Boophilus microplus: the pattern of bovine immunoglobulin isotype responses to high and low tick infestations

Cattle present variable levels of resistance to ticks and the immune correlates of these heritable phenotypes must be known in order to develop effective vaccines. The antibody responses to tick salivary antigens were examined in cattle of tick-susceptible (Holstein) and tick-resistant (Nelore) breeds. After heavy infestations, levels of IgG1 and IgG2 antibodies decreased in Holsteins and remained the same in Nelores. Conversely, levels of IgE antibodies increased in Holsteins. Different sizes of tick burdens modulated the IgG1 antibody response in a susceptible breed (Aberdeen): levels were higher than in controls in heavily infested animals, but not in those undergoing intermediary or minimal infestations. The three experimental groups presented similar levels of IgG2 antibodies. Levels of IgE antibodies were higher only in animals undergoing intermediate infestations. These results indicate that tick infestations suppress the IgG antibody response in susceptible breeds, that IgE antibodies are not protective, and that the dose of tick saliva modulates the isotype of host antibody responses.     

Involvement of AMP-activated protein kinase in fat depot-specific metabolic changes during starvation

The mechanisms controlling fat depot-specific metabolism are poorly understood. During starvation of mice, downregulation of lipogenic genes, suppression of fatty acid synthesis, and increases in lipid oxidation were all more pronounced in epididymal than in subcutaneous fat. In epididymal fat, relatively strong upregulation of uncoupling protein 2 and phosphoenolpyruvate carboxykinase genes was found. In mice maintained both at 20 and 30 degrees C, AMP-activated protein kinase was activated in epididymal but did not change in subcutaneous fat. Our results suggest that AMPK may have a role in the different response of various fat depots to starvation.     

Protein remodeling of the heart ventricles in hereditary hypertriglyceridemic rat: effect of ace-inhibition

The aim of this study was to determine whether protein remodeling of the heart ventricles and remodeling of the aorta were present in hereditary hypertriglyceridemic (hHTG) rats and whether treatment with the angiotensin-converting enzyme inhibitor, captopril could prevent these alterations. Three groups of rats were investigated in a four week experiment control Wistar /C/rats, hHTg rats, hHTg rats given captopril (100 mg/kg/day) (hHTg + CAP). In the hHTg group, the increased systolic blood pressure (SBP) was associated with hypertrophy of the LV and RV. Protein profile analysis revealed an enhancement of metabolic protein concentration in both ventricles. The concentration of total collagenous proteins was not changed in either ventricles. However, alterations in composition of cardiac collagen were detected, characterized by higher concentration of hydroxyproline in pepsin-insoluble fraction and lower concentration of hydroxyproline in pepsin soluble faction in the LV. Hypertrophy of aorta, associated with the reduction of nitric oxide dependent relaxation, was also present in hHTG rats. Captopril normalized SBP, reduced left ventricular hypertrophy (LVH), diminished metabolic protein concentration in both ventricles, and improved NO-dependent relaxation of the aorta. Furthermore, captopril partially reversed alterations in hydroxyproline concentration in soluble and insoluble collagenous fractions of the LV. We conclude that hypertrophy of both ventricles and the aorta are present in hHTG rats, along with protein remodeling of both ventricles. Captopril partially prevented left ventricular hypertrophy development and protein remodeling of the myocardium.     

Expression cloning screening of a unique and full-length set of cDNA clones is an efficient method for identifying genes involved in Xenopus neurogenesis

Functional screens, where a large numbers of cDNA clones are assayed for certain biological activity, are a useful tool in elucidating gene function. In Xenopus, gain of function screens are performed by pool screening, whereby RNA transcribed in vitro from groups of cDNA clones, ranging from thousands to a hundred, are injected into early embryos. Once an activity is detected in a pool, the active clone is identified by sib-selection. Such screens are intrinsically biased towards potent genes, whose RNA is active at low quantities. To improve the sensitivity and efficiency of a gain of function screen we have bioinformatically processed an arrayed and EST sequenced set of 100,000 gastrula and neurula cDNA clones, to create a unique and full-length set of approximately 2500 clones. Reducing the redundancy and excluding truncated clones from the starting clone set reduced the total number of clones to be screened, in turn allowing us to reduce the pool size to just eight clones per pool. We report that the efficiency of screening this clone set is five-fold higher compared to a redundant set derived from the same libraries. We have screened 960 cDNA clones from this set, for genes that are involved in neurogenesis. We describe the overexpression phenotypes of 18 single clones, the majority of which show a previously uncharacterised phenotype and some of which are completely novel. In situ hybridisation analysis shows that a large number of these genes are specifically expressed in neural tissue. These results demonstrate the effectiveness of a unique full-length set of cDNA clones for uncovering players in a developmental pathway.     

Synthesis of some diguanidino 1-methyl-2,5-diaryl-1H-pyrroles as antifungal agents

A series of novel 2,5-bis(guanidino-aryl)-1-methyl-1H-pyrroles 9a-h has been synthesized starting from 1-methyl-1H-pyrrole. The antifungal activities of compounds were evaluated by in vitro agar diffusion and broth dilution assay against Candida spp. and Aspergillus spp. Compound 9c from this series was found to be equipotent or more potent than fluconazole, whereas compound 9d was comparable to fluconazole against most of the tested strains.     

Light activation of channelrhodopsin-2 in excitable cells of Caenorhabditis elegans triggers rapid behavioral responses

For studying the function of specific neurons in their native circuitry, it is desired to precisely control their activity. This often requires dissection to allow accurate electrical stimulation or neurotransmitter application , and it is thus inherently difficult in live animals, especially in small model organisms. Here, we employed channelrhodopsin-2 (ChR2), a directly light-gated cation channel from the green alga Chlamydomonas reinhardtii, in excitable cells of the nematode Caenorhabditis elegans, to trigger specific behaviors, simply by illumination. Channelrhodopsins are 7-transmembrane-helix proteins that resemble the light-driven proton pump bacteriorhodopsin , and they also utilize the chromophore all-trans retinal, but to open an intrinsic cation pore. In muscle cells, light-activated ChR2 evoked strong, simultaneous contractions, which were reduced in the background of mutated L-type, voltage-gated Ca2+-channels (VGCCs) and ryanodine receptors (RyRs). Electrophysiological analysis demonstrated rapid inward currents that persisted as long as the illumination. When ChR2 was expressed in mechanosensory neurons, light evoked withdrawal behaviors that are normally elicited by mechanical stimulation. Furthermore, ChR2 enabled activity of these neurons in mutants lacking the MEC-4/MEC-10 mechanosensory ion channel . Thus, specific neurons or muscles expressing ChR2 can be quickly and reversibly activated by light in live and behaving, as well as dissected, animals.     

Structure of the major cytosolic glutathione S-transferase from the parasitic nematode Onchocerca volvulus

Onchocerciasis is a debilitating parasitic disease caused by the filarial worm Onchocerca volvulus. Similar to other helminth parasites, O. volvulus is capable of evading the host's immune responses by a variety of defense mechanisms, including the detoxification activities of the glutathione S-transferases (GSTs). Additionally, in response to drug treatment, helminth GSTs are highly up-regulated, making them tempting targets both for chemotherapy and for vaccine development. We analyzed the three-dimensional x-ray structure of the major cytosolic GST from O. volvulus (Ov-GST2) in complex with its natural substrate glutathione and its competitive inhibitor S-hexylglutathione at 1.5 and 1.8 angstrom resolution, respectively. From the perspective of the biochemical classification, the Ov-GST2 seems to be related to pi-class GSTs. However, in comparison to other pi-class GSTs, in particular to the host's counterpart, the Ov-GST2 reveals significant and unusual differences in the sequence and overall structure. Major differences can be found in helix alpha-2, an important region for substrate recognition. Moreover, the binding site for the electrophilic co-substrate is spatially increased and more solvent-accessible. These structural alterations are responsible for different substrate specificities and will form the basis of parasite-specific structure-based drug design investigations.     

Tentacles of in vitro-grown round-leaf sundew (<Emphasis Type="Italic">Drosera rotundifolia</Emphasis>L.) show induction of chitinase activity upon mimicking the presence of prey

Induction of plant-derived chitinases in the leaves of a carnivorous plant was demonstrated using aseptically grown round-leaf sundew (Drosera rotundifolia L.). The presence of insect prey was mimicked by placing the chemical inducers gelatine, salicylic acid and crustacean chitin on leaves. In addition, mechanical stirring of tentacles was performed. Chitinase activity was markedly increased in leaf exudates upon application of notably chitin. Application of gelatine increased the proteolytic activity of leaf exudates, indicating that the reaction of sundew leaves depends on the molecular nature of the inducer applied. In situ hybridization of sundew leaves with a Drosera chitinase probe showed chitinase gene expression in different cell types of non-treated leaves, but not in the secretory cells of the glandular heads. Upon induction, chitinase mRNA was also present in the secretory cells of the sundew leaf. The combined results indicate that chitinase is likely to be involved in the decomposition of insect prey by carnivorous plants. This adds a novel role to the already broad function of chitinases in the plant kingdom and may contribute to our understanding of the molecular mechanisms behind the ecological success of carnivorous plants in nutritionally poor environments.