Acute intermittent porphyria: fatal complications of treatment.

Acute neurovisceral attacks of porphyria can be life threatening. They are rare and notoriously difficult to diagnose clinically, but should be considered, particularly in female patients with unexplained abdominal pain, and associated neurological or psychiatric features or hyponatraemia. The diagnosis might be suggested by altered urine colour and can be confirmed by finding an elevated porphobilinogen concentration in fresh urine protected from light. Severe attacks require treatment with intravenous haem arginate and supportive management with safe drugs, including adequate analgesia. Intravenous glucose in water solutions are contraindicated as they aggravate hyponatraemia, which can prove fatal.

Family tracing to identify patients with familial hypercholesterolaemia: the second audit of the Department of Health Familial Hypercholesterolaemia Cascade Testing Project.

BACKGROUND: Family tracing is a method recognized to find new patients with familial hypercholesterolaemia (FH). We have implemented family tracing led by FH Nurses and have determined acceptability to patients, feasibility and costs. METHODS: Nurses were located at five National Health Service (NHS) Trusts; they identified FH patients and offered them family tracing. Responses and test results were recorded on a database and summarized on a family pedigree. RESULTS: The majority ( approximately 70%) of index cases participated; the proportion was lower when patients had been discharged from the clinics and in metropolitan areas. On average, 34% (range 13-50%) of relatives lived outside the catchment area of the clinics and could not attend the nurse-led FH clinics. Of the previously untested relatives, 76% who lived in the catchment area of the clinic came forward to be tested. One-third of the relatives who came forward for testing were children

Abundance and survival of microbial aerosols in the troposphere and stratosphere.

Bioaerosol transport in the atmosphere disperses microbial species between continents, affects human and plant health, and may influence hydrologic cycling. However, there have been few quantitative observations of bioaerosols at altitudes more than a few kilometers above the surface. Lack of data on bioaerosol distributions in the atmosphere has impeded efforts to assess the aerial dissemination of microbes and their vertical extent in the biosphere. In this study, a helium balloon payload system was used to sample microbial cells and dust particles in air masses as high as 38 km above sea level over three locations in the southwestern United States. The cell concentrations at altitudes between 3 and 29 km were highly similar (2-5 × 105 cells m-3) and approximately threefold lower than those observed in the convective boundary layer (CBL; 1 × 106 cells m-3), decreasing to 8 × 104 cells m-3 at 35-38 km. The detection of adenosine triphosphate (ATP) and recovery of bacteria possessing extreme tolerance to desiccation and shortwave ultraviolet radiation confirmed that certain microorganisms have the capacity to persist at lower altitudes of the stratosphere. Our data and related calculations provide constraints on the upper altitudinal boundary for microbial habitability in the biosphere.

Are patients with familial hypercholesterolaemia well managed in lipid clinics? An audit of eleven clinics from the Department of Health Familial Hypercholesterolaemia Cascade Testing project.

BACKGROUND: Familial hypercholesterolaemia (FH) is an autosomal co-dominant disorder which is relatively common, leads to high levels of LDL-cholesterol and if untreated to early coronary heart disease. An audit of current practice at National Health Service Trusts in England was undertaken to determine whether FH patients meet the diagnostic criteria for FH; are being offered appropriate advice and treatment; and to what extent their families are contacted and offered testing for the disorder. METHODS: Medical records of known FH patients (over 18 years of age and diagnosed before 31 December 2003) were accessed to obtain information on diagnosis, treatment and family tracing. RESULTS: The records of 733 FH patients were examined, 79% met the UK 'Simon Broome' register criteria for the diagnosis of definite or possible FH. Analyses showed that patients were usually offered appropriate advice and treatment, with 89% being on a statin. However, the audit indicated a high variability in family tracing between the sites, with significant differences in the frequency of inclusion of a family pedigree in the notes (range 1-71%, mean 35%); the general practitioner (GP) being advised that first-degree relatives should be tested (range 4-52%, mean 27%); and the proportion of relatives contacted and tested (range 6-50%, mean 32%). CONCLUSION: FH patients are well cared for in lipid clinics in England, are being given appropriate lifestyle advice and medication, but an increase in recording of LDL-cholesterol levels may lead to improvements in their management. Practice in family tracing appears to vary widely between clinics.

Bromocriptine inhibits pro-opiomelanocortin mRNA and ACTH precursor secretion in small cell lung cancer cell lines.

We have previously reported that a human small cell lung cancer (SCLC) cell line (COR L103) that expresses the proopiomelanocortin (POMC) gene and secretes ACTH precursor peptides is relatively resistant to glucocorticoid regulation. Using this model, we have now examined alternative regulatory mechanisms of the POMC gene and found that both the mRNA and ACTH precursor peptides were stimulated four- and two-fold, respectively, after 48 h incubation with db-cAMP. Next, we examined the dopamine agonist, bromocriptine, which acts predominantly through D2 receptors linked to adenyl cyclase to cause a reduction in intracellular cAMP. Bromocriptine suppressed cAMP levels and inhibited precursor peptide secretion within 24 h in a dose-dependent manner (0.15-15 microM). At the highest dose, peptide secretion was inhibited from 95 to 53 pmol/mg protein, and POMC mRNA was reduced by 50%, while beta-actin mRNA remained unchanged. This effect could not be mimicked by incubation of cells with the alpha-adrenergic antagonist, phenoxybenzamine, suggesting that the alpha-adrenergic effects of bromocriptine were not responsible for this observation. These cells also secrete estradiol, but the secretory rate was unaffected by bromocriptine, suggesting, with the beta-actin data, that the POMC inhibition was not a cytotoxic effect. No recovery in precursor peptide secretion was seen in a 48-h period after the removal of bromocriptine. However, when the postchallenge incubation was extended to 8 d, there was a recovery in secretory potential between day 3 and day 8 and normal growth kinetics in the 4 d after removal of the drug. In contrast to these findings, the mouse corticotroph cell line, AtT20, showed no response to bromocriptine, in keeping with reports that this agonist has no effect on anterior lobe corticotrophs. We conclude that bromocriptine effectively inhibits POMC expression in SCLC cells, and that this phenomenon might be of useful clinical application.

Direct measurement of the precursors of adrenocorticotropin in human plasma by two-site immunoradiometric assay.

An immunoradiometric assay (IRMA) for the direct measurement of the precursors of ACTH in unextracted human plasma has been developed and evaluated clinically in normal subjects and patients with disorders of the hypothalamic-pituitary-adrenal axis. The IRMA is based on an iodinated monoclonal antibody to ACTH and a monoclonal antibody to gamma MSH coupled to Sephacryl S300. The assay detects only peptides containing both epitopes, i.e. POMC (31K) and pro-ACTH (22K). The reference standard was partially purified POMC from culture medium of human corticotroph adenoma cells. The detection limit (greater than +2.5SD of the 0 standard) was 2.0 pmol/L and the within-assay coefficient of variation was less than 10% between 29 and 2600 pmol/L. Plasma concentrations of ACTH precursor peptides in 11 normal subjects sampled at 0930 h ranged from 5-34 pmol/L. The concentrations in the patient groups studied were: 260-2300 pmol/L in 5 patients with the ectopic ACTH syndrome associated with small cell lung cancer, less than 2.0-104 pmol/L in 10 patients with pituitary-dependent Cushing's disease, 23 pmol/L in a patient with Nelson's syndrome, and 3.0-230 pmol/L in 5 patients with Addison's disease. We conclude that this IRMA offers a simple and reliable method for measuring ACTH precursors in unextracted plasma. The proportionately greater elevation of ACTH precursors compared to ACTH in patients with the ectopic ACTH syndrome associated with small cell lung cancer but not in pituitary-dependent Cushing's syndrome, suggests that this assay may be clinically useful.

Differential release of proopiomelanocortin-derived peptides from the human pituitary: evidence from a panel of two-site immunoradiometric assays.

In humans, proopiomelanocortin (POMC) and the peptides derived from it have been individually identified in plasma under differing conditions. However, direct quantitative comparison has proved difficult because of the limitations of RIAs. Using a panel of monoclonal antibodies recognizing different regions of POMC, we have developed specific two-site immunoradiometric assays (IRMAs) for the ACTH precursors (POMC and pro-ACTH), ACTH, beta-lipotropin (beta LPH), beta-endorphin (beta EP), and the N-terminal POMC fragment (N-POC). We have quantified these peptides directly in plasma from normal subjects under basal conditions and in response to different regulatory factors. Basal levels of ACTH precursors, 5-40 pmol/L, were greater than or equal to ACTH, less than 0.9-11.3 pmol/L; N-POC, 5.6-16.8 pmol/L; beta LPH, 2.5-6.7 pmol/L; and beta EP less than or equal to 1.7 pmol/L. ACTH, N-POC, beta LPH, and beta EP levels increased in parallel in response to metyrapone (n = 8) and decreased in response to dexamethasone (n = 8), whereas ACTH precursor concentrations did not respond. After human CRH administration, peripheral concentrations of ACTH, N-POC, and beta LPH showed similar increments (median increment, 163%, 145%, and 172%, respectively; n = 6). POMC peptide responses to human CRH were also assessed in inferior petrosal sinuses draining the pituitary in 20 patients with pituitary-dependent Cushing's disease. In these patients, the increment in ACTH after CRH exceeded that in ACTH precursors by 4-fold (median, 459% and 96%). An increase in the ratios of ACTH/N-POC and ACTH/beta LPH was also apparent after CRH stimulation. The increment in beta EP after CRH always exceeded the increments in POMC and beta LPH. In summary, these data suggest that significant concentrations of ACTH precursors are present in the circulation of normal subjects, that ACTH precursors are not regulated in the same way as the processed POMC peptides, and that ACTH and beta EP are preferentially released from the pituitary in response to CRH.

Defective glucocorticoid regulation of proopiomelanocortin gene expression and peptide secretion in a small cell lung cancer cell line.

A human small cell lung cancer cell line (COR L103) that actively expresses the proopiomelanocortin (POMC) gene has been used as a model of extrapituitary ACTH-secreting tumors to investigate the phenomenon of resistence of ACTH production to glucocorticoids. After both short term (24 h) and long term (10 days) exposure to hydrocortisone at concentrations of 500 and 1000 nM, the accumulation of intracellular POMC mRNA, ACTH, and ACTH precursor peptides in the culture medium was not suppressed. These finding contrast with those in the pituitary corticotroph cell line AtT20, in which POMC mRNA, ACTH, and ACTH precursors were suppressed under the same conditions. Two other genes that are regulated by glucocorticoids in other cell types, the tyrosine amino transferase gene and the glucocorticoid receptor gene, were expressed in COR L103 cells. However, neither gene appeared to be regulated by hydrocortisone in this small cell lung cancer cell line. Further studies demonstrated that glucocorticoid receptor binding could be detected in the nucleus and cytoplasm, with a Kd of 5 X 10(-9) M. It is concluded that nonsuppression of POMC by glucocorticoids is probably part of a more global defect of glucocorticoid signaling in these cells, but that this defect lies distal to steroid binding in the nucleus.

Constitutive synthesis of 1,25-dihydroxyvitamin D3 by a human small cell lung cancer cell line.

One of 16 human small cell lung cancer cell lines examined was shown to synthesize a metabolite resembling 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. The NCI H82 line converted 25-hydroxyvitamin D3 (25OHD3) into a compound indistinguishable from 1,25-(OH)2D3 in 3 different high performance liquid chromatography systems. Electron impact mass spectra for the trimethylsilylethers of the metabolite and authentic 1,25-(OH)2D3 were indistinguishable. Binding to an anti-1,25-(OH)2D3 antibody was identical for the metabolite and authentic 1,25-(OH)2D3, whereas administration to rats in vivo caused equivalent stimulation of calcium transport measured in vitro in duodenal sacs. Activity of the H82 1 alpha-hydroxylase appears to be substrate dependent and is not stimulated by PTH, suggesting that it is similar to the enzyme expressed by activated macrophages and other cell types at extrarenal sites. Inhibition by ketoconazole indicates that, like the renal and extrarenal enzymes, the H82 enzyme is cytochrome P450 dependent. These data indicate that the H82 small cell lung cancer cell line constitutively expresses 25-hydroxyvitamin D3-1 alpha-hydroxylase and can synthesize 1,25-(OH)2D3.

Pro-opiomelanocortin gene expression and peptide secretion in human small-cell lung cancer cell lines.

Expression of the RNA coding for the ACTH-beta-lipotrophin precursor, pro-opiomelanocortin (POMC), has been demonstrated in five human small-cell lung cancer (SCLC) cell lines. Using Northern and slot-blot hybridization analysis of RNA and a bovine POMC cDNA as probe, the processed POMC RNA from SCLC cells was found to be approximately 1350 nucleotides in length, which is larger than that found in the normal human pituitary. Expression of the POMC gene was confirmed by measurement of ACTH precursors secreted by the cells, using a novel two-site immunoradiometric assay based on monoclonal antibodies, which directly quantifies both POMC and pro-ACTH but does not recognize ACTH. Levels of POMC in medium accumulated throughout the growth of the cells, in contrast to POMC RNA which showed a relatively constant level of expression. We conclude that human SCLC cell lines are valuable models for studying the aberrant expression and regulation of the human POMC gene.

Glucocorticoid inhibition of ACTH peptides: small cell lung cancer cell lines are more resistant than pituitary corticotroph adenoma cells.

In the normal pituitary, glucocorticoids are the principal negative regulatory of the pro-opiomelanocortin (POMC) gene which gives rise to the biologically active peptides ACTH and beta-endorphin. In Cushing's syndrome, ACTH-secreting pituitary tumours show a degree of glucocorticoid resistance, whilst ACTH-secreting extra-pituitary tumours have an even greater resistance to glucocorticoid excess. In an attempt to understand the mechanism of this phenomenon, we have compared the effects of glucocorticoids on POMC mRNA and peptide secretion in human and mouse corticotroph adenoma cells and in small cell lung carcinoma (SCLC) cells. ACTH precursor peptides were inhibited within 24 h by 25-50 nM hydrocortisone in primary cultures from a human corticotroph adenoma. In the mouse corticotroph adenoma cell line (AtT20), inhibition of both ACTH precursors and ACTH was not observed after 24 h but, by 10 days, glucocorticoids suppressed peptide levels with a concentration causing 50% inhibition of 50 nM hydrocortisone and maximal inhibition at 500 nM hydrocortisone. In marked contrast, there was no response to 500 nM hydrocortisone in the five SCLC cell lines (COR L103, COR L42, COR L24, COR L31, DMS 79) all of which secrete ACTH precursors. However, two of the five SCLC cell lines (COR L31 and DMS 79) were responsive to 1000 nM hydrocortisone. POMC mRNA, quantitated by slot-blot analysis, gave similar results for the five SCLC cell lines, implying that the abnormality may occur at the level of gene expression. When one of the three resistant cell lines (COR L103) was incubated with 2000 nM hydrocortisone or 2000 nM dexamethasone a clear suppression of precursor peptides and POMC mRNA was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of ACTH and ACTH precursor peptides secreted by human pituitary and lung tumour cells in vitro.

The molecular forms of ACTH secreted by established human small cell lung cancer (SCLC) cells and primary cultures derived from a bronchial carcinoid tumour, a pituitary adenoma and hyperplastic pituitary tissue have been characterized by Sephadex G-75 chromatography and quantified with two novel immunoradiometric assays for ACTH and ACTH precursor peptides. Pro-opiomelanocortin (POMC; Mr 31,000) and pro-ACTH (Mr 22,000) were secreted by all cell types. No smaller peptides were identified in the culture media from SCLC and bronchial carcinoid cells, implying a deficiency in the enzymes and/or intracellular organelles required for extensive POMC processing. A more heterogeneous profile of ACTH-containing peptides was produced by cells of pituitary origin, indicating more extensive proteolytic processing of POMC. However, the major peptide secreted by cells from a large aggressive pituitary adenoma was unprocessed POMC (Mr 31,000). These results suggest that both lung and pituitary cells in vitro retain their in-vivo pattern of POMC processing and provide valuable models in which to study the regulation of ACTH synthesis and secretion.

Biochemical diagnosis of phaeochromocytoma: two instructive case reports.

The biochemical features of two patients with phaeochromocytomas illustrate the inadvisability of depending on a single group of analytes for the diagnosis. The first case presented as a surgical emergency with retroperitoneal haemorrhage. Biochemical diagnosis was difficult since total 24 hour urinary free catecholamine excretion was within normal limits in two out of three samples, and only marginally raised in the third with an atypical preponderance of adrenaline. Plasma catecholamine concentrations were also normal. But urinary excretion of the catecholamine metabolites, metadrenaline and 4-hydroxy-3-methoxy mandelic acid (HMMA), was consistently raised. In contrast, the second patient presenting with headache and labile hypertension showed normal metabolite excretion in the face of grossly increased free noradrenaline excretion and raised plasma noradrenaline concentrations. It is therefore recommend that, as well as urinary free catecholamines, one group of their main metabolites, the 3-methoxy amines (normetadrenaline and metadrenaline) or HMMA, should routinely be measured whenever a phaeochromocytoma is suspected.

Thyroid function tests in patients with familial dysalbuminaemic hyperthyroxinaemia (FDH).

Two patients with familial dysalbuminaemic hyperthyroxinaemia (FDH) are described in whom the albumin variant resulted in raised total T4 levels, and artefactually raised free T4 using a 'single-step' technique employing an analogue of T4 as tracer. The first patient was clinically euthyroid and presented with relapse of schizophrenia and abnormal thyroid function tests (total T4 336 nmol/L, total T3 4.2 nmol/L, TSH 1.8 mU/L, free T4 73 pmol/L). These results led to diagnostic confusion and the patient was treated with a short course of anti-thyroid drugs. The second patient had signs and symptoms of thyrotoxicosis at her first visit but was clinically euthyroid 5 months later when she was 10 weeks pregnant. Thyroid function tests were total T4 259 nmol/L, total T3 3.6 nmol/L, TSH 3.8 mU/L, free T4 46 pmol/L. Further studies showed both patients to be biochemically euthyroid. A variant albumin was confirmed in both patients by a screening test for FDH and by reverse-flow electrophoresis. Family studies on 10 relatives of the first patient identified eight with FDH. A simple screening procedure for the indentification of FDH is described and the use of laboratory tests in suspected cases is discussed.

Hypophosphataemia in general practice patients.

We compared plasma phosphate concentrations in general practice patients and hospital inpatients and outpatients over an 8-month period. The distribution of results in all three groups was similar and 12-16% of results were at or below 0.8 mmol/L. In general practice patients, 8.3% of results from males and 12.1% from females were below the lower limit of their respective reference ranges. Eighteen of these patients (0.2% of results) had plasma phosphate concentrations < or = 0.4 mmol/L. On follow-up, only two of these patients had any attributable cause for their severe hypophosphataemia; in the remainder, it was unexpected and unexplained. Hypophosphataemia in outpatients and general practice patients is more common than has previously been appreciated. We present a strategy for further investigation of these patients.

Conditional probabilities and the grid test for schizophrenic thought disorder.

The grid test for schizophrenic thought disorder was administered to a sample of 149 psychiatric patients. This sample was drawn from a population with a known base rate of schizophrenia and an estimated base rate of thought disorder, thus allowing for the calculation of population specific cutting scores for the grid test. The grid test, using these data along with others from previous grid test studies, was then analysed for diagnostic efficiency using conditional probability formulae. The conclusion reached is that the grid test for schizophrenic thought disorder is unsatisfactory as a diagnostic instrument.

Detection of transient and persistent feline leukaemia virus infections.

A study was made of cats persistently or transiently viraemic with feline leukaemia virus (FeLV) following experimental oronasal infection. Cats of two ages were exposed to the virus. One group was infected when eight weeks old in the expectation that most of the cats would become persistently viraemic, and the second group when 16 weeks old, so that some would show signs of a transient infection and then recover. The periods following infection when virus was detectable in the blood and in the oropharynx were determined for each group. Three methods for detecting viraemia were compared: virus isolation, immunofluorescence on blood smears and an enzyme-linked immunosorbent assay (ELISA). There was good overall agreement among the three tests in detecting virus-positive cats. Virus was found sooner after infection by virus isolation than by the other methods, and virus appeared in the blood slightly sooner in cats which developed persistent viraemia than in transiently viraemic cats. Infectious FeLV was isolated from the oropharynx of all of the persistently viraemic cats, in most cases simultaneously with virus in the plasma. Virus was also isolated from the mouth of most transiently viraemic cats. Under field conditions such transient excretion of virus lasting only a few days would rarely be detected in a single sampling. This might explain how FeLV is maintained in free range urban cats in the absence of a large number of cats with persistent active FeLV infection. For routine diagnosis, immunofluorescence would appear to offer the best chance of differentiating transient and persistent infections by FeLV.