The participation of brain NO synthase in blood pressure control of adult spontaneously hypertensive rats

Increased blood pressure (BP) in genetic hypertension is usually caused by high activity of sympathetic nervous system (SNS) which is enhanced by central angiotensin II but lowered by central nitric oxide (NO). We have therefore evaluated NO synthase (NOS) activity as well as neuronal NOS (nNOS), inducible NOS (iNOS) and endothelial NOS (eNOS) protein expression in brainstem and midbrain of adult spontaneously hypertensive rats (SHR) characterized by enhanced sympathetic vasoconstriction. We also studied possible participation of brain NO in antihypertensive effects of chronic captopril treatment of adult SHR. NOS activity was increased in midbrain of SHR compared to Wistar-Kyoto (WKY) rats. This could be ascribed to enhanced iNOS expression, whereas nNOS expression was unchanged and eNOS expression was reduced in this brain region. In contrast, no significant changes of NOS activity were found in brainstem of SHR in which nNOS and iNOS expression was unchanged, but eNOS expression was increased. Chronic captopril administration lowered BP of adult SHR mainly by attenuation of sympathetic tone, whereas the reduction of angiotensin II-dependent vasoconstriction and the decrease of residual BP (amelioration of structural remodeling of resistance vessels) were less important. This treatment did not affect significantly either NOS activity or expression of any NOS isoform in the two brain regions. Our data do not support the hypothesis that altered brain NO formation contributes to sympathetic hyperactivity and high BP of adult SHR with established hypertension.     

Integrated view and comparative analysis of baseline protein expression in mouse and rat tissues

The increasingly large amount of proteomics data in the public domain enables, among other applications, the combined analyses of datasets to create comparative protein expression maps covering different organisms and different biological conditions. Here we have reanalysed public proteomics datasets from mouse and rat tissues (14 and 9 datasets, respectively), to assess baseline protein abundance. Overall, the aggregated dataset contained 23 individual datasets, including a total of 211 samples coming from 34 different tissues across 14 organs, comprising 9 mouse and 3 rat strains, respectively.In all cases, we studied the distribution of canonical proteins between the different organs. The number of canonical proteins per dataset ranged from 273 (tendon) and 9,715 (liver) in mouse, and from 101 (tendon) and 6,130 (kidney) in rat. Then, we studied how protein abundances compared across different datasets and organs for both species. As a key point we carried out a comparative analysis of protein expression between mouse, rat and human tissues. We observed a high level of correlation of protein expression among orthologs between all three species in brain, kidney, heart and liver samples, whereas the correlation of protein expression was generally slightly lower between organs within the same species. Protein expression results have been integrated into the resource Expression Atlas for widespread dissemination.     

Evaluation of DNA vaccination of spotted sand bass (Paralabrax maculatofasciatus) with two major outer-membrane protein-encoding genes from Aeromonas veronii

Genes encoding two major outer membrane proteins (OMPs) of the bacterial pathogen Aeromonas veronii, Omp38 and Omp48, were used to construct DNA vaccines. The protective effect of such vaccines against motile aeromonad septicaemia was evaluated in spotted sand bass (Paralabrax maculatofasciatus), an endemic species of the Mexican Northwest Pacific coast and a potential resource for the aquaculture industry. Weak protein expression, as determined by immunoblotting, was observed after transfection of eukaryotic cells with the DNA vaccines. Fish immunized with a single intramuscular injection of 20 microg of the omp38 and omp48 DNA vaccines showed slightly, but significantly elevated serum antibody levels 4 and 6 weeks after vaccination, compared to fish vaccinated with the control plasmid pcDNA3.1. Spotted sand bass vaccinated with the omp38 and omp48 DNA vaccines and challenged with A. veronii by intraperitoneal route recorded a relative percent survival (RPS) between 50 and 60%. Histopathological signs of motile aeromonad septicaemia were observed in around 40% of omp38 and omp48-vaccinated fish and 80% of pcDNA3.1-vaccinated control fish. The results indicate that P. maculatofasciatus vaccinated with a single dose of DNA plasmids encoding the major OMPs from A. veronii shows partial protection against infection and mortality by A. veronii experimental infection.     

L'entomophthoraleNeozygites fumosa pathogène de la Cochenille du manioc,Phenacoccus manihoti [Hom.: Pseudococcidae], en République populaire du Congo

Résumé Uniquement connue du sud des Etats-Unis d'Amérique, l'EntomophthoraleNeozygites fumosa (Speare) Remaudière & Keller a été retrouvée en République populaire du Congo, où elle attaque la Cochenille du manioc,Phenacoccus manihoti Matile-Ferrero [Hom.: Coccoidea Pseudococcidae]. De légères variations d'ordre biométrique sont mises en évidence au sein de l'espèce fongique. A ce jour, parmi les Entomophthorales mentionnées dans la littérature comme pathogènes de Cochenilles, seules deux espèces peuvent être retenues avec certitude:Neozygites fumosa etConidiobolus pseudococci Speare (Tyrrell & MacLeod), toutes deux pathogènes dePseudococcidae.

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Summary The entomophthoraceous fungus,Neozygites fumosa (Speare) Remaudière and Keller, which was only known from the south of the U.S.A., was found in the People's Republic of the Congo, on the Cassava mealybug,Phenacoccus manihoti Matile-Ferrero [Homoptera: Coccoidea, Pseudococcidae]. Slight biometrical variations appear within the fungal species. At the present time, among the Entomophthorales cited in the literature as parasitizing Coccoidea, two species only can be retained with certainty:Neozygites fumosa andConidiobolus pseudococci Speare (Tyrrell and MacLeod), both attackingPseudococcidae.

     

Apolipoprotein E Polymorphism in Hemodialyzed Patients and Healthy Controls

A possible association between end-stage renal disease (ESRD) and apolipoprotein E (APOE) polymorphism was found in some but not all studies. We have analyzed the APOE genotypes in 995 hemodialyzed patients (cases) and a sample of 6242 healthy individuals (controls) in the Czech Republic. There was a statistically significant difference in the frequency of APOE alleles between cases and controls, with more carriers of the APOE2 allele in ESRD patients (15.9%) than in controls (12.2%) (P = 0.005). The odds ratio of ESRD for the APOE2 allele, compared with APOE3E3 homozygotes, was 1.37 (95% confidence interval 1.13–1.67). The strength of the association increased with the time spent on hemodialysis: the odds ratio of all-cause ESRD in patients dialyzed for eight or more years was 1.27 (0.94–1.71), for 1–8 years 1.41 (1.09–1.81), and less than 1 year (nonsurvivors) 1.94 (0.88–4.18). This study suggests that the APOE2 allele is a possible genetic risk factor for all-cause ESRD in Caucasians.     

The Plasmodium serine‐type SERA proteases display distinct expression patterns and non‐essential in vivo roles during life cycle progression of the malaria parasite

Parasite proteases play key roles in several fundamental steps of the Plasmodium life cycle, including haemoglobin degradation, host cell invasion and parasite egress. Plasmodium exit from infected host cells appears to be mediated by a class of papain‐like cysteine proteases called ‘serine repeat antigens’ (SERAs). A SERA subfamily, represented by Plasmodium falciparum SERA5, contains an atypical active site serine residue instead of a catalytic cysteine. Members of this SERAser subfamily are abundantly expressed in asexual blood stages, rendering them attractive drug and vaccine targets. In this study, we show by antibody localization and in vivo fluorescent tagging with the red fluorescent protein mCherry that the two P. berghei serine‐type family members, PbSERA1 and PbSERA2, display differential expression towards the final stages of merozoite formation. Via targeted gene replacement, we generated single and double gene knockouts of the P. berghei SERAser genes. These loss‐of‐function lines progressed normally through the parasite life cycle, suggesting a specialized, non‐vital role for serine‐type SERAs in vivo. Parasites lacking PbSERAser showed increased expression of the cysteine‐type PbSERA3. Compensatory mechanisms between distinct SERA subfamilies may thus explain the absence of phenotypical defect in SERAser disruptants, and challenge the suitability to develop potent antimalarial drugs based on specific inhibitors of Plasmodium serine‐type SERAs.     

Prevention of high-fat diet-induced muscular lipid accumulation in rats by �� lipoic acid is not mediated by AMPK activation

Skeletal muscle triglyceride accumulation is associated with insulin resistance in obesity. Recently, it has been suggested that α lipoic acid (ALA) improves insulin sensitivity by lowering triglyceride accumulation in nonadipose tissues via activation of skeletal muscle AMP-activated protein kinase (AMPK). We examined whether chronic ALA supplementation prevents muscular lipid accumulation that is associated with high-fat diets via activation of AMPK. In addition, we tested if ALA supplementation was able to improve insulin sensitivity in rats fed low- and high-fat diets (LFD, HFD). Supplementing male Wistar rats with 0.5% ALA for 8 weeks significantly reduced body weight, both on LFD and HFD (−24% LFD+ALA vs. LFD, P < 0.01, and −29% HFD+ALA vs. HFD, P < 0.001). Oil red O lipid staining revealed a 3-fold higher lipid content in skeletal muscle after HFD compared with LFD and ALA-supplemented groups (P < 0.05). ALA improved whole body glucose tolerance (∼20% lower total area under the curve (AUC) in ALA supplemented groups vs. controls, P < 0.05). These effects were not mediated by increased muscular AMPK activation or ALA-induced improvement of muscular insulin sensitivity. To conclude, the prevention of HFD-induced muscular lipid accumulation and the improved whole body glucose tolerance are likely secondary effects due to the anorexic nature of ALA.     

Abstracts: Albany 2005, The 14th Conversation

Abstract

We report a cost efficient approach for amino-acid-type selective isotope labeling of proteins expressed in Leishmania tarentolae. The method provides an economically advantageous alternative to recently established protocol for isotopic labeling using expensive synthetic media. The method is based on cultivation of the L. tarentolae expression strain in a cheap complex medium supplemented with labeled amino acid(s). In this protocol, a labeled amino acid is deliberately diluted in the medium of undefined composition, which leads to a low-level isotope enrichment upon protein over-expression. The economic advantage of the protocol is achieved by avoiding large volumes of expensive synthetic medium. Decreased sensitivity of a NMR experiment due to low-level isotope enrichment is compensated by a five- to seven-fold increase of the yield of the recombinant protein in complex medium as compared to that in the synthetic medium. In addition, the decreased sensitivity can be compensated by using a higher magnetic field, cryo-detection system or higher number of transients during the NMR data acquisition. We show that enrichment as low as 5% does not compromise a NMR experiment and makes preparation of the recombinant proteins over- expressed in L. tarentolae economically viable. The method is demonstrated by selective labeling of the ～27 kDa enhanced green fluorescent protein (EGFP) with ¹⁵N-labeled valine.