Differential expression of odorant-binding proteins in the mandibular glands of the honey bee according to caste and age

Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) mediate both perception and release of chemical stimuli in insects. The genome of the honey bee contains 21 genes encoding OBPs and 6 encoding CSPs. Using a proteomic approach, we have investigated the expression of OBPs and CSPs in the mandibular glands of adult honey bees in relation to caste and age. OBP13 is mostly expressed in young individuals and in virgin queens, while OBP21 is abundant in older bees and is prevalent in mated queens. OBP14, which had been found in larvae, is produced in hive workers' glands. Quite unexpectedly, the mandibular glands of drones also contain OBPs, mainly OBP18 and OBP21. We have expressed three of the most represented OBPs and studied their binding properties. OBP13 binds with good specificity oleic acid and some structurally related compounds, OBP14 is better tuned to monoterpenoid structures, while OBP21 binds the main components of queen mandibular pheromone as well as farnesol, a compound used as a trail pheromone in the honey bee and other hymenopterans. The high expression of different OBPs in the mandibular glands suggests that such proteins could be involved in solubilization and release of semiochemicals.     

Proteome characterization of sea star coelomocytes--the innate immune effector cells of echinoderms

Sea star coelomic fluid is in contact with all internal organs, carrying signaling molecules and a large population of circulating cells, the coelomocytes. These cells, also known as echinoderm blood cells, are responsible for the innate immune responses and are also known to have an important role in the first stage of regeneration, i.e. wound closure, necessary to prevent disruption of the body fluid balance and to limit the invasion of pathogens. This study focuses on the proteome characterization of these multifunctional cells. The identification of 358 proteins was achieved using a combination of two techniques for protein separation (1-D SDS-PAGE followed by nanoLC and 2-D SDS-PAGE) and MALDI-TOF/TOF MS for protein identification. To our knowledge, the present report represents the first comprehensive list of sea star coelomocyte proteins, constituting an important database to validate many echinoderm-predicted proteins. Evidence for new pathways in these particular echinoderm cells are also described, and thus representing a valuable resource to stimulate future studies aiming to unravel the homology with vertebrate immune cells and particularly the origins of the immune system itself.     

Toll-Like Receptor 4 Engagement Drives Differentiation of Human and Murine Dendritic Cells from a Pro- into an Anti-Inflammatory Mode

The dendritic cell (DC) coordinates innate and adaptive immunity to fight infections and cancer. Our observations reveal that DCs exposed to the microbial danger signal lipopolysaccharide (LPS) in the presence of interferon-γ (IFN-γ) acquire a continuously changing activation/maturation phenotype. The DCs’ initial mode of action is pro-inflammatory via up-regulation among others of the signaling molecule interleukin (IL) 12, which polarizes IFN-γ secreting type 1 helper T-cells (Th1). Within 24 hours the same DC switches from the pro- into an anti-inflammatory phenotype. This is mediated by autocrine IL-10 release and secretion of soluble IL-2 receptor alpha (sIL-2RA) molecules. T-cells, when contacted with DCs during their anti-inflammatory phase loose their proliferative capacity and develop regulatory T-cell (Treg) -like anti-inflammatory functions indicated by IL-10 secretion and elevated FoxP3 levels. Studying the kinetics of IL-12 and IL-10 expression from LPS/IFN-γ activated myeloid DCs on a single cell level confirmed these observations. When T-cells are separated from DCs within 24 hours, they are spared from the anti-inflammatory DC activity. We conclude that, in addition to differentiation of DCs into distinct subsets, the observed sequential functional phases of DC differentiation permit the fine-tuning of an immune response. A better understanding of time-kinetic DC features is required for optimally exploiting the therapeutic capacity of DCs in cancer immune therapy.     

Antibody Repertoire in Paraneoplastic Cerebellar Degeneration and Small Cell Lung Cancer

The goal of this study is to determine whether patients with paraneoplastic cerebellar degeneration (PCD) and small-cell lung cancer (SCLC) have a specific repertoire of antibodies, if SOX1 antibodies (SOX1-ab) can predict the presence of SCLC, and if antibodies to cell surface antigens occur in this syndrome. Antibody analysis was done using immunohistochemistry on rat brain, immunoblot with recombinant antigens, screening of cDNA expression libraries, and immunolabeling of live neurons in 39 patients with PCD and SCLC. VGCC-ab were measured by RIA, and SOX1-ab, Hu-ab, and ZIC4-ab by immunoblot. Lambert-Eaton myastenic syndrome (LEMS) was present in 10 of 23 patients with electrophysiological studies. At least one antibody was detected in 72% of patients. The individual frequencies were: 49% SOX1-ab, 44% VGCC-ab, 31% Hu-ab, and 13% ZIC4-ab. SOX1-ab occurred in 76% of patients with VGCC-ab and 27% of those without VGCC-ab (p = 0.0036). SOX1-ab were not found in 39 patients with sporadic late-onset cerebellar ataxia, 23 with cerebellar ataxia and glutamic acid decarboxylase antibodies, and 73 with PCD and cancer types other than SCLC (31 without onconeural antibodies, 25 with Yo-ab , 17 with Tr-ab). Five patients (13%) had antibodies against unknown neuronal cell surface antigens but none of them improved with immunotherapy. One serum immunoreacted against the axon initial segment of neurons and another serum against ELKS1, a protein highly expressed in the cerebellum that interacts with the beta4-subunit of the VGCC. In conclusion, 72% of patients with PCD and SCLC had one or more antibodies that indicate the presence of this tumor. In these patients, VGCC-ab and SOX1-ab occur tightly associated. SOX1-ab are predictors of SCLC in ataxia patients with a specificity of 100% and sensitivity of 49%. Unlike limbic encephalitis with SCLC, antibodies to cell surface antigens other than VGCC-ab, are infrequent and do not predict response to treatment.     

Hyperglucagonemia precedes a decline in insulin secretion and causes hyperglycemia in chronically glucose-infused rats

Islet damage from glucose toxicity is implicated in the pathogenesis of type 2 diabetes, but the sequence of events leading to islet cell dysfunction and hyperglycemia remains unclear. To examine the early stages of islet pathology resulting from increased basal glucose loads, normal awake rats were infused with glucose continuously for 10 days. Plasma glucose and markers of islet and liver function were monitored throughout the infusion. After initial hyperglycemia, rats adapted to the infusion and maintained euglycemia for approximately 4 days. Continued infusion led to worsening hyperglycemia in just 5% of rats after 6 days, but 69% after 8 days and 89% after 10 days, despite unchanged basal and stimulated plasma insulin and C-peptide concentrations. In contrast, plasma glucagon concentrations increased fivefold. Endogenous glucose production (EGP) was appropriately suppressed after 4 days (2.8 ± 0.7 vs. 6.1 ± 0.4 mg·kg(-1)·min(-1) on day 0, P < 0.001) but tripled between days 4 and 8 (9.9 ± 1.7 mg·kg(-1)·min(-1), P < 0.01). Surprisingly, the increase in EGP was accompanied by increased mitochondrial phosphoenolpyruvate carboxykinase expression with appropriate suppression of the cytosolic isoform. Infusion of anti-glucagon antibodies normalized plasma glucose to levels identical to those on day 4 and ～300 mg/dl lower than controls. This improved glycemia was associated with a 60% reduction in EGP. These data support the novel concept that glucose toxicity may first manifest as α-cell dysfunction prior to any measurable deficit in insulin secretion. Such hyperglucagonemia could lead to excessive glucose production overwhelming the capacity of the β-cell to maintain glucose homeostasis.     

A virus-like particle-based Epstein-Barr virus vaccine

Epstein-Barr Virus (EBV) is an ubiquitous human herpesvirus which can lead to infectious mononucleosis and different cancers. In immunocompromised individuals, this virus is a major cause for morbidity and mortality. Transplant patients who did not encounter EBV prior to immunosuppression frequently develop EBV-associated malignancies, but a prophylactic EBV vaccination might reduce this risk considerably. Virus-like particles (VLPs) mimic the structure of the parental virus but lack the viral genome. Therefore, VLPs are considered safe and efficient vaccine candidates. We engineered a dedicated producer cell line for EBV-derived VLPs. This cell line contains a genetically modified EBV genome which is devoid of all potential viral oncogenes but provides viral proteins essential for the assembly and release of VLPs via the endosomal sorting complex required for transport (ESCRT). Human B cells readily take up EBV-based VLPs and present viral epitopes in association with HLA molecules to T cells. Consequently, EBV-based VLPs are highly immunogenic and elicit humoral and strong CD8+ and CD4+ T cell responses in vitro and in a preclinical murine model in vivo. Our findings suggest that VLP formulations might be attractive candidates to develop a safe and effective polyvalent vaccine against EBV.     

CD56brightCD16- killer Ig-like receptor- NK cells display longer telomeres and acquire features of CD56dim NK cells upon activation

Human NK cells can be divided into CD56(dim)CD16(+) killer Ig-like receptors (KIR)(+/-) and CD56(bright)CD16(-) KIR(-) subsets that have been characterized extensively regarding their different functions, phenotype, and tissue localization. Nonetheless, the developmental relationship between these two NK cell subsets remains controversial. We report that, upon cytokine activation, peripheral blood (PB)-CD56(bright) NK cells mainly gain the signature of CD56(dim) NK cells. Remarkably, KIR can be induced not only on CD56(bright), but also on CD56(dim) KIR(-) NK cells, and their expression correlates with lower proliferative response. In addition, we demonstrate for the first time that PB-CD56(dim) display shorter telomeres than PB- and lymph node (LN)-derived CD56(bright) NK cells. Along this line, although human NK cells collected from nonreactive LN display almost no KIR and CD16 expression, NK cells derived from highly reactive LN, efferent lymph, and PB express significant amounts of KIR and CD16, implying that CD56(bright) NK cells could acquire these molecules in the LN during inflammation and then circulate through the efferent lymph into PB as KIR(+)CD16(+) NK cells. Altogether, our results suggest that CD56(bright)CD16(-) KIR(-) and CD56(dim)CD16(+)KIR(+/-) NK cells correspond to sequential steps of differentiation and support the hypothesis that secondary lymphoid organs can be sites of NK cell final maturation and self-tolerance acquisition during immune reaction.     

Fatal neurological disease in scrapie-infected mice induced for experimental autoimmune encephalomyelitis

During the years or decades of prion disease incubation, at-risk individuals are certain to encounter diverse pathological insults, such as viral and bacterial infections, autoimmune diseases, or inflammatory processes. Whether prion disease incubation time and clinical signs or otherwise the pathology of intercurrent diseases can be affected by the coinfection process is unknown. To investigate this possibility, mice infected with the scrapie agent at both high and low titers were subsequently induced for experimental autoimmune encephalomyelitis, an immune system-mediated model of central nervous system (CNS) inflammation. We show here that co-induced mice died from a progressive neurological disease long before control mice succumbed to classical scrapie. To investigate the mechanism of the co-induced syndrome, we evaluated biochemical and pathological markers of both diseases. Brain and spleen PrP(Sc) levels in the dying co-induced mice were comparable to those observed in asymptomatic scrapie-infected animals, suggesting that co-induced disease is not an accelerated form of scrapie. In contrast, inflammatory markers, such as demyelination, immune cell infiltrates, and gliosis, were markedly increased in co-induced mouse spinal cords. Activated astrocytes were especially elevated in the medulla oblongata. Furthermore, PrP(sc) depositions were found in demyelinated white matter areas in co-induced mouse spinal cords, suggesting the presence of activated infected immune cells that infiltrate into the CNS to facilitate the process of prion neuroinvasion. We hypothesize that inflammatory processes affecting the CNS may have severe clinical implications in subjects incubating prion diseases.