BackgroundAstrocytes, which play an active role in chronic inflammatory diseases like multiple sclerosis, exist close to mast cells with which they share perivascular localization. We previously demonstrated the possibility that astrocytes and mast cells interact in vitro and in vivo. This study aimed to investigate the signaling pathways and the role for astrocytes in the interaction of astrocytes and mast cells.MethodsWe co-cultured human U87 glioblastoma (U87) and human mast cell-1 (HMC-1) cell lines, and mouse cerebral cortices-derived astrocytes and mouse bone marrow-derived mast cells (BMMCs). Intracellular Ca2+ ([Ca2+]i) was measured by confocal microscopy; CD40 siRNA by Silencer Express Kit; small GTPases by GTP-pull down assay; PKCs, MAPKs, CD40, CD40L, Jak1/2, STAT1, TNF receptor 1 (TNFR1) by Western blot; NF-κB and AP-1 by EMSA; cytokines by RT-PCR. An experimental allergic encephalomyelitis (EAE) model was induced using myelin oligodendrocyte glycoprotein (MOG) peptide and pertussis toxin in mice. Co-localization of TNFR1 and astrocytes in EAE brain tissues was determined by immunohistochemistry.ResultsEach astrocyte co-culture had increases in [Ca2+]i levels, release of cytokines and chemokines; activities of Rho-family GTPases, NF-κB/AP-1/STAT1727, and Jack1/2, STAT1701. These effects were inhibited by anti-CD40 antibody or CD40 siRNA, and signaling pathways for Jak1/2 were inhibited by anti-TNFR1 antibody. EAE score, expression of TNFR1, and co-localization of TNFR1 and astrocytes were enhanced in brain of the EAE model. Anti-CD40 antibody or 8-oxo-dG pretreatment reduced these effects in EAE model.ConclusionsThese data suggest that astrocytes activated by the CD40-CD40L interaction in co-culture induce inflammatory cytokine production via small GTPases, and the secreted cytokines re-activate astrocytes via Jak/STAT1701 pathways, and then release more cytokines that contribute to exacerbating the development of EAE. These findings imply that the pro-inflammatory mediators produced by cell-to-cell cross-talk via interaction of CD40-CD40L may be as a promising therapeutic target for neurodegenerative diseases like MS.

BACKGROUND AND PURPOSETransglutaminase 2 (TGase 2) expression is increased in inflammatory diseases, and TGase 2 inhibitors block these increases. We examined whether the R2 peptide inhibited the expression of TGase 2 in a mouse model of inflammatory allergic asthma. EXPERIMENTAL APPROACHC57BL/6 mice were sensitized and challenged by ovalbumin (OVA) to induce asthma. OVA-specific serum IgE and leukotrienes (LTs) levels were measured by enzyme-linked immunosorbent assay. Recruitment of inflammatory cells into bronchoalveolar lavage (BAL) fluid or lung tissues and goblet cell hyperplasia were assessed histologically. Airway hyperresponsiveness was determined in a barometric plethysmographic chamber. Expression of TGase 2, eosinophil major basic protein (EMBP), the adhesion molecule vascular cell adhesion molecule-1, Muc5ac and phospholipase A2 (PLA2) protein were determined by Western blot. Expression of mRNAs of Muc5ac, cytokines, matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) were measured by reverse transcriptase-polymerase chain reaction and nuclear factor-kB (NF-kB) by electrophoretic mobility shift assay. KEY RESULTSR2 peptide reduced OVA-specific IgE levels; the number of total inflammatory cells, macrophages, neutrophils, lymphocytes and eosinophils in BAL fluid and the number of goblet cells. Airway hyperresponsiveness, TGase 2 and EMBP levels, mRNA levels of interleukin (IL)-4, IL-5, IL-6, IL-8, IL-13, RANTES, tumour necrosis factor-a, and MMP2/9, Muc5ac, NF-kB activity, PLA2 activity and expressions, and LT levels in BAL cells and lung tissues were all reduced by R2 peptide. R2 peptide also restored expression of TIMP1/2. CONCLUSION AND IMPLICATIONSR2 peptide reduced allergic responses by regulating NF-kB/TGase 2 activity in a mouse model of allergic asthma. This peptide may be useful in the treatment of allergic asthma. AbbreviationsBAL, bronchoalveolar lavage; ELISA, enzyme-linked immunosorbent assay; EMBP, eosinophil major basic protein; EMSA, electrophoretic mobility shift assay; MMP, matrix metalloproteinase; NF-kB, nuclear factor-kappa B; OVA, ovalbumin; PAS, periodic acid-Schiff; PBS, phosphate buffered saline; PLA2, phospholipase A2; RT-PCR, reverse transcriptase-polymerase chain reaction; TGase 2, transglutaminase 2; TIMP, tissue inhibitor of matrix metalloproteinase BJP British Journal of Pharmacology

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