Dietary vasodilator and vitamin C/L-arginine/choline blend improve broiler feed efficiency during finishing and reduce woody breast severity at 6 and 7 wks.

Woody breast has become a considerable economic concern to the poultry industry. This myopathy presents rigid, pale breasts characterized by replacement of lean muscle protein with connective tissue, a result of hypoxia and oxidative stress in a metabolically starved muscle with inadequate circulation. Hence, the objectives were to supplement broiler diets with ingredients specifically aimed to improve circulation and oxidative status. About 1,344 male Ross 708 broilers were assigned to 1 of 4 diets: 1) a basal diet (control), 2) basal diet plus a blend of 0.2% supplemental L-arginine, 0.17% choline bitartrate, and 0.03% vitamin C (blend), 3) 0.1% vasodilator ingredient (vasodilator), or 4) 0.02% Astaxanthin ingredient (AsX). At d 14, 28, 42, and 49, performance outcomes were collected on all birds and serum from 16 broilers/diet (n = 64) was analyzed for creatine kinase and myoglobin. Once weekly beginning on d 28, a subset of 192 broilers were measured for breast width. On d 42 and 49, breast fillets from 16 broilers/diet (n = 64) were palpated for woody breast severity, weighed, and analyzed for compression force at 1-day postmortem and water-holding capacity at 2-day postmortem. mRNA was isolated from 15 breast fillets/timepoint for qPCR quantification of myogenic gene expression. Data were analyzed using Proc Mixed (SAS Version 9.4) with the fixed effect of diet. Feed conversion ratio was improved in the blend and vasodilator-fed birds d 42 to 49, each by over 2 points (P < 0.05). Breast width was increased in the control on d 42 compared to the vasodilator and AsX-fed broilers (P < 0.05). At d 42, there were 12% greater normal fillets in blend diet-fed birds and 13% more normal scores in vasodilator-fed birds at d 49 compared to the control. At d 49, myogenin expression was upregulated in the AsX diet compared to blend and control diets (P < 0.05), and muscle regulatory factor-4 expression was increased by 6.5% in the vasodilator diet compared to the blend and AsX diets (P < 0.05). Blend and vasodilator diets simultaneously improved feed efficiency in birds approaching market weight while reducing woody breast severity.

Dietary inositol-stabilized arginine silicate numerically reduced woody breast severity in male Ross 708 broilers without altering growth.

The woody breast (WB) myopathy is an unintended outcome of fast broiler chicken growth and high breast muscle yields. Myodegeneration and fibrosis in the living tissue are results of hypoxia and oxidative stress driven by lack of blood supply to muscle fibers. The study aim was to titrate a vasodilator ingredient, inositol-stabilized arginine silicate (ASI), as a feed additive to improve blood flow and ultimately, breast meat quality. A total of 1,260 male Ross 708 broilers were assigned to: 1) a control basal diet, or the control diet plus increasing ASI: 2) 0.025% ASI, 3) 0.05% ASI, 4) 0.10% ASI, or 5) 0.15% ASI. At d 14, 28, 42, and 49, growth performance was measured on all broilers and serum from 12 broilers/diet was analyzed for creatine kinase and myoglobin presence. On d 42 and 49, 12 broilers/diet were measured for breast width, then left breast fillets were excised, weighed, palpated for WB severity, and visually scored for degree of white striping (WS). At 1 d postmortem, 12 raw fillets/treatment underwent compression force analysis, and at 2d postmortem, the same fillets were analyzed for water-holding capacity. mRNA was isolated from 6 right breasts/diet at both d 42 and 49 for qPCR quantification of myogenic gene expression. Birds fed the lowest dose of 0.025% ASI had a 5-point/3.25% feed conversion ratio reduction compared to birds fed 0.10% ASI over wk 4 to 6 and reduced serum myoglobin at 6-wk of age compared to the control. Breasts from birds fed 0.025% ASI received 42% greater normal WB scores at d 42 compared to control fillets. At d 49, breasts from broilers fed 0.10 and 0.15% ASI received 33% normal WB scores. At d 49, 0.025% AS-fed broiler breasts showed no severe WS. Increased myogenin expression was observed in 0.05 and 0.10% ASI breast samples on d 42 and myoblast determination protein-1 expression was upregulated in breasts from birds fed 0.10% ASI on d 49 compared to the control. Therefore, a dietary inclusion of 0.025, 0.10, or 0.15% ASI was beneficial in reducing WB and WS severity and promoting muscle growth factor gene expression at age of harvest without diminishing bird growth or breast muscle yields.

Increased duration of laser environmental enrichment increased broiler physical activity and pen-wide movement without altering tibia measurements.

Inactivity and leg disorders negatively impact broiler welfare. Enrichment designed to increase barn complexity may encourage physical exercise. The study aim was to implement a second-generation laser enrichment device, previously shown to increase broiler activity, for extended periods of time and to measure behavior and tibia quality. A total of 1,360 Ross 708 broilers in 40 pens of 34 were assigned to laser enrichment or control (no laser enrichment) for 49 d. Seventy focal birds were randomly selected on d 0 for individual behavior analysis. Laser-enriched birds were exposed to 6-min laser periods 4 times daily. A 3-min novel object test was performed on all pens and tonic immobility was induced on 1 bird/pen on wk 1 and 6. Focal bird time budget and walking distance and pen-wide laser-following behavior and movement were obtained during laser periods d 0 to 8 and 1 d/wk through wk 7. Right tibias were collected from focal birds on d 49 for dual energy x-ray absorptiometry and bone breaking analysis. Time spent active during laser periods was increased on d 3, 6, and 8 and wk 2 to 3 in laser-enriched vs. control focal birds (P = 0.04). Time at the feeder was increased in laser-enriched focal birds on d 0, 3 to 4, and 8 and wk 2 and 4 (P < 0.01). Distance walked during laser periods was increased in laser-enriched focal birds on d 1, 3 to 5, 8, and wk 2 compared to the control (P < 0.01). Pen-wide movement was increased on d 0, 2, 4 to 8, and wk 1 to 5 and 7 in laser-enriched vs. control birds (P < 0.01). More laser-enriched broilers were within 25 cm of the novel object at 1 min 30 s than the control (P = 0.03), and latency to approach the novel object was reduced in both treatments at wk 6 compared to wk 1 (P < 0.01). Tonic immobility duration was increased by 123 s on wk 6 compared to wk 1, regardless of treatment (P < 0.01). Daily laser enrichment for extended periods increased bird activity without inducing fearfulness or altering tibia measures.

Breast muscle white striping and serum corticosterone reduced in broilers exposed to laser environmental enrichment.

Genetic selection for breast yields and fewer days to market has inadvertent effects on broiler meat quality. Woody breast (WB) and white striping (WS) are pectoralis major myopathies prevalent in commercial broilers. Effects of voluntary exercise on these disorders, specifically, are unknown. A second-generation laser enrichment device shown to induce activity in Ross 308 and 708 birds was implemented using 1,360 Ross 708 broilers randomly assigned to laser enrichment or control for 49 d. Laser-enriched birds were exposed to 6-min laser periods 4 times daily. Seventy focal birds were gait and contact dermatitis scored weekly. Blood was collected wk 5 to 7 from 56 broilers for serum corticosterone, myoglobin, and troponin. Seventy broilers were sampled for breast muscle width, fillet dimensions, and WB and WS at wk 6 and 7. One and 2-day postmortem, fillet compression force and water-holding capacity were measured. Serum corticosterone was reduced by up to 21% in laser-enriched birds wk 5 to 7 (P < 0.01). Serum myoglobin was increased in laser-enriched broilers by 5% on wk 5 (P < 0.01) but increased in control birds wk 6 to 7 by up to 13% (P < 0.01). Serum troponin was reduced in laser-enriched broilers by 9% at wk 5 (P < 0.01). Laser exposure increased breast width and fillet weight at d 42 by 1.08 cm (P < 0.05) and 30 g (P < 0.05). At d 49, fillet height was increased 0.42 cm in laser-enriched birds (P < 0.05). Laser enrichment reduced severe WS incidence at d 42 by 24% (P < 0.05) and on d 49 by 15% (P < 0.10). Severe WB score was numerically reduced by 11% in laser enrichment on d 42 and 18% on d 49 (P > 0.05). Water-holding capacity was improved in laser-enriched breasts (P < 0.01) and expression of myostatin and insulin-like growth factor 2 were increased on d 49 (P ≤ 0.01. Laser enrichment reduced markers of stress and muscle damage while improving breast muscle quality and is therefore a potential effective enrichment for commercial broilers.

Modeling cold curing of Pierce's disease in Vitis vinifera 'Pinot Noir' and 'Cabernet Sauvignon' grapevines in California.

Pierce's disease (PD) of Vitis vinifera grapevines is caused by the bacterium Xylella fastidiosa, a pathogen with a wide plant host range. Exposure of X. fastidiosa-infected plant tissue to cold temperatures has been shown to be effective at eliminating the pathogen from some plant hosts such as grapevines. This "cold curing" phenomenon suggests itself as a potential method for disease management and perhaps control. We investigated cold therapy of PD-affected 'Pinot Noir' and 'Cabernet Sauvignon' grapevine. In the fall, inoculated plants and controls of each cultivar were transported to each of four field sites in California (Foresthill, McLaughlin, Hopland, and Davis) that differed in the magnitude of cold winter temperatures. A model for progression of the elimination of plant disease in relation to temperature was conceptualized to be a temperature-duration effect, where temperatures below a particular threshold kill X. fastidiosa with increasing efficacy as the temperature decreases to some value <6?C. The temperature effect was modeled as a likelihood of a particular temperature killing the pathogen and is termed the ?killing index?. We developed a mathematical model for cold curing of grapevines inoculated with X. fastidiosa and calibrated the model with cold-curing data collected in a field study. Parameter estimation resulted in lowest sum of squared differences across all 10 trials to be low temperature below which the organism is killed (T(0)) = 6°C, number of hours to achieve 100% cure (N(100)) = 195 h, number of hours to achieve 10% cure (N(10)) = 20 h, and killing index (K(x)) = 0.45 for Pinot Noir and T(0) = 6°C, N(100) = 302 h, N(10) = 170 h, and K(x) = 0.41 for Cabernet Sauvignon. With the parameter estimates optimized by model calibration, the simulation model was effective at predicting cold curing in four locations during the experiment, although there were some differences between Hopland for Pinot Noir and Davis for Cabernet Sauvignon. Using historical temperature data, the model accurately predicted the known severity of PD in other grape-growing regions of California, suggesting that it may have utility in assessing the relative risk of developing PD in proposed new vineyard sites.

Algae-based feed ingredient protects intestinal health during Eimeria challenge and alters systemic immune responses with differential outcomes observed during acute feed restriction.

Compounds in microalgae-derived feed ingredients in poultry diets may improve intestinal physiology and immunity to protect against damage induced by physiological and pathogen challenges, but mechanisms are examined sparingly. The study objective was to evaluate changes to intestinal morphology, permeability, and systemic immunity in broilers fed a proprietary microalgae ingredient during 2 separate challenge studies. In study 1, two replicate 28 d battery cage trials used 200 Ross 308 broilers each (n = 400) fed a control diet ± 0.175% algae ingredient. Half of the birds were subjected to a 12 h feed restriction challenge and fluorescein isothiocyanate dextran (FITC-D) intestinal permeability assay on d 28. Study 2 used 800 broilers randomly assigned to the same dietary treatments and housed in floor pens for 42 d. At d 14, intestine and spleen samples were collected from 10 birds/ diet. Half of the remainder was orally inoculated with 10X Coccivac-B52 vaccine in a 2 × 2 factorial treatment design (diet and Eimeria inoculation). The FITC-D assay was conducted at 1, 3, 7, and 14 d post-inoculation (pi) while intestinal and spleen samples were collected at 3, 7, 14, and 28 dpi for histomorphology and flow cytometric immune cell assessment. Study 1 validated intestinal leakage via FITC-D absorbance induced by feed restriction but showed no algae-associated protective effects. In study 2, algae preserved intestinal integrity during coccidiosis (P = 0.04) and simultaneously protected jejunal villus height as early as 7dpi (P < 0.0001), whereas intestinal damage resolution in control birds did not occur until 14 dpi. Algae inclusion increased splenic T cells in unchallenged broilers at d 14 by 29.6% vs. control (P < 0.0001), specifically γδ T cell populations, without impacting performance (P < 0.03). During Eimeria challenge, splenic T cells in algae-fed birds did not show evidence of recruitment to peripheral tissues, while control birds showed a 16.7% reduction compared to their uninoculated counterparts from 3 to 7 dpi (P < 0.0001). This evidence suggests the algae ingredient altered the immune response in a manner that reduced recruitment from secondary lymphoid organs in addition to protecting intestinal physiology.

A novel environmental enrichment device improved broiler performance without sacrificing bird physiological or environmental quality measures.

Modern commercial broilers have been genetically selected for fast growth and heavy breast muscling, contributing to a top-heavy phenotype and increased leg lameness. A quick-growing phenotype coupled with poor leg health fosters inactivity. The objective of this study was to stimulate broiler movement using novel environmental enrichment and determine the impact of movement on production, leg health, and environmental parameters. A total of 1,200 Ross 308 broilers were housed in 40 pens with 30 birds/pen for 6 wk in 2 separate rooms (laser enrichment or control). Each enrichment device was mounted above 2 adjoining pens, projected 2 independent, randomly moving laser beams at the floor to stimulate innate predatory behavior, and was active 4 times daily in 4-min periods. Performance outcomes were calculated by pen and averaged per bird for each performance period and overall days 0 to 42. A total of 70 randomly selected focal birds were examined for breast blisters and footpad dermatitis each week and euthanized on day 42 for tibia quality measures. Air quality and litter moisture were sampled by week. Laser-enriched pens had greater average bird feed intake in starter (P < 0.001), grower (P = 0.004), finisher periods (P = 0.004), and overall days 0 to 42 (0.19 kg/bird; P = 0.0003). Average bird weight gain was also increased in enriched pens in each performance period: starter (P = 0.043), grower (P = 0.001), finisher (P < 0.001), and overall days 0 to 42 (0.24 kg/bird; P < 0.001). Enriched pens had improved feed conversion ratio (FCR) vs. control with a decrease of 3 FCR points in the grower (P = 0.031), 18 points in the finisher (P < 0.001), and 7 points overall (P < 0.001). Enriched pens had higher ADG during starter (P = 0.048), finisher (P < 0.001), and overall (5.7 g/bird/d; P < 0.001). No differences were found in breast blister, footpad dermatitis, tibia, air, or litter quality measures (P > 0.05). In summary, a novel enrichment device based on bird visual feeding and predatory instincts positively affected performance through decreased FCR and increased ADG without sacrificing external animal-based measures, tibia quality, or air or litter quality.

Can recognition of living things be selectively impaired?

Brain damage sometimes seems to impair recognition of living things, despite relatively preserved recognition of nonliving things. The most straightforward interpretation of this dissociation is that the recognition of living things depends on some specialized mechanisms that are not needed for the recognition of nonliving things. However, there are alternative interpretations of the dissociation in terms of the greater complexity or inter-item similarity of living things, or the more specific, within-category identifications that are usually required for living things. Surprisingly, the relevant tests to discriminate among these rival hypotheses have never been performed. We took the factors of visual complexity, inter-item similarity, specificity of identification, as well as others, into account in analyzing the visual recognition performance of two head-injured visual agnosic patients. In each case we found that recognition of living things was still disproportionately impaired when the effects of the other factors were accounted for.

T-cell receptor variable beta genes show differential expression in CD4 and CD8 T cells.

Studies in transgenic and inbred strains of mice have shown that the critical molecular interactions controlling positive selection involve major histocompatibility complex (MHC), T-cell receptor (TCR), and CD4 or CD8 coreceptor molecules. Correlations have been established between MHC gene products and the percentage of CD4 or CD8 T cells that express specific variable (V) beta-gene products as part of the alpha beta heterodimer. These studies have important implications regarding potential mechanisms of HLA-linked autoimmune diseases in humans. If similar interactions are required for positive selection in humans, one would predict that the TCR repertoire expressed by mature, peripheral blood CD4 and CD8 T cells would vary. To test this hypothesis the expression of specific TCR V beta-region genes by CD4 and CD8 T cells from healthy individuals was compared using both triple-color flow cytometry and polymerase chain reaction based experimental approaches. The results show that the TCR repertoire does vary as a function of CD4 and CD8 T-cell subsets. Among unrelated individuals certain V beta genes were consistently overrepresented in the CD4 population (V beta-5.1, -6.7a, and -18); some were skewed to the CD8 population (V beta-14) while others showed variable patterns (V beta-12 and -17). Deletion of entire V beta gene families was not observed suggesting that this is a rare event in humans. Attempts to correlate the expressed TCR repertoire in humans with HLA alleles will require consideration of these differences in expression as a function of subset.

Delays in insulin signaling towards glucose disposal in human skeletal muscle.

We explored whether the delay that occurs between a rise in plasma insulin and the increase of glucose disposal occurs before, at, or downstream of steps that are believed to be part of the insulin signaling cascade. Skeletal muscle biopsies were obtained from 16 nondiabetic subjects before, and 20 and 180 min after plasma insulin levels had been augmented in euglycemic hyperinsulinemic glucose clamps. Although plasma insulin had reached 98% of its final concentration within 10 min, insulin receptor kinase (IRK) activity, p85 associated with insulin receptor substrate-1 (IRS-1), IRS-1-associated phosphatidylinositol 3-kinase (PI3K) activity, and Thr(308)-protein kinase B (PKB) phosphorylation in the muscle biopsies at 20 min had reached only 60, 48, 34 and 47% respectively of those at 180 min. This suggests a delay before the level of IRK and little or no delay between IRK and PKB activation. The observation that glycogen synthase activity and glucose disposal at 20 min had both only reached 25% of the respective values at 180 min suggests an additional delay downstream of the investigated signaling steps.

Meropenem pharmacokinetics in a patient with multiorgan failure from Meningococcemia undergoing continuous venovenous hemodiafiltration.

Meropenem is a carbapenem antibiotic with a broad antibacterial spectrum of activity. Its main route of elimination is through the kidneys, with 63% of the drug excreted unchanged in the urine. Meropenem clearance is diminished in renal impairment; therefore, doses need to be adjusted in patients with varying degrees of renal function. An appropriate dose of meropenem for patients undergoing continuous venovenous hemodiafiltration (CVVHDF) is unknown. We evaluated the pharmacokinetics of meropenem in a patient with fulminant meningococcemia undergoing CVVHDF. Meropenem concentrations in serial venous, arterial, and ultrafiltrate samples after a 1 g intravenous dose were measured using high-performance liquid chromatography (HPLC). Meropenem clearance was found to be 129.36 mL/min and 141.29 mL/min for every 8- and 12-hour dosing, respectively. Trough levels were above the MIC90 for Neisseria meningitidis and most anaerobic pathogens. We recommend that meropenem 1 g intravenously every 12 hours be used as the initial dose in patients undergoing CVVHDF. Differences between meropenem clearance during CVVHDF and other forms of renal replacement therapy are discussed.

The amino acid substrate of bovine tyrosine hydroxylase.

Tyrosine hydroxylase catalyzes the tetrahydropterin-dependent hydroxylation of tyrosine to form 3,4-dihydroxyphenylalanine. Several nonphysiological aromatic amino acids have been examined as inhibitors and substrates for bovine adrenal tyrosine hydroxylase. The Ki values for para-substituted phenylalanines increase as the size of the substituent increases. For each A2 increase in surface area of the substituent, the free energy of binding becomes 50 cal more positive. Replacement of the phenyl ring with a pyridyl ring decreases the affinity about one order of magnitude. A number of these aromatic amino acids are also substrates for the enzyme. The KM values again increase in size with increasing size of the substituent, but the Vmax value is independent of the reactivity of the amino acid. The effect of size on binding is consistent with a tight interaction between the para position region of the substrate and the enzyme. The lack of a change in the Vmax value is consistent with the rate-limiting step in catalysis by bovine tyrosine hydroxylase being formation of the hydroxylating intermediate rather than hydroxylation of the amino acid. These results will be useful in designing mechanism-based inhibitors of catecholamine biosynthesis and establish that the mechanisms of rat and bovine tyrosine hydroxylase do not differ significantly.

Dewar benzene as a protecting group? Demonstration of photolithographic crystallization.

The concept of Dewar benzene as a supramolecular protecting group for solid-state aryl-aryl interactions is reported. Photoisomerization of 1,4,5,6-tetramethyl-bicyclo[2.2.0]hexa-2,5-diene-2,3-dicarboxylic acid dimethyl ester (1) to the corresponding benzene isomer proceeds with rapid crystal formation. Herein this property is applied to the photolithographic patterning of crystal domains on a surface. [structure: see text]

Arabidopsis thaliana LUP1 converts oxidosqualene to multiple triterpene alcohols and a triterpene diol.

The Arabidopsis thaliana LUP1 gene encodes an enzyme that converts oxidosqualene to pentacyclic triterpenes. Lupeol and beta-amyrin were previously reported as LUP1 products. Further investigation described here uncovered the additional products germanicol, taraxasterol, psi-taraxasterol, and 3,20-dihydroxylupane. These results suggest that the 80 known C(30)H(50)O compounds that are structurally consistent with being oxidosqualene cyclase products may be derived from fewer than 80 enzymes and that some C(30)H(52)O(2) compounds may be direct cyclization products of oxidosqualene.

Steric bulk at cycloartenol synthase position 481 influences cyclization and deprotonation.

Cycloartenol synthase converts oxidosqualene to the pentacyclic sterol precursor cycloartenol. An Arabidopsis thaliana cycloartenol synthase Ile481Val mutant was previously shown to produce lanosterol and parkeol in addition to its native product cycloartenol. Experiments are described here to construct Phe, Leu, Ala, and Gly mutants at position 481 and to determine their cyclization product profiles. The Phe mutant was inactive, and the Leu mutant produced cycloartenol and parkeol. The Ala and Gly mutants formed lanosterol, cycloartenol, parkeol, achilleol A, and camelliol C. Monocycles comprise most of the Gly mutant product, showing that an alternate cyclization route can be made the major pathway by a single nonpolar mutation.

Interleukin-12 p40 m-RNA expression in human kidney allograft biopsies.

Interleukin-12 (IL-12) is a heterodimeric cytokine implicated in the early differentiation of naive T-lymphocytes into the Th1 subset. IL-12 is important for induction of the cellular immune response against viruses, intracellular parasites and neoplasms. Its role in alloresponsiveness has not been fully elucidated. Preliminary data in the literature point toward the prevalence of Th1 lymphocytes in processes of allograft rejection. In attempt to further investigate the expression of this cytokine during episodes of cellular rejection of renal allografts, we searched for IL-12 message in human kidney allograft biopsies using the reverse transcriptase-polymerase chain reaction technique. Twenty-three allograft core biopsies from 19 patients were obtained percutaneously for clinical indications in 18 cases, and as part of an investigational protocol in five cases. A portion of the tissue was used for RNA extraction using the guanidium-thiocyanide phenol-chloroform method. Histology was performed on the remaining core material. Ten mg of total RNA were used for reverse transcription. PCR of the c-DNAs was done for 40 cycles using primers for the p40 subunit of IL-12 and GAPDH which was used as a control. PCR products were photographed after electrophoresis, transferred to a nylon membrane and hybridized with a radiolabelled cloned human IL-12 p40 1 kb c-DNA fragment. Autoradiographies were developed after 20-min exposure. All samples were run in triplicate. IL-12 p40 m-RNA was expressed in all 17 biopsies showing acute cellular rejection as well as in all three biopsies showing focal interstitial fibrosis. No message was found in the presence of normal allograft histology. This is the first in vivo report of IL-12 p40 subunit m-RNA expression during renal allograft rejection in humans. The role of this Th1 cytokine in the alloresponse deserves further investigation.

Interaction between cyclosporine and grapefruit juice requires long-term ingestion in stable renal transplant recipients.

STUDY OBJECTIVE: To examine the effect of the concurrent administration of increasing amounts of grapefruit juice, an inhibitor of drug metabolism, on the steady-state pharmacokinetics of cyclosporine. DESIGN: Open-label, three-period crossover, food-drug interaction study in stable renal transplant patients. SETTING: A university-affiliated clinical research center. PATIENTS: Sixteen stable renal transplant recipients. INTERVENTION: Cyclosporine was administered with 240 ml of water, 240 ml of grapefruit juice, or several 240-ml glasses of grapefruit juice, and serial blood samples were taken to estimate the effect of grapefruit juice on cyclosporine pharmacokinetics. MEASUREMENTS AND MAIN RESULTS: Grapefruit juice caused a significant increase in cyclosporine area under the curve, however, no significant effect was seen in other pharmacokinetic parameters. Grapefruit juice caused an increase in the 24-hour trough cyclosporine concentration, which may be of clinical significance if long-term ingestion of grapefruit juice is recommended. CONCLUSION: A drug interaction exists between cyclosporine and grapefruit juice, and it is likely at the level of intestinal drug absorption.

Renal replacement therapies.

Slow continuous renal replacement therapy is more hemodynamically stabilizing and is replacing conventional hemodialysis as the therapy of choice for acute renal failure in the intensive care unit. This article presents practical information, including basic terminology, basic physiology, technical aspects, and indications for and application of this technique.

Efficacy of an enteral 10-day methadone wean to prevent opioid withdrawal in fentanyl-tolerant pediatric intensive care unit patients.

OBJECTIVE: To demonstrate the efficacy of a 10-day, single daily dose, enteral methadone weaning protocol for the prevention of opioid withdrawal symptoms in pediatric patients after prolonged fentanyl exposure. DESIGN: Prospective, observational study. SETTING: Twenty-four-bed medical-surgical intensive care unit within a 222-bed pediatric teaching hospital. PATIENTS: Twenty-nine children, aged 1 day to 19.8 yrs, who received methadone to prevent opioid withdrawal after prolonged continuous fentanyl infusion exposure. INTERVENTIONS: Institution of a standardized methadone weaning protocol. MEASUREMENTS AND MAIN RESULTS: All 29 patients had received a continuous fentanyl infusion; duration of exposure was 14.5 +/- 9.2 days, cumulative fentanyl dose was 1.93 +/- 1.53 mg/kg, and peak fentanyl infusion was 9.6 +/- 4.3 microg/kg per hr. Twenty-five (86%) of 29 patients successfully completed the methadone wean in 10 days. Withdrawal complications that required a weaning delay were seen in three patients, and one patient expired before completion. Sixteen patients were discharged to complete their weaning schedule at home without incident. CONCLUSIONS: Opioid withdrawal symptoms in pediatric intensive care unit patients after prolonged fentanyl exposure can be avoided by using an enteral, 10-day, single daily dose methadone weaning protocol.

Cloning and characterization of the Dictyostelium discoideum cycloartenol synthase cDNA.

Cycloartenol synthase converts oxidosqualene to cycloartenol, the first carbocyclic intermediate en route to sterols in plants and many protists. Presented here is the first cycloartenol synthase gene identified from a protist, the cellular slime mold Dictyostelium discoideum. The cDNA encodes an 81-kDa predicted protein 50-52% identical to known higher plant cycloartenol synthases and 40-49% identical to known lanosterol synthases from fungi and mammals. The encoded protein expressed in transgenic Saccharomyces cerevisiae converted synthetic oxidosqualene to cycloartenol in vitro. This product was characterized by 1H and 13C nuclear magnetic resonance and gas chromatography-mass spectrometry. The predicted protein sequence diverges sufficiently from the known cycloartenol synthase sequences to dramatically reduce the number of residues that are candidates for the catalytic difference between cycloartenol and lanosterol formation.