Type I interferons drive MAIT cell functions against bacterial pneumonia.

Mucosal-associated invariant T (MAIT) cells are abundant in the lung and contribute to host defense against infections. During bacterial infections, MAIT cell activation has been proposed to require T cell receptor (TCR)-mediated recognition of antigens derived from the riboflavin synthesis pathway presented by the antigen-presenting molecule MR1. MAIT cells can also be activated by cytokines in an MR1-independent manner, yet the contribution of MR1-dependent vs. -independent signals to MAIT cell functions in vivo remains unclear. Here, we use Klebsiella pneumoniae as a model of bacterial pneumonia and demonstrate that MAIT cell activation is independent of MR1 and primarily driven by type I interferons (IFNs). During Klebsiella infection, type I IFNs stimulate activation of murine and human MAIT cells, induce a Th1/cytotoxic transcriptional program, and modulate MAIT cell location within the lungs. Consequently, adoptive transfer or boosting of pulmonary MAIT cells protect mice from Klebsiella infection, with protection being dependent on direct type I IFN signaling on MAIT cells. These findings reveal type I IFNs as new molecular targets to manipulate MAIT cell functions during bacterial infections.

Beyond allergic progression: From molecules to microbes as barrier modulators in the gut-lung axis functionality.

The "epithelial barrier hypothesis" states that a barrier dysfunction can result in allergy development due to tolerance breakdown. This barrier alteration may come from the direct contact of epithelial and immune cells with the allergens, and indirectly, through deleterious effects caused by environmental changes triggered by industrialization, pollution, and changes in the lifestyle. Apart from their protective role, epithelial cells can respond to external factors secreting IL-25 IL-33, and TSLP, provoking the activation of ILC2 cells and a Th2-biased response. Several environmental agents that influence epithelial barrier function, such as allergenic proteases, food additives or certain xenobiotics are reviewed in this paper. In addition, dietary factors that influence the allergenic response in a positive or negative way will be also described here. Finally, we discuss how the gut microbiota, its composition, and microbe-derived metabolites, such as short-chain fatty acids, alter not only the gut but also the integrity of distant epithelial barriers, focusing this review on the gut-lung axis.

CD1d-dependent rewiring of lipid metabolism in macrophages regulates innate immune responses.

Alterations in cellular metabolism underpin macrophage activation, yet little is known regarding how key immunological molecules regulate metabolic programs in macrophages. Here we uncover a function for the antigen presenting molecule CD1d in the control of lipid metabolism. We show that CD1d-deficient macrophages exhibit a metabolic reprogramming, with a downregulation of lipid metabolic pathways and an increase in exogenous lipid import. This metabolic rewiring primes macrophages for enhanced responses to innate signals, as CD1d-KO cells show higher signalling and cytokine secretion upon Toll-like receptor stimulation. Mechanistically, CD1d modulates lipid import by controlling the internalization of the lipid transporter CD36, while blocking lipid uptake through CD36 restores metabolic and immune responses in macrophages. Thus, our data reveal CD1d as a key regulator of an inflammatory-metabolic circuit in macrophages, independent of its function in the control of T cell responses.

Comparative metabolomics analysis of bronchial epithelium during barrier establishment after allergen exposure.

BACKGROUND: Several studies have shown a correlation between an altered metabolome and respiratory allergies. The epithelial barrier hypothesis proposes that an epithelial barrier dysfunction can result in allergic diseases development. Der p 1 allergen from house dust mite is a renowned epithelial barrier disruptor and allergy initiator due to its cysteine-protease activity. Here, we compared the metabolic profile of the bronchial epithelium exposed or not to Der p 1 during barrier establishment to understand its active role in allergy development. METHODS: Calu-3 cells were cultivated in air-liquid interface cultures and exposed to either Der p 1 or Ole e 1 allergens during barrier establishment. The comparative metabolomics analysis of apical and basolateral media were performed using liquid chromatography and capillary electrophoresis both coupled to mass spectrometry. RESULTS: We showed that epithelial barrier disruption by Der p 1 was associated with a specific metabolic profile, which was highly dependent on the state of the epithelium at the time of contact. Moreover, an apical-basolateral distribution of the metabolites was also observed, indicating a compartmentalization of the response with differential metabolic patterns. A number of metabolites were changed by Der p 1, mainly related to amino acids metabolism, such as L-arginine, L-kynurenine and L-methionine. CONCLUSION: This work is the first report on the metabolic response in human bronchial epithelial cells associated with cysteine-protease Der p 1 activity, which could contribute to allergy development. Moreover, it supports a reformulated epithelial barrier hypothesis that might help to explain allergies and their increasing prevalence.

Der p 1-based immunotoxin as potential tool for the treatment of dust mite respiratory allergy.

Immunotoxins appear as promising therapeutic molecules, alternative to allergen-specific-immunotherapy. In this work, we achieved the development of a protein chimera able to promote specific cell death on effector cells involved in the allergic reaction. Der p 1 allergen was chosen as cell-targeting domain and the powerful ribotoxin α-sarcin as the toxic moiety. The resultant construction, named proDerp1αS, was produced and purified from the yeast Pichia pastoris. Der p 1-protease activity and α-sarcin ribonucleolytic action were effectively conserved in proDerp1αS. Immunotoxin impact was assayed by using effector cells sensitized with house dust mite-allergic sera. Cell degranulation and death, triggered by proDerp1αS, was exclusively observed on Der p 1 sera sensitized-humRBL-2H3 cells, but not when treated with non-allergic sera. Most notably, equivalent IgE-binding and degranulation were observed with both proDerp1αS construct and native Der p 1 when using purified basophils from sensitized patients. However, proDerp1αS did not cause any cytotoxic effect on these cells, apparently due to its lack of internalization after their surface IgE-binding, showing the complex in vivo panorama governing allergic reactions. In conclusion, herein we present proDerp1αS as a proof of concept for a potential and alternative new designs of therapeutic tools for allergies. Development of new, and more specific, second-generation of immunotoxins following proDerp1αS, is further discussed.

Biophysical and biological impact on the structure and IgE-binding of the interaction of the olive pollen allergen Ole e 7 with lipids.

Ole e 7 allergen from Olea europaea pollen possesses a major clinical relevance because it produces severe symptoms, such as anaphylaxis, in allergic patients exposed to high olive pollen counts. Ole e 7 is a non-specific lipid transfer protein (nsLTP) characterized by the presence of a tunnel-like hydrophobic cavity, which may be suitable for hosting and, thus, transporting lipids -as it has been described for other nsLTPs-. The identification of the primary amino acid sequence of Ole e 7, and its production as a recombinant allergen, allowed characterizing its lipid-binding properties and its effect at air-liquid interfaces. Fluorescence and interferometry experiments were performed using different phospholipid molecular species and free fatty acids to analyse the lipid-binding ability and specificity of the allergen. Molecular modelling of the allergen was used to determine the potential regions involved in lipid interaction. Changes in Ole e 7 structure after lipid interaction were analysed by circular dichroism. Changes in the IgE binding upon ligand interaction were determined by ELISA. Wilhelmy balance measurements and fluorescence surfactant adsorption tests were performed to analyse the surface activity of the allergen. Using these different approaches, we have demonstrated the ability of Ole e 7 to interact and bind to a wide range of lipids, especially negatively charged phospholipids and oleic acid. We have also identified the protein structural regions and the residues potentially involved in that interaction, suggesting how lipid-protein interactions could define the behaviour of the allergen once inhaled at the airways.

Peptide Glycodendrimers as Potential Vaccines for Olive Pollen Allergy.

Olive pollen is one of the most important causes of respiratory allergy, with Ole e 1 being the most clinically relevant sensitizing allergen. Peptide-based vaccines represent promising therapeutic approaches, but the use of adjuvants is required to strengthen the weak immunogenicity of small peptides. We propose the use of dendrimeric scaffolds conjugated to the T cell immunodominant epitope of Ole e 1 (OE109-130) for the development of novel vaccines against olive pollen allergy. Four dendrimeric scaffolds containing an ester/ether with nine mannoses, an ester succinimidyl linker with nine N-acetyl-glucosamine units or nine ethylene glycol units conjugated to OE109-130 peptide were designed, and their cytotoxicity, internalization pattern, and immunomodulatory properties were analyzed in vitro. None of the dendrimers exhibited cytotoxicity in humanized rat basophil (RBL-2H3), human bronchial epithelial Calu-3, and human mast LAD2 cell lines. Confocal images indicated that mannosylated glycodendropeptides exhibited lower colocalization with a lysosomal marker. Moreover, mannosylated glycodendropeptides showed higher transport tendency through the epithelial barrier formed by Calu-3 cells cultured at the air-liquid interface. Finally, mannosylated glycodendropeptides promoted Treg and IL10+Treg proliferation and IL-10 secretion by peripheral blood mononuclear cells from allergic patients. Mannosylated dendrimers conjugated with OE109-130 peptide from Ole e 1 have been identified as suitable candidates for the development of novel vaccines of olive pollen allergy.

Human glutathione-S-transferase pi potentiates the cysteine-protease activity of the Der p 1 allergen from house dust mite through a cysteine redox mechanism.

Environmental proteases have been widely associated to the pathogenesis of allergic disorders. Der p 1, a cysteine-protease from house dust mite (HDM) Dermatophagoides pteronyssinus, constitutes one of the most clinically relevant indoor aeroallergens worldwide. Der p 1 protease activity depends on the redox status of its catalytic cysteine residue, which has to be in the reduced state to be active. So far, it is unknown whether Der p 1-protease activity could be regulated by host redox microenvironment once it reaches the lung epithelial lining fluid in addition to endogenous mite components. In this sense, Glutathione-S-transferase pi (GSTpi), an enzyme traditionally linked to phase II detoxification, is highly expressed in human lung epithelial cells, which represent the first line of defence against aeroallergens. Moreover, GSTpi is a generalist catalyst of protein S-glutathionylation reactions, and some polymorphic variants of this enzyme has been associated to the development of allergic asthma. Here, we showed that human GSTpi increased the cysteine-protease activity of Der p 1, while GSTmu (the isoenzyme produced by the mite) did not alter it. GSTpi induces the reduction of Cys residues in Der p 1, probably by rearranging its disulphide bridges. Furthermore, GSTpi was detected in the apical medium collected from human bronchial epithelial cell cultures, and more interesting, it increased cysteine-protease activity of Der p 1. Our findings support the role of human GSTpi from airways in modulating of Der p 1 cysteine-protease activity, which may have important clinical implications for immune response to this aeroallergen in genetically susceptible individuals.

Inducción de ovulación: conceptos básicos / Ovulation induction: basic concepts

La concepción sin éxito es uno de los trastornos crónicos más prevalentes que involucra adultos jóvenes. Desde el año 1978, cuando nació la primera bebé producto de técnicas de reproducción asistida, el manejo de la infertilidad femenina ha sido transformada por la Fertilizaciòn In Vitro. Los trastornos de la ovulación representan el 15% de las causas de infertilidad en las parejas y el 40% de infertilidad en las mujeres.