Combinatorial analysis of population dynamics, metabolite levels and malolactic fermentation in Saccharomyces cerevisiae/ Lachancea thermotolerans mixed fermentations.

The outcome of co- or sequential inoculation of Lachancea thermotolerans in winemaking remains unpredictable due to a lack of integrated data regarding the impact of grape juice composition on L. thermotolerans fermentation behaviour. Here, we investigate the impact of nitrogen composition on fermentation characteristics and aroma compound production in grape juice sequentially inoculated with commercial L. thermotolerans and S. cerevisiae strains. Subsequently, all treatments were subjected to malolactic fermentation (MLF) using two commercial strains of Oenococcus oeni. Addition of amino acids led to faster growth for S. cerevisiae fermentations, compared to the nitrogen-equivalent addition of diammonium phosphate (DAP). L. thermotolerans persistence in the mixed fermentations was significantly higher following DAP addition, with higher glycerol and lactic acid production. Interestingly, the lower total Nitrogen content in DAP-treated musts compared to other treatments did not alter the subsequent growth of S. cerevisiae. MLF was more similar between musts fermented with L. thermotolerans, regardless of nutrient regime, whereas significant differences in MLF completion times were observed for different nitrogen treatments in S. cerevisiae fermentations. Collectively, the data present an integrated view of the impact of nitrogen treatment on multispecies co-inoculation (growth kinetics and aromatic outcomes) and the downstream impact on MLF.

The energy content of barley fed to growing pigs: characterizing the nature of its variability and developing prediction equations for its estimation.

Currently, the pork industry attempts to formulate energy levels in swine diets to within a tolerance of 1.5%. This is difficult to achieve in practice when the energy content of primary ingredients fluctuates by up to 15%. This experiment was carried out to define the sources of variation in the energy content of barley and to develop a practical method to accurately estimate the DE and ME content of individual barley samples. Four samples of each of five covered barley varieties (AC Lacombe, B-1602, Bedford, Harrington, and Manley) were collected to obtain a range of quality within each variety. Five measurements were collected on each barley sample using 60 crossbred barrows in an apparent total tract digestibility study. The barrows, average BW of 35.3 kg, were housed in individual metabolism crates to facilitate separate collection of urine and feces. Five-day collection periods followed 5-d diet acclimation periods. Levels of total beta-glucan, ADF, CP, and starch (90% DM) in the 20 barley samples ranged from 2.7 to 4.5%, 4.5 to 9.2%, 10.8 to 15.1%, and 42.3 to 53.4%, respectively. The mean DE and ME content of the 20 samples were 2,934 and 2,857 kcal/kg (90% DM), respectively, and varied among samples by 15.2% (447 kcal). The complex structural cell wall carbohydrates seemed to have the greatest influence on the energy content of individual barley samples. The ADF fraction alone accounted for 85% of the total variation in energy content of the 20 samples. Converted into a prediction equation, DE = 3,526 - 92.8 x ADF (90% DM), the ADF content was used to estimate the DE content of barley with 85% accuracy. This experiment confirms the large variation in the energy content of barley, describes the factors that influence this variation, and presents equations based on chemical and(or) physical measurements that may be used to predict the DE and ME content of individual barley samples.

Influence of antigen challenge on platelet responsiveness in horses with chronic obstructive pulmonary disease.

A role for platelets in allergic airways disease has been postulated and changes in the responsiveness of circulating platelets have been demonstrated following antigen challenge of asthmatic human subjects. In this study agonist-induced aggregation of equine platelets in vitro has been compared before and after exposure of horses to a controlled hay and stray challenge. Prior to challenge the response of platelets, from horses with chronic obstructive pulmonary disease (COPD) and normal animals, to adenosine diphosphate (ADP) and platelet activating factor (PAF) did not differ. Five hours after initiation of the challenge, there was a modest but significant decrease in the response of platelets from the COPD horses to PAF, but not to ADP. Platelets from normal horses were not less sensitive to either agonist at this time. Twenty four hours after challenge the responses of platelets from COPD horses to both agonists were the same as pre-challenge values. These results demonstrate that antigen challenge alters the responsiveness of platelets from allergic horses to PAF and this change is suggestive of PAF release accompanying allergen exposure in the horse.

Allatostatin in hemocytes of the cockroach Diploptera punctata.

Allatostatins are neuropeptides that inhibit the production, by the corpora allata, of a major insect hormone, juvenile hormone. These peptides are produced by cells of the brain and ganglia as well as by midgut endocrine cells. Transport from these sites may contribute to the allatostatin content in the hemolymph (insect blood). Using a monoclonal antibody against Diploptera punctata allatostatin I (A-P-S-G-A-Q-R-L-Y-G-F-G-L-NH2) and in situ hybridization with a digoxigenin-labeled cRNA probe generated from a portion of the allatostatin gene, it is demonstrated that allatostatin is present in and synthesized by granular hemocytes of D. punctata. About 5% of the hemocytes react with anti-allatostatin antibody and a similar number hybridize with a cRNA probe that detects allatostatin-specific mRNA. Electron micrographs showed that allatostatin-immunoreactive material occurs in membrane-bound, uniformly dense granules that frequently fill fusiform-shaped cells. Allatostatin in cell and plasma fractions of hemolymph quantified by enzyme-linked immunosorbent assay and by bioassay for inhibition of juvenile hormone synthesis in vitro indicated that about equal quantities (0.1-0.2 fmol/microl) are present in cell and plasma fractions. The production of allatostatin by hemocytes suggests that allatostatins may function as regulatory peptides in hemolymph activities in addition to their other known functions.

Effects of platelet activating factor on the distribution of radiolabelled leucocytes and platelets in normal horses and asymptomatic horses with chronic obstructive pulmonary disease.

Antigen challenge is known to cause the recruitment of neutrophils to the lungs of horses with chronic obstructive pulmonary disease (COPD). To evaluate a possible role of platelet activating factor (PAF) in this process, the effects of PAF on the distribution of radiolabelled neutrophils were compared in normal horses and asymptomatic horses with COPD. Changes in lung function, heart rate and the distribution of platelets and eosinophils were also measured. PAF (5 ng kg-1 intravenously) caused immediate but transient increases in the number of radiolabelled neutrophils in the lungs and a concomitant decrease in the peripheral neutrophil count. The total numbers of circulating leucocytes and neutrophils were also significantly decreased by PAF. Rapid and reversible increases in heart rate, respiratory rate and pleural pressure were also observed. In separate experiments, the numbers of radiolabelled eosinophils and platelets in the lungs increased transiently after the administration of PAF. The responses to PAF were qualitatively and quantitatively similar in normal horses and asymptomatic COPD horses. The PAF receptor antagonist WEB2086 (3 mg kg-1 intravenously) inhibited the effects of PAF. These results suggest that PAF, if released in the lungs of horses with COPD during an antigen challenge, might contribute to the recruitment of leucocytes and the respiratory changes.

A study of the effect of a platelet activating factor (PAF) receptor antagonist on antigen challenge of horses with chronic obstructive pulmonary disease.

Antigen challenge involving exposure to straw and mouldy hay for 7 h produced lung function changes and neutrophil recruitment to the lungs in horses with chronic obstructive pulmonary disease (COPD). During the challenge, an increase in radiolabelled neutrophils in the lungs occurred, together with increased respiratory rate and pleural pressure. The role of platelet activating factor (PAF) in antigen-induced neutrophil accumulation, and increased pleural pressure and respiratory rate was investigated by administering the PAF receptor antagonist WEB 2086 to asymptomatic COPD horses prior to antigen challenge. WEB 2086 (3 mg/kg i.v.) did not affect antigen-induced changes in either neutrophil accumulation or respiratory function. These results suggest that PAF may not be an important mediator of the response to antigen in equine COPD.

Role of allatostatins in the regulation of juvenile hormone synthesis.

The identification of neuropeptides that inhibit juvenile hormone (JH) synthesis by the corpora allata (CA) has verified the existence of these allatostatins, which, from much experimental evidence, have long been postulated to occur. It also makes possible new approaches for studying the role of allatostatins in the regulation of JH synthesis. Allatostatins, localized immunocytochemically, occur in lateral neurosecretory cells of the brain that innervate the CA. Presumably their effect on the CA results from the release of allatostatins at these nerve endings. Allatostatins also occur in the hemolymph in cockroaches and have been shown to act on the CA through this pathway. The ability of allatostatins to inhibit CA depends not only on the concentration of the peptides but also on the sensitivity of the CA to them. Male Diploptera punctata were treated with JH analog following denervation of CA and implanted with a previtellogenic or vitellogenic ovary or injected with saline. Animals implanted with a vitellogenic ovary, compared to the previtellogenic ovary or saline, showed significantly increased JH synthesis by their CA and a reduced amount of allatostatin in the hemolymph. The denervated CA from these JH analog treated animals, following implantation with a previtellogenic and vitellogenic ovary, showed a tendency toward increased and decreased sensitivity, respectively, to a given dose of allatostatin in vitro compared to those from saline injected controls. Experiments such as these suggest that changes in release of allatostatins and in sensitivity of CA to them could be postulated to be major factors regulating JH synthesis in the cockroach.

Studies on the mechanisms involved in the inflammatory response in a reversed passive Arthus reaction in guinea-pig skin: contribution of neutrophils and endogenous mediators.

1. Mediators of inflammation can increase vascular permeability in at least two different ways: by acting directly on endothelial cells or, indirectly, through an incompletely understood mechanism, dependent on circulating neutrophils. Neutrophil-dependent oedema formation has been described in the skin of rabbits, rats, hamsters, mice and man. In contrast, we presented evidence in a previous study that local oedema formation induced by i.d. injection of chemoattractants in guinea-pig skin was neutrophil-independent. In the present study, we sought evidence of neutrophil-dependent oedema formation in immune-complex-mediated vasculitis, the reversed passive Arthus (RPA) reaction, in guinea-pig skin. We also investigated whether haemorrhage in the RPA reaction was neutrophil-dependent (as it is in other species) and the role of endogenous mediators of inflammation (prostaglandins, nitric oxide, histamine, PAF and leukotrienes) in contributing to the local inflammatory response. 2. In the RPA reaction, most oedema formation occurred over the first 60 min whereas 111In-neutrophil accumulation was still increasing from 60 to 240 min. The different kinetics of these two events suggested that they may be dissociated. 3. Oedema formation was partially inhibited by a long-acting PAF antagonist (UK-74,505) and an H1 histamine receptor antagonist (mepyramine) but not by a 5-lipoxygenase inhibitor (ZM 230487). A nitric oxide synthesis inhibitor (NG-nitro-L-arginine methyl ester, L-NAME) suppressed oedema formation by 68% whereas a cyclo-oxygenase inhibitor suppressed oedema by 27%. 4. 111In-neutrophil accumulation in the RPA reaction was partially suppressed by UK-74,505. In contrast, ZM 230487 was without effect at doses which abrogated arachidonic acid-induced 111In-neutrophil accumulation. 5. The anti-CD18 monoclonal antibody, (mAb) 6.5E F(ab')2, effectively inhibited 111In-neutrophil accumulation induced by PAF, zymosan-activated plasma (ZAP) and in the RPA reaction. However, oedema formation measured in the same sites was not altered. In contrast, oedema formation in the RPA reaction was partially suppressed by 6.5E whole mAb which was 2.5 times more potent than 6.5EF(ab')2 at inhibiting guinea-pig neutrophil adhesion to protein-coated plastic. Haemorrhage induced by PAF and in the RPA reaction was significantly inhibited by 6.5E F(ab')2 pretreatment.6. We conclude that in the RPA reaction in guinea-pig skin, oedema formation is partially neutrophil dependent as assessed by using an anti-CD18 mAb, whereas ZAP-induced oedema formation is neutrophil-independent. Haemorrhage was also dependent on neutrophil accumulation. In addition, our studies support a role for PAF in mediating both oedema formation and "'In-neutrophil accumulation in the RPA reaction. Endogenous release of histamine also appears to be important in mediating oedema formation suggesting that mast cells play a critical role in increases of vascular permeability in inflammatory reactions in guinea-pig skin. Moreover, our results confirm previous findings which suggest a dominant role for nitric oxide in maintaining cutaneous blood flow in the guinea-pig.

Duration of antigen-induced hyperresponsiveness in horses with allergic respiratory disease and possible links with early airway obstruction.

Antigen-induced airway hyperresponsiveness in allergic horses has previously been demonstrated when clinical signs of acute airway obstruction were apparent, as a consequence of exposure of animals to hay and straw for variable periods of time, and repeat measurements of hyperresponsiveness have been made no earlier than 1 week after challenge. In the present study airway responsiveness to methacholine has been measured in normal horses and allergic horses in clinical remission before and 24, 48 and 72 h after a hay and straw challenge of fixed, short, duration (7 h). Correlations between early increases in interpleural pressure and hyperresponsiveness have also been investigated. As in other studies, the mean airway responsiveness of groups of normal and allergic horses in clinical remission was not significantly different. The responsiveness to methacholine of allergic, but not normal, horses was increased after antigen challenge and was significantly greater than that of normal horses at 48 and 72 h after challenge (log PD8 cm: -0.77 +/- 0.28 vs. 0.27 +/- 0.14 at 48 h and -0.6 +/- 0.25 vs. 044 +/- 01 at 72 h; P < 0.05). There was also a significant correlation between interpleural pressure at the end of the 7-h challenge in allergic horses and the increase in responsiveness to methacholine at 24, 48 and 72 h. These results demonstrate that antigen induces an increase in airway responsiveness in allergic horses that persists for up to 3 days and which may be linked to the initial increase in interpleural pressure.

Early neutrophil but not eosinophil or platelet recruitment to the lungs of allergic horses following antigen exposure.

Previous studies have shown that bronchoalveolar lavage fluid from horses with allergic respiratory disease and showing clinical symptoms contains increased numbers of neutrophils. In some cases, the eosinophil count is also increased. In this study the time course of changes in lung function and the accumulation of radiolabelled leucocytes and platelets in the lungs of allergic and normal horses has been examined during a 7 hr allergen exposure. Antigen challenge had no effect on pleural pressure or the distribution of radiolabelled neutrophils, eosinophils or platelets in normal horses. In contrast, in 6/8 allergic horses, there was an increase in pleural pressure and neutrophil accumulation in the lungs, both of which were evident after 4-5 hr. However, during the 7 hr challenge period radiolabelled eosinophils were detected in the lungs of only 1/6 horses exhibiting an increase in pleural pressure and in 1/7 horses that failed to show a change in airway function despite a clinical history of allergic respiratory disease. Antigen challenge did not alter the distribution of radiolabelled platelets in the five allergic horses tested. These results demonstrate that increased pleural pressure is not accompanied by eosinophil or platelet accumulation in the lungs of horses with allergic respiratory disease following exposure to antigen. However, changes in airway function can be associated with neutrophil accumulation but can also take place in the absence of this cell recruitment. This raises the possibility that the presence of neutrophils in the lung is not a prerequisite for changes in lung function.

Consultation skills of young doctors: I--Benefits of feedback training in interviewing as students persist.

Thirty six young doctors who as medical students had been randomly allocated to either video feedback training or conventional teaching in interviewing skills during a psychiatry clerkship were reassessed five years later. Each doctor interviewed one patient with a psychiatric illness and two with a physical illness. Each interview was rated independently. Both groups had improved since the fourth year clerkship, but those given feedback training had maintained their superiority in the skills associated with accurate diagnosis. This superiority was as evident in their interviews with physically ill patients as it was with psychiatric patients. Both groups, however, still used "closed" questions and were more reluctant to cover psychosocial problems in physically ill patients. Those trained conventionally were clinically inadequate in both these aspects and in clarifying their patients' statements. Given these lasting benefits, all medical students should have feedback training in interviewing skills.

Consultation skills of young doctors: II--Most young doctors are bad at giving information.

Forty young doctors, half of whom had had feedback training in interviewing as students, were assessed five years later. Each interviewed three patients and after being given results of examination, investigations, and diagnosis and prognosis returned to discuss them with each patient for 10 minutes. These discussions were filmed on videotape and evaluated. There was no difference between the scores of interview trained and control doctors. Though most gave simple information on diagnosis and treatment, few mentioned investigations, aetiology, or prognosis. Very few obtained and took any account of patients' views or expectations of these matters. Some young doctors do discover for themselves how best to give patients information and advice, but most remain extremely incompetent. This is presumably because they get no training as students in this important aspect of clinical practice. This deficiency should be corrected, and competence tested before qualification to practise.

Training clinical teachers in psychiatry to teach interviewing skills to medical students.

To study the feasibility of training all clinical teachers in psychiatry to teach interviewing skills to medical students, 24 (unselected clinicians were assigned to one of four different training methods. They received either experiential or didactic instruction, and their initial teaching sessions were either supervised or unsupervised. A total of 287 medical students subsequently received feedback training from these teachers. While all students showed significant increases in skill after training, those taught by experientially trained teachers showed the greatest gains. Neither supervision nor the teachers' own interviewing skills exerted significant effects on students' performance. It is concluded that with only brief training unselected clinicians can become effective teachers of essential interviewing skills. Feedback training in such skills can, therefore, be incorporated into existing curricula without major disruption of other requirements.

The extracellular matrix of normal chick embryo fibroblasts: its effect on transformed chick fibroblasts and its proteolytic degradation by the transformants.

Extracellular matrix (ECM), prepared from chick embryo fibroblasts, contains fibronectin as the major structural protein along with collagen and other polypeptides as less abundant protein components. When Rous sarcoma virus-transformed chick embryo fibroblasts are cultured on the ECM in the presence of the tumor promoter tetradecanoyl phorbol acetate, the transformed cells lose their characteristic rounded morphology and align on and within the ECM fibrillar network. This restrictive aspect of ECM is only temporary, however, and with time (24-72 h) the transformed cells progressively degrade the ECM fibers and resume their rounded appearance. The matrix degradation can be monitored by employing biosynthetically radiolabeled ECM. The addition of purified chicken plasminogen to the Rous sarcoma virus-transformed chick embryo fibroblast cultures enhances the rate and extent of ECM degradation, due to the elevated levels in the transformed cultures of plasminogen activator. Plasminogen-dependent and -independent degradation of ECM has been characterized with regard to sensitivity to various natural and synthetic protease inhibitors and to the requirement of cell/ECM contact. Plasminogen-dependent degradation of ECM occurs rapidly when ECM and cells are in contact or separated, whereas plasminogen-independent degradation is greatly reduced when ECM and cells are separated, which suggests that cell surface-associated proteolytic enzymes are involved. A possible role in ECM degradation has been indicated for cysteine proteases, metallo enzymes, and plasminogen activator, the latter as both a zymogen activator and a direct catalytic mediator.

The teaching of interviewing skills: comparison of experienced and novice trainers.

During a clerkship in psychiatry thirty-six medical students were randomly allocated to one of three teachers who differed widely in their experience of teaching essential interviewing skills. Each teacher taught two groups of six medical students using videotape feedback and discussion of practice interviews. Independent raters who were blind to the teachers to whom the students had been assigned rate pre- and post-training interviews. All three teachers proved effective in teaching interviewing skills and it is concluded that most teachers could probably be taught to carry out this training.

Teaching skills in communication to medical students--a critical review of the methodology.

Over the past decade, there has been a substantial increase in papers concerned with training medical students in communication skills. In this paper, we consider what constitutes an adequate methodology for such research and whether recent papers meet this standard.