Eicosanoids mediate insect hemocyte migration

Hemocyte migration toward infection and wound sites is an essential component of insect defense reactions, although the biochemical signal mechanisms responsible for mediating migration in insect cells are not well understood. Here we report on the outcomes of experiments designed to test the hypotheses that (1) insect hemocytes are able to detect and migrate toward a source of N-formyl-Met-Leu-Phe (fMLP), the major chemotactic peptide from Escherichia coli and (2) that pharmaceutical modulation of eicosanoid biosynthesis inhibits hemocyte migration. We used primary hemocyte cultures prepared from fifth-instar tobacco hornworms, Manduca sexta in Boyden chambers to assess hemocyte migration toward buffer (negative control) and toward buffer amended with fMLP (positive control). Approximately 42% of negative control hemocytes migrated toward buffer and about 64% of positive control hemocytes migrated toward fMLP. Hemocyte migration was inhibited (by >40%) by treating hornworms with pharmaceutical modulators of cycloxygenase (COX), lipoxygenase and phospholipase A2 (PLA2) before preparing primary hemocyte cultures. The influence of the COX inhibitor, indomethacin, and the glucocorticoid, dexamethasone, which leads to inhibition of PLA2, was expressed in a dose-dependent way. The influence of dexamethasone was reversed by injecting arachidonic acid (precursor to eicosanoid biosynthesis) into hornworms before preparing primary hemocyte cultures. The saturated fatty acid, palmitic acid, did not reverse the inhibitor effect. These findings support both our hypotheses, first that insect hemocytes can detect and respond to fMLP, and second, that insect hemocyte migration is mediated by eicosanoids.     

Targeting of antigen to the herpesvirus entry mediator augments primary adaptive immune responses

Interactions between the herpesvirus entry mediator (HVEM) and the B- and T-lymphocyte attenuator (BTLA) inhibit B and T cell activation. HVEM-BTLA interactions are blocked by herpes simplex virus (HSV) glycoprotein D (gD) through binding of its N-terminal domain to the BTLA binding site of HVEM. In this study, we inserted viral antigens into the C-terminal domain of gD and expressed these antigens with plasmid or E1-deleted (replication-defective) adenovirus vectors. Viral antigens fused to gD induced T and B cell responses to the antigen that were far more potent than those elicited by the same antigen expressed without gD. The immunopotentiating effect required binding of the gD chimeric protein to HVEM. Overall, the studies demonstrate that targeting of antigen to the BTLA binding site of HVEM augments the immunogenicity of vaccines.     

Effects of the replacement of concentrate and fibre-rich hay by high-quality hay on chewing,rumination and nutrient digestibility in non-lactating Holstein cows

The aim of this study was to investigate the effects of high-quality hay with an elevated sugar content alone or with graded amounts of concentrate feed on chewing and ruminating activity, apparent total tract digestibility (ATTD) and ruminal pH at different time points after feeding in the free ruminal liquid (FRL) and the particle-associated ruminal liquid (PARL). Eight rumen cannulated non-lactating Holstein cows were arranged in a Latin square design in four experimental runs lasting 25 d each. The four diets tested were 60NQ (60% normal-quality hay + 40% concentrate), 60HQ (60% high-quality hay + 40% concentrate), 75HQ (75% high-quality hay + 25% concentrate) and 100HQ (100% high-quality hay). Normal and high-quality hays differed in sugar contents (11.3% vs. 18.7% in dry matter [DM]), neutral detergent fibre (NDF; 57.7% vs. 46.3% in DM), acid detergent fibre (ADF, 35.0% vs. 23.5% in DM) and crude protein (CP, 11.3% vs. 23.5% in DM). Data showed that ATTD of DM, CP, NDF and ADF was higher with the high-quality hay diets. Time spent eating was reduced with high-quality hay. However, time spent ruminating was longest in Group 100HQ. In all groups, ruminal pH of FRL and PARL decreased with time after the morning feeding. But 10 h later, pH of Group 100HQ was higher again compared with the other groups. Considering the average pH in FRL over all measured time points, cows in Groups 60NQ and 100HQ had higher pH values of 6.85 and 6.83, respectively. Regarding pH values in PARL, animals of Group 60NQ displayed the highest pH value (6.68), whereas the lowest value of 6.21 was found in Group 60HQ. Overall, results suggest that high-quality hay maintains the diet’s structural effectiveness by stimulating rumination and stabilising ruminal pH while greatly improving ATTD. However, the structural effectiveness of the high-quality hay gets impaired with increasing proportion of concentrate feed in the diet.     

Lidocaine block of cardiac sodium channels

Lidocaine block of cardiac sodium channels was studied in voltage-clamped rabbit purkinje fibers at drug concentrations ranging from 1 mM down to effective antiarrhythmic doses (5-20 μM). Dose-response curves indicated that lidocaine blocks the channel by binding one-to-one, with a voltage-dependent K(d). The half-blocking concentration varied from more than 300 μM, at a negative holding potential where inactivation was completely removed, to approximately 10 μM, at a depolarized holding potential where inactivation was nearly complete. Lidocaine block showed prominent use dependence with trains of depolarizing pulses from a negative holding potential. During the interval between pulses, repriming of I (Na) displayed two exponential components, a normally recovering component (τless than 0.2 s), and a lidocaine-induced, slowly recovering fraction (τ approximately 1-2 s at pH 7.0). Raising the lidocaine concentration magnified the slowly recovering fraction without changing its time course; after a long depolarization, this fraction was one-half at approximately 10 μM lidocaine, just as expected if it corresponded to drug-bound, inactivated channels. At less than or equal to 20 μM lidocaine, the slowly recovering fraction grew exponentially to a steady level as the preceding depolarization was prolonged; the time course was the same for strong or weak depolarizations, that is, with or without significant activation of I(Na). This argues that use dependence at therapeutic levels reflects block of inactivated channels, rather than block of open channels. Overall, these results provide direct evidence for the “modulated-receptor hypothesis” of Hille (1977) and Hondeghem and Katzung (1977). Unlike tetrodotoxin, lidocaine shows similar interactions with Na channels of heart, nerve, and skeletal muscle.     

A covariotide model explains apparent phylogenetic structure of oxygenic photosynthetic lineages

The aims of the work were (1) to develop statistical tests to identify whether substitution takes place under a covariotide model in sequences used for phylogenetic inference and (2) to determine the influence of covariotide substitution on phylogenetic trees inferred for photosynthetic and other organisms. (Covariotide and covarion models are ones in which sites that are variable in some parts of the underlying tree are invariable in others and vice versa.) Two tests were developed. The first was a contingency test, and the second was an inequality test comparing the expected number of variable sites in two groups with the observed number. Application of these tests to 16S rDNA and tufA sequences from a range of nonphotosynthetic prokaryotes and oxygenic photosynthetic prokaryotes and eukaryotes suggests the occurrence of a covariotide mechanism. The degree of support for partitioning of taxa in reconstructed trees involving these organisms was determined in the presence or absence of sites showing particular substitution patterns. This analysis showed that the support for splits between (1) photosynthetic eukaryotes and prokaryotes and (2) photosynthetic and nonphotosynthetic organisms could be accounted for by patterns arising from covariotide substitution. We show that the additional problem of compositional bias in sequence data needs to be considered in the context of patterns of covariotide/covarion substitution. We argue that while covariotide or covarion substitution may give rise to phylogenetically informative patterns in sequence data, this may not always be so.      

Characteristics and functions of Sendai virus-specific T-cell clones

Several murine Sendai virus-specific T-cell clones were characterized in vitro and in vivo. All T-cell clones were phenotypically Thy-1.2+, and most clones were Lyt-1+,2-; one T-cell clone was Lyt-1-,2-. Some of the clones proliferated in response to antigen presented on I region-compatible stimulator cells. Proliferation could be inhibited by monoclonal antibodies directed against class II antigens. Clones which proliferated in response to antigen secreted lymphokines which could be identified as Interleukin 2 and Interleukin 3. All of the clones tested in vivo induced a delayed-type hypersensitivity response in syngeneic mice challenged with antigens. Depending on the experimental conditions chosen, Interleukin 2-producing clones as well as non-Interleukin 2-producing clones mediated help for stimulation of cytolytic T lymphocytes.     

Adoptive transfer of delayed-type hypersensitivity to Sendai virus. I. Induction of two different subsets of T lymphocytes which differ in H-2 restriction as well as in the Lyt phenotype

This paper investigates kinetics and antigenic and genetic requirements for transfer of delayed-type hypersensitivity (DTH) reactions against Sendai virus. Kinetics of sensitization of effector cells are similar to those described in other virus systems. Maximum DTH response is elicited in the footpad 7 days after sensitization; maximal increase in footpad thickness is found approximately 24 hr after injection of virus and transfer of immune lymphocytes. DTH effector cells are Ig^? and are susceptible to treatment with anti-theta antibody and complement. After transfer of immune lymphocytes a DTH reaction can be elicited by infectious virus, uv light-inactivated virus or cell-cultured Sendai virus which lacks fusion capacity. Requirements of H-2 compatibility are dependent on the biological nature of the virus preparation used for challenge: High doses of infectious or uv light-inactivated Sendai virus require K, D, or IA region compatibility; low concentrations of infectious virus require K and/or D region compatibility, while cell-cultured fusion-negative Sendai virus requires IA region homology. At the level of induction K, D region-restricted DTH-effector T lymphocytes (Td) and cytolytic T lymphocytes (Tc) are stimulated to a greater extent by infectious virus than by uv light-inactivated virus. Furthermore, stimulation of these T lymphocytes is dependent on the route chosen for immunization: intravenous (iv) injection is superior to intraperitoneal (ip) injection; subcutaneous (sc) injection causes the lowest degree of sensitization. IA region-restricted Td lymphocytes are stimulated to comparable amounts by infectious or uv light-inactivated Sendai virus independent of the route of immunization. Td lymphocytes, which require IA region compatibility and Td lymphocytes which require K or D region compatibility can be distinguished by their Lyt phenotype, e.g., IA region-restricted Td lymphocytes are characterized by Lyt-1⁺2^? antigens, while K or D region-restricted Td lymphocytes are phenotypically Lyt-1(⁺)2⁺.     

Using ontologies to describe mouse phenotypes

The mouse is an important model of human genetic disease. Describing phenotypes of mutant mice in a standard, structured manner that will facilitate data mining is a major challenge for bioinformatics. Here we describe a novel, compositional approach to this problem which combines core ontologies from a variety of sources. This produces a framework with greater flexibility, power and economy than previous approaches. We discuss some of the issues this approach raises.     

Heart rate reduction by zatebradine reduces infarct size and mortality but promotes remodeling in rats with experimental myocardial infarction

The importance of heart rate for left ventricular remodeling and prognosis after myocardial infarction is not known. We examined the contribution of heart rate reduction by zatebradine, a direct sinus node inhibitor without negative inotropic effects on left ventricular function and dilatation, on mortality, energy metabolism, and neurohormonal changes in rats with experimental myocardial infarction (MI). Thirty minutes after left coronary artery ligation or sham operation, the rats were randomized to receive either placebo or zatebradine (100 mg x kg(-1) x day(-1) per gavage) continued for 8 wk. Mortality during 8 wk was 33.3% in the placebo and 23.0% in the zatebradine group (P < 0.05); MI size was 36 +/- 2% and 30 +/- 1% (means +/- SE, P < 0.05), respectively. Zatebradine improved stroke volume index in all treated rats but increased left ventricular volume in rats with small MI (2.43 +/- 0.10 vs. 1.81 +/- 0.10 ml/kg, P < 0.05) but not in rats with large MI (2.34 +/- 0.09 vs. 2.35 +/- 0.11 ml/kg, not significant). Zatebradine reduced left and right ventricular norepinephrine and increased left and right ventricular 3,4-dihydroxyphenyl ethylene glycol-to-norepinephrine ratio suggesting aggravation of cardiac sympathetic activation by zatebradine after MI. Creatine kinase and lactate dehydrogenase isoenzymes in rats with MI remained unchanged by zatebradine. Lowering heart rate per se reduces mortality and MI size in this model but induces adverse effects on left ventricular remodeling in rats with small MI.