Monocyte subsets involved in the development of systemic lupus erythematosus and rheumatoid arthritis.

AbstractMonocytes are evolutionally conserved innate immune cells that play essential roles for the protection of the host against pathogens and also produce several inflammatory cytokines. Thus, the aberrant functioning of monocytes may affect not only host defense but also the development of inflammatory diseases. Monocytes are a heterogeneous population with phenotypical and functional differences. Most recent studies have shown that monocytes are divided into three subsets, namely classical, intermediate and non-classical subsets, both in humans and mice. Accumulating evidence showed that monocyte activation is associated with the disease progression in autoimmune diseases, such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). However, it remains to be determined how monocytes contribute to the disease process and which subset is involved. In this review, we discuss the pathogenic role of monocyte subsets in SLE and RA on the basis of current studies by ourselves and others to shed light on the suitability of monocyte-targeted therapies in these diseases.

FcγRIIb on B Cells and Myeloid Cells Modulates B Cell Activation and Autoantibody Responses via Different but Synergistic Pathways in Lupus-Prone Yaa Mice.

C57BL/6 (B6).FcγRIIb-/- Yaa mice spontaneously develop lethal lupus nephritis. To define the cell type-specific role of FcγRIIb in Yaa-associated lupus, we established B cell- (CD19Cre Yaa), myeloid cell- (C/EBPαCre Yaa), and dendritic cell- (DC) (CD11cCre Yaa) specific FcγRIIb-deficient B6.Yaa mouse strains. CD19Cre Yaa mice developed milder lupus than B6.FcγRIIb-/- Yaa mice, indicating that FcγRIIb deficiency on B cells is not sufficient for the development of severe disease. Surprisingly, C/EBPαCre Yaa mice also showed autoantibody production and mild lupus similar to that in CD19Cre Yaa mice, whereas CD11cCre Yaa mice stayed disease free. These observations indicate that FcγRIIb deficiency in B cells and myeloid cells, but not DCs, contributes to the severe disease in B6.FcγRIIb-/- Yaa mice. Flow cytometric analysis showed that the frequency of peripheral Gr-1- but not Gr-1+ monocyte was increased in B6.FcγRIIb-/- Yaa and C/EBPαCre Yaa but not CD19Cre Yaa mice, suggesting a link between FcγRIIb deficiency on myeloid cells and the high frequency of Gr-1- monocytes. RNA sequencing revealed that compared with Gr-1+ monocytes, Gr-1- monocytes expressed higher levels of the B cell-stimulating cytokines BSF-3, IL-10, and IL-1ß, the DC markers CD11c, CD83, and Adamdec1, and the antiapoptotic factors Bcl2 and Bcl6. In conclusion, in Yaa-associated lupus nephritis, FcγRIIb on B cells and myeloid cells modulates B cell activation via different but synergistic pathways. Gr-1- monocytes are the most likely candidate myeloid cells involved.

Anti-CD11b antibody treatment suppresses the osteoclast generation, inflammatory cell infiltration, and autoantibody production in arthritis-prone FcγRIIB-deficient mice.

BACKGROUND: Previously we established an arthritis-prone FcγRIIB-deficient mouse strain (designated KO1). Anti-mouse CD11b mAb (5C6) has been reported to inhibit the recruitment of peripheral CD11b+ myelomonocytic cells from the blood to the inflammatory site. These cells include neutrophils and monocytes, both of which play important roles in the development of arthritis. Here we treated KO1 mice with 5C6 mAb in order to study its effect on arthritis development. METHODS: To evaluate the disease-preventive effect of 5C6, 4-month-old preclinical KO1 mice were divided into three groups: the first treated with 5C6 for 6 months, the second treated with normal rat IgG for 6 months, as a control, and the third left untreated. Arthritis severity and immunological abnormalities were compared among the groups, along with transcriptional levels of several important arthritis-related factors in ankle joints, spleen, and peripheral blood cells. RESULTS: The 5C6 treatment ameliorated arthritis in KO1 mice, showing decreases in inflammatory cell infiltration and osteoclast formation. Analysis of transcriptional levels in ankle joints revealed that compared with the two control groups, the 5C6-treated group showed downregulated expression of RANK, RANKL, MCP-1, RANTES, TNFα, and IL-6, and at the same time showed significantly up-regulated expression of the decoy receptor for RANKL, i.e. osteoprotegerin. In addition, the disease suppression was associated with the lower serum levels of autoantibodies, and the decreased frequencies of activated B cells and plasma cells. The expression levels of B cell activation/differentiation-related cytokines were suppressed in spleen and peripheral leukocytes of the 5C6-treated mice. Intriguingly, while untreated KO1 mice spontaneously developed marked monocytosis, the 5C6-treated mice showed the significantly down-regulated frequency of monocytes. CONCLUSIONS: The outcome of 5C6 treatment was complex, in which the 5C6-mediated disease-preventive effect is likely due on one hand to the decrease in the recruitment of inflammatory cells and osteoclast precursor monocytes from the periphery into the joints, and on the other hand to the suppression of B cell activation/maturation and of autoantibody production via the suppression of B cell stimulating cytokine production. The lower levels of these cytokines may be the secondary effect of the lower frequency of monocytes, since monocytes/macrophages are the major producers of these cytokines.

IL-6 signal blockade ameliorates the enhanced osteoclastogenesis and the associated joint destruction in a novel FcγRIIB-deficient rheumatoid arthritis mouse model.

OBJECTIVE: We earlier found that TNFα but not interleukin (IL)-17 is indispensable in the pathogenesis of spontaneously occurring rheumatoid arthritis (RA)-like disease in our newly established FcγRIIB-deficient C57BL/6 (B6) mouse model, designated KO1. Here, we examined the role of IL-6 in the pathogenesis of RA features in KO1, with particular reference to cartilage and bone destruction in arthritic joints. METHODS: To evaluate the preventive effect of MR16-1, a rat anti-mouse IL-6 receptor (IL-6R) mAb, 4-month-old preclinical KO1 mice were divided into three groups: the first treated with MR16-1 for 6 months, the second treated with normal rat IgG, as a control, and the third left untreated. The incidence and severity of arthritis, immunological abnormalities, and transcription levels of receptor activator of NF-κB ligand (RANKL), osteoprotegerin (OPG), and inflammatory cytokines/chemokines in ankle joint tissues were compared among the three groups. The therapeutic effect of MR16-1 was examined by treating 7-month-old KO1 mice in the early stages of arthritis for 2 months. RESULTS: Compared with the findings in the KO1 mice left untreated or treated with normal rat IgG, the development of arthritis was markedly suppressed in mice with MR16-1 treatment started from preclinical stages. The suppression was associated with the decrease in production of autoantibodies, rheumatoid factors (RF), and anti-cyclic citrullinated peptide (CCP). Histologically, marked synovitis, pannus formation, and cartilage and bone destruction associated with the increase in tartrate-resistant acid phosphatase (TRAP)-positive osteoclast generation were evident in the two control groups; however, these findings were virtually absent in MR16-1-treated mice. Real-time PCR analysis revealed that the up-regulated expression levels of MCP-1, IL-6, and TNFα, and the aberrantly high RANKL/OPG expression ratio in synovial joint tissues from the two control groups of mice with overt arthritis were significantly suppressed in MR16-1-treated mice. In mice with therapeutic MR16-1 treatment, there was no progression in arthritis score and the RANKL/OPG ratio in joint tissues was significantly suppressed. CONCLUSIONS: Administration of an anti-IL-6R mAb ameliorated spontaneously occurring RA-like disease features, indicating that IL-6, as well as TNFα, plays a pivotal role in the pathogenesis of RA in KO1 mice. Current studies showed that, in addition to the role in enhancing autoantibody production, IL-6 promotes synovial tissue inflammation and osteoclastogenesis, leading to the severe synovitis with pannus formation and the progressive cartilage and bone destruction in multiple joints.

Genome-wide genetic study in autoimmune disease-prone mice.

Mouse models of autoimmune diseases provide invaluable insights into the cellular and molecular bases of autoimmunity. Genetic linkage studies focusing on their abnormal quantitative phenotypes in relation to the loss of self-tolerance will lead to the identification of polymorphic genes that play pivotal roles in the genetic predisposition to autoimmunity. In this chapter, we first overview the basic concepts in the statistical genetics and then provide guides to genotyping microsatellite DNA markers and to quantitative trait loci mapping using a MAPMAKER program.

TNFα but not IL-17 is critical in the pathogenesis of rheumatoid arthritis spontaneously occurring in a unique FcγRIIB-deficient mouse model.

OBJECTIVE: TNFα and IL-17 have been shown to be the major inflammatory cytokines involved in the pathogenesis of rheumatoid arthritis (RA). Here, we examined the effect of these cytokines on spontaneously occurring RA in our newly established arthritis-prone FcγRIIB- deficient C57BL/6 (B6) mice, designated KO1, by introducing genetic deficiency of TNFα and IL-17 into KO1 mice. METHODS: KO1.TNFα(-/-) and KO1.IL-17(-/-) mice were established by crossing KO1 with TNFα-deficient and IL-17-deficient B6 mice, respectively. The incidence and severity of RA, cartilage and bone destruction, immunological abnormalities, and transcription levels of receptor activator of NF-κB ligand (RANKL)/osteoprotegerin (OPG) and inflammatory cytokines/chemokines in ankle joints were compared among KO1, KO1.TNFα(-/-), and KO1.IL-17(-/-) mice. RESULTS: The development of RA was completely inhibited in KO1.TNFα(-/-) mice. In contrast, KO1.IL-17(-/-) mice unexpectedly developed severe RA comparable to KO1. Compared with those in KO1 and KO1.IL-17(-/-) mice, frequencies of peripheral monocytes, known to be containing osteoclast precursors, were significantly decreased in KO1.TNFα(-/-) mice. Intriguingly, while RANKL expression levels in ankle joints did not differ among the three strains, OPG expression levels were drastically decreased in arthritis-prone, but not arthritis-free, mice. The expression levels of inflammatory cytokines/chemokines, such as MCP-1, IL-6, and TNFα, were up-regulated in arthritis-prone mice. CONCLUSION: TNFα is indispensable while IL-17 is dispensable in the pathogenesis of RA in KO1 mice. In this model, TNFα may contribute to the development of arthritis, through mediating the increase in frequencies of osteoclast precursors in circulation and their migration into the joints, and the decrease in OPG expression, leading to the up-regulated osteoclastogenesis associated with severe cartilage and bone destruction.

Nephrotoxic effect of subchronic exposure to S-(1,2-dichlorovinyl)-L-cysteine in mice.

The effect of subchronic exposure of S-(1,2-dichlorovinyl)-L-cysteine (DCVC), an active metabolite of trichloroethylene (TCE), was investigated in mice, as a part of mechanistic assessment of renal toxicity of TCE. To examine the subchronic effects of DCVC on kidney function, Balb/c male mice were administered DCVC orally and intraperitoneally once a week for 13 weeks at 1, 10 and 30 mg/kg (Main Study) and for 8 weeks at 30 mg/kg (PCR Study). At the terminal sacrifice, mice orally and intraperitoneally administered with 10 and 30 mg/kg showed significantly lower kidney weight and significantly higher blood urea nitrogen levels than the control group. Pathological examination revealed that a dose of 30 mg/kg delivered by both routes resulted in renal tubular degeneration characterized by tubular necrosis and interstitial fibrosis, and in degradation of the cortex. Degenerative changes were accompanied by the increased expression of tumor necrosis factor-α, interleukin-6 and cyclooxygenase-2 mRNAs in the kidney of mice treated with 30 mg/kg for 8 weeks. These pathohistological observations mostly corresponded to those in short-term toxicity studies on DCVC. DCVC might be a direct cause of renal toxicity, which is suggested from the aggravation in these symptoms with the dose increase.

Transgenic overexpression of G5PR that is normally augmented in centrocytes impairs the enrichment of high-affinity antigen-specific B cells, increases peritoneal B-1a cells, and induces autoimmunity in aged female mice.

To investigate signals that control B cell selection, we examined expression of G5PR, a regulatory subunit of the serine/threonine protein phosphatase 2A, which suppresses JNK phosphorylation. G5PR is upregulated in activated B cells, in Ki67-negative centrocytes at germinal centers (GCs), and in purified B220(+)Fas(+)GL7(+) mature GC B cells following Ag immunization. G5PR rescues transformed B cells from BCR-mediated activation-induced cell death by suppression of late-phase JNK activation. In G5PR-transgenic (G5PR(Tg)) mice, G5PR overexpression leads to an augmented generation of GC B cells via an increase in non-Ag-specific B cells and a consequent reduction in the proportion of Ag-specific B cells and high-affinity Ab production after immunization with nitrophenyl-conjugated chicken γ-globulin. G5PR overexpression impaired the affinity-maturation of Ag-specific B cells, presumably by diluting the numbers of high-affinity B cells. However, aged nonimmunized female G5PR(Tg) mice showed an increase in the numbers of peritoneal B-1a cells and the generation of autoantibodies. G5PR overexpression did not affect the proliferation of B-1a and B-2 cells but rescued B-1a cells from activation-induced cell death in vitro. G5PR might play a pivotal role in B cell selection not only for B-2 cells but also for B-1 cells in peripheral lymphoid organs.

Angiotensin II receptor antagonist reduces subsequent uterine arterial dysfunction in pregnant offspring of protein-restricted rat dams.

AIM: A low-protein diet (LPD) during pregnancy induces vascular dysfunction and hypertension in the offspring, prevented by administration of an angiotensin II type 1 (AT(1)) receptor antagonist in early life to the offspring. Whether such protection extends to subsequent pregnancy is unknown; we therefore hypothesized that administration of a specific AT(1) receptor antagonist (losartan) in early life to offspring of LPD dams would improve vascular dysfunction in their uterine arteries when they, in turn, were pregnant. METHODS: Pregnant rats were randomly divided into two dietary groups fed a control (C) or protein-restricted (R) diet throughout pregnancy. Between two and 10 weeks postnatally, female offspring (F(1)) were randomly assigned to drink either pure tap water (CO, RO) or water with losartan (CL, RL). Offspring were mated and killed on gestational day 19 or 20 in order to investigate uterine artery function. RESULTS: In pregnant offspring, vasoconstriction of the uterine arteries to phenylephrine (PE) and the thromboxane A2 mimetic U46619 was greater in RO than CO (F(1)). Responses to both antagonists were suppressed in RL (F(1)). Relaxation to sodium nitroprusside was increased in RO versus CO and suppressed in RL versus RO (F(1)). CONCLUSION: Administration of an AT(1) receptor antagonist to offspring during the suckling and juvenile period improves the uterine vascular dysfunction in pregnancy induced by prior maternal LPD during their development. Such treatment may contribute to decreasing the transmitted risks of maternal malnutrition from offspring to the subsequent generation.

Presumptive role of 129 strain-derived Sle16 locus in rheumatoid arthritis in a new mouse model with Fcγ receptor type IIb-deficient C57BL/6 genetic background.

OBJECTIVE: Fcγ receptor type IIb (FcγRIIb) is a major negative regulator of B cells, and the lack of FcγRIIb expression has been reported to induce systemic lupus erythematosus (SLE) in mice of the C57BL/6 (B6) genetic background. The 129 strain-derived Sle16 locus on the telomeric region of chromosome 1 including polymorphic Fcgr2b confers the predisposition to systemic autoimmunity when present on the B6 background. We undertook this study to examine the effect of the Sle16 locus on autoimmune disease in FcγRIIb-deficient B6 mice. METHODS: We established 2 lines of FcγRIIb-deficient B6 congenic mouse strains (KO1 and KO2) by selective backcrossing of the originally constructed FcγRIIb-deficient mice on a hybrid (129×B6) background into a B6 background. Although both lack FcγRIIb expression, the KO1 and KO2 strains carry different lengths of the 129 strain-derived telomeric chromosome 1 segment flanked to the null-mutated Fcgr2b gene; the KO1 strain carries a 129 strain-derived ∼6.3-Mb interval distal from the null-mutated Fcgr2b gene within the Sle16 locus, while this interval in the KO2 strain is of B6 origin. RESULTS: Unexpectedly, both strains failed to develop SLE; instead, the KO1 strain, but not the KO2 strain, spontaneously developed severe rheumatoid arthritis (RA) with an incidence reaching >90% at age 12 months. CONCLUSION: The current study shows evidence that the epistatic interaction between the Fcgr2b-null mutation and a polymorphic gene(s) in the 129 strain-derived interval located in the distal Sle16 locus contributes to RA susceptibility in a new mouse model with the B6 genetic background, although the participation of other genetic polymorphisms cannot be totally excluded.

Susceptibility loci for the defective foreign protein-induced tolerance in New Zealand Black mice: implication of epistatic effects of Fcgr2b and Slam family genes.

In contrast to normal mice, autoimmune-prone New Zealand Black (NZB) mice are defective in susceptibility to tolerance induced by deaggregated bovine γ globulin (DBGG). To examine whether this defect is related to the loss of self-tolerance in autoimmunity, susceptibility loci for this defect were examined by genome-wide analysis using the F(2) intercross of nonautoimmune C57BL/6 (B6) and NZB mice. One NZB locus on the telomeric chromosome 1, designated Dit (Defective immune tolerance)-1, showed a highly significant linkage. This locus overlapped with a locus containing susceptibility genes for autoimmune disease, namely Fcgr2b and Slam family genes. To investigate the involvement of these genes in the defective tolerance to DBGG, we took advantage of two lines of Fcgr2b-deficient B6 congenic mice: one carries autoimmune-type, and the other carries B6-type, Slam family genes. Defective tolerance was observed only in Fcgr2b-deficient mice with autoimmune-type Slam family genes, indicating that epistatic effects of both genes are involved. Thus, common genetic mechanisms may underlie the defect in foreign protein antigen-induced tolerance and the loss of self-tolerance in NZB mouse-related autoimmune diseases.

Vascular, renal and placental effects on pregnant offspring of protein-restricted rat dams.

AIM: Our previous study showed that a maternal low-protein diet induced hypertension and vascular dysfunction in rat offspring after day 175. In the present study, we hypothesized that these female offspring would develop hypertension in their own pregnancies even at ages less than 175 days because potential vascular dysfunction is exacerbated by the circulatory demands of pregnancy. MATERIAL AND METHODS: Wistar rats were fed an isocaloric diet containing either 18% (control group) or 9% (low-protein group) casein throughout pregnancy. The female offspring were fed standard chow and mated between days 70 and 125. At the end of pregnancy, blood pressure was measured, and the uterine arteries were dissected and investigated with a wire myograph. RESULTS: Placental weights were significantly lower in offspring of the low-protein group versus control. There were no significant differences in blood pressure. Renal expression of AT1 receptor mRNA was greater, and of AT2 receptor was less, in the low-protein group versus control. Vasoconstriction of uterine arteries to phenylephrine and U46619 was increased in the low-protein group, and vasodilatation to sodium nitroprusside was also increased. CONCLUSION: Low-protein diet induces vascular effects on female offspring in their pregnancy, in terms of increased uterine artery vasoconstriction. This may be compensated for by increased sensitivity to nitric oxide (NO), maintaining blood pressure normal in the face of the demands of pregnancy. Such renal and vascular effects, combined with placental size, may transmit risk of vascular dysfunction to subsequent generations.

FTY720 exerts a survival advantage through the prevention of end-stage glomerular inflammation in lupus-prone BXSB mice.

FTY720 is a novel investigational agent targeting the sphingosine 1-phosphate (S1P) receptors with an ability to cause immunosuppression by inducing lymphocyte sequestration in lymphoid organs. Systemic lupus erythematosus (SLE) is refractory autoimmune disease characterized by the production of a wide variety of autoantibodies and immune complex (IC)-mediated lupus nephritis. Among several SLE-prone strains of mice, BXSB is unique in terms of the disease-associated monocytosis in periphery and the reduced frequency of marginal zone B (MZ B) cells in spleen. In the present study, we examined the effect of FTY720 on lupus nephritis of BXSB mice. FTY720 treatment resulted in a marked decrease in lymphocytes, but not monocytes, in peripheral blood, and caused relocalization of marginal zone B (MZ B) cells into the follicle in the spleen. These changes did not affect the production of autoantibodies, thus IgG and C3 were deposited in glomeruli in FTY720-treated mice. Despite these IC depositions, FTY720-treated mice showed survival advantage with the improved proteinuria. Histological analysis revealed that FTY720 suppressed mesangial cell proliferation and inflammatory cell infiltration. These results suggest that FTY720 ameliorates lupus nephritis by inhibiting the end-stage inflammatory process following IC deposition in glomeruli.

Inhibitory IgG Fc receptor promoter region polymorphism is a key genetic element for murine systemic lupus erythematosus.

The autoimmune-type Fcgr2b with deletion polymorphism in AP-4-binding site in the promoter region is suggested to be one most plausible susceptibility gene for systemic lupus erythematosus (SLE). We previously found that there is a strong epistatic interaction between the autoimmune-type Fcgr2b polymorphism and Y chromosome-linked autoimmune acceleration (Yaa) mutation, thus severe SLE observed in BXSB males neither develops in BXSB females nor in the congenic BXSB.IIB(B6) males carrying wild C57BL/6-type Fcgr2b. Present studies examined whether the wild-type Fcgr2b could suppress SLE in mice carrying Yaa-unrelated SLE susceptibility genes. Comparison of disease features between SLE-prone (NZW x BXSB) F1 females and the congenic (NZW x BXSB.IIB(B6)) F1 females carrying wild-type Fcgr2b showed that, as compared with findings in the former, SLE features including activation/proliferation of not only B cells but also T cells and monocytes/macrophages were all inhibited in the latter. It was concluded that the autoimmune-type Fcgr2b promotes and the wild-type inhibits SLE through mechanisms that promote and suppress activation/proliferation of a wide variety of immune cells, respectively. Thus, the Fcgr2b polymorphism is a key genetic element for not only Yaa-related but also Yaa-unrelated lupus.

Ankylosing enthesitis associated with up-regulated IFN-gamma and IL-17 production in (BXSB x NZB) F(1) male mice: a new mouse model.

We found that in contrast to (BXSB x NZB) F(1) female mice that spontaneously develop severe systemic lupus erythematosus (SLE), male (BXSB x NZB) F(1) mice are not prone to SLE, but instead develop seronegative ankylosing enthesitis in ankle/tarsal joints only when caged in groups, with the incidence reaching 83% at 7 months of age. This ankylosis is microscopically characterized by a marked proliferation of fibroblast-like cells positive for bone morphogenetic protein (BMP)-2 in association with heterotropic formation of cartilages and bones in hyperplastic entheseal tissues and subsequent fusion of tarsal bones. Elevated potentials of popliteal lymph node T cells producing interleukin (IL)-17 and interferon (IFN)-gamma were significantly associated with joint ankylosis, suggesting the involvement of these cytokines in effector phase mechanisms of the disease, including up-regulated BMP signaling pathways. There was no difference in serum autoantibody levels between affected and unaffected mice. Parental BXSB and NZB strains of both sexes did not develop the disease even when caged in groups, indicating that the disease develops under the control of susceptibility genes derived from both parental strains. These results indicate that (BXSB x NZB) F(1) male mice are a suitable model for clarifying genetic, environmental and molecular mechanisms underlying ankylosing enthesitis and related diseases.

Centromeric interval of chromosome 4 derived from C57BL/6 mice accelerates type 1 diabetes in NOD.CD72b congenic mice.

The nonobese diabetic (NOD) mouse is a useful model of autoimmune type 1 diabetes exhibiting many similarities to human type 1 diabetes patients including the presence of auto-reactive T cells and pancreas-specific autoantiboies. Multiple Idd loci control the development of diabetes in NOD mice. CD72, a B cell membrane-bound glycoprotein carrying a C-type lectin-like domain, is an inhibitory co-receptor of the B cell antigen receptor (BCR) that negatively regulates BCR signaling. Among four known haplotypes of mouse CD72, NOD mice carry the CD72(c) haplotype, whereas most of the other inbred strains of mice carry either CD72(a) or CD72(b). In this study, we generated congenic NOD.CD72(b) mice that carry C57BL/6 (B6) mouse-derived centromeric chromosome 4 interval (24-45cM) surrounding the CD72(b) locus. Unexpectedly, NOD.CD72(b) mice were not protected from diabetes, but rather exhibited accelerated development of both insulitis and diabetes. Our result defines novel locus or loci in the vicinity of CD72 gene that negatively control diabetes, indicating that NOD disease is under complex genetic controls of not only Idd genes but also disease-resistant genes.

Aberrant genetic control of invariant TCR-bearing NKT cell function in New Zealand mouse strains: possible involvement in systemic lupus erythematosus pathogenesis.

Both suppressive and promoting roles of NKT cells have been reported in the pathogenesis of systemic lupus erythematosus (SLE). Herein, we found that although New Zealand mice have normal frequencies of NKT cells, their in vitro potential to produce IL-4 and IFN-gamma in response to alpha-galactosylceramide was remarkably impaired in New Zealand Black (NZB) mice prone to mild SLE, while production was highly up-regulated in nonautoimmune New Zealand White (NZW) mice and at intermediate levels in (NZB x NZW)F(1) mice, which are prone to severe SLE. Because this aberration is evident in young mice before disease onset, genetic mechanisms are thought to be involved. Genome-wide quantitative trait locus analysis and association studies revealed that a locus linked to D11Mit14 on chromosome 11 may be involved in the difference in cytokine-producing potential between NZB and NZW NKT cells. Additionally, (NZB x NZW)F(1) x NZB backcross progeny with the NZW genotype for D11Mit14 showed significantly increased frequencies of age-associated SLE phenotypes, such as high serum levels of IgG, IgG anti-DNA Abs, and lupus nephritis. In coculture studies, alpha-galactosylceramide-stimulated NKT cells from NZW and (NZB x NZW)F(1) mice, but not from NZB mice, showed significantly enhanced Ig synthesis by B cells. These findings suggest that the D11Mit14-linked NZW locus may contribute to the development of SLE in (NZB x NZW)F(1) mice through a mechanism that up-regulates NKT cell function. Thus, this NZW allele may be a candidate of the NZW modifiers that act to promote (NZB x NZW)F(1) disease.

CD3 and immunoglobulin G Fc receptor regulate cerebellar functions.

The immune and nervous systems display considerable overlap in their molecular repertoire. Molecules originally shown to be critical for immune responses also serve neuronal functions that include normal brain development, neuronal differentiation, synaptic plasticity, and behavior. We show here that FcgammaRIIB, a low-affinity immunoglobulin G Fc receptor, and CD3 are involved in cerebellar functions. Although membranous CD3 and FcgammaRIIB are crucial regulators on different cells in the immune system, both CD3epsilon and FcgammaRIIB are expressed on Purkinje cells in the cerebellum. Both CD3epsilon-deficient mice and FcgammaRIIB-deficient mice showed an impaired development of Purkinje neurons. In the adult, rotarod performance of these mutant mice was impaired at high speed. In the two knockout mice, enhanced paired-pulse facilitation of parallel fiber-Purkinje cell synapses was shared. These results indicate that diverse immune molecules play critical roles in the functional establishment in the cerebellum.

IL-4Ralpha polymorphism in regulation of IL-4 synthesis by T cells: implication in susceptibility to a subset of murine lupus.

To thoroughly understand the role of IL-4 in the pathogenesis of systemic lupus erythematosus (SLE), a prototypic antibody-mediated systemic autoimmune disease, we examined the potential of in vitro IL-4 production by anti-CD3 mAb-stimulated splenic T cells in SLE model of NZB, BXSB and related mouse strains. Unexpectedly, both SLE-prone NZB and BXSB mice had a limited potential to produce IL-4, while disease-free NZW mice had a high potential. Levels in (NZB x NZW) F1 and (NZW x BXSB) F1 were in between. Genome-wide search for quantitative trait loci (QTL) controlling this variation identified a single significant QTL in the vicinity of IL-4Ralpha gene on chromosome 7. Sequence analysis of IL-4Ralpha cDNA revealed that there are 17 nucleotide substitutions resulting in eight amino acid changes between NZB and NZW strains. BXSB showed the identical sequence, as did NZB. Thus, it was suggested that the NZW-type polymorphism controls a high potential and the NZB/BXSB-type polymorphism controls a low potential for IL-4 production by T cells. Linkage studies using NZW x (NZW x BXSB) F1 male and (NZB x NZW) F1 x NZW female back-cross mice revealed that the BXSB/NZB-type IL-4Ralpha polymorphism significantly linked to BXSB, but not to (NZB x NZW) F1 lupus. Thus, the low IL-4-producing phenotype appears to predispose to SLE in BXSB, but not NZB-related strains, suggesting that the role of IL-4 in the pathogenesis may differ between certain subsets of SLE, even if they show similar disease phenotypes.

Requirement of tryptophan hydroxylase during development for maturation of sensorimotor gating.

Deficits in sensorimotor gating, a function to focus on the most salient stimulus, could lead to a breakdown of cognitive integrity, and could reflect the "flooding" by sensory overload and cognitive fragmentation seen in schizophrenia. Sensorimotor gating emerges at infancy, and matures during childhood. The mechanisms that underlie its development are largely unclear. Here, we screened the mouse genome, and found that tryptophan hydroxylase (TPH) is implicated in the maturation of sensorimotor gating. TPH, an enzyme involved in the biosynthesis of serotonin, proved to be required only during the weaning period for maturation of sensorimotor gating, but was dispensable for its emergence. Proper serotonin levels during development underlie the mature functional architecture for sensorimotor gating via appropriate actin polymerization. Thus, maintaining proper serotonin levels during childhood may be important for mature sensorimotor gating in adulthood.