Ryuichi Fujisawa

Intervention of Thymus and Activation-Regulated Chemokine Attenuates the Development of Allergic Airway Inflammation and Hyperresponsiveness in Mice

Thymus- and activation-regulated chemokine (TARC; CCL17) is a lymphocyte-directed CC chemokine that specifically chemoattracts CC chemokine receptor 4-positive (CCR4+) Th2 cells. To establish the pathophysiological roles of TARC in vivo, we investigated here whether an mAb against TARC could inhibit the induction of asthmatic reaction in mice elicited by OVA. TARC was constitutively expressed in the lung and was up-regulated in allergic inflammation. The specific Ab against TARC attenuated OVA-induced airway eosinophilia and diminished the degree of airway hyperresponsiveness with a concomitant decrease in Th2 cytokine levels. Our results for the first time indicate that TARC is a pivotal chemokine for the development of Th2-dominated experimental allergen-induced asthma with eosinophilia and AHR. This study also represents the first success in controlling Th2 cytokine production in vivo by targeting a chemokine.

Human B Cells Immortalized with Epstein-Barr Virus Upregulate CCR6 and CCR10 and Downregulate CXCR4 and CXCR5

ABSTRACT Compared to peripheral blood resting B cells, Epstein-Barr virus (EBV)-immortalized B cells consistently express CCR6 and CCR10 at high levels and CXCR4 and CXCR5 at low levels. Accordingly, these cells vigorously responded to the ligands of CCR6 and CCR10 but not to those of CXCR4 and CXCR5. In a human EBV-negative B-cell line, BJAB, stable expression of EBNA2 upregulated CCR6, while stable expression of EBNA2 as well as LMP1 downregulated CXCR4. On the other hand, upregulation of CCR10 or downregulation of CXCR5 was not induced in BJAB by stable expression of EBNA2 or LMP1. Thus, these changes may be due to a plasmablast-like stage of B-cell differentiation fixed by EBV immortalization. EBV-infected B cells in infectious mononucleosis are known to avoid germinal centers and accumulate under the mucosal surfaces. EBV-associated opportunistic lymphomas also tend to occur in extranodal sites. These preferred sites of in vivo localization are consistent with the unique profile of chemokine receptor expression exhibited by EBV-immortalized B cells.

Establishment of Monoclonal Anti-Retroviral gp70 Autoantibodies from MRL/lpr Lupus Mice and Induction of Glomerular gp70 Deposition and Pathology by Transfer into Non-Autoimmune Mice

ABSTRACT Several strains of mice, including MRL/MpJ mice homozygous for the Fas mutant lpr gene (MRL/lpr mice), F1 hybrids of New Zealand Black and New Zealand White mice, and BXSB/MpJ mice carrying a Y-linked autoimmune acceleration gene, spontaneously develop immune complex-mediated glomerulonephritis. The involvement of the envelope glycoprotein gp70 of an endogenous xenotropic virus in the formation of circulating immune complexes and their deposition in the glomerular lesions have been demonstrated, as has the pathogenicity of various antinuclear, antiphospholipid, and rheumatoid factor autoantibodies. In recent genetic linkage studies as well as in a study of cytokine-induced protection against nephritis development, the strongest association of serum levels of gp70–anti-gp70 immune complexes, rather than the levels of antinuclear autoantibodies, with the development and severity of glomerulonephritis has been demonstrated, suggesting a major pathogenic role of anti-gp70 autoantibodies in the lupus-prone mice. However, the pathogenicity of anti-gp70 autoantibodies has not yet been directly tested. To examine if anti-gp70 autoantibodies induce glomerular pathology, we established from unmanipulated MRL/lpr mice hybridoma clones that secrete monoclonal antibodies reactive with endogenous xenotropic viralenv gene products. Upon transplantation, a high proportion of these anti-gp70 antibody-producing hybridoma clones induced in syngeneic non-autoimmune and severe combined immunodeficiency mice proliferative or wire loop-like glomerular lesions. Furthermore, deposition of gp70 in glomeruli and pathological changes were observed after intravenous injection of representative clones of purified anti-gp70 immunoglobulin G, demonstrating pathogenicity of at least some anti-gp70 autoantibodies.

N-Terminal Cleavage Fragment of Glycosylated Gag Is Incorporated into Murine Oncornavirus Particles

ABSTRACT Glycosylated Gag (Glycogag) is a transmembrane protein encoded by murine and feline oncornaviruses. While the protein is dispensible for virus replication, Glycogag-null mutants of a neurovirulent murine oncornavirus are slow to spread in vivo and exhibit a loss of pathogenicity. The function of this protein in the virus life cycle, however, is not understood. Glycogag is expressed at the plasma membrane of infected cells but has not been detected in virions. In the present study we have reexamined this issue and have found an N-terminal cleavage fragment of Glycogag which was pelleted by high-speed centrifugation and sedimented in sucrose density gradients at the same bouyant density as virus particles. Its association with virions was confirmed by velocity sedimentation through iodixanol, which effectively separated membrane microvesicles from virus particles. Furthermore, the apparent molecular weight of the virion-associated protein was different from that of the protein extracted from the plasma membrane, suggesting some level of specificity or selectivity of incorporation.

Induction of microthrombotic thrombocytopenia in normal mice by transferring a platelet‐reactive, monoclonal anti‐gp70 autoantibody established from MRL/lpr mice: an autoimmune model of thrombotic thrombocytopenic purpura

MRL/MpJ‐lpr/lpr (MRL/lpr) mice spontaneously develop immune complex‐mediated glomerulonephritis and thrombocytopenia. Although the presence of cross‐reactive anti‐phospholipid antibodies in sera of MRL/lpr mice has been demonstrated, possible relationships between detected autoantibodies and the development of thrombocytopenia have not been elucidated. Recent genetic analyses in a few different strains of lupus‐prone mice have pointed out a close correlation between autoantibodies reactive with endogenous retroviral env gene product, gp70, and the development and severity of glomerulonephritis. In the process of establishing possibly nephritogenic anti‐gp70 autoantibody‐producing hybridoma cells from MRL/lpr mice, we identified an IgG2a‐producing anti‐gp70 hybridoma clone that induced microvascular intraluminal platelet aggregation, thrombocytopenia, and amenia upon transplantation into syngeneic non‐autoimmune mice. This and two other anti‐gp70 antibodies bound onto the surface of mouse platelets, and purified IgG2a of the anti‐gp70 autoantibody induced glomerular lesions with characteristics of thrombotic thrombocytopenic purpura when injected into non‐autoimmune mice. The pathogenic anti‐gp70 autoantibody specifically precipitated a platelet protein with an approximate relative molecular mass of 40 000.