krn-7000 and Bronchial-Hyperreactivity

Despite their increasing use in autoimmune, inflammatory, and allergic conditions, the mechanism of action of i.v. Igs (IVIg) is poorly understood. On the basis of the critical role of invariant NKT (iNKT) cells in allergic airway inflammation (AAI) and their constitutive expression of the low-affinity IgG receptor FcγRIIIA, we surmised that IVIg targets iNKT cells to exert their anti-inflammatory effect. We found that IVIg treatment significantly inhibited AAI in OVA-sensitized C57BL/6 mice and downregulated α-galactosylceramide-induced iNKT cell activation and cytokine production. Allergic responses were restored in iNKT cell-deficient mice by transferring iNKT cells from PBS- but not from IVIg-treated mice, suggesting that IVIg acts directly on activated iNKT cells that have a critical role in AAI. The inhibitory effects of IVIg on both iNKT cell activation/function and OVA-driven AAI were lost in FcγRIIIA(-/-) mice. Our data unravel an FcγRIIIA-dependent inhibitory effect of IVIg on activated iNKT cells that confers protection in AAI.

Topics: Adoptive Transfer; Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Bronchial Hyperreactivity; Cytokines; Galactosylceramides; Immunoglobulins, Intravenous; Inflammation Mediators; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Natural Killer T-Cells; Ovalbumin; Receptors, IgG; Respiratory Hypersensitivity; Spleen

Vitamin D receptor (VDR) deficiency (knockout [KO]) results in a failure of mice to generate an airway hyperreactivity (AHR) response on both the BALB/c and C57BL/6 background. The cause of the failed AHR response is the defective population of invariant NKT (iNKT) cells in the VDR KO mice because wild-type (WT) iNKT cells rescued the AHR response. VDR KO mice had significantly fewer iNKT cells and normal numbers of T cells in the spleen compared with WT mice. In BALB/c VDR KO mice, the reduced frequencies of iNKT cells were not apparent in the liver or thymus. VDR KO and WT Th2 cells produced similar levels of IFN-γ and IL-5. On the BALB/c background, Th2 cells from VDR KO mice produced less IL-13, whereas on the C57BL/6 background, Th2 cells from VDR KO mice produced less IL-4. Conversely, VDR KO iNKT cells were defective for the production of multiple cytokines (BALB/c: IL-4, IL-5, and IL-13; C57BL/6: IL-4 and IL-17). Despite relatively normal Th2 responses, BALB/c and C57BL/6 VDR KO mice failed to develop AHR responses. The defect in iNKT cells as a result of the VDR KO was more important than the highly susceptible Th2 background of the BALB/c mice. Defective iNKT cell responses in the absence of the VDR result in the failure to generate AHR responses in the lung. The implication of these mechanistic findings for human asthma requires further investigation.

Topics: Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cells, Cultured; Galactosylceramides; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Natural Killer T-Cells; Pneumonia; Receptors, Calcitriol; Species Specificity; Th2 Cells

The prevalence of asthma continues to increase in westernized countries, and optimal treatment remains a significant therapeutic challenge. Recently, CD1d-restricted invariant NKT (iNKT) cells were found to play a critical role in the induction of airway hyperreactivity (AHR) in animal models and are associated with asthma in humans. To test whether iNKT cell-targeted therapy could be used to treat allergen-induced airway disease, mice were sensitized with OVA and treated with di-palmitoyl-phosphatidyl-ethanolamine polyethylene glycol (DPPE-PEG), a CD1d-binding lipid antagonist. A single dose of DPPE-PEG prevented the development of AHR and pulmonary infiltration of lymphocytes upon OVA challenge, but had no effect on the development of OVA-specific Th2 responses. In addition, DPPE-PEG completely prevented the development of AHR after administration of alpha-galactosylceramide (alpha-GalCer) intranasally. Furthermore, we demonstrate that DPPE-PEG acts as antagonist to alpha-GalCer and competes with alpha-GalCer for binding to CD1d. Finally, we show that DPPE-PEG completely inhibits the alpha-GalCer-induced phosphorylation of ERK tyrosine kinase in iNKT cells, suggesting that DPPE-PEG specifically blocks TCR signaling and thus activation of iNKT cells. Because iNKT cells play a critical role in the development of AHR, the inhibition of iNKT activation by DPPE-PEG suggests a novel approach to treat iNKT cell-mediated diseases such as asthma.

Topics: Allergens; Animals; Antigens, CD1d; Binding, Competitive; Bronchial Hyperreactivity; Cell Line; Disease Models, Animal; Female; Galactosylceramides; Humans; Immunosuppressive Agents; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Natural Killer T-Cells; Ovalbumin; Phosphatidylethanolamines; Polyethylene Glycols

Invariant NKT cells (iNKT cells) play a pivotal role in the development of allergen-induced airway hyperresponsiveness (AHR) and inflammation. However, it is unclear what role they play in the initiation (sensitization) phase as opposed to the effector (challenge) phase. The role of iNKT cells during sensitization was examined by determining the response of mice to intratracheal transfer of OVA-pulsed or OVA-alpha-galactosylceramide (OVA/alphaGalCer)-pulsed bone marrow-derived dendritic cells (BMDCs) prior to allergen challenge. Wild-type (WT) recipients of OVA-BMDCs developed AHR, increased airway eosinophilia, and increased levels of Th2 cytokines in bronchoalveolar lavage fluid, whereas recipients of OVA/alphaGalCer BMDCs failed to do so. In contrast, transfer of these same OVA/alphaGalCer BMDCs into IFN-gamma-deficient (IFN-gamma(-/-)) mice enhanced the development of these lung allergic responses, which was reversed by exogenous IFN-gamma treatment following OVA-BMDC transfer. Further, Jalpha18-deficient recipients, which lack iNKT cells, developed the full spectrum of lung allergic responses following reconstitution with highly purified WT liver or spleen iNKT cells and transfer of OVA-BMDCs, whereas reconstituted recipients of OVA/alphaGalCer BMDCs failed to do so. Transfer of iNKT cells from IFN-gamma(-/-) mice restored the development of these responses in Jalpha18-deficient recipients following OVA-BMDC transfer; the responses were enhanced following OVA/alphaGalCer BMDC transfer. iNKT cells from these IFN-gamma(-/-) mice produced higher levels of IL-13 in vitro compared with WT iNKT cells. These data identify IFN-gamma as playing a critical role in dictating the consequences of iNKT cell activation in the initiation phase of the development of AHR and airway inflammation.

Topics: Adoptive Transfer; Allergens; Animals; Bone Marrow Cells; Bronchial Hyperreactivity; Cells, Cultured; Coculture Techniques; Dendritic Cells; Female; Galactosylceramides; Inflammation Mediators; Interferon-gamma; Intubation, Intratracheal; Ligands; Liver; Mice; Mice, Inbred C57BL; Mice, Knockout; Natural Killer T-Cells; Ovalbumin; Respiratory Hypersensitivity

T-bet(-/-) mice have been shown to have a profound deficiency in the ability to generate invariant NKT (iNKT) cells in the periphery due to a halt in terminal maturation, but despite this deficiency, T-bet(-/-) mice develop spontaneous airway hyperreactivity (AHR) and airway inflammation. Because in some situations the development of AHR requires the presence of iNKT cells, we sought to more clearly understand how AHR develops in T-bet(-/-) mice by examining T-bet(-/-) mice in several distinct mouse models of asthma, including spontaneous, OVA-induced and alpha-galactosylceramide (alpha-GalCer)-induced AHR. Surprisingly, we found that administration of alpha-GalCer, which very specifically activates iNKT cells, greatly increased the AHR response in the T-bet(-/-) mice. Moreover, in T-bet(-/-) mice, spontaneous AHR as well as AHR induced with OVA or alpha-GalCer were all eliminated by blocking CD1d, the restricting element of iNKT cells, using an anti-CD1d-blocking mAb. Although the number of the iNKT cells in T-bet(-/-) mice was reduced compared with that in wild-type mice, the remaining iNKT cells produced primarily IL-4 and IL-13, and only minimal amounts of IFN-gamma. We conclude therefore that the AHR that develops in T-bet(-/-) mice is dependent on the presence of iNKT cells, and that whereas T-bet(-/-) have reduced numbers of iNKT cells, these are sufficient for the development of AHR.

Topics: Animals; Antibodies, Blocking; Antigens, CD1d; Bronchial Hyperreactivity; Disease Models, Animal; Epitopes, T-Lymphocyte; Galactosylceramides; Lymphopenia; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Natural Killer T-Cells; Ovalbumin; T-Box Domain Proteins

Activated natural killer T (NKT) cells produce a broad range of cytokines, including IL-4 and IFN-gamma, that determine immunomodulatory functions in various animal models. In this report, we show that a well-known proinflammatory cytokine, IL-17 is also produced by a distinct population of NKT cells upon TCR stimulation. Administration of alpha-galactosylceramide (alpha-GalCer), a strong agonist of NKT cells, induces rapid IL-17 production by a small population of NKT cells, mostly belonging to a population different from that of IL-4- and IFN-gamma-producing NKT cells. IL-17-producing NKT cells showed unresponsiveness after stimulation of alpha-GalCer as conventional NKT cells. During airway inflammation induced by pulmonary activation of NKT cells with alpha-GalCer, IL-17 contributes to the infiltration of neutrophils into the airway but has no effect on airway hyperreactivity (AHR). These results indicate that TCR stimulation induces IL-17 expression by a novel population of NKT cells and may help to explain diverse NKT cell functions.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Female; Flow Cytometry; Galactosylceramides; Interleukin-17; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Neutrophil Infiltration; Neutrophils; Receptors, Antigen, T-Cell; Reverse Transcriptase Polymerase Chain Reaction; RNA; T-Lymphocyte Subsets

Allergic asthma is a multifaceted syndrome consisting of eosinophil-rich airway inflammation, bronchospasm, and airway hyper-responsiveness (AHR). Using a mouse model of allergic asthma, we previously reported that invariant NKT (iNKT) cells increase the severity of this disease. Herein, we demonstrate that a single i.v. injection of alpha-galactosylceramide (alpha-GalCer), 1 h before the first airway allergen challenge of OVA-sensitized mice, abrogates elicitation of AHR, airway eosinophilia, IL-4 and IL-5 production in bronchoalveolar lavage fluid, and specific anti-OVA IgE antibodies. Further, alpha-GalCer administered intranasally also strongly inhibited the major symptoms of asthma in sensitized and challenged mice. Alpha-GalCer treatment induces iNKT cell accumulation in the lungs, and shifts their cytokine profile from pro-asthmatic IL-4 to a protective IFN-gamma production. The role of IFN-gamma from iNKT cells in protection was shown by adoptive transfer of sorted iNKT cells from OVA-sensitized and alpha-GalCer-treated mice which protected immunized recipients from manifesting asthma by an IFN-gamma-dependent pathway. Our findings demonstrate for the first time that alpha-GalCer administered locally inhibits asthma symptoms, even in predisposed asthmatic mice, through an iNKT cell- and IFN-gamma-dependent pathway.

Topics: Administration, Intranasal; Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Flow Cytometry; Galactosylceramides; Hypersensitivity; Injections, Intravenous; Interferon-alpha; Killer Cells, Natural; Male; Mice; Ovalbumin