Eukaryotic inhibitors or activators elicit responses to chemosensory compounds by ruminal isotrichid and entodiniomorphid protozoa.

Our objectives were to evaluate potential signaling pathways regulating rumen protozoal chemotaxis using eukaryotic inhibitors potentially coordinated with phagocytosis as assessed by fluorescent bead uptake kinetics. Wortmannin (inhibitor of phosphoinositide 3-kinase), insulin, genistein (purported inhibitor of a receptor tyrosine kinase), U73122 (inhibitor of phospholipase C), and sodium nitroprusside (Snp, nitric oxide generator, activating protein kinase G) were preincubated with mixed ruminal protozoa for 3h before assessing uptake of fluorescent beads and chemosensory behavior to glucose, peptides, and their combination; peptides were also combined with guanosine triphosphate (GTP; a chemorepellent). Entodiniomorphids were chemoattracted to both glucose and peptides, but chemoattraction to glucose was increased by Snp and wortmannin without effect on chemoattraction to peptides. Rate of fluorescent bead uptake by an Entodinium caudatum culture decreased when beads were added simultaneously with feeding and incubated with wortmannin (statistical interaction). Wortmannin also decreased the proportion of mixed entodiniomorphids consuming beads. Isotrichid protozoa exhibited greater chemotaxis to glucose but, compared with entodiniomorphids, were chemorepelled to peptides. Wortmannin increased chemotaxis by entodiniomorphids but decreased chemotaxis to glucose by isotrichids. Motility assays documented that Snp and wortmannin decreased net swimming speed (distance among 2 points per second) but not total swimming speed (including turns) by entodiniomorphids. Wortmannin decreased both net and total swimming behavior in isotrichids. Results mechanistically explain the isotrichid migratory ecology to rapidly take up newly ingested sugars and subsequent sedimentation back to the ventral reticulorumen. In contrast, entodiniomorphids apparently integrate cellular motility with feeding behavior to consume small particulates and thereby stay associated and pass with the degradable fraction of rumen particulates. These results extend findings from aerobic ciliate models to explain how rumen protozoa have adapted physiology for their specific ecological niches.

Dimensions of wisdom perception across twelve countries on five continents.

Wisdom is the hallmark of social judgment, but how people across cultures recognize wisdom remains unclear-distinct philosophical traditions suggest different views of wisdom's cardinal features. We explore perception of wise minds across 16 socio-economically and culturally diverse convenience samples from 12 countries. Participants assessed wisdom exemplars, non-exemplars, and themselves on 19 socio-cognitive characteristics, subsequently rating targets' wisdom, knowledge, and understanding. Analyses reveal two positively related dimensions-Reflective Orientation and Socio-Emotional Awareness. These dimensions are consistent across the studied cultural regions and interact when informing wisdom ratings: wisest targets-as perceived by participants-score high on both dimensions, whereas the least wise are not reflective but moderately socio-emotional. Additionally, individuals view themselves as less reflective but more socio-emotionally aware than most wisdom exemplars. Our findings expand folk psychology and social judgment research beyond the Global North, showing how individuals perceive desirable cognitive and socio-emotional qualities, and contribute to an understanding of mind perception.

Male reproductive system defects and subfertility in a mutant mouse model of oculodentodigital dysplasia.

Oculodentodigital dysplasia (ODDD) is a dysmorphogenesis syndrome resulting from mutations in the GJA1 gene encoding the gap junction protein, connexin43 (CX43). In the testis this connexin localizes between Leydig cells, Sertoli cells and between Sertoli cells and germ cells. It is essential for Sertoli cell differentiation and spermatogenesis. This study explored male fertility in Gja1(Jrt) /+ mice which carry a dominant mutation that causes an amino acid substitution (G60S) in CX43. Gja1(Jrt) /+ mice mimic the phenotype of ODDD. Immunodetection methods revealed a reduction of both total CX43 and CX43 in membrane plaques in mutant testes. Correspondingly, intercellular coupling along the tubules was diminished as revealed by fluorescent dye transfer. Light and electron microscopy revealed loss of germ cells and sloughing of germ cells into the tubular lumen. There were also irregularities in size and shape of Sertoli cell nuclei. Analyses of cauda epididymal sperm indicated significant decreases in sperm count and sperm velocity parameters associated with sperm vigour, and significantly lower sperm head movement parameters associated with progressiveness. A significant decrease was also observed in total per cent motility. These results further confirm a critical role for CX43 in spermatogenesis and sperm maturation and support the possibility of subfertility in ODDD human males.

Operation of a continuous flow liquid helium magnetic microscopy cryostat as a closed cycle system.

We demonstrate successful operation of a continuous flow liquid helium magnetic cryostat (Oxford Instruments, Microstat MO) in closed cycle operation using a modular cryocooling system (ColdEdge Technologies, Stinger). For the system operation, we have developed a custom gas handling manifold and we show that despite the lower cooling power of the cryocooler with respect to the nominal cryostat cooling power requirements, the magnetic cryostat can be operated in a stable manner. We provide the design of the gas handling manifold and a detailed analysis of the system performance in terms of cooling times, magnetic field ramping rates, and vibrations at the sample. Base temperatures can be reached within 10 h while the superconducting magnet can be energized at a ramping rate of 0.5 T/min. Vibrations are measured interferometrically and show amplitudes with a root mean square on the order of 5 nm, permitting the use of the system for sensitive magnetic microscopy experiments.

Model assisted startup of anaerobic digesters fed with thermally hydrolysed activated sludge.

This paper presents the use of the IWA ADM1 to predict and interpret results from two full-scale anaerobic digesters fed with thermal hyrolysate (waste activated sludge with a long upstream sludge age) from a Cambi hydrolysis process operating at 165°C and 6 bar-g. The first digester was fed conventionally-though intermittently, while the second was heavily diluted through a substantial component of the evaluation period (110 days). There were a number of important outcomes-related to both model application, and model predictions. Input and inert COD: mass ratio was very important, and was considerably higher than the 1.42 g g⁻¹ used for biomass throughout the IWA activated sludge and anaerobic digestion models. Input COD: VS ratio was 1.6 g g⁻¹, and inert COD: VS ratio was 1.7 g g⁻¹. The model succeeded on a number of levels, including effective prediction of important outputs (degradability, gas flow and composition, and final solids), clarification of the substantial data scatter, prediction of recovery times during operationally poor periods, and cross-validation of the results between digester 1 and digester 2. Key failures in model performance were related to an early incorrect assumption of the COD: VS ratio of 1.42 g g⁻¹, and intermittent high acetate levels, most likely caused by inhibition, and rapid acclimatisation to ammonia. The acute free ammonia limit was found to be 0.008 M NH(3)-N, while the chronic inhibition constant (K(I,NH3,ac)) was 0.007 ± 0.001 M NH3-N. Overall, this is a complex system, and application of the model added significant confidence to the initial operational decisions during an aggressive startup on an atypical feed.

Double deletion of Panx1 and Panx3 affects skin and bone but not hearing.

Pannexins (Panxs), large-pore channel forming glycoproteins, are expressed in a wide variety of tissues including the skin, bone, and cochlea. To date, the use of single knock-out mouse models of both Panx1 and Panx3 have demonstrated their roles in skin development, bone formation, and auditory phenotypes. Due to sequence homology between Panx1 and Panx3, when one Panx is ablated from germline, the other may be upregulated in a compensatory mechanism to maintain tissue homeostasis and function. To evaluate the roles of Panx1 and Panx3 in the skin, bone, and cochlea, we created the first Panx1/Panx3 double knock-out mouse model (dKO). These mice had smaller litters and reduced body weight compared to wildtype controls. The dKO dorsal skin had decreased epidermal and dermal area as well as decreased hypodermal area in neonatal but not in older mice. In addition, mouse skull shape and size were altered, and long bone length was decreased in neonatal dKO mice. Finally, auditory tests revealed that dKO mice did not exhibit hearing loss and were even slightly protected against noise-induced hearing damage at mid-frequency regions. Taken together, our findings suggest that Panx1 and Panx3 are important at early stages of development in the skin and bone but may be redundant in the auditory system. KEY MESSAGES: Panx double KO mice had smaller litters and reduced body weight. dKO skin had decreased epidermal and dermal area in neonatal mice. Skull shape and size changed plus long bone length decreased in neonatal dKO mice. dKO had no hearing loss and were slightly protected against noise-induced damage.

Evaluation, selection and initial performance of a large scale centralised biosolids facility at Oxley Creek Water Reclamation Plant, Brisbane.

Recent upgrades to Brisbane City Council's Water Reclamation Plants (WRP) for improved nutrient removal has led to a significant increase in sludge production, and loss in potential anaerobic degradability. This increase in sludge production brought forth an economic driver for an improved, integrated biosolids handling strategy. The solution adopted by Brisbane Water, was a centralised thermal hydrolysis facility at Oxley Creek WRP to process the waste activated sludges generated at a total of five WRPs in the region. The thermal hydrolysis process uses high temperature and pressure to make non-degradable material more readily digestible by anaerobic bacteria and hence achieve greater efficiency in the overall anaerobic digestion process. Dewaterability is also improved, as the thermal hydrolysis process releases some of the bound water from the waste activated sludge. The viability of this solution is linked primarily to the reduction in mass and volume of biosolids which leads to lower transportation and disposal costs. After four months of operation, the Oxley Creek thermal hydrolysis plant has resulted in a 70% reduction in biosolids quantities (bulk volume) and disposal costs. The process is currently being optimised. Savings to date have reached about AUD 80,000 per month. Technical and operational issues which emerged during commissioning are also described.

Phenotypic characterisation of cell populations in the brains of horses experimentally infected with West Nile virus.

BACKGROUND: West Nile virus (WNV), a mosquito borne member of the Flaviviridae, is one of the most commonly diagnosed agents of viral encephalitis in horses and people worldwide. OBJECTIVES: A cassette of markers for formalin-fixed paraffin-embedded tissue and an archive of tissues from experimental infections in the horse were used to investigate the equine neuroimmune response to WNV meningoencephalomyelitis to phenotype the early response to WNV infection in the horse. STUDY DESIGN: Quantitative analysis using archived tissue from experimentally infected horses. METHODS: The thalamus and hindbrain from 2 groups of 6 horses were compared and consisted of a culture positive tissues from WNV experimentally horses, in the other, normal horses. Formalin-fixed paraffin-embedded tissue from the thalamus and hindbrain were immunolabeled for microglia, astrocytes, B cells, macrophages/neutrophils, CD3+ T cells. Fresh frozen tissues were immunolabeled for CD4+ and CD8+ T lymphocyte cell markers. Cell counts were obtained using a computer software program. Differences, after meeting assumptions of abnormality, were computed using a general linear model with a Tukey test (P<0.05) for pairwise comparisons. RESULTS: In WNV-challenged horses, Iba-1+ microglia, CD3+ T lymphocyte and MAC387+ macrophage staining were significantly increased. The T cell response for the WNV-challenged horses was mixed, composed of CD4+ and CD8+ T lymphocytes. A limited astrocyte response was also observed in WNV-challenged horses, and MAC387+ and B cells were the least abundant cell populations. MAIN LIMITATIONS: The results of this study were limited by a single collection time post-infection. Furthermore, a comprehensive analysis of cellular phenotypes is needed for naturally infected horses. Unfortunately, in clinical horses, there is high variability of sampling in terms of days post-infection and tissue handling. CONCLUSIONS: The data show that WNV-challenged horses recruit a mixed T cell population at the onset of neurologic disease.

A brief introduction to the military workplace culture.

BACKGROUND: Military culture and workplace are areas of interest for researchers across disciplines. However, few publications on military culture exist. OBJECTIVE: The purpose of this article is to introduce general concepts regarding the structure and culture of the United States Military and discuss how this creates challenges for reintegrating into the civilian world. METHOD: Topics that will be covered in this article include an overview of the Department of Defense (DoD) and Department of Veterans Affairs (VA), socialization to military culture, the unique features of the military as a workplace, the cultural experiences of military personnel reintegrating back into the community, and the challenges faced by military members and their spouses. RESULTS: The provided information on military culture will expand military cultural competency so that civilian employers can enhance their ability to create supportive workplaces for veterans and military spouses during times of transition and reintegration. DISCUSSION: The unique characteristics of the military culture should be understood by those who work with or plan to work with military populations.

The mouse Na+-K+-ATPase gamma-subunit gene (Fxyd2) encodes three developmentally regulated transcripts.

The Na(+)-K(+)-ATPase is understood to function as a hetero-oligomer of alpha- and beta-subunits, but a third subunit, gamma, has been proposed to influence the enzyme's catalytic function. Recently, two variants of the gamma-subunit have been described in kidney, raising the possibility of multiple gamma-subunits with diverse functions. We now report the cloning and sequencing of the mouse gamma-subunit gene (Fxyd2). Analysis of the structure of the gene shows that it encodes three mRNAs that have distinct NH(2)-terminal (extracellular) encoding sequences but common transmembrane and COOH-terminal-encoding sequences resulting from differential splicing and, probably, alternate promoter usage. The three mRNAs have tissue-specific expression patterns. The existence of three different extracellular domains of the gamma-variants and how they may interact with the sodium pump to alter its cation transport properties must now be taken into account for future understanding of the modulation of the Na(+)-K(+)-ATPase by its gamma-subunit.

A 19F NMR study of the interaction of 3-fluoro-4-demethoxydaunomycin with the hexanucleotide d(TCCGGA)2.

19F NMR spectroscopy has been used to study the binding of the fluorinated anthracycline 3-fluoro-4-demethoxydaunomycin with the hexanucleotide d(TCCGGA)2. In the spectrum of the 1:1 anthracycline-d(TCCGGA)2 complex four resonances of approximately equal intensity were observed. This indicated that 4-demethoxydaunomycin intercalated at all possible sites with similar affinity. This suggests that the specific high affinity binding sites that are observed in anthracycline-DNA footprinting experiments are strongly regulated by the local DNA conformation.

Cardiovascular and hormonal responses to liquid food challenge in idiopathic Parkinson's disease, multiple system atrophy, and pure autonomic failure.

We investigated the effect of a balanced liquid meal on blood pressure (BP) and heart rate (with patients supine and during head-up tilt), and on levels of plasma catecholamines, glucose, and insulin, in patients with idiopathic Parkinson's disease (IPD), multiple system atrophy (MSA), pure autonomic failure (PAF), and in healthy subjects (controls). After food, supine BP fell in IPD, but to a greater extent in MSA and PAF. In controls, BP was unchanged. Head-up tilt did not lower BP in IPD and controls, but there was a postprandial fall to lower levels in both MSA and PAF. Plasma norepinephrine levels rose in IPD pre- and postprandially during tilt, but were unchanged in MSA and PAF. These data suggest that in IPD, food causes a smaller fall in supine BP than in MSA and PAF. In IPD, as in controls, food does not induce or unmask postural hypotension, unlike in MSA and PAF, in which BP falls to even lower levels. There are therefore differences in the responses to food ingestion between these groups. This may be of value in separation of these disorders at an early stage.

Effects of the peptide release inhibitor, octreotide, on daytime hypotension and on nocturnal hypertension in primary autonomic failure.

OBJECTIVE: To investigate the effects of the somatostatin analogue octreotide, which inhibits the release of various peptides, on 24-h ambulatory blood pressure profiles in subjects with primary (idiopathic) autonomic failure due to sympathetic denervation, and in particular to determine whether octreotide reduces daytime hypotension and whether it causes or accentuates nocturnal hypertension. SUBJECTS AND METHODS: Eighteen subjects with primary autonomic failure, confirmed by detailed physiological and biochemical autonomic tests, were studied in a randomized manner on two occasions, with and without octreotide treatment (1 mu g/kg body weight subcutaneously, twice a day at 0800 and 1800 h). Blood pressure was measured using the SpaceLabs 90207 system. This was connected at 0900 h with programmed recordings at 30-min intervals until 2300 h and at 60-min intervals until the next morning. There were additional subject-initiated recordings after 5 min each of lying, sitting and standing four times during the day, while sitting after lunch at noon and while standing following walking in the evening. Additional analyses included calculation of cumulative sum (cusum)-derived parameters and construction of cusum plots. RESULTS: After octreotide treatment, the overall mean daytime systolic/diastolic blood pressure (mmHg) was raised (123 +/- 2/77 +/- 1 without treatment versus 128 +/- 2/79 +/- 1 with treatment). There was a reduction in postural (supine versus standing: from 96 +/- 3/62 +/- 3 without treatment to 106 +/- 5/67 +/- 4 with treatment), postprandial (107 +/- 3/65 +/- 2 to 122 +/- 5/75 +/- 4) and exertion-induced (96 +/- 5/61 +/- 5 to 113 +/- 6/71 +/- 5) hypotension. Symptoms of hypotension were reduced by octreotide. Nocturnal blood pressure was lower after octreotide (139 +/- 3/84 +/- 1 versus 129 +/- 3/78 +/- 2). Analyses with the cusum technique further demonstrated blood pressure recovery during the day, with a reduction in the magnitude of change at night after octreotide treatment. CONCLUSIONS: In primary autonomic failure, 24-h ambulatory blood pressure profiles and cusum analyses indicate that octreotide has beneficial effects in reducing postural, postprandial and exertion-induced hypotension, without causing or increasing nocturnal hypertension.

Second-generation peptidomimetic inhibitors of protein farnesyltransferase demonstrating improved cellular potency and significant in vivo efficacy.

The synthesis and evaluation of analogues of previously reported farnesyltransferase inhibitors, pyridyl benzyl ether 3 and pyridylbenzylamine 4, are described. Substitution of 3 at the 5-position of the core aryl ring resulted in inhibitors of equal or less potency against the enzyme and decreased efficacy in a cellular assay against Ras processing by the enzyme. Substitution of 4 at the benzyl nitrogen yielded 26, which showed improved efficacy and potency and yet presented a poor pharmacokinetic profile. Further modification afforded 30, which demonstrated a dramatically improved pharmacokinetic profile. Compounds 26 and 29 demonstrated significant in vivo efficacy in nude mice inoculated with MiaPaCa-2, a human pancreatic tumor-derived cell line.

A human-mouse hybridoma which secretes monoclonal thyroglobulin autoantibody with properties similar to those of the donor patient's serum autoantibody.

Human monoclonal antibodies produced by Epstein Barr (EB) virus transformation and/or cell fusion are frequently IgM antibodies which tend to cross react with a range of antigens and often bear little relationship to the highly specific IgG antibodies associated with human autoimmune disease. By fusing EB virus transformed B lymphocytes from a Hashimoto patient with a mouse myeloma line and selecting for synthesis of IgG class thyroglobulin (Tg) antibody, we have developed a hybridoma (VB/5) secreting Tg antibody of IgG2 subclass and lambda light chain type which has the characteristics of a monoclonal antibody on isoelectric focussing. The antibody has a high affinity for human Tg and recognises Tg from other primates but not non-primate Tg. However, it does not react with human thyroid peroxidase or a panel of other autoantigens. In terms of affinity constant, functional affinity and affinity heterogeneity, the antibody closely resembles the IgG2 lambda Tg antibody present in the serum of the Hashimoto patient whose B lymphocytes were used to develop the hybridoma. In addition to providing a useful reference standard for Tg antibody IgG subclass assays, VB/5 antibody and the hybridoma line provide a valuable starting point for detailed studies of Tg autoantibodies and the genes coding for the variable regions of their heavy and light chains.

A radioimmunoassay for the regulatory allylic steroid, 3 alpha-hydroxy-4-pregnen-20-one (3 alpha HP).

The allylic steroid, 3 alpha-hydroxy-4-pregnen-20-one (3 alpha HP), found in gonadal and brain tissues by radiotracer and chemical methods, had been shown to play a role in gametogenesis, gonadotropin secretion and brain excitability. Since no simple assay was available, a radioimmunoassay for 3 alpha HP was developed using [3H]3 alpha HP and an antiserum raised against 3 alpha HP-20-CMO conjugated to bovine serum albumin. The specificity of the assay for the 3 alpha allylic configuration of 3 alpha HP was confirmed by examining 32 other steroids; cross-reaction with steroids containing different configurations (including metabolites of 3 alpha HP such as progesterone) was less than 0.9%. A Scatchard plot indicated a Ka of 1.56 X 10(9) M-1. Inter- and intra-assay coefficients of variation were 13.1 and 4.5%, respectively. The sensitivity of the assay was 6 pg and the 50% intercept of the standard curve was approx. 123 pg. The measurement by RIA of 3 alpha HP from standard solutions and HPLC purified tissue extracts was confirmed qualitatively and quantitatively by GC/MS methods. The RIA method was employed to determine 3 alpha HP levels in cultured Sertoli cells and in serum of intact and ovariectomized adult rats. Although for most uses, chromatography would not be necessary, two possible methods are presented to enable the separation of 3 alpha HP from other interfering steroids prior to RIA.

Symptoms associated with orthostatic hypotension in pure autonomic failure and multiple system atrophy.

The symptoms caused by or relating to orthostatic hypotension (over 20 mmHg systolic blood pressure) were evaluated using a questionnaire in 72 patients with primary chronic autonomic failure, 32 of whom had pure autonomic failure (PAF, and 40 multiple system atrophy (MSA). The most common posturally related symptoms were dizziness (84% PAF, 83% MSA), syncope (91% PAF, 45% MSA), visual disturbances (75% PAF, 53% MSA) and suboccipital/paracervical 'coat-hanger' neck pain (8 l% PAF, 53% MSA). Chest pain occurred mainly in patients with PAF (44% PAF, 13% MSA). Improvement occurred with sitting or lying flat. Non-specific symptoms (weakness, lethargy and fatigue) were common in both groups (91% PAF, 85% MSA); six patients (one PAF, five MSA) had these symptoms only. Postural symptoms (mainly dizziness and neck pain) were worse in the morning and with warm temperature, straining, exertion, arm movements and food ingestion; they were more common in PAF. Compensatory autonomic symptoms, such as palpitations and sweating, did not occur in either group. In conclusion, orthostatic hypotension caused symptoms of cerebral hypoperfusion (syncope, dizziness and visual disturbances); neck pain, presumably due to muscle hypoperfusion, also occurred frequently. These symptoms were exacerbated by various factors in daily life and were relieved by returning to the horizontal. Non-specific symptoms (such as fatigue) also were common. In MSA, despite substantial orthostatic hypotension, fewer patients had syncope, visual disturbance and neck pain; the reasons for this are unclear. Lack of these features does not exclude the need to assess and investigate orthostatic hypotension and possible autonomic failure.

Properties of methicillin-resistant Staphylococcus aureus colonizing patients in a burns unit.

One hundred cultures of methicillin-resistant Staphylococcus aureus (MRSA) were isolated from patients in a Regional Burns Unit between December 1984 and May 1985. These organisms produced large amounts of beta-lactamase which readily hydrolyzed flucloxacillin but they were sensitive to teicoplanin, dicloxacillin and cephalothin at 37.5 degrees C. The MRSA strains did not differ from methicillin-sensitive isolates in sensitivity to unsaturated fatty acids, survival in serum and plasma or desiccation. However, each culture of this strain was negative or only weakly-positive for bound coagulase and cell bound protein A. Few (eight out of 44) cultures contained plasmids and the resistance to four antibacterials was not transferable in mixed cultures. No attempt was made to isolate patients colonized with MRSA which were rarely isolated elsewhere in the hospital.

Suppression of spermatogenesis without inhibition of steroidogenesis by a 1,2,3-trihydroxypropane solution.

This study presents the first evidence that the non-hormonal, biological substance, 1,2,3- trihydroxypropane (THP) acts as a selective and potent antispermatogenic agent without any apparent toxic or endocrine side effects. The studies were done on 119 rats which were injected intratesticularly (50-200 microliter) with either sterile filtered distilled water (Control) or water/THP (3:7; Treated) and histological, biochemical and fertility determinations were made up to 11 weeks after the injections. The results show that within one week of a single injection with THP, the weights of testes and epididymides are significantly less than those of the Controls and the reduced weights persist for at least 11 weeks. The seminiferous tubules are largely depleted of spermatogenetic cells by 2 weeks and they remain devoid of dividing germ cells for the 11 week period. The Leydig cells have a normal appearance and histochemically show the same 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) and 3 beta-HSD activities as Controls. In vitro studies of the testes showed that the activities of the steroid enzymes 20 alpha-HSD, 17 beta-HSD, 17 alpha-hydroxylase and C17-20-lyase and the production of testosterone and androstenedione were not altered by the THP treatment. Similarly, serum levels of testosterone, LH, and FSH, measured by RIA, and weights of the prostate and seminal vesicles were the same as in Controls. Treated males showed the same degree of sexual behaviour and mating frequency as the Controls, but after the 3rd mating were 100% infertile for the duration of the experiments. The total number of sperm in epididymides of Treated rats was reduced by 99.99% after the 3rd mating.(ABSTRACT TRUNCATED AT 250 WORDS)