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Background Bortezomib (Bz) is a proteasome inhibitor that directly targets antibody-producing plasma cells. We recently reported the first randomized control trial that evaluated the effects of Bz in patients with systemic lupus erythematosus (SLE). In that study, we demonstrated that Bz treatment is associated with many adverse reactions in patients with refractory disease. In the present study, we examine the therapeutic and toxic effects of Bz on MRL/MpJ-lpr/lpr (MRL/lpr) mice with severe disease activity. Methods Female MRL/lpr mice at 10 and 14 weeks of age were treated with phosphate buffered saline (PBS) (n = 19), Bz (750 μg/kg twice weekly) (n = 27), or cyclophosphamide (Cyc) (1 mg/body, once in 2 weeks) (n = 20). Cellular subsets, serum immunoglobulin, anti-double-stranded DNA (anti-dsDNA) antibody titer, and a pathological index of glomerulonephritis were then analyzed at 22 weeks of age. Survival curves of the 10-week-old and 14-week-old Bz-treated groups were compared. Blood counts, creatinine, liver enzymes, and serum cytokine levels were measured 1 week after Bz treatment. Gene expression profiling of spleens from Bz and Cyc treatment mice were compared with those from control mice. Results The anti-dsDNA antibody levels were significantly higher in 14-week-old than in 10-week-old mice, indicating a higher disease activity at 14 weeks. A significant decrease in the number of splenic cells and glomerulonephritis index was observed in Bz-treated and Cyc-treated mice. Bz, but not Cyc, significantly decreased serum immunoglobulin and anti-dsDNA antibody titer levels. Survival curve analysis revealed a significantly higher mortality rate in 14-week-old than in 10-week-old Bz-treated and control groups. Following two injections of Bz, serum IL-6 and TNF-α levels were significantly more elevated in 14-week-old than in 10-week-old mice. Potentially immunogenic molecules, such as heat shock proteins, were characteristically upregulated in spleens of Bz-treated but not Cyc-treated mice. Conclusions In spite of its therapeutic effect, Bz treatment had more toxic effects associated with increased proinflammatory cytokine levels in mice with a higher disease activity. Understanding the mechanism of the toxicity and developing preventive strategies against it is important for the safe clinical application of Bz in human SLE. Electronic supplementary material The online version of this article (doi:10.1186/s13075-017-1397-7) contains supplementary material, which is available to authorized users.

The aryl hydrocarbon receptor (AHR) is responsible for susceptibility to its ligand-dependent responses. However, the effect of non-AHR factors is less clear. To explore the non-AHR factors, we used two mouse strains with different AHR genetic variants, namely C3H/lpr and MRL/lpr strains with Ala and Val as the 375th amino acid residue, respectively. To assess the contribution of AHR alone, COS-7 cells transiently expressing AHR from each strain were treated with 6-formylindolo[3,2-b]carbazole (FICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and xenobiotic-responsive element (XRE)-driven reporter gene activities were measured. FICZ-EC50 values for the C3H/lpr and MRL/lpr AHR-mediated transactivation were 0.023 and 0.046 nM, respectively, indicating a similar susceptibility in both AHR genotypes. In contrast, C3H/lpr AHR was fourfold more sensitive to TCDD than MRL/lpr AHR. By a pull-down assay using a XRE-containing PCR product as bait and the hepatic nuclear extracts of both FICZ-treated mouse strains, we identified two interacting proteins as heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP-A2) and its splicing variant (hnRNP-A2b). Immunoprecipitation assays demonstrated the AHR interaction with hnRNP-A2/B1. When hnRNP-A2 was co-expressed with the MRL/lpr or C3H/lpr AHR in COS-7, FICZ treatment decreased EC50 to about threefold in both AHR genotypes, compared with EC50 in AHR alone. Similarly, hnRNP-A2b co-expression also lowered the FICZ-EC50 values. In TCDD-treated COS-7, responses depended on the AHR genotype; while no change in TCDD-EC50 was observed for C3H/lpr AHR when hnRNP-A2 was co-expressed, the value was reduced to nearly tenfold for MRL/lpr AHR. Co-transfection with hnRNP-A2b attenuated the AHR sensitivity to TCDD. In conclusion, the hnRNP-A2/B1 interacting with AHR may be a modulator of the AHR ligand sensitivity.

Dioxins cause various toxic effects through the aryl hydrocarbon receptor (AHR) in vertebrates, with dramatic species and strain differences in susceptibility. Although inbred mouse strains C3H/HeJ-lpr/lpr (C3H/lpr) and MRL/MpJ-lpr/lpr (MRL/lpr) are known as dioxin-sensitive and dioxin-resistant mice, respectively, the molecular mechanism underlying this difference remains unclear. The difference in the hepatic proteome of the two mouse strains treated with vehicle or 2,3,7,8-tetrabromodibenzo-p-dioxin (TBDD) was investigated by a proteomic approach of two-dimensional electrophoresis (2-DE) coupled with matrix-assisted laser desorption/ionization time-of-flight/time-of-flight tandem mass spectrometry (MALDI-TOF/TOF). To confirm the strain-difference in response to TBDD treatment, cytochrome P450 (CYP) 1A1 and 1A2 protein levels were measured in both strains. A dose of 10 µg/kg body weight of TBDD induced hepatic CYP1A1 and CYP1A2 expression in both strains, but the expression levels of both CYP1A proteins were higher in C3H/lpr mice than in MRL/lpr mice, supporting that C3H/lpr mice are more sensitive to dioxins than MRL/lpr mice. Proteins that were more induced or suppressed by TBDD treatment in C3H/lpr mice were successfully identified by 2-DE and MALDI-TOF/TOF, including proteins responsible for AHR activation through production of endogenous ligands such as aspartate aminotransferase, indolethylamine N-methyltransferase, and aldehyde dehydrogenases, as well as proteins reducing oxidative stress, such as superoxide dismutase and peroxiredoxins. Taken together, our results provide insights into the molecular mechanism underlying the high dioxin susceptibility of the C3H/lpr strain, in which AHR activation by TBDD is more prompted by the production of endogenous ligands, but the adaptation to oxidative stress is also acquired.

Angiogenic homeostasis is maintained by a balance between vascular endothelial growth factor (VEGF) and Notch signaling in endothelial cells (ECs). We screened for molecules that might mediate the coupling of VEGF signal transduction with down-regulation of Notch signaling, and identified B-cell chronic lymphocytic leukemia/lymphoma6-associated zinc finger protein (BAZF). BAZF was induced by VEGF-A in ECs to bind to the Notch signaling factor C-promoter binding factor 1 (CBF1), and to promote the degradation of CBF1 through polyubiquitination in a CBF1-cullin3 (CUL3) E3 ligase complex. BAZF disruption in vivo decreased endothelial tip cell number and filopodia protrusion, and markedly abrogated vascular plexus formation in the mouse retina, overlapping the retinal phenotype seen in response to Notch activation. Further, impaired angiogenesis and capillary remodeling were observed in skin-wounded BAZF−/− mice. We therefore propose that BAZF supports angiogenic sprouting via BAZF-CUL3-based polyubiquitination-dependent degradation of CBF1 to down-regulate Notch signaling.

Atherothrombosis is the primary pathophysiology that underlies ischemic cerebral infarction. Osteopontin (OPN) is produced in atherosclerotic lesions and is cleaved by activated thrombin. We hypothesized that the rupture or damage of an unstable atherosclerotic plaque increases plasma levels of thrombin-cleaved OPN (trOPN). This study included 90 patients who received carotid angioplasty with stenting (CAS), 23 patients with essential hypertension (EHT) and 10 patients who were treated with carotid endarterectomy (CEA). The CAS patient group included 36 patients that had pre- and post-operative blood tests, diffusion-weighted imaging (DWI) using cerebral MRIs and estimated thrombus debris within the protection device. Immunohistochemistry of CEA specimens revealed that trOPN was detected around intra-plaque vessels. The highest tertile of plasma trOPN levels in CAS patients was higher than trOPN levels in EHT patients. Post-operative trOPN levels were significantly higher in symptomatic compared with asymptomatic patients (P=0.003). New ipsilateral DWI-positive patients revealed higher post-operative trOPN levels (P=0.003) and a higher grade of thrombi (P

Almost 70 years after the description of 'collagen disease' by P. Klemperer et al., it is still controversial whether the diversity and similarity of pathological manifestations among the collagen diseases depends on ambiguity in diagnosis or is an intrinsic quality of the collagen diseases themselves. A genome wide analysis of the MRL mouse models of collagen disease may shed some light on the complex pathological manifestations. Study of the susceptibility loci to each type ofcollagen disease (such as glomerulonephritis, vasculitis, arthritis, sialoadenitis and dacryoadenitis) in the mice, revealed that these lesions developed because of a cumulative effect of multiple gene loci, none of which can induce the related phenotype alone. This may indicate that collagen disease develops in 'a polygenic system', as proposed by K. Mather in 1949. Each lesion in the mice developed because of an additive effect of the polygenes, which is also, in part, hierarchical. Some of the polygenes seemed to be common to those in other collagen diseases as well. Some of the positional candidate genes involved an allelic polymorphism in the coding or promoter regions, thus possibly causing a qualitative or quantitative difference in their function, respectively. As a result, a particular combination of the polygenes with such an allelic polymorphism may thus play a critical role in leading the cascade reaction to developing collagen disease, and also the regular variation in the pathological manifestations. We herein describe this as a polygene network of collagen disease.

Objective To study the genetic contribution of major histocompatibility complex class I polypeptide-related sequence A (MICA), important in natural killer (NK) cell function, in patients with systemic lupus erythematosus (SLE). Methods Japanese patients with SLE (n = 716), those with rheumatoid arthritis (RA) (n = 327), and healthy control subjects (n = 351) were genotyped for the Val129Met polymorphism (rs1051792) and transmembrane (TM) alanine-encoding GCT repeats, termed A4, A5, A5.1, A6, and A9, in the MICA gene. Recombinant human MICA-GST fusion proteins were tested on the NK cell line NK92MI for the expression of NK group 2, member D (NKG2-D), NK cell–mediated cytotoxicity, and interferon-γ (IFNγ) production. Results The MICA129Met allele, TMA9 allele, and 129Met/Met genotype were positively associated with SLE (corrected P [Pcorr] = 0.01 and odds ratio [OR] 1.3, Pcorr = 0.003 and OR 1.6, and Pcorr = 0.02 and OR 1.8, respectively), while the MICA129Val allele was negatively associated with SLE (Pcorr = 0.01, OR 0.8). The MICA129Met;A9 haplotype was also associated with SLE (Pcorr = 0.0006, OR 1.8), and there was an additive genetic effect between the MICA129Met;A9 haplotype and HLA–DRB1*15:01. When NK92MI cells were incubated in vitro with recombinant human disease-associated 129Met;A9 (the combination of polymorphisms at 129Met and TMA9), expression of NKG2-D on NK92MI cells and cytotoxicity of the NK cells were inhibited, but production of IFNγ from NK92MI cells was enhanced. Conclusion The MICA polymorphism is genetically associated with SLE, and MICA appears to contribute to the pathogenesis of SLE by modulating NK cell function.

Synovial hemangioma has been reported to be relatively rare, and usually occurs in childhood and adolescence. However, there are a few reports of the disease in infants. In this report, we diagnosed synovial hemangioma in a 3-year-old girl who had swelling and pain in her left knee. Gadolinium-enhanced magnetic resonance imaging revealed abnormal intensity in an intra-articular lesion. We performed arthroscopy, and arrived at a final diagnosis based on a scopic biopsy. Synovial hemangioma should be considered as a possible diagnosis in infants with swelling and pain in the knee.

Fractalkine (Fkn) is expressed on injured endothelial cells and is a membrane-bound chemokine that attracts cells expressing its receptor, CX3CR1, including CD16+monocytes (CD16+Mos). To clarify the role played by Fkn in the development of glomerular lesions in lupus nephritis, we examined Fkn expression and CD16+Mo accumulation induced in experimental C.B-17/Inc-scid/scid (SCID) lupus model mice by injection of IgG3-producing hybridoma clones obtained from MRL/lpr mice. Glomerular Fkn expression and accumulation of CD16+Mos were semiquantitatively evaluated using laser capture microdissection and real-time PCR. Injection of the 2B11.3 and 7B6.8 clones induced formation of glomerular proliferative and wire-loop lesions, respectively. Immunohistological analysis of the localization of Fkn and CD16+Mos revealed that Fkn expression and CD16+Mo accumulation were markedly elevated in glomerular lesions induced by 2B11.3, whereas no elevation was detected in those induced by 7B6.8. In addition, to examine the contribution of glomerular Fkn to the development of proliferative lesions, L cells producing an Fkn antagonist (Fkn-AT) were transplanted into SCID mice exhibiting proliferative lupus nephritis (DPLN) induced by 2B11.3. Notably, transplantation of the Fkn-AT-producing cells was functionally and histologically protective against this DPLN. Taken together, our findings suggest that Fkn and CD16+Mo accumulation are partially associated with the severity and diversity of histology of lupus nephritis.

Autoimmune diseases are a heterogeneous group of diseases characterized by immune reactions against either a major or a limited number of the bodies own autoantigens, causing inflammation and damage to tissues and organs. Thus, identification of autoantigens is an important first step to understanding autoimmune diseases. Here we demonstrate a simple screening method for identification of autoantigens reacting with patient serum antibodies by combination of an N-terminal biotinylated protein library (BPL), produced using a wheat cell-free protein production system, and a commercially available luminescence system. Optimization studies using well-characterized autoantigens showed specific interactions between N-terminal biotinylated proteins and antibody that were sensitively detected under homogeneous reaction conditions. In this optimized assay, 1 μL of the translation mixture expressing the biotinylated proteins produced significant luminescence signal by addition of diluted serum between 1:500 and 1:10 000 in 25 μL of reaction volume. For the BPL construction, 214 mouse genes, consisting of 103 well-known autoantigens and 111 genes in the mouse autoimmune susceptibility loci, and the sera of MRL/lpr mouse were used as an autoimmune model. By this screening method, 25 well-known autoantigens and 71 proteins in the loci were identified as autoantigen proteins specifically reacting with sera antibodies. Cross-referencing with the Gene Ontology Database, 26 and 38 of autoantigen proteins were predicted to have nuclear localization and identified as membrane and/or extracellular proteins. The immune reaction of six randomly selected proteins was confirmed by immunoprecipitation and/or immunoblot analyses. Interestingly, three autoantigen proteins were recognized by immunoprecipitation but not by immunoblot analysis. These results suggest that the BPL-based method could provide a simple system for screening of autoantigen proteins and would help with identification of autoantigen proteins reacting with antibodies that recognize folded proteins, rather than denatured or unfolded forms., We developed a novel autoantigen protein screening method by combination of an N-terminal biotinylated protein library (BPL), produced using a wheat cell-free system, and a luminescence system. By this screening method, proteins in autoimmune susceptibility loci were identified as autoantigen proteins specifically reacting with sera antibodies. A BPL-based method would help identify autoantigen proteins reacting with antibodies that recognize folded proteins, rather than denatured or unfolded forms.

A cumulative effect of the susceptibility genes with polymorphic alleles may be responsible for rheumatoid arthritis (RA). The objective of this study was to clarify whether susceptibility to RA is under the control of common allelic loci between two different RA models induced by extrinsic and intrinsic factors, collagen-induced arthritis (CIA) in DBA/1 mice and arthritis in MRL/Mp (MRL) mice associated with the Fas deficient mutant gene, Fas(lpr), respectively. CIA was examined in mice of parental DBA/1 and MRL, (MRL x DBA/1) F1 and (MRL x DBA/1) F2 progenies. In genome-wide screening of the severity in the F2 using microsatellite markers, significant linkage was observed on chromosomes 5 and 17 at map position of D5Mit259 and H-2, respectively, associated with DBA/1 alleles, while there was no loci associated with arthritis of MRL-Fas(lpr) mice previously identified. In a quantitative trait locus (QTL) analysis, the locus on chromosome 5 showed the highest peak at map position 35 cM (LOD score 6.0). This study may indicate that the arthritis induced by extrinsic and intrinsic factors is under the control of a different combination of susceptibility genes with common and different alleles, possibly simulating the genetic heterogeneity of RA.

Signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) is an adaptor molecule containing a Src homology 2 (SH2) domain. SAP is expressed in T cells and natural killer (NK) cells and binds to the cytoplasmic domains of SLAM family receptors, resulting in the subsequent recruitment of Fyn. The SAP (SH2D1A) gene is located on the X chromosome and is responsible for X-linked lymphoproliferative disease, characterized by higher susceptibility to Epstein-Barr virus infection. The SAP-mediated signal is not only essential for the development of NKT cells, i.e. unconventional CD1d-restricted T cells with invariant Valpha14 T cell receptors, but also for the regulation of the function of NK cells and conventional T cells. The role of SAP-mediated signaling in the induction of autoimmune diseases has been analyzed using animal models such as lupus, hepatitis, and graft-versus-host disease and is considered important in their pathogenesis in humans. In this review we highlight the current findings on SAP-mediated signaling in hematopoietic cells and discuss its importance in autoimmune diseases and immunological disorders.

Enzyme-linked immunosorbent assays (ELISA) have been widely used to determine quantitatively autoantibodies. However, the processes for the purification and immobilization of antigens in conventional ELISA methods include multiple steps, which have hampered the application for screening of autoantibodies. Here, we have developed a novel ELISA system using the plates pre-coated with glutathione casein to capture recombinant proteins fused to N-terminal glutathione S-transferase (GST). The GST-fused proteins were synthesized with the wheat germ cell-free protein production system. Thus, the present system combined the GST-capture ELISA with the cell-free protein production system, which allowed immobilization of the recombinant proteins with one-step purification. Using this ELISA method, we determined whether rheumatoid factors (RF), which have been considered as one of the representative disease-specific autoantibodies for rheumatoid arthritis (RA), were genetically associated with severity of arthritis in a mouse model for RA, MRL/Mp-lpr/lpr (MRL/lpr). GST-fused human IgG1-Fc (GST-Fc), synthesized with the robotic protein synthesizer, were used as reactants for RF. Serum samples for RF were prepared from 11 lines of a recombinant inbred mouse strain, MXH/lpr, which was established from intercrosses between MRL/lpr and non-arthritic C3H/HeJ-lpr/lpr (C3H/lpr) strains, composed of a different genomic recombination derived from the parental strains in each line. A correlation of RF titers with the severity of the arthritis in these lines was not significant, indicating genetic dissociation of RF from arthritis and that RF is not necessarily required for the development of RA. The present method may provide high-throughput screening for determining the disease-specific autoantibodies in autoimmune diseases.

Problem statement: Mast cells develop from bone marrow-derived progenitor cells and are distributed in the skin or mucosa where they play proinflammatory roles in the first line of defense. Since some tumors in humans and experimental animals exhibited infiltration of increased mast cells, we investigated the contribution of mast cells to the override of tumor rejection. Approach: MRL/N-1 cells are malignant fibrous histiocytoma-like cells established from the spleen of a Fas ligand (FasL)-deficient MRL/Mp-FasLgld/gld (MRL/gld) mouse and are implantable in Fas-deficient MRL/Mp-Faslpr/lpr (MRL/lpr) mice. MRL/N-1 cells were implanted in MRL/gld, MRL/lpr and MRL/+mice after antibody treatments or with mast cells or macrophages and the tumor growth was observed. Results: MRL/N-1 cells were rejected by Fas-intact syngeneic MRL/+ mice in CD8+ T cell-mediated manner. This rejection was inhibited by the co-implanted mast cells. MRL/N-1 cells transfected with FasL were rejected by MRL/+ and MRL/gld mice. Conclusion: Mast cells abrogate the rejection of MRL/N-1 tumor cells and that this tumor rejection is mediated by CD8+ T cells and dependent on host Fas-FasL axis.

Based on the hypothesis that the complex pathological and immunological manifestations of rheumatoid arthritis (RA) and the related diseases are under the control of multiple gene loci with allelic polymorphism, a recombinant congenic mouse strain was prepared between an MRL/Mp-lpr/lpr (MRL/lpr) strain, which develops arthritis resembling RA, and a non-arthritic strain C3H/HeJ-lpr/lpr (C3H/lpr). In MRL/lpr x (MRL/lpr x C3H/lpr) F1 mice, the mice developing severe arthritis were selected based on joint swelling to further continue intercrosses, and then an McH-lpr/lpr-RA1 (McH/lpr-RA1) strain was established and its histopathological phenotypes of joints and autoimmune traits were analyzed. Arthritis in McH/lpr-RA1 mice developed at a higher incidence by 20 weeks of age, compared with that in the MRL/lpr mice, who had severe synovitis (ankle, 60.3%; knee, 65.1%), and also fibrous and fibrocartilaginous lesions of articular ligamenta resembling enthesopathy (ankle, 79.4%; knee, 38.1%), resulting in ankylosis. The lymphoproliferative disorder was less, and serum levels of IgG and IgG autoantibodies including anti-dsDNA and rheumatoid factor were lower than those of both MRL/lpr and C3H/lpr strains. McH/lpr-RA1 mice may provide a new insight into the study of RA regarding the common genomic spectrum of seronegative RA and enthesopathy.

The dendritic cell immunoreceptor (official gene symbol Clec4a2, called Dcir here) is a C-type lectin receptor expressed mainly in dendritic cells (DCs) that has a carbohydrate recognition domain in its extracellular portion and an immunoreceptor tyrosine-based inhibitory motif, which transduces negative signals into cells, in its cytoplasmic portion. We found high Dcir expression in the joints of two mouse rheumatoid arthritis models. Because the structural characteristics of Dcir suggest that it may have an immune regulatory role, and because autoimmune-related genes are mapped to the DCIR locus in humans, we generated Dcir-/- mice to learn more about the pathological roles of this molecule. We found that aged Dcir-/- mice spontaneously develop sialadenitis and enthesitis associated with elevated serum autoantibodies. Dcir-/- mice showed a markedly exacerbated response to collagen-induced arthritis. The DC population was expanded excessively in aged and type II collagen-immunized Dcir-/- mice. Upon treatment with granulocyte-macrophage colony-stimulating factor, Dcir-/- mouse-derived bone marrow cells (BMCs) differentiated into DCs more efficiently than did wild-type BMCs, owing to enhanced signal transducer and activator of transcription-5 phosphorylation. These observations indicate that Dcir is a negative regulator of DC expansion and has a crucial role in maintaining the homeostasis of the immune system.

Aging readily affects immune system under the influence of environmental and/or intrinsic factors while accelerating the development of various immune disorders including autoimmune diseases. Little is known about molecular and cellular mechanisms connecting between immune senescence and development of autoimmune diseases. Here, we first show strain-specific and aging-sensitive onset of B-cell abnormality in a lupus-prone MRL/Mp.Fas(lpr) (MRL/lpr) strain of mice. This abnormality was characterized by the regression of B lymphopoiesis in the bone marrow of this strain. We next examined the association between the B-cell regression and onset of autoimmune diseases in aged (MRL/lpr x C3H/He.Fas(lpr)) F2 mice, in which pathologic phenotypes, such as glomerulonephritis, vasculitis, sialoadenitis and arthritis, variously developed. We also searched whole genome to identify genetic loci linked to the B-cell regression by using the same F2 mice. The B-cell regression manifested in the spleen of F2 mice was retrospectively evaluated by reverse transcriptase-based PCR quantification. The results demonstrated that the onset of autoimmune diseases in the F2 mice was not associated with the aging-sensitive B-cell regression. The genetic study identified a significant locus responsible for the B-cell regression in the vicinity of D5Mit233 (29 cM). This is first evidence for the presence of a genetic locus that affects B lymphopoiesis in an aging-sensitive manner.

Genomic recombination events, including VDJ recombination (VDJR) and class-switch recombination (CSR), are indispensable for the adaptation and progression of the acquired immune system. These processes are completed by orderly, temporal onsets of the gene rearrangements along with B-cell differentiation. The presence of various premature transcripts of immunoglobulin heavy chain (IgH) alleles has been demonstrated during B-cell ontogeny. These include D(H)-J(H) (DJ)-mu, J(H)-mu, and sterile transcripts of C(H). Since these transcripts can be detected during the onset of VDJR and CSR, their presence is believed to reflect a structural change in the genome, favoring VDJR and CSR. This report presents evidence of persistent DJ transcription and onset of CSR on an unproductive IgH allele in peripheral tissues. Nucleotide sequence analysis revealed that these transcripts showed DJ-gamma (Dgamma) configuration and that characteristics of the variable region were essentially the same as those of the DJ-mu transcript previously described. It was noted that the small intestine abundantly expresses Dgamma transcripts with gamma2b and gamma1 isotypes of the IgH constant region. The present findings indicate the onset of CSR preceding V(H) to DJ joining in an unproductive IgH allele of the peripheral B cell and the specificity for the gut-associated condition for B-cell differentiation in the small intestine.

Vascular cell adhesion molecule-1 (VCAM-1, CD106) is important in leukocyte trafficking and its increased expression is associated with a number of chronic inflammatory diseases, including rheumatoid arthritis (RA). A soluble form of VCAM-1 (sVCAM-1) is generated by shedding of the membrane-bound molecule. The concentration of sVCAM-1 is increased in the sera of RA patients, but its pathological role has not been elucidated. The effect of sVCAM-1 relative to protection or aggravation of disease on the development of spontaneous arthritis was examined in an animal model of RA, namely MRL-Fas(lpr) mice (which display a disease resembling human RA), by generation of sVCAM-1 transgenic MRL-Fas(lpr) mice. Transgenic MRL-Fas(lpr) mice that expressed sVCAM-1 had higher incidence and increased severity of arthritis associated with higher levels of serum IgG rheumatoid factor compared with non-transgenic MRL-Fas(lpr) mice. These results suggest that sVCAM-1 plays an arthritogenic role in the development of inflammatory arthritis in MRL-Fas(lpr) mice and may present an important target for therapeutic strategy of RA.

Sex-related differences (SrD) are a general characteristic of human autoimmune diseases. There is an increasing body of evidence that suggests a link between sex-related hormones and autoimmune onsets. Here, through a genetic approach using a lupus mouse model, we attempted to show the involvement of genetic factors in the development of SrD in autoimmune diseases. Using MRL/lpr x (MRL/lpr x C57BL/6.Fas(lpr))F1 (MBN2) mice, the whole genome was searched to identify linkage loci to autoimmune phenotypes inherited from a lupus MRL/Mp.Fas(lpr) (MRL/lpr) strain of mice, which exhibits glomerulonephritis, splenomegaly and antinuclear autoantibody. The genome-wide association study confirmed four linkage loci on chromosomes 4, 7, 13, and 17. Furthermore, differential analyses performed using male and female groups of MBN2 mice revealed that two loci located on chromosomes 4 (41-72 cM, MRL/lpr allele) and 7 (4-21 cM, B6/lpr allele) were male specific and suppressed autoimmune phenotypes. Notably, the sum effect of the two loci adequately explained a range of SrD developed in the MBN2 mice. Our present findings suggest the presence of a male-predominant mechanism underlying the development of SrD in autoimmunity, depending on the effects of autosomal loci under an undefined male-specific condition.

Background Fractalkine (Fkn) is a chemokine that affects cells expressing its receptor, CX3CR1, including CD16-positive (CD16 + ) monocytes/macrophages (CD16 + Mos). The relationship of levels of glomerular Fkn expression and infiltration by CD16 + Mos with the severity and diversity of glomerular lesions in human lupus nephritis is not known. Study Design Retrospective cross-sectional analysis of variables observed in biopsy specimens. Settings & Participants 88 patients with systemic lupus erythematosus. Predictor Histological class and severity of lupus nephritis according to the International Society of Nephrology/Renal Pathology Society and clinicopathologic factors. Outcomes Outcome variables are assays related to the degree of glomerular Fkn expression and CD16 + Mo infiltration. Measurements Immunohistological grading of Fkn staining, number of CD16 + Mos, and messenger RNA levels of Fkn and CD16 in glomeruli. Results Patients with proliferative lupus nephritis (class III and IV glomeruli) showed significantly greater glomerular Fkn expression and CD16 + Mo counts than those with other classes. Infiltrating CD16 + Mos within glomeruli expressed CX3CR1. Moreover, glomerular Fkn expression significantly correlated with the histopathologic activity index and CD16 + Mo counts, and CD16 + Mo counts significantly correlated with serum levels of blood urea nitrogen, complement (CH 50 ), and anti-DNA antibody; estimated glomerular filtration rate; and urinary protein excretion. Glucocorticoid therapy had a tendency to decrease both glomerular Fkn expression and CD16 + Mo counts. Limitations Only frozen biopsy specimens (from 49 patients) were analyzed for the evaluation of glomerular Fkn expression. Conclusion Disease activity and proliferative glomerular lupus nephritis lesions are associated with the glomerular Fkn expression and CD16 + Mo accumulation.

Seasonal changes in the infection state of marine birnavirus (MABV) in Japanese flounder Paralichthys olivaceus and in rearing sea water are described. Sea water and 10–11 healthy fish were sampled monthly from April 2002 to February 2003. The MABV genome was detected throughout the year in >80% of the fish examined at each sampling. The virus was isolated from the liver, kidney, and spleen, but not from the brain. The detection rate in each organ increased from April to October, and then decreased. Detection of virus antigen by the indirect fluorescent antibody technique also showed that the virus was present from spring to autumn (Jun–September) in the liver, kidney, and spleen, but not the brain. Sequence analysis of the MABV genome at the VP2-NS region revealed two specific mutations compared to the standard yellowtail strain (Y-6). It is suggested that the infection state of MABV in Japanese flounder changes to a latent or persistent infection after autumn. MABV was detected in sea water between September and February, suggesting that virus particles in the environment are relatively higher during cool seasons.

Article, INTERNAL MEDICINE. 46(21):1815-1819(2007)

Objective: To identify the genetic loci regulating the incidence and severity of renal autoimmune vasculitis developed in murine lupus. Methods: Vasculitis of renal arteries was histopathologically evaluated in MRL/Mp- Fas lpr (MRL/lpr), C57BL/6- Fas lpr (B6/lpr), (MRL/lpr×B6/lpr) F 1 , and MRL/lpr×(MRL/lpr×B6/lpr) F 1 backcross mice. Using genomic DNA samples of the backcross mice, genome-wide scans, association studies, and linkage analyses were carried out based on genotypes of polymorphic microsatellite markers. Correlations of vasculitis grade and levels of various autoantibodies were also evaluated. Results: Two recessive susceptibility loci of the MRL allele were identified on chromosomes 4 and 1, which had previously been defined as the autoimmune related loci termed Arvm1 and Sle-1/Nba2 , respectively. The former was epistatic to the latter in a female specific manner. The titre of antinuclear autoantibody (ANA) in IgG class, but not ANA in IgM class or anti-dsDNA in either IgG or IgM class, correlated significantly with vasculitis grade. Conclusions: The present loci have been reported in previous studies using a different set of murine strains, suggesting that they are of importance in the development of autoimmune vasculitis in murine models. The concomitance of autoimmune vasculitis and IgG ANA suggests a shared genetic factor regulating these traits.

Objective Crescent formation in the renal glomerulus is a typical manifestation of progressive glomerulopathy associated with fatal renal failure; therefore, its prevention is of clinical importance. Little is known about the pathogenic mechanism for crescent formation. This study was undertaken in an attempt to identify the events that are critical for crescent formation in immune complex crescentic glomerulonephritis (CGN) by analyzing a novel mutant strain of mice. Methods A spontaneous mutant strain of mice was isolated from the autoimmune-prone strain EOD, which stably develops fatal CGN. The mutant phenotypes were assessed histopathologically, hematologically, and immunologically. The mutation was searched for with positional cloning using microsatellite markers. Results Compared with wild-type EOD (WT-EOD) mice, mutant EOD (mut-EOD) mice showed marked improvement in CGN in conjunction with an improvement in spontaneous mortality. In WT-EOD mice, an inverse correlation between blood urea nitrogen concentration and blood platelet count and massive accumulation of platelets in the glomerulus were evident, suggesting that an accumulation of platelets in the glomerulus contributes to the progression of CGN. The mutant platelets showed an abnormal aggregation in response to collagen and thrombin, associated with a bleeding tendency in mut-EOD mice. Genetic analysis revealed a deleterious mutation in the cappuccino gene (cno), which encodes a protein that belongs to a complex called the biogenesis of lysosome-related organelle complex 1 and is profoundly involved in platelet function. Morphologic examination revealed a partial defect in dense body formation in the δ-granule of platelets. Conclusion The present findings suggest that platelet functions have a critical role in crescent formation in autoimmune GN.

Objective IK cytokine has been isolated as a factor that inhibits interferon-γ (IFNγ)–induced expression of class II major histocompatibility complex (MHC) antigens. Aberrant expression of class II MHC antigens has reportedly been recognized in the target organs of autoimmune diseases and been associated with disease activity. In this study, we investigated whether IK cytokine can ameliorate the progression of lupus nephritis in MRL/lpr mice. Methods A truncated IK analog was prepared and transfected into a nonmetastatic fibroblastoid cell line, and then injected subcutaneously into MRL/lpr mice at ages 8 weeks (before the onset of lupus nephritis) and 12 weeks (at the early stage of the disease). Results An IK cytokine, when it was translated from methionine at position 316, acted as a secretory protein. This truncated IK cytokine (tIK) reduced IFNγ-induced class II MHC expression in various cells through decreased expression of class II MHC transcription activator. Treatment of MRL/lpr mice with tIK significantly reduced renal damage as compared with control mice. A significant decrease in macrophage and T cell infiltration was found in the kidneys of tIK-treated mice, resulting in decreased production of IFNγ and interleukin-2. Mice treated with tIK also showed significant reduction of anti-DNA antibodies and circulating immune complexes. A specific reduction of class II MHC expression was observed on B cells and monocytes as well as in the kidney. Conclusion We prepared a potent IK analog and demonstrated its ability to ameliorate the progression of lupus nephritis. This agent may therefore provide a new therapeutic approach for lupus nephritis.

Background: The spontaneous onset of macroscopic arthropathy in the ankle of the particular F 1 mice descended from two Fas -deficient strains of mice; a mutant substrain of MRL/Mp. Fas lpr (MRL/rpl) and C3H/He. Fas lpr (C3H/lpr) was recently observed. Aim: To histopathologically characterise and genetically interpret the unique inheritance mode of disease in this arthropathy model. Methods: MRL/rpl, C3H/lpr, (MRL/rpl × C3H/lpr; MC) F 1 , (C3H/lpr × MRL/rpl; CM) F 1 and MCF 2 mice were bred under specific pathogen-free conditions. Histopathological grade of arthropathy was determined at 6 months by examination under a light microscope. To search for a linkage locus to the arthropathy, the whole genome of selected 48 male MCF 2 mice with 71 polymorphic microsatellite markers was scanned, followed by quantitative trait locus analysis. Results: The incidence of microscopically defined arthropathy in the male and female MCF 1 groups was 100% and 19.4%, respectively. No incidence was observed in the parental strains, MRL/rpl and C3H/lpr, and in CMF 1 mice. In the MCF 1 mice, the arthropathy mainly affected the ankle joints and was histopathologically characterised by marked entheseal proliferation with chondrocytic differentiation and ossification in the ankle joints, the manifestations similar to ankylosing enthesitis reported previously. An MRL/rpl-derived autosomal dominant susceptibility locus was mapped in the distal of D7Mit68 (60 cM) to the ankylosis onset. Conclusion: The MCF 1 mice stably develop spontaneous ankylosing disorders in the ankle, with a male predominance. The unique inheritance mode of ankylosis is possibly interpreted by the genetic interaction between the autosomal dominant locus and a Y-linked locus.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in phosphoinositide turnover and is involved in a variety of physiological functions. We analyzed PLCdelta1 knockout mice and found that PLCdelta1 is required for the maintenance of skin homeostasis. However, there were no remarkable abnormalities except hair loss and runting in PLCdelta1 knockout mice, even though PLCdelta1 is broadly distributed. Here, we report that mice lacking both PLCdelta1 and PLCdelta3 died at embryonic day 11.5 (E11.5) to E13.5. PLCdelta1/PLCdelta3 double-knockout mice exhibited severe disruption of the normal labyrinth architecture in the placenta and decreased placental vascularization, as well as abnormal proliferation and apoptosis of trophoblasts in the labyrinth area. Furthermore, PLCdelta1/PLCdelta3 double-knockout embryos supplied with a normal placenta by the tetraploid aggregation method survived beyond E14.5, clearly indicating that the embryonic lethality is caused by a defect in trophoblasts. On the basis of these results, we conclude that PLCdelta1 and PLCdelta3 are essential in trophoblasts for placental development.

We report a rare case demonstrating the relationship between secondary amyloidosis and uterine leiomyosarcoma. A 59-year-old female with high fever was referred to our hospital. Laboratory tests revealed increased white blood cells, eosinophilia and an accelerated erythrocyte sedimentation rate. Endoscopic examination of the stomach and colon revealed amyloid deposits in their mucosa. The patient showed no symptoms that suggested amyloidosis. No other organs or tissues were surveyed for amyloid deposition. Ga scintigraphy, computed tomography and magnetic resonance imaging suggested necrotic infectious leiomyoma of the uterus, which was considered to be the cause of the fever. The patient underwent total hysterectomy. The histological diagnosis of the mass revealed a low-grade uterine leiomyosarcoma with necrosis. Amyloid deposits in the gastric mucosa disappeared 1 year after the operation. In this case, amyloid deposition was detected by endoscopic biopsy before clinical manifestations. The deposition was reversible and was successfully treated. Thus, it is logical and useful to undertake endoscopic mucosa biopsy to check for amyloid deposition in patients with systemic inflammation, whose serum amyloid A protein level has been high for several months. In addition, peripheral eosinophilia was also detected in this case. Although eosinophilia associated with malignant tumor has been recognized, it is an uncommon occurrence.

Summary Accumulation of plasma cells in the synovium is one of the diagnostic hallmarks in the histopathological manifestations of rheumatoid arthritis (RA). This seems to be prominent even prior to significant B cell infiltration and/or formation of lymphoid follicles in the synovium. To clarify the mechanism of early plasma cell accumulation, we examined in situ expression of chemokines and their receptors using synovial targeting biopsy specimens, which were obtained under arthroscopy from early RA patients. By immunohistochemical staining, plasma cells were found to express a chemokine receptor CXCR3, while synovial fibroblasts in the synovial sublining regions expressed its ligand, Mig/CXCL9. By reverse transcription-polymerase chain reaction (RT-PCR), using targeted lesions of synovial tissues obtained by laser capture microdissection, expression levels of Mig/CXCL9 in the synovial sublining regions were remarkably high and were likely to be associated with interferon (IFN)-γ expression. Furthermore, cultured synovial fibroblasts were confirmed to produce Mig/CXCL9 upon stimulation with IFN-γ. Our results indicate that in the early stage of RA, plasma cells expressing CXCR3 may be recruited directly from the circulation into the synovial sublining regions by its ligand, Mig/CXCL9, produced by synovial fibroblasts.

Objective Lupus nephritis is characterized by immune complex deposition and inflammatory cell infiltration into the renal glomeruli. Local generation of chemokines and the presence of chemokine receptors on the infiltrating cells may be involved in this process. Fractalkine (Fkn)/CX3CL1 and its receptor, CX3CR1, form one such chemokine system. We therefore undertook this study to investigate whether Fkn antagonist inhibits the initiation and progression of lupus nephritis in MRL/lpr mice. Methods NH2-terminally truncated Fkn/CX3CL1 analogs were transfected into a nonmetastatic fibroblastoid cell line, MRL/N-1, and injected subcutaneously into MRL/lpr mice. Results Fkn analogs truncated by ≥4 amino acid residues from the N-terminus failed to induce chemotaxis and calcium influx by CX3CR1-expressing cells. Of these, the most potent antagonist (Fkn-AT) lacked the 4 N-terminal amino acid residues. Fkn expression in the glomerulus was significantly increased in 12-week-old MRL/lpr mice. Expression was localized predominantly in the glomerular endothelial cells, but was occasionally observed in the mesangial cells and, to a lesser extent, in the interstitial microvasculature. Inoculation of MRL/lpr mice with Fkn-AT before the onset or during the early stages of lupus nephritis significantly reduced glomerular hypercellularity, glomerulosclerosis, crescent formation, and vasculitis compared with control mice. This seemed to be due to a marked reduction in macrophage accumulation. In contrast, Fkn antagonist did not affect pneumonitis, sialadenitis, lymphadenopathy, or splenomegaly. Conclusion We prepared a novel potent Fkn antagonist and demonstrated its ability to delay the initiation and ameliorate the progression of lupus nephritis. This agent may therefore provide a new therapeutic approach to lupus nephritis.

Potentially, autoimmune diseases develop from a combination of multiple genes with allelic polymorphisms. An MRL/Mp-Fas(lpr) (/) (lpr) (MRL/lpr) strain of mice develops autoimmune diseases, including lupus nephritis, but another lpr strain, C3H/HeJ-Fas(lpr) (/) (lpr) (C3H/lpr) does not. This indicates that MRL polymorphic genes are involved in the development of the diseases. By quantitative trait loci (QTL) analysis using 527 of the (MRL/lpr x C3H/lpr)F(2) mice, we identified a novel locus for susceptibility to lupus nephritis at map position D5Mit115 on chromosome 5, the same alias of the osteopontin (Opn) gene (LOD score =4.0), susceptible in the MRL allele. In functional analyses of the MRL and C3H Opn alleles using synthetic osteopontin (OPN) made with a new method "cell-free system" with wheat germ ribosomes, the MRL-OPN induced higher expression and production of immunoglobulins as well as cytokines including TNF-alpha, IL-1beta and IFN-gamma in splenocytes and/or macrophages than that of the C3H allele. These findings suggest that allelic polymorphism of OPN causes the functional differences in antibody production and macrophage activation between MRL and C3H strains, possibly involved in the development of lupus nephritis.

SUMMARY MRL/Mp mice bearing the Fas deletion mutant gene, lpr (MRL/lpr), spontaneously develop polyarthritis, sialoadenitis and dacryoadenitis, resembling rheumatoid arthritis (RA), and also corneal involvement such as keratopathy and scleritis, which is a major complication in RA patients. In this study, we found that the expression levels of IL-1β and MMP-1 mRNAs in cornea were high in both MRL/lpr and MRL/Mp-+/+ strains of mice at an age younger than when they develop any inflammatory lesions. This was not true of other inbred strains, even those bearing the lpr gene, and also not of (NZB × NZW) F1 lupus mice. There was no significant difference in the expression of IL-1α and TGFβ in cornea in these strains. Using crosses between MRL/lpr and C3H/HeJ-lpr/lpr (C3H/lpr) mice, at least the expression of IL-1β was found to be under the control of the MRL genetic background, likely with a recessive mode of inheritance. Considering that IL-1β in cornea was detected particularly in the epithelial layer, the high expression of IL-1β in cornea is most likely involved in the genetic predisposition for corneal involvement and possibly also for arthritis in an MRL strain of mice.

A 37-year-old man was admitted to our hospital because of fever, polyarthralgia, and subcutaneous tumors. There was swelling of the bilateral wrists and ankles, and subcutaneous tumors over the bilateral elbow joints. Despite his complaints of multiple symptoms, clinical investigations failed to reveal any abnormality. Although laboratory parameters improved rapidly after steroid therapy, the symptoms remained unchanged, and there was an enormous discrepancy between the laboratory data and his symptoms. A biopsy specimen from one of the subcutaneous tumors revealed foreign-body granuloma associated with a foreign body fragment. Because the nursing staff later discovered that the patient had been carrying out self-injection, a diagnosis of Munchausen syndrome was made. Munchausen syndrome should be included in the differential diagnosis of rheumatic diseases.

MRL/MpTn-gld/gld (MRL/gld) mice, which are deficient in a functional Fas ligand (FasL), spontaneously develop autoimmune diseases involving both lethal glomerulonephritis and systemic arteritis, while MRL/Mp-+/+ (MRL/+) and C3H/HeJ-gld/gld (C3H/gld) do not. To determine the cells responsible for the development of glomerulonephritis and arteritis, we transferred bone marrow cells from MRL/gld mice to undiseased MHC-compatible gld/gld or +/+ mice. In bone marrow irradiation chimeras, MRL/gld bone marrow cells were transferred to lethally irradiated MRL/+ or C3H/HeJ-+/+ (C3H/+) mice, and both recipients developed glomerulonephritis associated with hypergammaglobulinemia without causing graft-versus-host (GVH)-like diseases. However, a striking difference between them was that MRL/+ recipients developed arteritis, but C3H/+ recipients did not. In bone marrow mixed chimeras formed by transferring MRL/gld bone marrow cells to unirradiated mice, the MRL/gld bone marrow cells induced glomerulonephritis in C3H/gld mice, but not in C3H/+ and MRL/+ mice. These results indicate that bone marrow cells from MRL/gld mice can cause glomerulonephritis in mice, even in those with a C3H background, possibly if they survive longer by escaping from Fas-mediated apoptosis, while the development of arteritis requires the MRL genetic background in the re-cipients. This is the first report of the transfer of arteritis in lupus mice to undiseased recipients.

Objective To clarify the mode of inheritance of the tissue distribution of vasculitis in MRL/Mp-lpr/lpr (MRL/lpr) lupus-prone mice and to identify the susceptibility loci. Methods Vasculitis in individual MRL/lpr, C3H/HeJ-lpr/lpr (C3H/lpr), (MRL/lpr × C3H/lpr)F1, and (MRL/lpr × C3H/lpr)F2 intercross mice was analyzed by histopathologic grading of main branches of the aorta and of medium-sized arteries in the lower limbs. Genomic DNA samples from F2 intercross mice were examined by simple sequence-length polymorphism analysis, and the polymorphic microsatellite markers highly associated with vasculitis in each tissue were determined as vasculitis susceptibility loci. Results A susceptibility locus with significant linkage to vasculitis of main branches of the aorta was mapped on chromosome 4 at D4Mit213 (map position 13.3cM) selectively in males, while vasculitis of medium-sized arteries in the lower limbs was mapped to different chromosomes: at D8Mit31 on chromosome 8 (map position 33.0) selectively in females and at D5Mit36 on chromosome 5 (map position 65.0). All of these were different from the previously defined loci governing susceptibility to vasculitis involving the kidneys. Conclusion Systemic vasculitis in MRL/lpr mice is genetically controlled with cumulative effects of multiple gene loci, each of which has tissue specificity.

A Wnt coreceptor low-density lipoprotein receptor-related protein 5 (LRP5) plays an essential role in bone accrual and eye development. Here, we show that LRP5 is also required for normal cholesterol and glucose metabolism. The production of mice lacking LRP5 revealed that LRP5 deficiency led to increased plasma cholesterol levels in mice fed a high-fat diet, because of the decreased hepatic clearance of chylomicron remnants. In addition, when fed a normal diet, LRP5-deficient mice showed a markedly impaired glucose tolerance. The LRP5-deficient islets had a marked reduction in the levels of intracellular ATP and Ca 2+ in response to glucose, and thereby glucose-induced insulin secretion was decreased. The intracellular inositol 1,4,5-trisphosphate (IP3) production in response to glucose was also reduced in LRP5−/− islets. Real-time PCR analysis revealed a marked reduction of various transcripts for genes involved in glucose sensing in LRP5−/− islets. Furthermore, exposure of LRP5+/+ islets to Wnt-3a and Wnt-5a stimulates glucose-induced insulin secretion and this stimulation was blocked by the addition of a soluble form of Wnt receptor, secreted Frizzled-related protein-1. In contrast, LRP5-deficient islets lacked the Wnt-3a-stimulated insulin secretion. These data suggest that Wnt/LRP5 signaling contributes to the glucose-induced insulin secretion in the islets.

The interaction between OX40 and OX40 ligand (OX40L) is suggested to provide T cells with an effective costimulatory signals during T cell-APC interaction. To examine the in vivo effect of constitutive OX40/OX40L interaction during immune regulation, we report the establishment of OX40L-transgenic (OX40L-Tg) mice that constitutively express OX40L on T cells. Markedly elevated numbers of effector memory CD4+ T cells, but not CD8+ T cells, were observed in the secondary lymphoid organs of OX40L-Tg mice. Upon immunization with keyhole limpet hemocyanin in the absence of adjuvant, profound T cell proliferative responses and cytokine productions were seen in the OX40L-Tg mice as compared with wild-type mice. Furthermore, in OX40L-Tg mice administrated with superantigen, this constitutive OX40/OX40L interaction on CD4+ T cells completely prevented normal in vivo clonal T cell deletion. Interestingly, OX40L-Tg mice on the C57BL/6 background spontaneously developed interstitial pneumonia and inflammatory bowel disease that was accompanied with a significant production of anti-DNA Ab in the sera. Surprisingly, these diseases were not evident on the OX40L-Tg mice on the BALB/c strain. However, such inflammatory diseases were successfully reproducible in recombination-activating gene (RAG)2-deficient mice upon transfer of OX40L-Tg CD4+ T cells. Blockade of OX40/OX40L interaction in the recipient RAG2-deficient mice completely prevented disease development. The present results orchestrated in this study indicate that OX40/OX40L interaction may be a vital link in our understanding of T cell-mediated organ-specific autoimmunity.

SUMMARYIn this study we established a new animal model for exploring the pathogenesis of autoimmune pancreatitis. We have found previously that MRL/Mp-+/+(MRL/+) mice develop pancreatitis spontaneously by an autoimmune mechanism but only when they are more than 34 weeks old. Because this disease might be a model of multi-factorial diseases controlled by genetic and environmental factors, beginning at 6 weeks old, we injected polyinosinic:polycytidylic acid (poly I:C) into MRL/+ mice and in addtion, into MRL/Mp mice bearing the Fas deletion mutant gene, lpr (MRL/lpr). Poly I:C induced chronic severe pancreatitis in all the MRL/+ mice and to a lesser extent in the MRL/lpr mice by 18 weeks of age. There was no pancreatitis in control mice of both strains at the same age. Other than chronic pancreatitis, no severe autoimmune diseases were observed in MRL/+ mice. Immunohistochemical examinations revealed predominant infiltration of CD4+ T cells and Mac-2+ activated macrophages in the pancreatic lesions. Splenic expression of the mRNAs for TNF-α and IL-10, which is known to suppress the development of pancreatitis, were increased in both strains of mice. These findings suggest that an MRL strain of mice treated with poly I:C might be a good model for developing new approaches to the study of the pathogenesis of autoimmune pancreatitis.

Circulating leukocytes, particularly neutrophils and monocytes, are important effector cells in the induction of many forms of glomerulonephritis. Adhesion molecules, especially selectins, are also thought to be critical for the development of this disease. We examined the possible suppressive effect of soluble E-selectin on the development of experimental lupus nephritis induced by the injection of a hybridoma clone (2B11.3) derived from an MRL/MpJ-lpr/lpr lupus mouse. This clone produces IgG3 antibodies that induce severe proliferative glomerulonephritis resembling lupus nephritis when injected into normal mice. Transgenic mice with a soluble E-selectin gene were injected intraperitoneally with the hybridoma cells and histopathologically examined on day 15. As a result, the development of glomerulonephritis was significantly suppressed. This suppression was characterized by fewer inflammatory cell infiltrates, compared with non-transgenic litter mates, despite the fact that there were no remarkable differences in immunoglobulin deposits or expression of E-selectin between the two groups. These findings suggest that by controlling inflammatory cell infiltration, soluble E-selectin plays a preventative role in the development of a particular type of lupus nephritis.

Rheumatoid arthritis (RA) is a systemic disorder characterized by synovial inflammation and subsequent destruction and deformity of synovial joints. The articular lesions start with synovitis, focal erosion of unmineralized cartilage, and then culminate in the destruction of subarticular bone by pannus tissue. Periarticular osteopenia and systemic osteoporosis follow as late complications of RA. Osteoclasts, specialized cells that resorb bone, play a central role in developing these osteolytic lesions. To elucidate the mechanism of osteoclastogenesis and bone destruction in autoimmune arthritis, we investigated the expression of RANK ligand (RANKL), RANK, and osteoprotegerin (OPG) mRNA in a mouse type II collagen-induced arthritis (CIA) model by in situ hybridization. The results indicated that most of the TRAP-positive mono- and multinucleated cells in the inflamed and proliferating synovium and in the pannus were RANK-positive authentic osteoclasts and their precursors. In the inflamed synovium and pannus of the mouse CIA model, synovial fibroblastic cells around these RANK-positive cells were strongly positive for RANKL. Moreover, RANKL-positive osteoblasts on the endosteal bone surface, at a distance from the affected synovial joints, increased significantly in the mouse CIA model prior to periarticular osteopenia and systemic osteoporosis. These data indicated that the RANKL-RANK system plays an important role for osteoclastogenesis in both local and systemic osteolytic lesions in autoimmune arthritis, and can therefore be a good target for therapeutic intervention.