Monoclonal antibody produced against partially purified sporozoite antigen inhibits invasion of cells by sporozoites of avian Eimeria species

Previous studies showed that molecules of in vitro-cultured primary turkey kidney cells bound to 23-, 40-, and 60- to 65-kDa antigens of sodium dodecyl sulfate-solubilized sporozoites of Eimeria adenoeides. Similar binding to antigens of three other species of avian Eimeria, E. tenella, E. acervulina, and E. meleagrimitis, is now reported. Strips containing the most avidly bound sporozoite antigen (approximately 40 kDa) were excised from the sodium dodecyl sulfate-polyacrylamide gels on which E. adenoeides antigens had been electrophoretically separated. The strips were homogenized and injected into mice to produce hybridoma cell lines. Twelve cell lines secreting monoclonal antibodies (McAb) that reacted with E. adenoeides sporozoites were detected. One of these McAb, H11C3, reacted with structures in the anterior tip of sporozoites of E. adenoeides and five other species of avian Eimeria. When included in the inoculation medium, this McAb significantly inhibited invasion of cultured kidney cells by sporozoites of E. adenoeides and E. tenella. In contrast, when the sporozoites were pretreated with McAb H11C3 and then washed free of the antibody, no inhibition of invasion was observed.     

Group B streptococci inhibit the chemotactic activity of the fifth component of complement

Infection with group B streptococci (GBS) is associated with a poor acute inflammatory response in which neutrophils fail to localize at the site of invasion. In the present studies, we have examined the effects of group B streptococci on C-derived chemotactic activity in human serum. Fresh human serum was activated to form C5a and C5adesarg by incubation with zymosan. The activated serum was then incubated with group B organisms, centrifuged, and the supernatants tested for chemotactic activity for human polymorphonuclear leukocytes. Group B organisms caused a dose-dependent decrease in C-dependent chemotactic activity. The degree of inhibition was profound with 1 X 10(9) bacteria/ml (10% of control). Experiments indicated that significant chemotactic factor inactivation occurred within 2 min of exposure to GBS organisms, while maximal inhibition occurred after 30 min incubation. A number of different strains of GBS of types I, II, and III possessed inhibitory activity. In contrast, group D streptococci, Staphylococcus aureus, Escherichia coli and Klebsiella pneumoniae failed to inhibit the C-derived chemotactic activity in human serum. Group A streptococci that were M protein positive also inactivated C-dependent chemotactic activity in serum, as previously reported. The inhibitory activity of the GBS strains could be abolished by heat or trypsin treatment but not by neuraminidase, pronase, or pepsin. C5a levels in zymosan-activated serum as measured by RIA were not decreased after incubation with an inhibitory strain suggesting that absorption was not involved. SDS-PAGE analysis revealed that group B streptococci degrade the C5a molecule, increasing its electrophoretic mobility by removing a fragment with a m.w. of approximately 650 Da. Thus, one of the reasons for the poor inflammatory response at the site of GBS infection may reside in the ability of these pathogens to inactivate C-derived inflammatory mediators. The GBS C5a-ase activity probably serves as an additional virulence factor for these organisms contributing to the poor inflammatory response characteristic of group B streptococcal infection.     

Interactions between frequency-dependent and vertical transmission in host-parasite systems.

We investigate host-pathogen dynamics and conditions for coexistence in two models incorporating frequency-dependent horizontal transmission in conjunction with vertical transmission. The first model combines frequency-dependent and uniparental vertical transmission, while the second addresses parasites transmitted vertically via both parents. For the first model, we ask how the addition of vertical transmission changes the coexistence criteria for parasites transmitted by a frequency-dependent horizontal route, and show that vertical transmission significantly broadens the conditions for parasite invasion. Host-parasite coexistence is further affected by the form of density-dependent host regulation. Numerical analyses demonstrate that within a host population, a parasite strain with horizontal frequency-dependent transmission can be driven to extinction by a parasite strain that is additionally transmitted vertically for a wide range of parameters. Although models of asexual host populations predict that vertical transmission alone cannot maintain a parasite over time, analysis of our second model shows that vertical transmission via both male and female parents can maintain a parasite at a stable equilibrium. These results correspond with the frequent co-occurrence of vertical with sexual transmission in nature and suggest that these transmission modes can lead to host-pathogen coexistence for a wide range of systems involving hosts with high reproductive rates.     

Efficacy of the anticaspase agent zVAD-fmk on post-thaw viability of canine spermatozoa

Cryopreservation protocols for gametes are constantly improved with the aim of increasing the post-thaw viability of gametes. It is becoming clear that stress, resulting from cryopreservation, reduces cell numbers by apoptosis. Apoptosis, or programmed cell death, is a gene-activated event that occurs as a normal consequence of development and as a result of cellular stress. Apoptosis is mediated by the family of cysteine-dependent asparate-specific proteases (caspases). The present study was designed to test the hypothesis that addition of an anticaspase (zVAD-fmk) that has inhibitory properties against caspases and apoptosis to semen extenders and to the thaw medium would increase post-thaw viability of canine spermatozoa. Extenders were added in a two-step process. A dose of 100 microM caspase inhibitor was used. Four groups (n=6 for each) were composed based on the presence or absence of the caspase inhibitor: Group I (control), no caspase inhibitor in the extender or the thaw medium; Group II, caspase inhibitor in the thaw medium; Group III, caspase inhibitor in Extender II; and Group IV, caspase inhibitor in both Extender II and the thaw medium. Post-thaw motility, plasma membrane integrity, and acrosome status were investigated. The addition of caspase inhibitor to Extender II or to the thaw medium failed to improve the parameters that were studied. The results suggest that this caspase inhibitor may not be beneficial to the post-thaw motility of canine spermatozoa if used at this concentration.     

Caveolin-1: a critical regulator of lung injury

Caveolin-1 (cav-1), a 22-kDa transmembrane scaffolding protein, is the principal structural component of caveolae. Cav-1 regulates critical cell functions including proliferation, apoptosis, cell differentiation, and transcytosis via diverse signaling pathways. Abundant in almost every cell type in the lung, including type I epithelial cells, endothelial cells, smooth muscle cells, fibroblasts, macrophages, and neutrophils, cav-1 plays a crucial role in the pathogenesis of acute lung injury (ALI). ALI and its severe form, acute respiratory distress syndrome (ARDS), are responsible for significant morbidity and mortality in intensive care units, despite improvement in ventilation strategies. The pathogenesis of ARDS is still poorly understood, and therapeutic options remain limited. In this article, we summarize recent data regarding the regulation and function of cav-1 in lung biology and pathology, in particular as it relates to ALI. We further discuss the potential molecular and cellular mechanisms by which cav-1 expression contributes to ALI. Investigating the cellular functions of cav-1 may provide new insights for understanding the pathogenesis of ALI and provide novel targets for therapeutic interventions in the future.     

Autophagic proteins: New facets of the oxygen paradox

Oxygen (O 2), while essential for aerobic life, can also cause metabolic toxicity through the excess generation of reactive oxygen species (ROS). Pathological changes in ROS production can originate through the partial reduction of O 2 during mitochondrial electron transport, as well as from enzymatic sources. This phenomenon, termed the oxygen paradox, has been implicated in aging and disease, and is especially evident in critical care medicine. Whereas high O 2 concentrations are utilized as a life-sustaining therapeutic for respiratory insufficiency, they in turn can cause acute lung injury. Alveolar epithelial cells represent a primary target of hyperoxia-induced lung injury. Recent studies have indicated that epithelial cells exposed to high O 2 concentrations die by apoptosis, or necrosis, and can also exhibit mixed-phenotypes of cell death (aponecrosis). Autophagy, a cellular homeostatic process responsible for the lysosomal turnover of organelles and proteins, has been implicated as a general response to oxidative stress in cells and tissues. This evolutionarily conserved process is finely regulated by a complex interplay of protein factors. During autophagy, senescent organelles and cellular proteins are sequestered in autophagic vacuoles (autophagosomes) and subsequently targeted to the lysosome, where they are degraded by lysosomal hydrolases, and the breakdown products released for reutilization in anabolic pathways. Autophagy has been implicated as a cell survival mechanism during nutrient-deficiency states, and more generally, as a determinant of cell fate. However, the mechanisms by which autophagy and/or autophagic proteins potentially interact with and/or regulate cell death pathways during high oxygen stress, remain only partially understood.     

Uncoating of clathrin-coated vesicles in presynaptic terminals: roles for Hsc70 and auxilin.

We have examined the roles of Hsc70 and auxilin in the uncoating of clathrin-coated vesicles (CCVs) during neuronal endocytosis. We identified two peptides that inhibit the ability of Hsc70 and auxilin to uncoat CCVs in vitro. When injected into nerve terminals, these peptides inhibited both synaptic transmission and CCV uncoating. Mutation of a conserved HPD motif within the J domain of auxilin prevented binding to Hsc70 in vitro and injecting this mutant protein inhibited CCV uncoating in vivo, demonstrating that the interaction of auxilin with Hsc70 is critical for CCV uncoating. These studies establish that auxilin and Hsc70 participate in synaptic vesicle recycling in neurons and that an interaction between these proteins is required for CCV uncoating.     

Integrated system approach to dark fermentative biohydrogen production for enhanced yield,energy efficiency and substrate recovery

The challenges of climate change, dwindling fossil reserves, and environmental pollution have fuelled the need to search for clean and sustainable energy resources. The process of biohydrogen has been highlighted as a propitious alternative energy of the future because it has many socio-economic benefits such as non-polluting features, the ability to use diverse feedstocks including waste materials, the process uses various microorganisms, and it is the simplest method of producing hydrogen. However, the establishment of a biohydrogen driven economy has been hindered by low process yields due to the accumulation of inhibitory products. Over the past few years, various optimization methods have been used in literature. Among these, integration of bioprocesses is gaining increasing prominence as an effective approach that could be used to achieve a theoretical yield of 4 mol H₂ mol^?1 glucose. In batch integrated systems, dark fermentation is used as a primary process for conversion of substrates into biohydrogen, carbon dioxide, and volatile fatty acids. This is followed by a secondary anaerobic process for further biohydrogen conversion efficiency. This review discusses the current challenges facing scale-up studies in dark fermentation process. It elucidates the potential of batch integrated systems in biohydrogen process development. Furthermore, it explores the various integrated fermentation techniques that are employed in biohydrogen process development. Finally, the review concludes with recommendations on improvement of these integrated processes for enhanced biohydrogen yields which could pave a way for the establishment of a large-scale biohydrogen production process.     

Deficiency in the c-Jun NH2-terminal kinase signaling pathway confers susceptibility to hyperoxic lung injury in mice

Hyperoxia generates an oxidative stress in the mouse lung, which activates the major stress-inducible kinase pathways, including c-Jun NH2-terminal kinase (JNK). We examined the effect of Jnk1 gene deletion on in vivo responses to hyperoxia in mice. The survival of Jnk1-/- mice was reduced relative to wild-type mice after exposure to continuous hyperoxia. Jnk1-/- mice displayed higher protein concentration in bronchoalveolar lavage (BAL) fluid and increased expression of heme oxygenase-1, a stress-inducible gene, after 65 h of hyperoxia. Contrary to other markers of injury, the leukocyte count in BAL fluid of Jnk1-/- mice was markedly diminished relative to that of wild-type mice. The decrease in BAL leukocyte count was not associated with any decrease in lung myeloperoxidase activity at baseline or after hyperoxia treatment. Pretreatment with inhaled lipopolysaccharide increased BAL neutrophil content and extended hyperoxia survival time to a similar extent in Jnk1-/- and wild-type mice. Associated with increased mortality, Jnk1-/- mice had increased pulmonary epithelial cell apoptosis after exposure to hyperoxia compared with wild-type mice. These results indicate that JNK pathways participate in adaptive responses to hyperoxia in mice.