Identification of alkaptonuria in the general population: a United Kingdom experience describing the challenges, possible solutions and persistent barriers.

Progress in research into rare diseases is challenging. This paper discusses strategies to identify individuals with the rare genetic disease alkaptonuria (AKU) within the general population. Strategies used included a questionnaire survey of general practitioners, a dedicated website and patient network contact, targeted family screening and medical conference targeting. Primary care physicians of the UK were targeted by a postal survey that involved mailing 11,151 UK GPs; the response rate was 18.2%. We have identified 75 patients in the UK with AKU by the following means: postal survey (23), targeted family screening (11), patient networks and the website (41). Targeting medical conferences (AKU, rare diseases, rheumatology, clinical biochemistry, orthopaedics, general practitioners) did not lead to new identification in the UK but helped identify overseas cases. We are now aware of 626 patients worldwide including newly identified non-UK people with AKU in the following areas: Slovakia (208), the rest of Europe (including Turkey) (79), North America (including USA and Canada) (110), and the rest of the world (154). A mechanism for identifying individuals with AKU in the general population-not just in the UK but worldwide-has been established. Knowledge of patients with AKU, both in the UK and outside, is often confined to establishing their location in a particular GP practice or association with a particular medical professional. Mere identification, however, does not always lead to full engagement for epidemiological research purposes or targeting treatment since further barriers exist.

Micronutrient concentrations in respiratory syncytial virus and human metapneumovirus in Yemeni children.

BACKGROUND: Acute respiratory infections (ARI) cause significant childhood mortality. Nutritional homeostasis, particularly micronutrient levels, is important in modulating response to infection. More information is required regarding micronutrient levels in ARI viral infections, especially newly identified viruses such as human metapneumovirus (HMPV). AIM: To describe zinc, copper, selenium and vitamins A and E concentrations in children with respiratory syncytial virus (RSV) and/or HMPV in relation to levels of C-reactive protein (CRP). METHODS: The presence of RSV/HMPV in nasopharyngeal aspirates (NPA) was identified in 246 children using RTPCR. Zinc, copper, selenium and vitamins A and E concentrations were measured using inductive coupled plasma mass spectrometry and high performance liquid chromatography. RESULTS: 183 children had RSV, 39 had HMPV and 24 were co-infected. Zinc concentrations were lower in children with HMPV than in children with RSV or RSV/HMPV co-infection. Copper concentrations were lower in children with RSV than in children with RSV/HMPV or HMPV and zinc/copper ratios were lower in children with HMPV/RSV or RSV than in children with HMPV alone. Retinol and a alpha-tocopherol were lower in children with RSV than in children with HMPV. Most children had low selenium concentrations. Children with RSV and raised CRP (>5 mg/L) had higher copper and lower zinc/copper ratios than those with low CRP (< or =5 mg/L). Children with HMPV and raised CRP had higher copper and lower zinc concentrations than children with low CRP. Children with RSV/HMPV and raised CRP had higher copper concentrations. Children with RSV/HMPV and raised CRP had higher a alpha-tocopherol concentrations. CONCLUSION: The profiles of micronutrients differ in children with RSV and HMPV and are confounded by CRP. These results may guide strategies for micronutrient supplementation in ARI.

An architectural understanding of natural sway frequencies in trees.

The relationship between form and function in trees is the subject of a longstanding debate in forest ecology and provides the basis for theories concerning forest ecosystem structure and metabolism. Trees interact with the wind in a dynamic manner and exhibit natural sway frequencies and damping processes that are important in understanding wind damage. Tree-wind dynamics are related to tree architecture, but this relationship is not well understood. We present a comprehensive view of natural sway frequencies in trees by compiling a dataset of field measurement spanning conifers and broadleaves, tropical and temperate forests. The field data show that a cantilever beam approximation adequately predicts the fundamental frequency of conifers, but not that of broadleaf trees. We also use structurally detailed tree dynamics simulations to test fundamental assumptions underpinning models of natural frequencies in trees. We model the dynamic properties of greater than 1000 trees using a finite-element approach based on accurate three-dimensional model trees derived from terrestrial laser scanning data. We show that (1) residual variation, the variation not explained by the cantilever beam approximation, in fundamental frequencies of broadleaf trees is driven by their architecture; (2) slender trees behave like a simple pendulum, with a single natural frequency dominating their motion, which makes them vulnerable to wind damage and (3) the presence of leaves decreases both the fundamental frequency and the damping ratio. These findings demonstrate the value of new three-dimensional measurements for understanding wind impacts on trees and suggest new directions for improving our understanding of tree dynamics from conifer plantations to natural forests.

B-type natriuretic peptide (BNP) attenuates the L-type calcium current and regulates ventricular myocyte function.

A fundamental question in physiology is how hormones regulate the functioning of a cell or organ. It was therefore the aim of this study to investigate the effect(s) of BNP-32 on calcium handling by ventricular myocytes obtained from the rat left ventricle. We specifically tested the hypothesis that BNP-32 decreased the L-type calcium current (I(Ca,L)). Perforated patch clamp technique was used to record I(Ca,L) and action potential (AP) in voltage and current clamp mode, respectively. Myocyte shortening was measured using a photodiode array edge-detection system and intracellular calcium transients were measured by fluorescence photometry. Western blotting was used to determine the relative change in the expression of proteins. At the concentrations tested, BNP-32 significantly decreased cell shortening in a dose-dependent manner; increased the phase II slope of the AP by 53.0%; increased the APD(50) by 16.9%; reduced the I(Ca,L) amplitude with a 22.9% decrease in the peak amplitude and reduced Ca(2+)-dependent inactivation; increased the V(1/2) activation of the L-type calcium channel by 51.1% and decreased V(1/2) inactivation by 31.8%; and, intracellular calcium transient amplitude was significantly decreased by 32.0%, whereas the time to peak amplitude and T(1/2) were both significantly increased by 38.7% and 89.4% respectively. Sarcoplasmic reticulum Ca(2+)-ATPase (SERCA2a) protein expression was reduced by BNP-32. These data suggest that BNP-32 regulates ventricular myocyte function by attenuating I(Ca,L), altering the AP and reducing SERCA2a activity and/or expression. This study suggests a novel constitutive mechanism for the autocrine action of BNP on the L-type calcium channel in ventricular myocytes.

Hypoadrenalism is not associated with early mortality during tuberculosis treatment in Malawi.

SETTING: In the developing world, early mortality within 1 month of commencing tuberculosis (TB) treatment is high, particularly with human immunodeficiency virus (HIV) co-infection. In Malawi, 40% of those who die do so in the first month of treatment. Reasons remain unclear and may include delayed diagnosis, opportunistic infections, immune restoration inflammatory syndrome (IRIS) or malnutrition. One possible contributing factor is underlying hypoadrenalism associated with TB-HIV, exacerbated by rifampicin (RMP) induction of P450 and glucocorticoid metabolism. OBJECTIVE: To assess the prevalence of hypoadrenalism in TB patients before and after commencement of TB treatment, and relationship with early mortality. DESIGN: Prospective descriptive study assessing hypoadrenalism before and after anti-tuberculosis treatment, HIV status and outcome up to 3 months post-treatment. RESULTS: Of 51 patients enrolled, 29 (56.9%) were female (median age 32 years, range 18-62). Of 43 patients HIV-tested, 38 (88.3%) were HIV-positive and 15.7% died within the first month. At 3 months, 11 (21.6%) were known to have died. Adequate cortisol levels were found in 49/51 (95.9%) before commencing RMP. Neither of the two with reduced response died. All 34 patients revealed adequate cortisol responses at 2 weeks. CONCLUSION: No evidence of hypoadrenalism was found in this first study to assess adrenal function and outcome of anti-tuberculosis treatment.

Nutrition in clinical practice-the refeeding syndrome: illustrative cases and guidelines for prevention and treatment.

The refeeding syndrome is a potentially lethal complication of refeeding in patients who are severely malnourished from whatever cause. Too rapid refeeding, particularly with carbohydrate may precipitate a number of metabolic and pathophysiological complications, which may adversely affect the cardiac, respiratory, haematological, hepatic and neuromuscular systems leading to clinical complications and even death. We aimed to review the development of the refeeding syndrome in a variety of situations and, from this and the literature, devise guidelines to prevent and treat the condition. We report seven cases illustrating different aspects of the refeeding syndrome and the measures used to treat it. The specific complications encountered, their physiological mechanisms, identification of patients at risk, and prevention and treatment are discussed. Each case developed one or more of the features of the refeeding syndrome including deficiencies and low plasma levels of potassium, phosphate, magnesium and thiamine combined with salt and water retention. These responded to specific interventions. In most cases, these abnormalities could have been anticipated and prevented. The main features of the refeeding syndrome are described with a protocol to anticipate, prevent and treat the condition in adults.

Effect of micronutrient supplementation on mood in nursing home residents.

One third of older people in nursing and/or residential homes have significant symptoms of depression. In younger people, deficiencies in selenium, vitamin C and folate are associated with depression. This study examines the association between micronutrient status and mood before and after supplementation. The objective was to determine whether the administration of selenium, vitamin C and folate improved mood in frail elderly nursing home residents. Mood was assessed using the Hospital Anxiety and Depression rating scale (HAD), and Montgomery-Asberg Depression Rating Scale (MADRS). Micronutrient supplementation was provided for 8 weeks in a double-blinded randomised controlled trial. Significant symptoms of depression (29%) and anxiety (24%) were found at baseline. 67% of patients had low serum concentrations of vitamin C, but no-one was below the reference range for selenium. Depression was significantly associated with selenium levels, but not with folate or vitamin C levels. No individual with a HAD depression score of >or=8, had selenium levels >1.2 microM. In those patients with higher HAD depression scores, there was a significant reduction in the score and a significant increase in serum selenium levels after 8 weeks of micronutrient supplementation. Placebo group scores were unchanged. This small study concluded that depression was associated with low levels of selenium in frail older individuals. Following 8 weeks of micronutrient supplementation, there was a significant increase in selenium levels and improved symptoms of depression occurred in a subgroup.

Defining malnutrition: A plea to rethink.

In a recent consensus report in Clinical Nutrition the undernourished category of malnutrition was proposed to be defined and diagnosed on the basis of a low BMI or unintentional weight loss combined with low BMI or FFMI with certain cut off points. The definition was endorsed by ESPEN despite recent endorsement of a very different definition. The approach aims to assess whether nutritional intake is sufficient but is imprecise because a low BMI does not always indicate malnutrition and individuals with increasing BMI's may have decreasing FFM's. The pathophysiology of individuals, considered to be malnourished in rich countries and in areas with endemic malnutrition, results predominantly from deficient nutrition combined with infection/inflammation. Both elements jointly determine body composition and function and consequently outcome of disease, trauma or treatment. When following the consensus statement only an imprecise estimate is acquired of nutritional intake without knowing the impact of inflammation. Most importantly, functional abilities are not assessed. Consequently it will remain uncertain how well the individual can overcome stressful events, what the causes are of dysfunction, how to set priorities for treatment and how to predict the effect of nutritional support. We therefore advise to consider the pathophysiology of malnourished individuals leading to inclusion of the following elements in the definition of malnutrition: a disordered nutritional state resulting from a combination of inflammation and a negative nutrient balance, leading to changes in body composition, function and outcome. A precise diagnosis of malnutrition should be based on assessment of these elements.

Micronutrients in health and disease.

Micronutrients play a central part in metabolism and in the maintenance of tissue function. An adequate intake therefore is necessary, but provision of excess supplements to people who do not need them may be harmful. Single micronutrient deficiency states are comparatively easily recognised and treated. Subclinical deficiency, often of multiple micronutrients, is more difficult to recognise, and laboratory assessment is often complicated by the acute phase response. Clinical benefit is most likely in those people who are severely depleted and at risk of complications, and is unlikely if this is not the case. There is little evidence for supplements leading to a reduction in the incidence of infections in the elderly population, in coronary artery disease, or in malignant disease. The best evidence for benefit is in critical illness, and in children in developing countries consuming a deficient diet. More clinical trials are required with good clinical outcomes to optimise intake in prevention and treatment of disease.

Influence of whole body protein turnover rate on resting energy expenditure in patients with cancer.

Whole body protein turnover and resting energy expenditure are measured simultaneously in weight stable and weight losing patients with lung (n = 22) or colorectal cancer (n = 38). These results were compared with those from weight stable and weight losing non-cancer controls (n = 22). Rates of whole body protein turnover were calculated from the plateau isotopic enrichment of urinary ammonia and urea following a primed, continuous, 24-h infusion of [15N]glycine. Resting energy expenditure was measured by indirect calorimetry. All groups of cancer patients had significantly elevated rates of whole body protein turnover (P less than 0.05) and synthesized, on average, 1.9 g/kg/day more protein compared with weight stable non-cancer controls. In contrast, the resting energy expenditure of cancer patients and controls was similar. Moreover, there was no correlation between individual rates of whole body protein turnover. Thus, although cancer patients had rates of whole body protein turnover which were 50-70% greater than controls, this did not result in a measurable increase in resting energy expenditure. The assumption that elevation of whole body protein turnover or resting energy expenditure causes weight loss in cancer patients must be an oversimplification. An acute phase protein response was observed in the majority of cancer patients. Although the presence of such an inflammatory response did not correlate with the rate of whole body protein turnover, the role of inflammatory mediators in the pathogenesis of disturbed protein metabolism in cancer patients merits further investigation.

Poly(A)-binding RNAs from nuclei and polysomes of BHK-21 cells.

The presence in nuclei and cytoplasm of cultured BHK-21/C13 cells (baby hamster fibroblasts) of RNA species with high affinity for poly(A) was detected using either a Millipore-poly(A)-binding assay or columns of poly(A)-Sepharose 4B. The nuclear species which labels rapidly is metabolically unstable and appears to be a specific subclass of heterogenous nuclear RNA. Digestion with T1 RNAase gives rise to a low level (1%) of oligonucleotide fragments of a disperse size range which are relatively rich in uridylate residues. The cytoplasmic species with affinity for poly(A) is similar in size to polyadenylated messenger RNAs and also associates with polysomes. However it appears to be distinct from the polyadenylated messenger RNAs by virtue of an unusual base composition and relative metabolic instability.

Cancer cachexia: influence of systemic ketosis on substrate levels and nitrogen metabolism.

The aim of this study was to determine whether a ketogenic diet could decrease nitrogen losses in cachectic cancer patients and at the same time reduce the supply of glucose for tumor energy metabolism. Five patients with malignant disease and severe weight loss (mean 32%) were fed via a fine bore nasogastric tube. A normal diet was given for 6 d and this was followed by 7 d of an isonitrogenous, isocaloric, ketogenic diet. Both diets were well tolerated. At 7 d the mean ketone body concentration in the blood of patients fed the ketogenic diet was 1.21 +/- 0.33 mM. This ketosis was associated with a significant reduction of the concentration in blood of glucose, lactate, and pyruvate (p less than 0.05). There was, however, no significant alteration in host N balance or whole-body protein synthesis, degradation, or turnover rates. Whether the change from glucose- to fat-derived energy substrates might reduce tumor growth rates in the long term remains to be determined.

Trace element supplementation modulates pulmonary infection rates after major burns: a double-blind, placebo-controlled trial.

Infections remain the leading cause of death after major burns. Trace elements are involved in immunity and burn patients suffer acute trace element depletion after injury. In a previous nonrandomized study, trace element supplementation was associated with increased leukocyte counts and shortened hospital stays. This randomized, placebo-controlled trial studied clinical and immune effects of trace element supplements. Twenty patients, aged 40 +/- 16 y (mean +/- SD), burned on 48 +/- 17% of their body surfaces, were studied for 30 d after injury. They consumed either standard trace element intakes plus supplements (40.4 micromol Cu, 2.9 micromol Se, and 406 micromol Zn; group TE) or standard trace element intakes plus placebo (20 micromol Cu, 0.4 micromol Se, and 100 micromol Zn; group C) for 8 d. Demographic data were similar for both groups. Mean plasma copper and zinc concentrations were below normal until days 20 and 15, respectively (NS). Plasma selenium remained normal for group TE but decreased for group C (P < 0.05 on days 1 and 5). Total leukocyte counts tended to be higher in group TE because of higher neutrophil counts. Proliferation to mitogens was depressed compared with healthy control subjects (NS). The number of infections per patient was significantly (P < 0.05) lower in group TE (1.9 +/- 0.9) than in group C (3.1 +/- 1.1) because of fewer pulmonary infections. Early trace element supplementation appears beneficial after major burns; it was associated with a significant decrease in the number of bronchopneumonia infections and with a shorter hospital stay when data were normalized for burn size.

Biochemical changes associated with severe trauma.

The effects of different intravenous nutritional regimens on a number of biochemical indices of nutritional status were studied during the 8-day period following severe trauma. The inclusion of large amounts of amino acids (high nitrogen (N) was shown to greatly improve N balance over an isocaloric regimen containing no amino acids (O g N). The concentration of serum albumin, transferrin, prealbumin, and retinol-binding protein all fell during the study period in both patient groups, whereas the serum concentrations of acute phase reactants and of ribonuclease increased in the two groups. The sum of plasma levels of branched-chain amino acids and the essential amino acids was increased to a greater extent in the high N group. These amino acid totals and the ratio of glycine/valine showed a significant correlation with N balance in this group. Despite the marked difference in N balance, 3-methylhistidine excretion was increased but equal in the two nutritional groups, suggesting an increased rate of muscle protein breakdown in both groups, which appears not to be influenced by amino acid nutrition. It is concluded that N balance can be significantly improved in the immediate posttrauma period by provision of amino acids together with energy substrates. None of the biochemical variables measured, with the exception of plasma levels of essential amino acids, reflected these marked differences in N balance.

Assessment of nutritional status and in vivo immune responses in alcoholic liver disease.

Nutritional status and in vivo immune responses were investigated in 30 patients with alcoholic liver disease who were drinking heavily up until emergency hospital admission. Investigations were performed on admission and after 2 wk abstention and adequate hospital diet. No relationship was found between the severity of liver disease revealed histologically and the recent quantity or total duration of alcohol intake, inadequacy of diet, or nutritional status. Skin anergy was more common in those patients with cirrhosis but did not relate to depletion in circulating T lymphocytes, poor nutritional status, or to the direct effect of alcohol toxicity. Acute alcohol toxicity did, however, produce extensive and rapidly reversible metabolic and cellular changes including reduction in serum potassium, magnesium and phosphate and depletion of all circulating lymphocyte subpopulations.

Rapid, sensitive detection of plasma IL-1 by extraction and bioassay.

A method for separating IL-1 from plasma inhibitors by silica extraction has been developed and coupled to a highly sensitive bioassay using the LBRM TG6 cell line and the I1-2 dependent HT2A cell line. Using this assay we have detected IL-1 activity in plasma from patients undergoing elective surgery.

Effect of glycoprotein and protein hormones on human meningioma cell proliferation in vitro.

Speculation that meningiomas are subject to female hormone influence is supported by their higher incidence in women and reports of exacerbation of symptoms during pregnancy and the luteal phase of the menstrual cycle. Previous reports have concentrated on the effects of the steroid hormones oestradiol and progesterone on meningioma proliferation. In this study we have investigated the roles of the glycoproteins LH, FSH and human chorionic gonadotrophin (hCG), and the protein hormones prolactin (PRL) and human placental lactogen (hPL) on the proliferation of human meningiomas in vitro. The three glycoproteins had an inhibitory effect on meningioma proliferation ranging from 5.0-50.0%, 10.0-63.0% and 2.4-34.0% at the highest concentrations of LH (25 mIU/ml), FSH (15 mIU/ml) and hCG (30 IU/ml) respectively. Cultures were also treated with PRL (100 and 200 ng/ml) and hPL (5 and 10 ng/ml) and the protein hormones had a stimulatory effect on cell proliferation of 12.0-55.5% and 11.4-73.6% when treated with 200 ng/ml PRL and 10 micrograms/ml hPL respectively. Our data suggest that increasing levels of the protein hormones PRL and hPL, falling levels of hCG and the absence of LH and FSH in the second and third trimesters of pregnancy may play a role in the acceleration of meningioma growth in these stages of pregnancy.

Increased interleukin 6 concentrations in the absence and presence of HIV-1 infection in haemophilia.

AIMS: To measure IL-6 concentrations in asymptomatic HIV-1 antibody positive and negative haemophilic patients and to correlate these with IgG concentrations and CD4 positive cell numbers. METHODS: IL-6 concentrations were measured by bioassay in stored serum from a prospective cohort of haemophilic patients in whom immunoglobulins and T cell subsets had been determined. Values of IL-6 and immunoglobulins were also correlated with severity of liver disease. RESULTS: IL-6 concentrations were raised in haemophilic patients independent of HIV-1 antibody status. In HIV-1 antibody positive patients there was no correlation between IL-6 concentrations and CD4 positive cell numbers, but there was a correlation with IgG (r = 0.4; p = 0.05). In HIV-1 antibody negative patients there were no significant correlations. CONCLUSIONS: Haemophilic patients have increased IL-6 concentrations; in HIV-1 positive patients this is independent of the decline in CD4 cell count. The raised concentrations do not correlate with the clinical severity of liver disease.

Thiamine deficiency in the critically ill.

There have been recent reports of hospitalised patients developing clinical thiamine deficiency, combined with much debate on the optimal supplementation of thiamine for the parenterally fed patient, particularly in the intensive therapy environment. We performed a retrospective study on 158 patients admitted to the Intensive Care Unit who required nutritional support. Patients who survived had significantly higher body thiamine status than those who died (p less than 0.01). There was no difference between serum albumin concentrations of the two groups. Twenty percent of the patients had biochemical evidence of thiamine deficiency and the mortality rate in these patients was 72% as compared with 50% mortality overall. Follow-up results suggest that current levels of thiamine supplementation are insufficient for critically ill intravenously fed patients. We suggest that patients be given a loading dose of 50-250 mg thiamine on admission to the Intensive Care Unit.

Influence of selenium supplements on the post-traumatic alterations of the thyroid axis: a placebo-controlled trial.

OBJECTIVE: To investigate whether early selenium (Se) supplementation can modify the post-traumatic alterations of thyroid hormone metabolism, since the first week after trauma is characterised by low plasma Se and negative Se balances. DESIGN: Prospective, placebo-controlled randomised supplementation trial. SETTING: Surgical ICU in a tertiary university hospital. PATIENTS: Thirty-one critically ill trauma patients aged 42 +/- 16 years (mean +/- SD), with severe multiple injury (Injury Severity Score 30 +/- 7). INTERVENTION: Supplementation during the first 5 days after injury with either Se or placebo. The selenium group was further randomised to receive daily 500 microg Se, with or without 150 mg alpha-tocopherol (AT) and 13 mg zinc supplements. The placebo group received the vehicle. Circulating Se, AT, zinc, and thyroid hormones were determined on D0 (= day 0, admission), D1, D2, D5, D10, and D20. RESULTS: Plasma Se, low on D0, normalised from D1 in the selenium group; total T4 and T3 increased more and faster after D2 (P = 0.04 and 0.08), reverse T3 rising less between D0 and D2 (P = 0.05). CONCLUSIONS: Selenium supplements increased the circulating Se levels. Supplementation was associated with modest changes in thyroid hormones, with an earlier normalisation of T4 and reverse T3 plasma levels. The addition of AT and zinc did not produce any additional change.