Splenic phagocytes promote responses to nucleosomes in (NZB x NZW) F1 mice

Autoantigen presentation to T cells is crucial for the development of autoimmune disease. However, the mechanisms of autoantigen presentation are poorly understood. In this study, we show that splenic phagocytes play an important role in autoantigen presentation in murine lupus. Nucleosomes are major autoantigens in systemic lupus erythematosus. We found that nucleosome-specific T cells were stimulated dominantly in the spleen, compared with lymph nodes, lung, and thymus. Among splenic APCs, F4/80(+) macrophages and CD11b(+)CD11c(+) dendritic cells were strong stimulators for nucleosome-specific T cells. When splenic phagocytes were depleted in (NZB x NZW) F(1) (NZB/W F(1)) mice, nucleosome presentation in the spleen was dramatically suppressed. Moreover, depletion of splenic phagocytes significantly suppressed anti-nucleosome Ab and anti-dsDNA Ab production. Proteinuria progression was delayed and survival was prolonged in phagocyte-depleted mice. The numbers of autoantibody- secreting cells were decreased in the spleen from phagocyte-depleted mice. Multiple injections of splenic F4/80(+) macrophages, not those of splenic CD11c(+) dendritic cells, induced autoantibody production and proteinuria progression in NZB/W F(1) mice. These results indicate that autoantigen presentation by splenic phagocytes including macrophages significantly contributes to autoantibody production and disease progression in lupus-prone mice.     

Immunologic abnormality in NZB/NZW F1 mice. Thymus-independent occurrence of B cell abnormality and requirement for T cells in the development of autoimmune disease, as evidenced by an analysis of the athymic nude individuals

Both NZB nu/+ and NZW nu/+ mice were microbially clean by cesarean section. The (NZB x NZW)F1 hybrid (NZB/W) nu/nu mice and nu/+ littermates were then generated by mating of NZB nu/+ with NZW nu/+mice under specific pathogen-free conditions. The female NZB/W F1 nu/nu mice did not develop autoimmune kidney disease, whereas all of nu/+ female littermates mice exhibited proteinuria and died of renal failure with a 50% survival time of 35 wk. Namely, nude mice had no signs of proteinuria up to the time of their death caused by other diseases rather than glomerulonephritis, and their mean survival time was greater than 45 wk. Nude mice had also no anti-ssDNA antibody in their serum. However, splenic B cells of NZB/W nude mice exhibited hyper-responsiveness to both LPS and B151-TRF2, a T cell-derived polyclonal B cell-stimulation factor, and produced large numbers of Ig-secreting cells and anti-TNP plaque-forming cells as well as anti-ssDNA antibody comparable to the nu/+ littermate mice. Interestingly, thymus-engrafted NZB/W nude mice developed autoimmune disease exemplified by the induction of anti-ssDNA antibody and proteinuria at approximately the same time as their nu/+ littermates. These results indicate that the B cell hyper-responsiveness found in NZB/W mice is apparently determined by the T cell-independent process, and T cells are obligatorily required for the development of autoimmune disease in NZB/W mice.     

Ig-reactive CD4+CD25+ T cells from tolerized (New Zealand Black x New Zealand White)F1 mice suppress in vitro production of antibodies to DNA

We have recently shown that tolerogenic administration of an artificial peptide (pConsensus) that is based on sequences within the V(H) regions of several murine anti-dsDNA Ig delays appearance of autoantibodies in female (New Zealand Black (NZB) x New Zealand White (NZW))F(1) (NZB/W F(1)) mice and significantly prolongs their survival. The aim of this study was to characterize the T cell population(s) involved in pConsensus-induced down-regulation of autoimmune responses in tolerized NZB/W F(1) mice. Using MHC class II dimers loaded with tolerogenic peptide, we found that pCons favored expansion of peptide-reactive CD4(+)CD25(+) regulatory T cells (T(R)) that inhibited in vitro production of anti-dsDNA Ab-forming cells. Suppression by T(R) was abrogated by the presence in culture of Ab to glucocorticoid-induced TNFR family member 18 or to TGFbeta latency-associated protein. These findings suggest possible relevance of Ag specificity in the mechanism of T(R)-mediated immune tolerance to Ig-derived peptides in NZB/W F(1) mice.     

Very low-dose tolerance with nucleosomal peptides controls lupus and induces potent regulatory T cell subsets

We induced very low-dose tolerance by injecting lupus prone (SWR x NZB)F1 (SNF1) mice with 1 mug nucleosomal histone peptide autoepitopes s.c. every 2 wk. The subnanomolar peptide therapy diminished autoantibody levels and prolonged life span by delaying nephritis, especially by reducing inflammatory cell reaction and infiltration in kidneys. H4(71-94) was the most effective autoepitope. Low-dose tolerance therapy induced CD8+, as well as CD4+ CD25+ regulatory T (Treg) cell subsets containing autoantigen-specific cells. These adaptive Treg cells suppressed IFN-gamma responses of pathogenic lupus T cells to nucleosomal epitopes at up to a 1:100 ratio and reduced autoantibody production up to 90-100% by inhibiting nucleosome-stimulated T cell help to nuclear autoantigen-specific B cells. Both CD4+ CD25+ and CD8+ Treg cells produced and required TGF-beta1 for immunosuppression, and were effective in suppressing lupus autoimmunity upon adoptive transfer in vivo. The CD4+ CD25+ T cells were partially cell contact dependent, but CD8+ T cells were contact independent. Thus, low-dose tolerance with highly conserved histone autoepitopes repairs a regulatory defect in systemic lupus erythematosus by generating long-lasting, TGF-beta-producing Treg cells, without causing allergic/anaphylactic reactions or generalized immunosuppression.     

Suppression of disease in New Zealand Black/New Zealand White lupus-prone mice by adoptive transfer of ex vivo expanded regulatory T cells

An increasing number of studies indicate that a subset of CD4(+) T cells with regulatory capacity (regulatory T cells; T(regs)) can function to control organ-specific autoimmune disease. To determine whether abnormalities of thymic-derived T(regs) play a role in systemic lupus erythematosus, we evaluated T(reg) prevalence and function in (New Zealand Black x New Zealand White)F(1) (B/W) lupus-prone mice. To explore the potential of T(regs) to suppress disease, we evaluated the effect of adoptive transfer of purified, ex vivo expanded thymic-derived T(regs) on the progression of renal disease. We found that although the prevalence of T(regs) is reduced in regional lymph nodes and spleen of prediseased B/W mice compared with age-matched non-autoimmune mice, these cells increase in number in older diseased mice. In addition, the ability of these cells to proliferate in vitro was comparable to those purified from non-autoimmune control animals. Purified CD4(+)CD25(+)CD62L(high) B/W T(regs) were expanded ex vivo 80-fold, resulting in cells with a stable suppressor phenotype. Adoptive transfer of these exogenously expanded cells reduced the rate at which mice developed renal disease; a second transfer after treated animals had developed proteinuria further slowed the progression of renal disease and significantly improved survival. These studies indicate that thymic-derived T(regs) may have a significant role in the control of autoimmunity in lupus-prone B/W mice, and augmentation of these cells may constitute a novel therapeutic approach for systemic lupus erythematosus.     

Antigen-specific therapy of murine lupus nephritis using nucleosomal peptides: tolerance spreading impairs pathogenic function of autoimmune T and B cells

In the (SWR x NZB)F1 mouse model of lupus, we previously localized the critical autoepitopes for nephritogenic autoantibody-inducing Th cells in the core histones of nucleosomes at aa positions 10-33 of H2B and 16-39 and 71-94 of H4. A brief therapy with the peptides administered i.v. to 3-mo-old prenephritic (SWR x NZB)F1 mice that were already producing pathogenic autoantibodies markedly delayed the onset of severe lupus nephritis. Strikingly, chronic therapy with the peptides injected into 18-mo-old (SWR x NZB)F1 mice with established glomerulonephritis prolonged survival and even halted the progression of renal disease. Remarkably, tolerization with any one of the nucleosomal peptides impaired autoimmune T cell help, inhibiting the production of multiple pathogenic autoantibodies. However, cytokine production or proliferative responses to the peptides were not grossly changed by the therapy. Moreover, suppressor T cells were not detected in the treated mice. Most interestingly, the best therapeutic effect was obtained with nucleosomal peptide H416-39, which had a tolerogenic effect not only on autoimmune Th cells, but autoimmune B cells as well, because this peptide contained both T and B cell autoepitopes. These studies show that the pathogenic T and B cells of lupus, despite intrinsic defects in activation thresholds, are still susceptible to autoantigen-specific tolerogens.     

Effects of Cystamine on antioxidant activities and regulatory T cells in lupus‐prone mice

Attenuated antioxidant activities, irregular cytokines expressions and reduced regulatory T cells, are strongly associated with the pathogenesis of systemic lupus erythematosus (SLE). Despite the well‐established beneficial effects of cystamine on lupus‐prone mice, the extent to which cystamine contributes to antioxidant activity and the reduction of regulatory T cells has seldom been investigated. Therefore, this study elucidates how cystamine affects anti‐oxidant activities in NZB/W F1 mice by performing assays of Glutathione (GSH), 1,1‐diphenyl‐2‐ picryl‐hydrazyl (DPPH) and malondialdehyde thiobarbituric acid (MDA). In addition, investigations of the effects of cystamine on CD4⁺/CD25⁺ regulatory T cells and interleukin‐6 (IL6)/STAT‐3 signalling were performed with flow cytometry and immunoblots. Experimental results reveal more significantly reduced MDA and increased GSH and DPPH in NZB/W F1 mice receiving cystamine than in those mice receiving PBS. Meanwhile, CD4⁺/CD25⁺ regulatory T cells more significantly increase in NZB/W F1 mice receiving cystamine than in those mice receiving PBS, accompanied by significantly reduced IL‐6/phosphorylated STAT‐3 expression. The above findings suggest the beneficial effects of cystamine in terms of increasing antioxidant activities and CD4⁺/CD25⁺ regulatory T cells in lupus‐prone mice by suppressing IL‐6/STAT3 signalling.     

Transferred B cells from autoimmune NZB/N mice fail to activate T suppressor cells

T-cell-mediated suppression of the antibody response of autoimmune NZB/N mice to Type III pneumococcal polysaccharide (SSS-III) can readily be induced in situ by priming with a subimmunogenic dose of SSS-III; however, the transfer of either "young" (8 weeks old) or "old" (42 weeks old) SSS-III-primed B cells, which activates suppressor T cells in normal BALB/cByJ mice, fails to induce suppression of the antibody response in recipient NZB/N mice, regardless of the number of cells transferred or the time interval between transfer and immunization. Transfer of 51Cr-labeled B cells demonstrated that syngeneic primed B cells home to the spleens of NZB/N mice in somewhat lower numbers than in BALB/cByJ mice, although the differences observed may not be sufficient to explain the complete absence of activation of suppressor T cells. These findings suggest that B cells from autoimmune NZB/N mice are unable to activate T suppressor cells upon transfer; this disorder in a normal regulatory mechanism may be important in the pathogenesis of disease.     

Class-specific regulation of anti-DNA antibody synthesis and the age-associated changes in (NZB x NZW)F1 hybrid mice

Investigations of regulatory helper and suppressor T cells in the in vitro anti-DNA antibody synthesis in NZB x NZW (B/W) F1 hybrid mice were initiated by the development of an in vitro system in which G10-passed B cells from B/W F1 mice were cocultured with mitomycin C-treated T cells in the presence of Con A and either in the presence or in the absence of LPS. It was revealed that each IgG and IgM anti-DNA antibody synthesis was under the regulation of separate L3T4+ helper and Ly-2+ suppressor T cells. The function of these class-specific regulatory T cells was age-dependent. Although the helper effect of L3T4+ T cells on IgG antibody synthesis increased, the effect of L3T4+ T cells on IgM antibody production decreased in B/W F1 mice with aging. The IgG anti-DNA antibody production in the cocultures of L3T4+ T cells and B cells was suppressed by addition of Ly-2+ T cells from young but not aged B/W F1 mice, whereas the production of IgM anti-DNA antibodies was suppressed by Ly-2+ T cells from aged but not young B/W F1 mice. We also found that although IgM anti-DNA antibody-producing B cells were already present in 2-mo-old mice, B cells producing IgG antibodies under the influence of L3T4+ T cells appeared in mice at 7 mo of age. These data clearly indicate that separate class-specific regulatory T cells are involved in the production of IgM and IgG anti-DNA antibodies and that the total serum level of the antibodies is reflected by both their age-associated changes and the generation of antibody-forming B cells in B/W F1 mice.     

Mechanism of action of transmembrane activator and calcium modulator ligand interactor-Ig in murine systemic lupus erythematosus

B cell-activating factor belonging to the TNF family (BAFF) blockade prevents the onset of disease in systemic lupus erythematosus (SLE)-prone NZB/NZW F(1) mice. To determine the mechanism of this effect, we administered a short course of TACI-Ig with and without six doses of CTLA4-Ig to 18- to 20-wk-old NZB/NZW F(1) mice and evaluated the effect on B and T cell subsets and on anti-dsDNA Ab-producing B cells. Even a brief exposure to TACI-Ig had a beneficial effect on murine SLE; CTLA4-Ig potentiated this effect. The combination of TACI-Ig and CTLA4-Ig resulted in a temporary decrease in serum IgG levels. However, after cessation of treatment, high titers of IgG anti-dsDNA Abs appeared in the serum and IgG Abs deposited in the kidneys. Despite the appearance of pathogenic autoantibodies, the onset of proteinuria was markedly delayed; this was associated with prolonged depletion of B cells past the T1 stage, a decrease in the size of the spleen and lymph nodes, and a decrease in the absolute number of activated and memory CD4(+) T cells. TACI-Ig treatment normalized serum levels of IgM that are markedly elevated in NZB/W F(1) mice; this appeared to be due to a prolonged effect on the ability of the splenic microenvironment to support short-lived IgM plasma cells. Finally, a short course of combination TACI-Ig and CTLA4-Ig prolonged life and even reversed proteinuria in aged NZB/W F(1) mice, suggesting that BAFF blockade may be an effective therapeutic strategy for active SLE.     

Tolerance defects in New Zealand Black and New Zealand Black X New Zealand White F1 mice

The susceptibility of autoimmune NZB and (NZB X NZW)F1 mice to the induction of tolerance by monomeric BSA was compared with several normal mouse strains. Unresponsiveness in T and B lymphocyte compartments was probed by challenging with DNP8BSA and measuring anti-DNP and anti-BSA antibodies separately. Tolerance induced by monomeric BSA was carrier specific, and there was no evidence of epitope-specific suppression. Normal NZW, NFS, and B10.D2 mice were easily rendered tolerant with monomeric BSA and did not produce anti-DNP or anti-BSA antibodies after challenge with DNP8BSA. By contrast, the lack of anti-DNP antibody response in similarly treated NZB mice was dependent on the dose of monomeric BSA, indicating that the helper T cells were partially resistant to tolerance induction. NZB mice treated with a high dose of monomeric BSA produced anti-BSA, but not anti-DNP, antibodies after immunization. Thus, the anti-carrier B cells in NZB mice may have been primed by monomeric BSA. The presence of the xid gene on the NZB background rendered the mice susceptible to induction of tolerance, suggesting that the tolerance defect in NZB mice involves the B cell compartment. This abnormal antibody response was a dominant trait: (NZB X NFS)F1 and (NZB X B10.D2)F1 mice had the same characteristics as NZB mice. These F1 hybrids do not develop autoimmune disease, indicating that resistance to experimental tolerance induction expressed at a B cell level may not be sufficient for disease development. In contrast to NZB and other NZB F1 hybrids, (NZB X NZW)F1 hybrids treated with monomeric BSA and challenged with DNP8BSA responded to both DNP and BSA. The contribution of a B cell defect to the tolerance abnormality of (NZB X NZW)F1 mice was examined by analyzing the effect of the xid gene on the progeny of (NZB.xid X NZW)F1 mice. Unlike the effect of the xid gene on NZB mice, both phenotypically normal heterozygous female and phenotypically xid hemizygous male mice produced anti-DNP and anti-BSA antibodies after tolerance induction and immunization, demonstrating that a major helper T cell abnormality was present in (NZB X NZW)F1 mice. The (NZW X B10.D2)F1 hybrid was rendered tolerant by this procedure, indicating that the helper T cell defect (NZB X NZW)F1 mice may have resulted from gene complementation with the NZB mice contributing partial resistance of T helper cells to tolerance induction.(ABSTRACT TRUNCATED AT 400 WORDS)     

Immunotherapy with costimulatory dendritic cells to control autoimmune inflammation

Costimulation-deficient dendritic cells (DCs) prevent autoimmune disease in mouse models. However, autoimmune-prone mice and humans fail to control expansion of peripheral autoreactive effector memory T cells (T(EMs)), which resist immunoregulation by costimulation-deficient DCs. In contrast, activation of DC costimulation may be coupled with regulatory capacity. To test whether costimulatory DCs control T(EMs) and attenuate established autoimmune disease, we used RelB-deficient mice, which have multiorgan inflammation, expanded peripheral autoreactive T(EMs), and dysfunctional Foxp3(+) regulatory T cells (Tregs) cells and conventional DCs. T(EMs) were regulated by Foxp3(+) Tregs when costimulated by CD3/CD28-coated beads or wild-type DCs but not DCs deficient in RelB or CD80/CD86. After transfer, RelB and CD80/CD86-sufficient DCs restored tolerance and achieved a long-term cure of autoimmune disease through costimulation of T(EM) and Foxp3(+) Treg IFN-γ production, as well as induction of IDO by host APCs. IDO was required for regulation of T(EMs) and suppression of organ inflammation. Our data challenge the paradigm that costimulation-deficient DCs are required to regulate established autoimmune disease to avoid T(EM) activation and demonstrate cooperative cross-talk between costimulatory DCs, IFN-γ, and IDO-dependent immune regulation. IFN-γ and IDO activity may be good surrogate biomarkers measured against clinical efficacy in trials of autoimmune disease immunoregulation.     

Germinal Center B Cell Depletion Diminishes CD4+ Follicular T Helper Cells in Autoimmune Mice

Background

Continuous support from follicular CD4⁺ T helper (Tfh) cells drives germinal center (GC) responses, which last for several weeks to produce high affinity memory B cells and plasma cells. In autoimmune Sle1 and NZB/W F1 mice, elevated numbers of Tfh cells persist, promoting the expansion of self-reactive B cells. Expansion of circulating Tfh like cells have also been described in several autoimmune diseases. Although, the signals required for Tfh differentiation have now been well described, the mechanisms that sustain the maintenance of fully differentiated Tfh are less understood. Recent data demonstrate a role for GC B cells for Tfh maintenance after protein immunization.

Methods and Finding

Given the pathogenic role Tfh play in autoimmune disease, we explored whether B cells are required for maintenance of autoreactive Tfh. Our data suggest that the number of mature autoreactive Tfh cells is controlled by GC B cells. Depletion of B cells in Sle1 autoimmune mice leads to a dramatic reduction in Tfh cells. In NZB/W F1 autoimmune mice, similar to the SRBC immunization model, GC B cells support the maintenance of mature Tfh, which is dependent mainly on ICOS. The CD28-associated pathway is dispensable for Tfh maintenance in SRBC immunized mice, but is required in the spontaneous NZB/W F1 model.

Conclusion

These data suggest that mature Tfh cells require signals from GC B cells to sustain their optimal numbers and function in both autoimmune and immunization models. Thus, immunotherapies targeting B cells in autoimmune disease may affect pathogenic Tfh cells.     

Expansion and hyperactivity of CD1d-restricted NKT cells during the progression of systemic lupus erythematosus in (New Zealand Black x New Zealand White)F1 mice

CD1d-restricted NKT cells expressing invariant TCR alpha-chain rearrangements (iNKT cells) have been reported to be deficient in humans with a variety of autoimmune syndromes and in certain strains of autoimmune mice. In addition, injection of mice with alpha-galactosylceramide, a specific glycolipid agonist of iNKT cells, activates these T cells and ameliorates autoimmunity in several different disease models. Thus, deficiency and reduced function in iNKT cells are considered to be risk factors for the development of such diseases. In this study we report that the development of systemic lupus erythematosus in (New Zealand Black (NZB) x New Zealand White (NZW))F(1) mice was paradoxically associated with an expansion and activation of iNKT cells. Although young (NZB x NZW)F(1) mice had normal levels of iNKT cells, these expanded with age and became phenotypically and functionally hyperactive. Activation of iNKT cells in (NZB x NZW)F(1) mice in vivo or in vitro with alpha-galactosylceramide indicated that the immunoregulatory role of iNKT cells varied over time, revealing a marked increase in their potential to contribute to production of IFN-gamma with advancing age and disease progression. This evolution of iNKT cell function during the progression of autoimmunity may have important implications for the mechanism of disease in this model of systemic lupus erythematosus and for the development of therapies using iNKT cell agonists.     

Autoimmunity stimulated by adoptively transferred dendritic cells is initiated by both alphabeta and gammadelta T cells but does not require MyD88 signaling

Vaccination of nonautoimmune prone mice with syngeneic dendritic cells (DC) readily induces anti-DNA autoantibodies but does not trigger systemic disease. We observed that anti-DNA autoantibody generation absolutely required alphabeta T cells and that gammadelta T cells also contributed to the response, but that regulatory T cells restrained autoantibody production. Although both NZB/W F(1) mice and DC vaccinated C57/BL6 mice produced autoantibodies against dsDNA, vaccinated mice had higher levels of Abs against H1 histone and lower levels of antinucleosome Abs than NZB/W F(1) mice. Despite a 100-fold increase in IL-12 and Th1 skewing to a foreign Ag, OVA, synergistic TLR activation of DC in vitro failed to augment anti-DNA Abs or promote class switching beyond that induced by LPS alone. TLR stimulation was not absolutely required for the initial loss of B cell tolerance because anti-DNA levels were similar when wild-type (WT) or MyD88-deficient DC were used for vaccination or WT and MyD88-deficient recipients were vaccinated with WT DC. In contrast, systemic administration of LPS, augmented anti-DNA Ab levels and promoted class switching, and this response was dependent on donor DC signaling via MyD88. LPS also augmented responses in the MyD88-deficient recipients, suggesting that LPS likely exerts its effects on both transferred DC and host B cells in vivo. These results indicate that both the alphabeta and gammadelta subsets are necessary for promoting autoantibody production by DC vaccination, and that although TLR/MyD88 signaling is not absolutely required for initiation, this pathway does promote augmentation, and Th1-mediated skewing, of anti-DNA autoantibodies.     

Responses of B cells from autoimmune mice to IL-5

Three strains of mice (NZB/W F1 X NZW (NZB/W), BXSB, and MRL/Mp-lpr/lpr (MRL/lpr] develop an autoimmune disease that is clinically and immunologically similar to human SLE. A characteristic of these mice is polyclonal B cell hyperactivity. To explore whether this may be related to hyper-responsiveness to B cell stimulatory factors, we investigated the proliferative and secretory responses of B cells from these mice to semi-purified natural and rIL-5, a major regulator of B cell development in the mouse. As this lymphokine stimulates growth and differentiation of activated B cells, attention was focused on in vivo-activated B cell populations, obtained from the interface of 50/65% Percoll density gradients, from normal or autoimmune mice. This cell population from NZB/W mice secreted IgM and incorporated [3H]TdR at significantly higher levels in response to IL-5, and was more sensitive to IL-5, than a comparable population from several normal murine strains. NZB/W female and male mice displayed heightened responses to IL-5, indicating that this is characteristic of the strain in general and is not associated with the accelerated severe disease of the females. Small resting B cells from NZB/W and normal mice were insensitive to IL-5 stimulation. In contrast to NZB/W mice, no difference was observed in the magnitude of either proliferative or Ig secretory responses between in vivo-activated B cell populations from autoimmune BXSB and MRL/lpr or normal mice. Thus, B cell hyper-responsiveness to IL-5 is a characteristic of NZB/W mice but not of two other lupus-prone murine strains. As one unique feature of NZB/W mouse B cells compared to normal and other autoimmune B cells is an elevated proportion of Ly-1+ B cells, the possibility of IL-5 hyper-responsiveness being associated with this B cell subpopulation was investigated. Fluorescence-activated cell sorter sorted Ly-1+ and Ly-1- B cells both responded to IL-5, however Ly-1+ B cells consistently showed a higher stimulation index in both proliferative and Ig secretory responses to this lymphokine.     

Atorvastatin inhibits autoreactive B cell activation and delays lupus development in New Zealand black/white F1 mice

Systemic lupus erythematosus is a multisystem autoimmune disease characterized by a wide range of immunological abnormalities that underlie the loss of tolerance. In this study we show that administration of atorvastatin to lupus-prone NZB/W F(1) mice resulted in a significant reduction in serum IgG anti-dsDNA Abs and decreased proteinuria. Histologically, the treatment was associated with reduced glomerular Ig deposition and less glomerular injury. Disease improvement was paralleled by decreased expression of MHC class II on monocytes and B lymphocytes and reduced expression of CD80 and CD86 on B lymphocytes. Consequent upon this inhibition of Ag presentation, T cell proliferation was strongly impaired by atorvastatin in vitro and in vivo. A significant decrease in MHC class II expression was also observed in the target organ of lupus disease (i.e., the glomerulus). Serum cholesterol in atorvastatin-treated lupus mice fell to the level found in young NZB/W mice before disease onset. This is the first demonstration that atorvastatin can delay the progression of a spontaneous autoimmune disease and may specifically benefit patients with systemic lupus erythematosus.     

Immature dendritic cells suppress collagen-induced arthritis by in vivo expansion of CD49b+ regulatory T cells

Dendritic cells (DCs) are specialized APCs with an important role in the initiation and regulation of immune responses. Immature DCs (iDCs) reportedly mediate tolerance in the absence of maturation/inflammatory stimuli, presumably by the induction of regulatory T cells. In this study, we show for the first time that repetitive iDC injections trigger the expansion of a novel regulatory population with high immunomodulatory properties, able to protect mice from collagen-induced arthritis. These regulatory T cells are characterized by the expression of the CD49b molecule and correspond to a CD4+ alpha-galactosylceramide/CD1d-nonrestricted T cell population producing IL-10. Adoptive transfer of < 10(5) TCRbeta+ CD49b+ cells isolated from the liver of iDCs-vaccinated mice, conferred a complete protection against arthritis. This protection was associated with an attenuation of the B and T cell response associated with a local secretion of IL-10. Thus, together these data demonstrate that iDCs can expand and activate a novel regulatory population of CD49b+ T cells, with high immunosuppressive potential able to mediate protection against a systemic autoimmune disease.     

Cellular basis of in vitro anti-DNA antibody production: evidence for T cell dependence of IgG-class anti-DNA antibody synthesis in the (NZB X NZW)F1 hybrid

An in vitro system was designed to measure anti-DNA antibody synthesis, and the cellular basis of this autoantibody production in NZB X NZW (B/W)F1 (B/W F1) mice was analyzed. The spleen cells from old B/W F1 mice contained a number of B cells that spontaneously produced anti-DNA antibodies of both IgM and IgG classes in the absence of stimulants, thereby demonstrating that these B cells had been activated in vivo. These activated B cells could be removed by Sephadex G-10 column (G-10) filtration. Such G-10-passed, homogeneously small B cells were activated by the stimulant lipopolysaccharide (LPS) and produced both IgM and IgG class anti-DNA antibodies. The G-10-passed cells contained both B and T cells, and the cytotoxic treatment of the cells with monoclonal antibodies to T cells, anti-Thy-1 and anti-L3T4, abolished the LPS-induced IgG class, but not IgM class, anti-DNA antibody syntheses. Thus, the LPS-induced production of IgG class anti-DNA antibodies in B/W F1 mice is regulated by T cells. Reconstitution experiments revealed the requirement of T-B cell contact but not of the proliferative response of T cells. Moreover, there was no apparent adherent cell requirement. Such IgG class anti-DNA antibodies were produced only by spleen cells from old B/W F1 mice, but not from young B/W F1, NZB, NZW, and C57BL/6 mice. Like IgM class anti-DNA antibodies, LPS-induced synthesis of polyclonal IgM was T cell-independent. Only a slight reduction in the polyclonal IgG synthesis was observed after the G-10-passed cells had been treated with anti-Thy-1 antibody plus complement. This study should facilitate investigation of cell to cell interactions in the formation of autoantibodies and their correlations to immunologic abnormalities in autoimmune disease.     

Reversal of advanced murine lupus in NZB/NZW F1 mice by treatment with monoclonal antibody to L3T4

Murine lupus in NZB/NZW F1 (B/W) mice can be prevented by weekly treatment with monoclonal antibodies (MAb) to L3T4 (on "helper/inducer" T cells) if treatment is begun prior to the onset of clinical illness. To determine whether anti-L3T4 could reverse as well as prevent murine lupus, we monitored a cohort of 30 B/W females until age 7 mo, when severe autoimmune disease was established, and then we examined the effects of weekly treatment with MAb to L3T4. The rate of target cell clearance by MAb was considerably slower in old B/W mice than it was in young B/W mice or in normal (BALB/c and C57BL/6) mice. Nonetheless, treatment with anti-L3T4 depleted 90% of circulating L3T4+ cells over 3 mo. In treated mice, the concentration of anti-DNA antibodies fell by 80%, renal insufficiency was reversed, and 1 yr survival was 75% compared to 17% in controls. These findings indicate that L3T4+ cells play an important role in perpetuating murine lupus in B/W mice even after severe disease is present. Because the L3T4 antigen in mice is homologous to the Leu-3/T4 (CD4) antigen in humans, these findings suggest that treatment with CD4 MAb may be effective in people with systemic lupus erythematosus.