Extracellular RNA facilitates hypoxia‐induced leukocyte adhesion and infiltration in the lung through TLR3‐IFN‐γ‐STAT1 signaling pathway

Endogenous ligands released from dying cells, including extracellular RNA (eRNA), cause TLR activation, which is associated with inflammation and vascular diseases. However, the importance of this response in acute hypoxia (AH) remains unexplored. Here, we observed eRNA‐mediated TLR3 activation during exposure of mice to AH in the absence of exogenous viral stimuli. RNaseA treatment diminished AH‐induced expression of IFN and cell adhesion molecules (CAMs) and myeloid cell infiltration in the lung, and TLR3 gene silencing or neutralization with antibodies markedly attenuated AH‐ or poly I:C‐induced IFN and CAM expression and leukocyte adhesion (LA) and myeloid cell infiltration in the lung. However, RNaseA treatment or TLR3 gene silencing failed to alter AH‐induced cell death and proliferation in lung vasculature. Furthermore, IFN‐γ—but not IFN‐α—regulated AH‐induced CAM expression and LA. Treatment with RNaseA, TLR3 siRNA, neutralizing antibodies, or a STAT1 inhibitor substantially decreased AH‐ and poly I:C‐induced STAT1 phosphorylation, CAM expression, and myeloid cell infiltration, suggesting a central role for STAT1 phosphorylation in AH‐induced LA and infiltration. We conclude that eRNA activates TLR3 and facilitates, through in vivo IFN‐γ?STAT1 signaling, AH‐induced leukocyte infiltration in the lung. Thus, RNaseA might provide a therapeutic alternative for patients with lung diseases.     

ORMDL3/GSDMB genotype as a risk factor for early-onset adult asthma is linked to total serum IgE levels but not to allergic sensitization

BackgroundAn orosomucoid-like 3 (ORMDL3)/gasdermin B (GSDMB) gene locus on chromosome 17q is consistently associated with childhood-onset asthma, which is highly atopic. As some evidence suggests the relationship between asthma and allergic sensitization reflects asthma patient susceptibility to augmented IgE responses driven by common environmental allergens rather than an increased asthma risk after allergen exposure, we aimed to determine any relationships between this locus region and childhood-onset adult asthma with regard to serum total IgE levels or allergic sensitization.MethodsWe conducted a case–control association study using three independent Japanese populations (3869 total adults) and analyzed the ORs for association of rs7216389, an expression quantitative trait locus for ORMDL3/GSDMB, with adult asthma according to onset age. Additionally, associations between the rs7216389 genotype and total serum IgE levels or allergic sensitization was examined.ResultsRs7216389 was associated with both childhood-onset adult asthma (OR for asthmatic patients afflicted at the age of 10 years or younger = 1.61, p = 0.00021) and asthmatic patients with higher levels of total serum IgE (OR for asthmatic patients with IgE ≥1000IU/mL = 1.55, p = 0.0033). In both healthy controls and in the combined healthy and asthmatic individuals, rs7216389 was correlated with increased total serum IgE levels (p < 0.0005), but not allergic sensitization (p > 0.1).ConclusionsORMDL3/GSDMB is an important susceptibility gene for childhood-onset adult asthma in Japanese populations and this association is linked to elevated total serum IgE levels but not to allergic sensitization.     

Characterization of binding properties of ICAM-1 peptides to LFA-1: inhibitors of T-cell adhesion

In the present study, we characterized the binding site of two intercellular adhesion molecule-1-derived cyclic peptides, cIBC and cIBR, to the LFA-1 on the surface of T cells. These peptides had been able to inhibit LFA-1/intercellular adhesion molecule-1 signal by blocking the signal-2 of immune synapse. Both peptides prefer to bind to the closed form of LFA-1 I-domain, indicating that two peptides act as allosteric inhibitors against intercellular adhesion molecule-1. Binding site mapping using monoclonal antibodies proposes that cIBC binds to around residues 266-272 of LFA-1 I-domain where this site is adjacent to the metal ion-dependent adhesion site. On the other hand, cIBR binds to the pocket called L-site where is distant from metal ion-dependent adhesion site. Cross-inhibition mapping between two peptides show that cIBR could inhibit the binding of cIBC but not vice versa, suggesting that cIBR has some properties that allow this peptide bind to more than one site. Structural comparison between cIBC and cIBR reveals that cIBR is more flexible than cIBC, allowing this peptide bind to exposed region, such as cIBC-binding site as well as cramped pocket like L-site. Our findings are important for understanding the selectivity of cIBC and cIBR peptides; thus, they can be conjugated with drugs and transported specifically to the target.     

Transactivation of the ICAM-1 gene by CD30 in Hodgkin's lymphoma

The ICAM-1/LFA-1 complex mediates cell-cell interaction. ICAM-1 is overexpressed in Hodgkin/Reed-Sternberg (H/RS) cells, and serum levels of its soluble form are higher in Hodgkin's lymphoma (HL) patients than in controls. There are no data, however, regarding the regulation of expression of ICAM-1 in H/RS cells. CD30 was identified in H/RS cells of HL and has attracted much interest as a molecular marker of HL. To analyze ICAM-1 expression in H/RS cells, we examined the expression of ICAM-1, LFA-1, CD30 and CD30L in HL-derived cell lines. All cell lines expressed ICAM-1 and CD30, but not LFA-1 or CD30L. CD30 induced ICAM-1 expression. Analysis of the ICAM-1 promoter showed the importance of NF-kappaB binding site for CD30-induced ICAM-1 gene expression. Coexpression of IkappaB, IKK, NIK and TRAF dominant-negative constructs with CD30 inhibited CD30-induced activation of ICAM-1 promoter, suggesting that CD30 induces ICAM-1 via NF-kappaB signalling. The ICAM-1 promoter was activated by the C-terminal region of CD30, which activated NF-kappaB signalling. A decoy CD30 lacking the cytoplasmic region inhibited ICAM-1 promoter activity in HL cell lines. Thus, in H/RS cells, ligand-independent activation of CD30 signalling activates NF-kappaB and this leads to constitutive ICAM-1 expression, suggesting a link between 2 well known phenotypic characteristics of HL, CD30 and ICAM-1 overexpression.     

Blood-derived macrophages infiltrate the retina and activate Muller glial cells under experimental choroidal neovascularization

Inflammation is a major mechanism in the pathogenesis of age-related macular degeneration, the most important cause of blindness in the elderly. Previous studies have focused on the role of macrophages in regulating the growth of pathological new vessels over the retina, called choroidal neovascularization (CNV). However, no research has been done to evaluate the role of inflammation as a mechanism of vision loss and retinal degeneration in the retina underlying CNV. In other neuropathological conditions, hematogenous macrophages and/or resident microglia contribute to neurodegeneration. We have combined laser-induced CNV in mice and bone marrow transplantation with GFP-labeled bone marrow to determine the relative role of recruited blood-derived macrophages versus resident microglia in the retina associated with CNV. Using these chimeric mice, we have found that many GFP-labeled cells infiltrated the retina underlying CNV but not the retina unaffected by CNV. Immunostaining for the cell adhesion molecules VCAM 1, ICAM 1, and PECAM was strongly upregulated in retinal blood vessels under CNV. All GFP-labeled cells were immunoreactive for the macrophage marker F4/80. Most (70%) of the F4/80 immunoreactive cells were GFP-labeled under CNV. The density of resident microglia did not increase. Most GFP-labeled cells were found in close proximity to activated Muller cells. Depleting circulating macrophages with clodronic acid diminished the density of F4/80 immunoreactive cells as well as the density of pERK immunoreactive Muller cells in the retina under CNV. Thus, recruitment of blood-derived macrophages more than resident microglia seems to be associated with CNV.     

Soluble adhesion molecules in young children with signs of beta-cell autoimmunity--prospective follow-up from birth

BACKGROUND: This study aimed at evaluating the relationship between the circulating concentrations of soluble intercellular adhesion molecule-1 (sICAM-1) and sL-selectin and the appearance of beta-cell autoimmunity, and at assessing whether these molecules could assist in the identification of environmental factors implicated in the immune process damaging the pancreatic beta-cells. METHODS: Serum levels of soluble adhesion molecules were measured with enzyme-linked immunosorbent assays over the first 2 years of life in 65 children seroconverting to positivity for autoantibodies and 65 control children, all with HLA-conferred susceptibility to type 1 diabetes (T1D). RESULTS: The total integrated concentrations of soluble adhesion molecules were comparable between the two groups. The autoantibody-positive children tended to have higher sL-selectin concentrations during the 3-month seroconversion (SC) period than did the control children during the corresponding period (P = 0.07), the difference being significant (P = 0.03) after excluding subjects with signs of a concurrent enterovirus infection. Autoantibody-positive children had higher concentrations of sL-selectin in the 3-month period when an enterovirus infection was detectable than did the control children (P = 0.018). No significant difference could, however, be seen after excluding the children with concomitant seroconversion to autoantibody positivity. CONCLUSIONS: Elevated concentrations of sL-selectin are temporally associated with seroconversion to autoantibody positivity suggesting that leukocyte activation might coincide with the appearance of beta-cell autoimmunity. Early-onset progressive beta-cell autoimmunity, on the other hand, is not reflected in overall increased concentrations of soluble adhesion molecules in the peripheral circulation during the first 2 years of life in children carrying increased HLA-conferred disease susceptibility. Enterovirus infections (EVIs) are not independently associated with increased circulating sL-selectin concentrations in young children with enhanced HLA-conferred susceptibility to T1D.