A tetravalent dengue nanoparticle stimulates antibody production in mice

Background

Dengue is a major public health problem worldwide, especially in the tropical and subtropical regions of the world. Infection with a single Dengue virus (DENV) serotype causes a mild, self-limiting febrile illness called dengue fever. However, a subset of patients experiencing secondary infection with a different serotype progresses to the severe form of the disease, dengue hemorrhagic fever/dengue shock syndrome. Currently, there are no licensed vaccines or antiviral drugs to prevent or treat dengue infections. Biodegradable nanoparticles coated with proteins represent a promising method for in vivo delivery of vaccines.

Findings

Here, we used a murine model to evaluate the IgG production after administration of inactivated DENV corresponding to all four serotypes adsorbed to bovine serum albumin nanoparticles. This formulation induced a production of anti-DENV IgG antibodies (p < 0.001). However, plaque reduction neutralization assays with the four DENV serotypes revealed that these antibodies have no neutralizing activity in the dilutions tested.

Conclusions

Our results show that while the nanoparticle system induces humoral responses against DENV, further investigation with different DENV antigens will be required to improve immunogenicity, epitope specicity, and functional activity to make this platform a viable option for DENV vaccines.     

Determining resistance to Pseudomonas syringae in tomato, a comparison of different molecular markers

Pseudomonas syringae pv. tomato (Pst) is the causal agent of bacterial speck disease in tomato. Resistance to Pst is determined by Pto, a single resistance gene that belongs to a multi-gene family clustered on chromosome 5. Pst-resistant phenotypes in cultivated tomato are determined by a semi-dominant allele of Solanum pimpinellifolium, which was introgressed into Solanum lycopersicum in the past century. Seed companies, which are continuously interested in producing resistant varieties, can benefit from genetic markers closely linked to the Pto locus in breeding programs based on marker-assisted selection. In this research, three sequence characterised amplified region markers have been developed for identification of resistant and susceptible genotypes of Solanum lycopersicum. A cleaved amplified polymorphic sequence marker has been adapted to a real-time polymerase chain reaction platform using high-resolution melting curve analysis. Application to a tomato population for breeding programs is described. Advantages and disadvantages of the different markers are discussed.     

New heteroaryl carbamates: Synthesis and biological screening in vitro and in mammalian cells of wild-type and mutant HIV-protease inhibitors

New heteroaryl HIV-protease inhibitors bearing a carbamoyl spacer were synthesized in few steps and high yield, from commercially available homochiral epoxides. Different substitution patterns were introduced onto a given isopropanoyl-sulfonamide core that can have either H or benzyl group. The in vitro inhibition activity against recombinant protease showed a general beneficial effect of both carbamoyl moiety and the benzyl group, ranging the IC₅₀ values between 11 and 0.6?nM. In particular, benzofuryl and indolyl derivatives showed IC₅₀ values among the best for such structurally simple inhibitors. Docking analysis allowed to identify the favorable situation of such derivatives in terms of number of interactions in the active site, supporting the experimental results.The inhibition activity was also confirmed in HEK293 mammalian cells and was maintained against protease mutants. Furthermore, the metabolic stability was comparable with that of the commercially available inhibitors.     

Valproate Administered after Traumatic Brain Injury Provides Neuroprotection and Improves Cognitive Function in Rats

Background

Traumatic brain injury (TBI) initiates a complex series of neurochemical and signaling changes that lead to pathological events including neuronal hyperactivity, excessive glutamate release, inflammation, increased blood-brain barrier (BBB) permeability and cerebral edema, altered gene expression, and neuronal dysfunction. It is believed that a drug combination, or a single drug acting on multiple targets, may be an effective strategy to treat TBI. Valproate, a widely used antiepileptic drug, has a number of targets including GABA transaminase, voltage-gated sodium channels, glycogen synthase kinase (GSK)-3, and histone deacetylases (HDACs), and therefore may attenuate a number of TBI-associated pathologies.

Methodology/Principal Findings

Using a rodent model of TBI, we tested if post-injury administration of valproate can decrease BBB permeability, reduce neural damage and improve cognitive outcome. Dose-response studies revealed that systemic administration of 400 mg/kg (i.p.), but not 15, 30, 60 or 100 mg/kg, increases histone H3 and H4 acetylation, and reduces GSK-3 activity, in the hippocampus. Thirty min post-injury administration of 400 mg/kg valproate improved BBB integrity as indicated by a reduction in Evans Blue dye extravasation. Consistent with its dose response to inhibit GSK-3 and HDACs, valproate at 400 mg/kg, but not 100 mg/kg, reduced TBI-associated hippocampal dendritic damage, lessened cortical contusion volume, and improved motor function and spatial memory. These behavioral improvements were not observed when SAHA (suberoylanilide hydroxamic acid), a selective HDAC inhibitor, was administered.

Conclusion/Significance

Our findings indicate that valproate given soon after TBI can be neuroprotective. As clinically proven interventions that can be used to minimize the damage following TBI are not currently available, the findings from this report support the further testing of valproate as an acute therapeutic strategy.     

Metal regulation of siderophore synthesis in Pseudomonas aeruginosa and functional effects of siderophore-metal complexes.

Pseudomonas aeruginosa synthesizes two siderophores, pyochelin and pyoverdin, characterized by widely different structures, physicochemical properties, and affinities for Fe(III). Titration experiments showed that pyochelin, which is endowed with a relatively low affinity for Fe(III), binds other transition metals, such as Cu(II), Co(II), Mo(VI), and Ni(II), with appreciable affinity. In line with these observations, Fe(III) and Co(II) at 10 microM or Mo(VI), Ni(II), and Cu(II) at 100 microM repressed pyochelin synthesis and reduced expression of iron-regulated outer membrane proteins of 75, 68, and 14 kDa. In contrast, pyoverdin synthesis and expression of the 80-kDa receptor protein were affected only by Fe(III). All of the metals tested, except Mo(VI), significantly promoted P. aeruginosa growth in metal-poor medium; Mo(VI), Ni(II), and Co(II) were more efficient as pyochelin complexes than the free metal ions and the siderophore. The observed correlation between the affinity of pyochelin for Fe(III), Co(II), and Mo(VI) and the functional effects of these metals indicates that pyochelin may play a role in their delivery to P. aeruginosa.     

Proteolytic and partial sequencing studies of the bifunctional dihydrofolate reductase-thymidylate synthase from Daucus carota

The bifunctional dihydrofolate reductase-thymidylate synthase (DHFR-TS) of Daucus carota has been further characterized as regards molecular weight, amino acid composition, protease digestion and microsequencing of proteolytic peptides. Data reported in this paper demonstrate that the carrot protein has a calculated M _r of 124000 thus indicating that, contrarily to what has previously been suggested, it occurs as a dimer of identical subunits. Results of partial amino acid microsequencing show the presence of sequences highly homologous with those of the active sites of both DHFR and TS from other organisms confirming, at the structural level, the bifunctional nature of the carrot protein. As in the case of Leishmania tropica DHFR-TS, incubation of the carrot protein with V8 protease led to a rapid loss of TS activity while retaining that of DHFR. However the pattern of proteolysis did not allow to establish whether the sequence of domains is DHFR-TS as in Leishmania, or vice versa. Low homology of other amino acid sequences, as judged by computer analysis, and absence of common epitopes indicate an apparent divergence between carrot and leishmanian proteins.     

Effect of bisacodyl and cascara on growth of aberrant crypt foci and malignant tumors in the rat colon.

Laxatives abuse has been associated with an increased risk for colon cancer. However, little is known about laxatives long-term carcinogenic potential in experimental studies. The present study was designed to investigate the effects of bisacodyl (4.3 and 43 mg/kg) and cascara (140 and 420 mg/kg) on azoxymethane (AOM)-induced aberrant crypt foci (ACF) and tumors. Animals, divided in 10 groups were treated with AOM and laxatives (alone or in combination) for 13 weeks. At the end of treatment animals were killed and the colon removed and analysed for the determination of ACF and tumors. Bisacodyl (4.3 and 43 mg/kg), given alone, did not induce the development of colonic ACF and tumors. Bisacodyl (4.3 mg/kg) coupled with AOM increased the number of crypt per focus, but not the number of tumors. Bisacodyl (43 mg/kg) significantly increased the number of crypt per focus and tumors. Cascara (140 and 420 mg/kg) did not induce the development of colonic ACF and tumors and did not modify the number of AOM-induced ACF and tumors. The results of the present study indicate a possible promoting effect of bisacodyl on rat colon carcinogenesis (especially at higher doses) and absence of any promoting or initiating activity of a laxative and diarrhoeal dose of cascara.     

A rapid method for measuring ATP + ADP + AMP in marine sediment

In this report, I describe a method for rapid measurement of total adenylate (ATP + ADP + AMP) in marine sediment samples for estimating microbial biomass. A simple ‘boil and dilute’ method is described here, whereby adding boiled MilliQ water to sediments increases the detection limit for ATP + ADP + AMP up to 100-fold. The lowered detection limit of this method enabled the detection ATP + ADP + AMP in relatively low-biomass sub-seafloor sediment cores with 10⁴ 16S rRNA gene copies per gram. Concentrations of ATP + ADP + AMP correlated with 16S rRNA gene concentrations from bacteria and archaea across six different sites that range in water depth from 1 to 6000 m indicating that the ATP + ADP + AMP method can be used as an additional biomass proxy. In deep sea microbial communities, the ratio of ATP + ADP + AMP concentrations to 16S rRNA genes >1 m below seafloor was significantly lower compared to communities in the upper 30 cm of sediment, which may be due to reduced cell sizes and or lower ATP + ADP + AMP concentrations per cell in the deep sea sub-seafloor biosphere. The boil and dilute method for ATP + ADP + AMP is demonstrated here to have a detection limit sufficient for measuring low biomass communities from deep sea sub-seafloor cores. The method can be applied to frozen samples, enabling measurements of ATP + ADP + AMP from frozen sediment cores stored in core repositories from past and future international drilling campaigns.