operatively in terms of mortality, functional outcome (pain and mobility), change in dependency or social status. We conclude that in the East Dorset health authority this combined approach has resulted in a significant reduction in the length of inpatient stay without any other apparent effect on patient outcome.

Characterisation, density, and distribution of kainate receptors in normal and Alzheimer's diseased human brain.

The specific binding of [3H]kainic acid was investigated in membrane preparations from human parietal cortex obtained postmortem. Saturation studies revealed that binding occurred to a single population of sites with a KD of 15 nM and a Bmax of 110 fmol/mg of protein. The kinetically determined dissociation constant for these sites agreed well with that obtained from saturation analyses. Pharmacological characterisation of these sites gave a profile consistent with those reported for kainate receptor sites in animal brain. The integrity of kainate receptors was studied in several brain regions from six patients who had died of Alzheimer's disease and from six closely matched control subjects. No change in either the affinity or the number of kainate receptors was seen in any of the regions studied, despite the loss of neocortical and hippocampal glutamatergic terminals in the Alzheimer's diseased brains, as previously reported.

Osteoporosis, falls, and age in fracture of the proximal femur.

The relative importance of osteoporosis and risk of falling in causing hip fracture is not known. Femoral neck bone mass was measured in 708 elderly people, all of whom had fallen and injured a hip. Below 75 years of age there was a steep increase in relative risk of fracture with reduced bone mass, measured by Singh grade. Above that age the increase in risk was small, and neuromuscular responses which protect the skeleton against trauma may be more important than bone mass.

Does radiolabelled diphosphonate retention reflect bone metabolism in the elderly?

The 24-h whole-body retention of diphosphonates (WBR), a useful measure of bone metabolism in young subjects, was measured in 27 elderly subjects (19 from home and eight in-patients). WBR rose with increasing age (r = 0.55, P less than 0.01) and was negatively correlated with renal function (r = -0.60, P less than 0.01). In five subjects given vitamin D, WBR did not fall. The measurement of WBR in the elderly has to be assessed in the light of an accurate assessment of renal function and so is rarely clinically useful.

Differential alterations in striatal acetylcholine function in rats during 12 months' continuous administration of haloperidol, sulpiride, or clozapine.

Rats were treated continuously for 12 months with therapeutically equivalent doses of either haloperidol (1.4-1.6 mg/kg/day), sulpiride (102-109 mg/kg/day), or clozapine (24-27 mg/kg/day). After treatment for 3 and 12 months with haloperidol or clozapine but not sulpiride, striatal acetylcholine levels were increased. Striatal choline acetyltransferase activity was not altered by any drug treatment. Vmax for striatal acetylcholinesterase activity during the course of 12 months' treatment with haloperidol or clozapine, but not with sulpiride, tended to increase; Km was not altered by any drug treatment. Bmax for specific striatal [3H]quinuclidinyl benzilate binding was not altered by haloperidol or sulpiride treatment but was transiently elevated after 6 months of clozapine treatment, thereafter returning to control levels. Kd was not altered by any drug treatment. These findings indicate that alterations in striatal acetylcholine content caused by chronic treatment with some but not all neuroleptics are due to changes in cholinergic neuronal activity rather than neurotransmitter synthesis or destruction. The effects of haloperidol but not those of clozapine may be related to the emergence of functional striatal dopamine receptor supersensitivity. Since haloperidol (which is associated with a high prevalence of tardive dyskinesias) but not clozapine (which is not) had similar effects on striatal cholinergic function, the latter may not be related to the emergence of tardive dyskinesias during chronic therapy.