The murine homolog (Mph) of human herpesvirus entry protein B (HveB) mediates entry of pseudorabies virus but not herpes simplex virus types 1 and 2

A mouse member of the immunoglobulin superfamily, originally designated the murine poliovirus receptor homolog (Mph), was found to be a receptor for the porcine alphaherpesvirus pseudorabies virus (PRV). This mouse protein, designated here murine herpesvirus entry protein B (mHveB), is most similar to one of three related human alphaherpesvirus receptors, the one designated HveB and also known as poliovirus receptor-related protein 2. Hamster cells resistant to PRV entry became susceptible upon expression of a cDNA encoding mHveB. Anti-mHveB antibody and a soluble protein composed of the mHveB ectodomain inhibited mHveB-dependent PRV entry. Expression of mHveB mRNA was detected in a variety of mouse cell lines, but anti-mHveB antibody inhibited PRV infection in only a subset of these cell lines, indicating that mHveB is the principal mediator of PRV entry into some mouse cell types but not others. Coexpression of mHveB with PRV gD, but not herpes simplex virus type 1 (HSV-1) gD, inhibited entry activity, suggesting that PRV gD may interact directly with mHveB as a ligand that can cause interference. By analogy with HSV-1, envelope-associated PRV gD probably also interacts directly with mHveB during viral entry.     

Potential Role of Cerebral Glutathione in the Maintenance of Blood-Brain Barrier Integrity in Rat

Using the model of glutathione (GSH) depletion, possible role of GSH in the maintenance of blood-brain barrier (BBB) integrity was evaluated in rats. Administration (ip) of GSH depletors, diethyl maleate (DEM, 1–4 mmol/kg), phorone (2–3 mmol/kg) and 2-cyclohexene-1-one (CHX, 1 mmol/kg), to male adults was found to deplete brain and liver GSH and increase the BBB permeability to micromolecular tracers (sodium fluorescein and [¹⁴C]sucrose) in a dose-dependent manner at 2h. However, BBB permeability to macromolecular tracers such as horseradish peroxidase and Evan's blue remained unaltered. It was also shown that observed BBB permeability dysfunction was associated with brain GSH depletion. A lower magnitude of BBB increase in rat neonates, as compared to adults, indicated a possible bigger role of GSH in the BBB function of mature brain. The treatment with N-acetylcysteine, methionine and GSH provided a partial to full protection against DEM-induced brain (microvessel) GSH depletion and BBB dysfunction; however, the treatment with -tocopherol, ascorbic acid and turmeric were not effective. Our studies showed that cerebral GSH plays an important role in maintaining the functional BBB integrity.     

Synthesis, structural properties and insulin-enhancing potential of bis(quercetinato)oxovanadium(IV) conjugate

An oxovanadium complex of quercetin (2), exhibits highly potent insulin-enhancing activity in streptozotocin-induced diabetic mice. It also mimics mitogenic potential of insulin as evaluated by [H(3)]thymidine uptake assay making an effective, orally active insulin-enhancing agent for the treatment of both type 1 and type 2 diabetes without any noticeable toxic effects.     

The excised heat-shock domain of alphaB crystallin is a folded, proteolytically susceptible trimer with significant surface hydrophobicity and a tendency to self-aggregate upon heating

The lens protein, alpha-crystallin, is a molecular chaperone that prevents the thermal aggregation of other proteins. The C-terminal domain of this protein (homologous to domains present in small heat-shock proteins) is implicated in chaperone function, although the domain itself has been reported to show no chaperone activity. Here, we show that the domain can be excised out of the intact alphaB polypeptide and recovered directly in pure form through the transfer of CNBr digests of whole lens homogenates into urea-containing buffer, followed by dialysis-based refolding of digests under acidic conditions and a single gel-filtration purification step. The folded (beta sheet) domain thus obtained is found to be (a) predominantly trimeric, and to display (b) significant surface hydrophobicity, (c) a marked tendency to undergo degradation, and (d) a tendency to aggregate upon heating, and on exposure to UV light. Thus, the twin 'chaperone' features of multimericity and surface hydrophobicity are clearly seen to be insufficient for this domain to function as a chaperone. Since alpha-crystallin interacts with its substrates through hydrophobic interactions, the hydrophobicity of the excised domain indicates that separation of domains may regulate function; at the same time, the fact is also highlighted that surface hydrophobicity is a liability in a chaperone since heating strengthens hydrophobic interactions and can potentially promote self-aggregation. Thus, it would appear that the role of the N-terminal domain in alpha-crystallin is to facilitate the creation of a porous, hollow structural framework of >/=24 subunits in which solubility is effected through increase in the ratio of exposed surface area to buried volume. Trimers of interacting C-terminal domains anchored to this superstructure, and positioned within its interior, might allow hydrophobic surfaces to remain accessible to substrates without compromising solubility.     

Isolation of etiological agent of hydropericardium syndrome in chicken embryo liver cell culture and its serological characterization

The virus causing hydropericardium syndrome was isolated in chicken embryo liver (CEL) cell culture from livers obtained from naturally infected broilers. The cytopathic effects characterized by rounding and degeneration of cells were visible 36 hr post infection in first passage. At 4th passage level, the infectivity titre was 5.24 log10 TCID50/ml. In May-Grunwald and Giemsa stained cells, basophilic intranuclear inclusions ('bird eye' inclusion), typical of aviadenovirus infection, were observed. The specificity of inclusion was confirmed by indirect immunofluorescence. Various serological tests, such as agar gel precipitation test, counter immuno electrophoresis, micro serum neutralization test and enzyme linked immunosorbent assay were also standardized to confirm the isolation of etiological agent of hydropericardium syndrome in CEL cell culture and to diagnose the disease in poultry.     

Studies on fermentative production of squalene

Fermentative production of squalene under anaerobic conditions using commercially available compressed baker's yeast (Saccharomyces cerevisiae), and a strain of Torulaspora delbrueckii isolated from molasses was studied. Yield of squalene from S. cerevisiae and T. delbrueckii were found to be 41.16 and 237.25 g g^–1 respectively, dry weight of yeast cells. Isolation and purification of squalene from the lipid extracts obtained by cell lysis of either strain were achieved chromatographically. The purified squalene was characterized spectroscopically against an authentic standard.     

Influence of sodium ion on heavy metal-induced inhibition of light-regulatd proton efflux and active carbon uptake in the cyanobacterium Anabaena flos-aquae

The light-induced proton efflux and active carbon uptake are inhibited by mercury and cadmium ions in Anabaena flos-aquae. The inhibitory effects of these heavy metal ions are reversed by 40 mM concentration of sodium. Here we report that light-induced proton efflux is sodium-dependent which leads to a characteristic enhancement in the rate of photosynthetic oxygen generation and carbon fixation. A low concentration (10 M) of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) significantly inhibited the rate of oxygen generation while 10 M carbonyl cyanide-m-chlorophenylhydrazone (CCCP) completely blocked the oxygen generation activity in the organism. The chlorophyll-a fluorescence yield indicates that little fluorescence quenching occurred in the absence of sodium ion. Increasing the extracellular sodium ion accelerated both the initial rate and the extent of fluorescence quenching. These results support the assumption that metal-induced inhibition of the photosynthetic machinery may be mediated by the movement of protons.     

Metabolic fate of [14C]-ethanol into endothelial cell phospholipids including platelet-activating factor, sphingomyelin and phosphatidylethanol

The metabolic fate of ethanol into the phospholipid pool of calf pulmonary artery endothelial cells was studied. [14C]-ethanol was incorporated into various endothelial cell phospholipids including phosphatidylethanol (PEth), which may represent a substantial fraction in microdomains of membrane phospholipids. The incorporation into phospholipids was reduced in the presence of pyrazole and cyanamide, inhibitors of ethanol metabolism. Wortmannin, the phosphatidylinositol 3-kinase inhibitor, increased [14C]-PEth formation. [3H]-acetate was also incorporated into endothelial cell phospholipids but in a different pattern. Distribution of [3H]-acetate and [14C]-ethanol into the fatty acyl moiety versus the glycerophosphoryl backbone of the phospholipids was also different. Stimulation of the endothelial cells with ATP increased [3H]-acetate incorporation into platelet-activating factor (PAF) and ethanol decreased it. Ethanol exposure increased ATP-stimulated [3H]-acetate incorporation into sphingomyelin. However, ATP had no effect on the incorporation of [14C]-ethanol into any phospholipids. The results suggest that the two precursors contribute to a separate acetate pool and that the sphingomyelin cycle may be sensitized in ethanol-treated cells. Thus, metabolic conversions of ethanol into lipids and the effect of ethanol on specific lipid mediators, e.g PAF, PEth and sphingomyelin, may be critical determinants in the altered responses of the endothelium in alcoholism.     

Assessment of peroxidase isozyme marker-based model for cross identifications in hybrids (F(1)) of urdbean [ Vigna mungo (L.) Hepper

Four hybrids (4 F₁s) were chosen out of crosses in the urdbean [Vigna mungo (L.) Hepper, 2n = 22] having contrasting morphological characters. Zymograms for isozyme peroxidase were drawn from the patterns obtained from parents and their respective F₁ hybrids on the basis of relative similarities to parental bands. The selfed or crossed nature of hybrid pods was determined from the zymograms and their analysis. The number of bands and their intensities gave an idea about the extent of crossing in F₁ populations. Genetic identity (I) values were indicative of their selfed nature. Dendrograms were constructed on the basis of genetic identity values to display the relative similarities between the populations. Analysis was based on individual pods to confirm their hybrid or selfed nature. Possible use of this technique for identification of F₁ pods and elimination of selfed pods might be implemented to shorten the breeding operations during crossing.     

Exuberant inflammation in nicotinamide adenine dinucleotide phosphate-oxidase-deficient mice after allogeneic marrow transplantation

We have shown that NO and superoxide (O-*2)contribute to donor T cell-dependent lung dysfunction after bone marrow transplantation (BMT) in mice. We hypothesized that inhibiting superoxide production during inducible NO synthase induction would suppress oxidative/nitrative stress and result in less severe lung injury. Irradiated mice lacking the phagocytic NADPH-oxidase (phox(-/-)), a contributor to superoxide generation, were conditioned with cyclophosphamide and given donor bone marrow in the presence or absence of inflammation-inducing allogeneic spleen T cells. On day 7 after allogeneic BMT, survival, weight loss, and indices of lung injury between phox(-/-) and wild-type mice were not different. However, the majority of macrophages/monocytes from phox(-/-) mice given donor T cells produced fewer oxidants and contained less nitrotyrosine than cells obtained from T cell-recipient wild-type mice. Importantly, suppressed oxidative stress was associated with marked infiltration of the lungs with inflammatory cells and was accompanied by increased bronchoalveolar lavage fluid levels of the chemoattractants monocyte chemoattractant protein-1 and macrophage-inflammatory protein-1alpha and impaired clearance of recombinant mouse macrophage-inflammatory protein-1beta from the circulation. Furthermore, cultured macrophages/monocytes from NADPH-deficient mice produced 3-fold more TNF-alpha compared with equal number of cells from NADPH-sufficient mice. The high NO production was not modified during NADPH-oxidase deficiency. We conclude that phox(-/-) mice exhibit enhanced pulmonary influx of inflammatory cells after BMT. Although NO may contribute to increased production of TNF-alpha in phox(-/-) mice, the data suggest that NADPH-oxidase-derived oxidants have a role in limiting inflammation and preventing lung cellular infiltration after allogeneic transplantation.