Immune responses in hibernating little brown myotis (Myotis lucifugus) with white-nose syndrome.

White-nose syndrome (WNS) is a fungal disease responsible for decimating many bat populations in North America. Pseudogymnoascus destructans (Pd), the psychrophilic fungus responsible for WNS, prospers in the winter habitat of many hibernating bat species. The immune response that Pd elicits in bats is not yet fully understood; antibodies are produced in response to infection by Pd, but they may not be protective and indeed may be harmful. To understand how bats respond to infection during hibernation, we studied the effect of Pd inoculation on the survival and gene expression of captive hibernating Myotis lucifugus with varying pre-hibernation antifungal antibody titres. We investigated gene expression through the transcription of selected cytokine genes (Il6, Il17a, Il1b, Il4 and Ifng) associated with inflammatory, Th1, Th2 and Th17 immune responses in wing tissue and lymph nodes. We found no difference in survival between bats with low and high anti-Pd titres, although anti-Pd antibody production during hibernation differed significantly between infected and uninfected bats. Transcription of Il6 and Il17a was higher in the lymph nodes of infected bats compared with uninfected bats. Increased transcription of these cytokines in the lymph node suggests that a pro-inflammatory immune response to WNS is not restricted to infected tissues and occurs during hibernation. The resulting Th17 response may be protective in euthermic bats, but because it may disrupt torpor, it could be detrimental during hibernation.

A high throughput in vitro analytical approach to screen for oxidative stress potential exerted by nanomaterials using a biologically relevant matrix: human blood serum.

Limited studies have shown that selected nanomaterials (NMs) impart various forms of toxicity in biological systems; however, a common metric to screen for potential toxicity is needed. This study optimized and utilized a 'Ferric reducing ability of serum (FRAS)' assay as a screening tool to quantitate the degree of oxidative damage induced by NMs on human blood serum. Antioxidants in blood protect against oxidative damage caused by free radicals via chemical quenching and will decrease when exposed to oxidatively stressful materials. Using this approach, the antioxidant capacity of NM treated serum was significantly decreased by nano-silver, a series of nano-carbon blacks, fullerene soot, and nano-TiO(2) (anatase, p<0.05), but not with nano-alumina, fullerite, purified fullerene, fine TiO(2) (rutile) and Min-U-Sil 5. Particle surface area and not biological particle size was highly associated with the degree of oxidative stress observed. This approach appears responsive to multiple determinants of oxidative damage, including particle chemistry, surface area and impurities, and may be a valid screening method to determine oxidative damage imparted by nanomaterials.

Micro-CT evaluation of rheumatoid arthritis mouse model disease progression: Manual tracings versus semi-automated routines.

PURPOSE: The primary goal of this study was to demonstrate the value of micro-CT imaging in a rheumatoid arthritis (RA) mouse model. The secondary goal was to assess whether manual correction of the articular surface regions of interest (ROI) identification of the semi-automated methods may result in more effective assessment of bone volume and density loss. MATERIALS AND METHODS: Collagen-induced arthritis (CIA) was induced in six DBA/1J mice at 12 weeks of age and three other DBA/1J identical mice served as controls. Micro-CT images were acquired at baseline and at four, seven, and nine weeks post-induction. Disease was monitored via ROI analysis, and ROIs were first generated using semi-automated techniques. These ROIs were manually manipulated so that a variety of edge irregularities were corrected. Effort was focused on the proximal and distal humerus and the distal femur. ROI volume and density were calculated, and data were compared. A histologic analysis of the study mice was also performed after the last time frame. RESULTS: There was a significant difference between the volume data comparison between the manually manipulated data and the semi-automated routine data across all time frames and across both humeri and femurs. There was no significant difference in densities calculated in Hounsfield units across any of the time frames, humeri or femurs, except for one time frame. CONCLUSION: Our findings suggest that the manual correction technique of semi-automated data can be used to quantify and evaluate bone volume, density, and joint surface architecture changes in a RA mouse model.

Constitutive expression of cat-86 associated with a change in the transcription start point.

The translational attenuation regulatory model suggests a mechanism that can explain the induction of cat-86 by chloramphenicol (Cm). In this model, Cm serves to stall a ribosome at a specific site in a leader region of cat-86 transcripts. The stalled ribosome is thought to destabilize a downstream region of RNA secondary structure that normally sequesters the cat-86 ribosome-binding site (RBS-3). Three mutations in codon 4 of the cat-86 leader have been identified which result in constitutive cat expression. Each of the three mutations generates a likely -10 promoter sequence in the leader. Twenty nucleotides (nt) upstream is the wild-type sequence, 5'-TTGAAA, which differs from the consensus sigA -35 domain by only a single nt. The transcription start point from the resulting mutant promoter is within the DNA region that normally specifies the RNA secondary structure that sequesters cat-86 RBS-3. Thus, the basis for the constitutive phenotype is the absence of the RNA secondary structure in the transcripts driven by the promoter generated through mutagenesis of leader codon 4.

Studies of cholesterol and bile acid metabolism, and early atherogenesis in hamsters fed GT16-239, a novel bile acid sequestrant (BAS).

The purpose of this study was to compare the efficacy of GT16-239, an alkylated, cross-linked poly(allylamine) bile acid sequestrant with cholestyramine on cholesterol and bile acid metabolism, and early aortic atherosclerosis in hypercholesterolemic male F1B Golden Syrian hamsters. In this controlled study, 42 hamsters were divided into six groups and were fed a chow-based hypercholesterolemic diet supplemented with a 10% oil blend (55% coconut/45% corn), 0.1% cholesterol (w/w) (control) and either 0.9 or 1.2% cholestyramine or 0.2, 0.4 or 0.6% GT16-239 for 13 weeks. Laboratory analyses included evaluating plasma lipoprotein cholesterol and triglyceride concentrations, hepatic HMG-CoA reductase and 7 alpha-hydroxylase activities, fecal excretion of bile acids and neutral sterols, hepatic cholesterol concentrations, and early atherosclerosis (aortic fatty streak area). Relative to the control diet, the 0.6% GT16-239 versus the 1.2% cholestyramine significantly inhibited the elevation of plasma lipoprotein total cholesterol (TC) (-69% vs -40%), high density lipoprotein-cholesterol (HDL-C) (-49% vs -30%), and non-HDL-C (-81 vs -48%) concentrations; increased the activities of both HMG-CoA reductase (1492% vs 62%) and 7 alpha-hydroxylase (175% vs 86%); lowered the concentration of hepatic cholesteryl ester (-94% vs -59%); increased fecal cholesterol concentration (+28% vs -10%); and decreased aortic fatty streak area (-100% vs -86%). Unexpected findings of this comparison were increased fecal concentrations of cholic acid (533%) and chenodeoxycholic acid (400%) and the reduction in lithocholic acid (-50%) in the 0.6% GT16-239 compared to the 1.2% cholestyramine group. In summary, GT16-239 had a greater impact on cholesterol metabolism and early atherosclerosis in hypercholesterolemic hamsters than cholestyramine.

Diurnal variation in sensory and pain thresholds correlated with mood states.

Twenty-two healthy volunteers were tested for diurnal variations in six mood states and in the sensory and pain thresholds. Cutaneous electrical stimulation was used to detect sensory and pain thresholds and a Mood Questionnaire was used to assess mood states. Both sensory and pain thresholds were found to be significantly lower in the afternoon than in the morning. The detection and pain thresholds were not related to the mood states.

Goals in hemiplegia care.

Physicians should assume leadership and a greater role in the post-stroke management of the hemiplegic patient, so that the recipient of care becomes an active participant in the treatment team. The patient should be fully aware of his condition, and partake in establishment of the goals together with subsequent evaluation of accomplishments and progress. Some short-term and some longer-term goals are outlined as a guide. A method for performance scoring is suggested. These performance ratings may assist in establishing priorities for discharge planning and continuation of care. They also may permit more accurate assessment and documentation of the patient's capacities and the efficacy of the various approaches to adequate care.

Regulation of plasma lipoprotein levels by dietary triglycerides enriched with different fatty acids.

Saturated vegetable oils (coconut, palm, and palm kernel oil) containing predominantly saturated fatty acids, lauric (12:0) or myristic (14:0 and palmitic (16:0), raise plasma total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) levels in animals and humans, presumably by decreasing LDL receptor activity and/or increasing LDL-C production rate. Although stearic acid (18:0) is chemically a saturated fatty acid, both human and animal studies suggest it is biologically neutral (neither raising nor lowering) blood cholesterol levels. Although earlier studies indicated that medium chain fatty acids (8:0-10:0) were also thought to be neutral, more recent studies in animals and humans suggest otherwise. Unsaturated vegetable oils such as corn, soybean, olive, and canola oil, by virtue of their predominant levels of either linoleic acid (18:2) or oleic acid (18:1), are hypocholesterolemic, probably as a result of their ability to upregulate LDL receptor activity and/or decrease LDL-C production rate. Whether trans fatty acids such as trans oleate (t18:1), in hydrogenated products such as margarine, are hypercholesterolemic remains controversial. Studies in humans suggest that their cholesterol-raising potential falls between the native nonhydrogenated vegetable oil and the more saturated dairy products such as butter. Assessment of the magnitude of the cholesterolemic response of trans 18:1 is difficult because in most diet studies its addition is often at the expense of cholesterol-lowering unsaturated fatty acids, making an independent evaluation almost impossible.

Mosquitoes (Diptera: Culicidae) captured in the Iquitos area of Peru.

A mosquito capture program was initiated to study mosquito species and their potential for arboviral transmission in the Peruvian Amazon. More than 35,000 mosquitoes of 13 different genera and at least 25 species were captured in urban and sylvan sites in the Iquitos area. These findings represent the first published list of Peruvian mosquitoes since 1971 and the first such list from the Peruvian Amazon.

Apple juice prevents oxidative stress and impaired cognitive performance caused by genetic and dietary deficiencies in mice.

Increased oxidative stress contributes to the decline in cognitive performance during normal aging and in neurodegenerative conditions such as Alzheimer s disease. Dietary supplementation with fruits and vegetables that are high in antioxidant potential have in some cases compensated for dietary and/or genetic deficiencies that promote increased oxidative stress. Herein, we demonstrate that apple juice concentrate, administered ad libitum in drinking water, can compensate for the increased reactive oxygen species and decline in cognitive performance in maze trials observed when normal and transgenic mice lacking apolipoprotein E are deprived of folate and vitamin E. In addition, we demonstrate that this protective effect is not derived from the sugar content of the concentrate.

Effectiveness of permethrin-treated military uniform fabric against human body lice.

Military uniform fabric patches treated with permethrin were evaluated against natural and laboratory strains of human body lice, Pediculus humanus, L. Permethrin-treated fabric was toxic to body lice on contact and quickly affected feeding behavior and the likelihood of disease transmission, even when washed up to 20 times. The use of permethrin-treated clothing offers a new passive approach in human louse control not previously feasible. Military personnel wearing permethrin-treated uniforms, therefore, can expect significant and long-term protection from lice and louse-borne diseases in endemic areas.

Postanesthesia care of the cocaine abuser.

The use of cocaine is increasing in the United States. An increasing number of cocaine users will be admitted to the PACU, requiring nurses to be knowledgeable about this hazardous drug. Cocaine produces harmful effects on the central nervous, respiratory, and cardiovascular systems. Nursing process can be used to guide the patient's care. Changes in physical, mental, and emotional status must be assessed often. Multiple nursing diagnoses are appropriate for this patient in the PACU. Maintaining airway and alleviating pain are nursing interventions that challenge the PACU nurse. The goal of nursing care is to provide a safe, quiet, and comfortable recovery from anesthesia. Rehabilitation is the long-range goal for the patient who needs expert nursing care, a therapeutic environment, and understanding.

The cis-effect of a nascent peptide on its translating ribosome: influence of the cat-86 leader pentapeptide on translation termination at leader codon 6.

Inducible cat genes from Gram-positive bacteria are regulated by translation attenuation. The inducer chloramphenicol stalls a ribosome at a specific site in the leader of cat transcripts; this destabilizes a downstream stem-loop structure that normally sequesters the ribosome-binding site for the cat structural gene. The five-amino-acid peptide MVKTD that is synthesized when a ribosome has translated to the leader induction site is an inhibitor of peptidyl transferase in vitro. Thus, the peptide may be the in vivo determinant of the site of ribosome stalling. Here we provide evidence that the leader pentapeptide can exert a cis-effect on its translating ribosome in vivo. Converting leader codon 6 to the ochre codon results in expression of cat-86 in the absence of inducer. We term this autoinduction. Autoinduction is abolished by mutations that change the amino-acid sequence of the leader peptide but have no, or little, effect on the sequence of nucleotides at the leader stall site. In contrast, four nucleotide changes within the leader site occupied by the stalled ribosome that result in synonymous codon replacements do not diminish autoinduction. Our evidence indicates that the cat-86 leader pentapeptide can alter the function of its translating ribosome.

Sample preparation for gas chromatography with electron capture detection: determination of total and free thyroxin in serum.

Relatively clean gas chromatography with electron capture detection (GC-ECD) chromatograms are obtained for both total and free thyroxin (T4) in serum by improving sample preparation. This is based on establishing a sequence of steps that cumulatively overcome two classes of interference: those present in the initial sample and those introduced by the procedure. The main source for the latter contaminants is the derivatization step, a problem that was largely overcome by employing HPLC after this step. Also it is helpful to use ion-exchange columns early in the procedure under fast-flow conditions with intermediate flows of air to speed up and enhance their reliability. The work establishes some guidelines for future applications of GC-ECD to the determination of sub-nanogram analytes requiring derivatization, an area in which GC-ECD has been remiss in the past. As a side benefit, total T4 in serum is determined by HPLC for the first time with uv detection.

Pain clinic #14. Fibromyalgia and myofascial pain: either, neither, or both?

Musculoskeletal pain syndromes and the accompanying physical and emotional sequelae are frequent reasons for self-medication and physician visits. Most musculoskeletal pain syndromes are self-limited, periodically flaring up and subsiding. Fibrositis and myofascial pain syndromes, which affect a significant number of patients with musculoskeletal pain, should be clinically differentiated to reduce unnecessary work-ups and improve patient management at reduced costs. The more common pain locations are reviewed, and a cost-effective, comprehensive approach to management is outlined.

Ribosome regulation by the nascent peptide.

Studies of bacterial and eukaryotic systems have identified two-gene operons in which the translation product of the upstream gene influences translation of the downstream gene. The upstream gene, referred to as a leader (gene) in bacterial systems or an upstream open reading frame (uORF) in eukaryotes, encodes a peptide that interferes with a function(s) of its translating ribosome. The peptides are therefore cis-acting negative regulators of translation. The inhibitory peptides typically consist of fewer than 25 residues and function prior to emergence from the ribosome. A biological role for this class of translation inhibitor is demonstrated in translation attenuation, a form or regulation that controls the inducible translation of the chloramphenicol resistance genes cat and cmlA in bacteria. Induction of cat or cmlA requires ribosome stalling at a particular codon in the leader region of the mRNA. Stalling destabilizes an adjacent, downstream mRNA secondary structure that normally sequesters the ribosome-binding site for the cat or cmlA coding regions. Genetic studies indicate that the nascent, leader-encoded peptide is the selector of the site of ribosome stalling in leader mRNA by cis interference with translation. Synthetic leader peptides inhibit ribosomal peptidyltransferase in vitro, leading to the prediction that this activity is the basis for stall site selection. Recent studies have shown that the leader peptides are rRNA-binding peptides with targets at the peptidyl transferase center of 23S rRNA. uORFs associated with several eukaryotic genes inhibit downstream translation. When inhibition depends on the specific codon sequence of the uORF, it has been proposed that the uORF-encoded nascent peptide prevents ribosome release from the mRNA at the uORF stop codon. This sets up a blockade to ribosome scanning which minimizes downstream translation. Segments within large proteins also appear to regulate ribosome activity in cis, although in most of the known examples the active amino acid sequences function after their emergence from the ribosome, cis control of translation by the nascent peptide is gene specific; nearly all such regulatory peptides exert no obvious trans effects in cells. The in vitro biochemical activities of the cat/cmla leader peptides on ribosomes and rRNA suggest a mechanism through which the nascent peptide can modify ribosome behavior. Other cis-acting regulatory peptides may involve more complex ribosomal interactions.

Erythromycin induces expression of the chloramphenicol acetyltransferase gene cat-86.

The plasmid gene cat-86 specifies chloramphenicol-inducible chloramphenicol acetyltransferase in Bacillus subtilis. This gene, like the erythromycin-inducible erm genes, is regulated by translational attenuation. Here we show that cat-86 is also inducibly regulated by erythromycin. cat-86 does not confer resistance to erythromycin.

Peptidyl transferase inhibition by the nascent leader peptide of an inducible cat gene.

The site of ribosome stalling in the leader of cat transcripts is critical to induction of downstream translation. Site-specific stalling requires translation of the first five leader codons and the presence of chloramphenicol, a sequence-independent inhibitor of ribosome elongation. We demonstrate in this report that a synthetic peptide (the 5-mer) corresponding to the N-terminal five codons of the cat-86 leader inhibits peptidyl transferase in vitro. The N-terminal 2-, 3-, and 4-mers and the reverse 5-mer (reverse amino acid sequence of the 5-mer) are virtually without effect on peptidyl transferase. A missense mutation in the cat-86 leader that abolishes induction in vivo corresponds to an amino acid replacement in the 5-mer that completely relieves peptidyl transferase inhibition. In contrast, a missense mutation that does not interfere with in vivo induction corresponds to an amino acid replacement in the 5-mer that does not significantly alter peptidyl transferase inhibition. Our results suggest that peptidyl transferase inhibition by the nascent cat-86 5-mer peptide may be the primary determinant of the site of ribosome stalling in the leader. A model based on this concept can explain the site specificity of ribosome stalling as well as the response of induction to very low levels of the antibiotic inducer.