Decrease in Formalin-Inactivated Respiratory Syncytial Virus (FI-RSV) Enhanced Disease with RSV G Glycoprotein Peptide Immunization in BALB/c Mice

Respiratory syncytial virus (RSV) is a high priority target for vaccine development. One concern in RSV vaccine development is that a non-live virus vaccine would predispose for enhanced disease similar to that seen with the formalin inactivated RSV (FI-RSV) vaccine. Since a mAb specific to RSV G protein can reduce pulmonary inflammation and eosinophilia seen after RSV infection of FI-RSV vaccinated mice, we hypothesized that RSV G peptides that induce antibodies with similar reactivity may limit enhanced disease after subunit or other non-live RSV vaccines. In support of this hypothesis, we show that FI-RSV vaccinated mice administered RSV G peptide vaccines had a significant reduction in enhanced disease after RSV challenge. These data support the importance of RSV G during infection to RSV disease pathogenesis and suggest that use of appropriately designed G peptide vaccines to reduce the risk of enhanced disease with non-live RSV vaccines merits further study.     

Curcuma longa Extract Associated with White Pepper Lessens High Fat Diet-Induced Inflammation in Subcutaneous Adipose Tissue

Background

Supra-nutritional doses of curcumin, derived from the spice Curcuma longa, have been proposed as a potential treatment of inflammation and metabolic disorders related to obesity. The aim of the present study was to test whether Curcuma longa extract rich in curcumin and associated with white pepper (Curcuma-P®), at doses compatible with human use, could modulate systemic inflammation in diet-induced obese mice. We questioned the potential relevance of changes in adiposity and gut microbiota in the effect of Curcuma-P® in obesity.

Methodology/Principal Findings

Mice were fed either a control diet (CT), a high fat (HF) diet or a HF diet containing Curcuma longa extract (0.1 % of curcumin in the HF diet) associated with white pepper (0.01 %) for four weeks. Curcumin has been usually combined with white pepper, which contain piperine, in order to improve its bioavailability. This combination did not significantly modify body weight gain, glycemia, insulinemia, serum lipids and intestinal inflammatory markers. Tetrahydrocurcumin, but not curcumin accumulated in the subcutaneous adipose tissue. Importantly, the co-supplementation in curcuma extract and white pepper decreased HF-induced pro-inflammatory cytokines expression in the subcutaneous adipose tissue, an effect independent of adiposity, immune cells recruitment, angiogenesis, or modulation of gut bacteria controlling inflammation.

Conclusions/Significance

These findings support that nutritional doses of Curcuma longa, associated with white pepper, is able to decrease inflammatory cytokines expression in the adipose tissue and this effect could be rather linked to a direct effect of bioactive metabolites reaching the adipose tissue, than from changes in the gut microbiota composition.     

Barth syndrome: Cellular compensation of mitochondrial dysfunction and apoptosis inhibition due to changes in cardiolipin remodeling linked to tafazzin (TAZ) gene mutation

Cardiolipin is a mitochondrion-specific phospholipid that stabilizes the assembly of respiratory chain complexes, favoring full-yield operation. It also mediates key steps in apoptosis. In Barth syndrome, an X chromosome-linked cardiomyopathy caused by tafazzin mutations, cardiolipins display acyl chain modifications and are present at abnormally low concentrations, whereas monolysocardiolipin accumulates. Using immortalized lymphoblasts from Barth syndrome patients, we showed that the production of abnormal cardiolipin led to mitochondrial alterations. Indeed, the lack of normal cardiolipin led to changes in electron transport chain stability, resulting in cellular defects. We found a destabilization of the supercomplex (respirasome) I + III₂ + IV_n but also decreased amounts of individual complexes I and IV and supercomplexes I + III and III + IV. No changes were observed in the amounts of individual complex III and complex II. We also found decreased levels of complex V. This complex is not part of the supercomplex suggesting that cardiolipin is required not only for the association/stabilization of the complexes into supercomplexes but also for the modulation of the amount of individual respiratory chain complexes. However, these alterations were compensated by an increase in mitochondrial mass, as demonstrated by electron microscopy and measurements of citrate synthase activity. We suggest that this compensatory increase in mitochondrial content prevents a decrease in mitochondrial respiration and ATP synthesis in the cells. We also show, by extensive flow cytometry analysis, that the type II apoptosis pathway was blocked at the mitochondrial level and that the mitochondria of patients with Barth syndrome cannot bind active caspase-8. Signal transduction is thus blocked before any mitochondrial event can occur. Remarkably, basal levels of superoxide anion production were slightly higher in patients' cells than in control cells as previously evidenced via an increased protein carbonylation in the taz1Δ mutant in the yeast. This may be deleterious to cells in the long term. The consequences of mitochondrial dysfunction and alterations to apoptosis signal transduction are considered in light of the potential for the development of future treatments.     

An assessment of ADAMs in bone cells: absence of TACE activity prevents osteoclast recruitment and the formation of the marrow cavity in developing long bones

ADAMs (A Disintegrin And Metalloprotease domain) are metalloprotease-disintegrin proteins that have been implicated in cell adhesion, protein ectodomain shedding, matrix protein degradation and cell fusion. Since such events are critical for bone resorption and osteoclast recruitment, we investigated whether they require ADAMs. We report here which ADAMs we have identified in bone cells, as well as our analysis of the generation, migration and resorptive activity of osteoclasts in developing metatarsals of mouse embryos lacking catalytically active ADAM 17 [TNFalpha converting enzyme (TACE)]. The absence of TACE activity still allowed the generation of cells showing an osteoclastic phenotype, but prevented their migration into the core of the diaphysis and the subsequent formation of marrow cavity. This suggests a role of TACE in the recruitment of osteoclasts to future resorption sites.     

In vivo impact of CpG1826 oligodeoxynucleotide on CD8 T cell primary responses and survival

CpG oligodeoxynucleotide (ODN) promotes maturation of APCs in vivo and induces strong type 1 T cell responses in mice. In this study, we have investigated the ability of CpG1826 to modulate peptide-specific CD8 T cell responses in a context where CD4 T cells are likely to play a minor role. The effects of CpG1826 were evaluated in a system where a population of NP68-specific F5 TCR transgenic CD8 T cells is diluted into a polyclonal host following adoptive transfer into C57BL/10 syngeneic recipients. Using this approach, we found that CpG1826 enhanced the ability of F5 CD8 T cells to undergo multiple divisions in vivo, to express IFN-gamma ex vivo, and to up-regulate memory-associated cell surface markers such as CD122 (IL-2Rbeta) and Ly-6C. Moreover, CpG1826 greatly increased in vivo cytotoxic activity. Using tetramer detection, we found that CpG1826 promoted long-term survival of Ag-specific CD8 T cells after immunization while no NP68-specific cells were detected when the cognate peptide was injected alone. These results indicate that CpG1826 acts as an adjuvant which increases CD8 T cell effector responses and promotes long-term survival of NP68 peptide-specific cells in vivo. They also suggest that this adjuvant can modulate CD8 T cell responses in a system which is likely to be independent of CD4 T cell help.     

Evaluation of a high-density oligonucleotide array for characterization of grlA, grlB, gyrA and gyrB mutations in fluoroquinolone resistant Staphylococcus aureus isolates

Using high-density oligonucleotide array technology, 30 Staphylococcus aureus strains were studied for the presence of mutations in genes involved in fluoroquinolone resistance: grlA, gyrA, grlB and gyrB. For the two most important genes, gyrA and grlA, correlation with sequencing reached 95.1%. If all genes were considered, correlation was 88.8%.     

Fish can show emotional fever: stress-induced hyperthermia in zebrafish

Whether fishes are sentient beings remains an unresolved and controversial question. Among characteristics thought to reflect a low level of sentience in fishes is an inability to show stress-induced hyperthermia (SIH), a transient rise in body temperature shown in response to a variety of stressors. This is a real fever response, so is often referred to as ‘emotional fever’. It has been suggested that the capacity for emotional fever evolved only in amniotes (mammals, birds and reptiles), in association with the evolution of consciousness in these groups. According to this view, lack of emotional fever in fishes reflects a lack of consciousness. We report here on a study in which six zebrafish groups with access to a temperature gradient were either left as undisturbed controls or subjected to a short period of confinement. The results were striking: compared to controls, stressed zebrafish spent significantly more time at higher temperatures, achieving an estimated rise in body temperature of about 2–4°C. Thus, zebrafish clearly have the capacity to show emotional fever. While the link between emotion and consciousness is still debated, this finding removes a key argument for lack of consciousness in fishes.     

Synthesis and GABAA receptor activity of 2,19-sulfamoyl analogues of allopregnanolone

The synthesis of new analogues of allopregnanolone with a bridged sulfamidate ring over the β-face of ring A has been achieved from easily available precursors, using an intramolecular aziridination strategy. The methodology also allows the synthesis of 3α-substituted analogues such as the 3α-fluoro derivative. GABA_A receptor activity of the synthetic analogues was evaluated by assaying their effect on the binding of [³H]flunitrazepam and [³H]muscimol. The 3α-hydroxy-2,19-sulfamoyl analogue and its N-benzyl derivative were more active than allopregnanolone for stimulating binding of [³H]flunitrazepam. For the binding of [³H]muscimol, both synthetic analogues and allopregnanolone stimulated binding to a similar extent, with the N-benzyl derivative exhibiting a higher EC₅₀. The 3α-fluoro derivative was inactive in both assays.     

Infectivity of Leishmania mexicana Is Associated with Differential Expression of Protein Kinase C-Like Triggered during a Cell-Cell Contact

Mammalian host cell invasion by Leishmania is a complex process in which various parasite and host cell components interact, triggering the activation of signaling cascades in both cells. Little is known regarding PKC biological functions in Leishmania sp. during parasite-macrophage interaction. PKC-like enzyme was first identified in homogenates and membrane fraction of L. mexicana stationary promastigotes by immunoblot. PKC-like enzyme activity was then detected in cell homogenates but also on intact promastigotes showing for the first time the presence of an ecto-PKC dependent on Ca²⁺/phosphatidylserine for activation. This ecto-PKC was activated with phorbol myristate acetate (PMA) and inhibited by RO-32-0432, a selective PKCαβ_Iε bisindolylmaleimide inhibitor. Interestingly, the Leishmania PKC- activity was higher in the infective stationary than in non-infective logarithmic stage. Then, promastigotes at different stages of time proliferation curve were used in order to identify the role of PKC-like during macrophage invasion. After attachment to macrophages, PKC-like is over-expressed in promastigotes at the 6^th culture day but also at the 4^th day of culture corresponding to the maximal infection capacity. An antibody microarray for MAPK and PKC corroborate the Leishmania PKC-like over-expression during contact with macrophages. Pretreatment with RO-32-0432 inhibitor reduced the number of infected macrophages and the parasite burden. These data suggest for the first time a direct link between PKC expression level and infectivity, and provide evidence that PKC-like plays a critical role in attachment and in the internalization steps involved in the invasion process.