Increased circulating CD11b+CD11c+ dendritic cells (DC) in aged BWF1 mice which can be matured by TNF-alpha into BLC/CXCL13-producing DC

Dendritic cells (DC) play a pivotal role in regulating immune responses. We previously reported aberrant high production of B lymphocyte chemoattractant (BLC/CXCL13) by DC in aged BWF1 mice, amurine model for systemic lupus erythematosus (SLE). We describe here that CD11b+CD11c+ cells were markedly increased in the peripheral blood (PBL-DC) in aged BWF1, but not in similarly aged NZB or NZW mice. Part of PBL-DC showed a typical dendritic morphology and expressed MHC class II molecules, and had a weak, but significant antigen-presenting ability in mixed lymphocytereaction. PBL-DC were chemoattracted to several chemokines in vitro including secondary lymphoid tissue chemokine (SLC), liver and activation-regulated chemokine (LARC), RANTES, macrophage inflammatory protein-1alpha, whereas splenic mature DC from aged BWF1 mice were preferentially chemoattracted towards SLC. BLC production was induced when PBL-DC were cultured in the presence of TNF-alpha for 3 days. BLC expression was also induced in bone marrow-derived DC when they were differentiated into mature DC in the presence of TNF-alpha and IL-1beta, while both IFN-alpha and IFN-gamma failed to induce BLC expression in bone marrow-derived DC. Since TNF-alpha expression is increased in aged BWF1 mice, DC recruitment in the circulation and maturation into BLC-producing DC by TNF-alpha may play a pivotal role in the development of systemic autoimmune diseases.     

Differences between CD8+ T cells in lupus-prone (NZB x NZW) F1 mice and healthy (BALB/c x NZW) F1 mice may influence autoimmunity in the lupus model

Immunization with portions of a murine antibody to DNA induced Ig peptide-reactive peripheral CD8+ inhibitory T (Ti) cells in non-autoimmune (BALB/c x NZW) F1 (CWF1) mice. Those Ti suppressed in vitro production of IgG anti-DNA by lymphocytes from MHC-matched, lupus-prone (NZB x NZW) F1 (BWF1) mice, primarily via secretion of transforming growth factor-beta (TGF-beta). However, splenic CD8+ cells from immunized BWF1 mice failed to suppress anti-DNA. Therefore, BWF1 mice were studied for defects in peripheral CD8+ T cells. The potential to suppress autoimmunity mediated by activated CD4+ helper T and B cells in BWF1 mice was assessed. As BWF1 mice aged, peripheral CD8+ T cells expanded little; fewer than 10% displayed surface markers of activation and memory. In contrast, quantities of splenic CD4+ T and B cells increased; high proportions displayed activation/memory markers. In old compared to young BWF1 mice, splenic cell secretion of two cytokines required for generation of CD8+ T effectors, IL-2 and TGF-beta, was decreased. Immunizing BWF1 mice activated peptide-reactive CD8+ T cells, but their number was decreased compared to young BWF1 or old normal mice. While peptide-reactive splenic CD8+ T cells from immunized BWF1 mice did not survive in short-term cultures, similar CD8+ T cell lines from immunized CWF1 mice expanded and on transfer into BWF1 mice delayed autoimmunity and prolonged survival. Therefore, CD8+ T cells in old BWF1 mice are impaired in expansion, acquisition of memory, secretion of cytokine, and suppression of autoimmunity. Understanding these defects might identify targets for therapy in systemic lupus erythematosus.     

L-selectin and alpha 4 beta 7 integrin homing receptor pathways mediate peripheral lymphocyte traffic to AKR mouse hyperplastic thymus.

Before the development of thymic lymphoma, AKR mice undergo a striking lymphoid hyperplasia of the thymic medulla. We have previously shown that there is a marked increase in traffic of B and T lymphocytes from the periphery into the preneoplastic, hyperplastic thymuses of these mice, in contrast to the scant traffic of such cells to normal thymuses. The traffic of lymphocytes to lymph nodes and Peyer's patches is controlled in part by the interaction of lymphocyte adhesion molecules called homing receptors with their tissue-selective endothelial ligands known as vascular addressins. We have investigated the roles of homing receptors and vascular addressins in the traffic of lymphocytes to the AKR hyperplastic thymus. We demonstrate that development of hyperplasia is accompanied by an increase in the number of thymic medullary blood vessels with high endothelial venule morphology and expression of the peripheral node addressin (PNAd) and the mucosal addressin (MAdCAM-1). In vitro and in vivo functional assays show that the addressin/homing receptor pairs PNAd/L-selectin and MAdCAM-1/alpha 4 beta 7 are involved in lymphocyte traffic to the hyperplastic thymus. These results indicate that molecular adhesion mechanisms involved in tissue-selective migration of lymphocytes to peripheral lymph node and to mucosal lymphoid tissues play a role in the recruitment of B and T lymphocytes to the AKR thymus and thus in the pathogenesis of thymic hyperplasia.     

Systemic autoimmunity in BAFF-R-mutant A/WySnJ strain mice

Systemic lupus erythematosis is an autoimmune disease of unknown etiology. Lupus pathology is thought to reflect autoantibody-mediated damage due to a failure of B lymphocyte tolerance. Since excessive B cell-activating factor belonging to the TNF family (BAFF) expression correlates with human and murine lupus, and BAFF signals B cell survival through BAFF-R, it is believed that excessive BAFF-R signaling can subvert B cell tolerance and facilitate lupus development. Here we report the unexpected finding that BAFF-R-mutant A/WySnJ mice develop a lupus-like syndrome. These mice carry the B cell maturation defect-1 (Bcmd-1) mutant allele of the Baffr gene. Bcmd-1 causes premature B cell death and profound B cell deficiency. Despite having 90% fewer splenic B cells than normal mice, A/WySnJ mice had an 18-fold increased frequency of splenocytes secreting IgM antibodies to dsDNA, and increased amounts of circulating IgM and IgG to dsDNA by 9 months of age. By age 11 months, most A/WySnJ mice displayed renal pathology characteristic of lupus, including proteinuria as well as periodic acid-Schiff-positive deposits and glomerular capillary bed destruction. Importantly, we genetically linked this autoimmunity to Bcmd-1, since congenic AW.Baffr(+/+) mice carrying a wild-type allele developed none of these phenotypes. Our data provide the first evidence linking altered BAFF-R signaling to the development of B cell-mediated autoimmunity.     

Increased Foxp3+ CD4+ Regulatory T Cells with Intact Suppressive Activity but Altered Cellular Localization in Murine Lupus

Foxp3⁺ CD4⁺ regulatory T (T_reg) cells play a pivotal role in the maintenance of dominant self tolerance. Understanding how the failures of immune control by T_reg cells are involved in autoimmune diseases is important for the development of effective immunotherapies. In the present study, we analyzed the characteristics of endogenous T_reg cells in (NZB × NZW) F1 (BWF1) mice, a murine model of systemic lupus erythematosus. Unexpectedly, T_reg number and frequency in aged BWF1 mice with developing lupus nephritis were increased, not decreased, and in vitro suppressive activity in lymphoid organs was intact. In addition, T_reg cells trafficked to target organs because cells were present in the kidney and lung. T_reg cells of aged BWF1 mice exhibited altered localization within lymph organs, however, and an altered phenotype, with higher expression levels of chemokine receptors and activation markers, suggesting a highly activated cellular state. Notably, the expression levels of co-stimulatory molecules were also markedly enhanced in the T_reg cells of aged BWF1 mice. Furthermore, T_reg cells of BWF1 mice did not show any suppressive effects on antibody production in vitro. Taken together, we conclude that T_reg cells in BWF1 mice are not predisposed to functional incompetence but rather are present in a highly activated state.     

Apoptosis and the thymic microenvironment in murine lupus.

The thymus of New Zealand black (NZB) mice undergoes premature involution. In addition, cultured thymic epithelial cells from NZB mice undergo accelerated preprogrammed degeneration. NZB mice also have distinctive and well-defined abnormalities of thymic architecture involving stromal cells, defined by staining with monoclonal antibodies specific for the thymic microenvironment. We took advantage of these findings, as well as our large panel of monoclonal antibodies which recognize thymic stroma, to study the induction of apoptosis in the thymus of murine lupus and including changes of epithelial architecture. We studied NZB, MRL/lpr, BXSB/Yaa, C3H/gld mice and BALB/c and C57BL/6 as control mice. Apoptosis was studied both at basal levels and following induction with either dexamethasone or lipopolysaccharide (LPS). The apoptotic cells were primarily found in the thymic cortex, and the frequency of apoptosis in murine lupus was less than 20% of controls. Moreover, all strains of murine lupus had severe abnormalities of the cortical network. These changes were not accentuated by dexamethasone treatment in cultured thymocytes. However, the thymus in murine lupus was less susceptible to LPS-induced apoptosis than control mice. Finally we note that the number of thymic nurse cells (TNC) was lowest in NZB mice. Our findings demonstrate significant abnormalities in the induction of apoptosis and the formation of TNC-like epithelial cells in SLE mice, and suggest that the abnormalities of the thymic microenvironment have an important role in the pathogenesis of murine lupus.     

Elevated Production of B Cell Chemokine CXCL13 is Correlated with Systemic Lupus Erythematosus Disease Activity

Introduction  

B lymphocyte chemoattractant (BLC/CXCL13), a CXC chemokine, is involved in B1 and B2 cell trafficking for the activation of autoreactive T helper (Th) cells and autoantibody production in target organs during the development of lupus. CXCL13 can induce the trafficking of CXCR5+ T lymphocyte subset designated as follicular helper T lymphocytes (T_FH) which are specifically involved in autoantibody production.     

Adoptive transfer of autoimmune splenic dendritic cells to lupus-prone mice triggers a B lymphocyte humoral response

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by increased autoantibody production that leads to multiple tissue injuries. Dendritic cells (DCs) are important orchestrators of immune responses and key components in fine-tuning the balance between tolerance and immunity. However, their role in autoimmune disorders such as SLE remains uncertain. We analyzed the contribution of DCs in triggering SLE by adoptively transferring splenic DCs from aged autoimmune [NZB×NZW]F1 (BWF1) mice to young healthy BWF1 mice. We observed that the transfer of DCs from autoimmune mice to pre-autoimmune mice induced high autoantibody titers in the serum of recipient mice. Moreover, autoimmune DCs from aged BWF1 mice were crucial for the expansion and differentiation of plasmablasts and CD5⁺ B cells or B1-like cells in the peripheral blood, and spleen of recipient BWF1 mice, a phenomenon that is observed in autoimmune BWF1 mice. On the other hand, DCs from aged BWF1 mice participated in the expansion and differentiation of DCs and IFN-γ-producing T cells. These results reveal that DCs from autoimmune BWF1 mice exhibit functional and phenotypic characteristics that allow them to trigger B cell hyperactivation, as well as DC and T cell expansion and differentiation, thereby promoting an exacerbated humoral response in lupus-prone mice.     

The perivascular space as a path of hematopoietic progenitor cells and mature T cells between the blood circulation and the thymic parenchyma

It is known that selected populations of lymphoid cells migrate into and from the adult thymus through blood vessels at the cortico-medullary junction and in the medulla. Here, we show that in the perivascular spaces (PVS) of mice surrounding large blood vessels, CD117-positive hematopoietic progenitor cells, CD4 single-positive (SP) and CD8SP T cells are located. However, developing thymocytes, CD25-positive cells and CD4 and CD8 double-positive cells, are not detectable in the PVS. After intravenous (i.v.) injection of CD117-positive bone marrow (BM) cells from C57BL/6 mice into non-irradiated RAG2 mutant mice i.v., donor-derived cells first preferentially migrate into the PVS within 30 min, and then the number of donor-derived cells in the thymic parenchyma increases. Likewise, newly developed mature T cells in the thymic parenchyma of RAG2 mutant mice transferred with wild-type BM cells migrate to the PVS, before leaving the thymus to the circulation. Accumulation of mature T cells was observed after treatment with sphingosine-1 phosphate receptor agonist FTY720 not only in the medulla but also in the thymic PVS. These results suggest that the PVS is a transit pathway for progenitor cells to immigrate into the thymus and for mature T cells to emigrate from the thymus.     

BLC (CXCL13) is expressed by different dendritic cell subsets in vitro and in vivo

Dendritic cells (DC) attract both T and B lymphocytes to induce an efficient antigen-specific immune response. Recently, it was shown that na?ve T cells are attracted to DC by dendritic cell chemokine 1 (DC-CK1, CCL18). The potent B lymphocyte chemoattractant BLC (CXCL13) was previously shown to be essential for homing of lymphocytes into secondary lymphoid organs and for the development of B cell follicles. As the cells that produce BLC are largely unknown and BLC could be a candidate chemokine for the recruitment of B cells to DC, we analyzed different DC subsets for expression of BLC. Here we demonstrate that monocyte-derived DC as well as activated blood DC indeed express and secrete BLC. Interestingly, ligation of the CD40 molecule down-regulated BLC expression in monocyte-derived DC. Staining of tonsilar sections indicated that BLC is expressed by follicular dendritic cells and germinal center dendritic cells (GCDC) in vivo. Real-time quantitative PCR confirmed the expression of BLC in isolated GCDC. Since both B cells and activated T cells express the receptor for BLC, our findings implicate an important role for BLC in establishing the interaction of DC with T cells and B cells. Furthermore, CD40/CD40 ligand interactions could modulate this process by down-regulating the expression of BLC.     

Changes in the cytokine profile of lupus-prone mice (NZB/NZW)F1 induced by Plasmodium chabaudi and their implications in the reversal of clinical symptoms

We have previously observed that aged lupus-prone (NZB/NZW)Fl (BWF1) mice when infected with Plasmodium chabaudi show an improvement in their clinical lupus-like symptoms. In order to study the mechanisms involved in the long-lasting protective effect of the P. chabaudi infection in lupus-prone mice we analysed specific aspects of the cellular response, namely the profiles of cytokine mRNA expression and cytokine secretion levels in old BWF1 mice, in comparison with uninfected age-matched BWF1 mice and infected or uninfected BALB/c mice. Two months after infection, cells from BWF1 mice were stimulated with concanavalin A (Con A) and demonstrated a recovery of T cell responsiveness that reached the levels obtained with BALB/c cells. Old BWF1 mice showed high levels of interferon-gamma (IFN-gamma) and IL-5 production and correspondingly low levels of IL-2 and IL-4 secretion before infection with P. chabaudi. Infection did not modify the IFN-gamma levels of BWF1 T cells, whereas it considerably increased the secretion of the Th2-related cytokines IL-4, IL-5 and IL-10. In addition, only BWF1 T cells showed increased mRNA expression of tumour necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta). This counter-regulatory cytokine network of infected BWF1 mice may be involved in the improvement of their lupus symptoms. The results of our investigations using the complex model of P. chabaudi infection can be extended and, by using more restricted approaches, it may be possible to explain the multiple regulatory defects of lupus-prone mice.     

Traffic of mature lymphocytes into the mouse thymus

The thymus is not generally considered to participate in bi-directional peripheral lymphocyte recirculation. We have demonstrated the entry of peripheral lymph node T cells using transfers between Thy-1 congenic mice. These peripheral T cells that enter the thymus bear an essentially medullary (or peripheral) phenotype and on section stains are confined to the medulla where they constitute 0.2-0.3% of the cells. These cells are remarkable for their frequent expression of the peripheral node homing receptor MEL-14 and for their persistence. Using transferred fluorescent labeled cells we also identify the entry of peripheral B cells into the thymus. Possible roles for peripheral B and T cells in the thymus include participation in thymocyte maturation or selection, the generation of thymic pathology and reactions to thymic pathologic processes.     

Bone marrow-thymus axis in senescence

This presentation offers a brief review of the bone marrow-thymus axis in senescence, a putatitive model for thymocyte differentiation, and recent results of our work on the status of pre-thymic stem cells in aged mice. The data presented here provide further evidence for a thymus endocrine influence on the bone marrow stem cells, specifically lymphocyte precursors. It has been pstulated that the thymic hormones may act on lymphocyte precursors in the bone marrow and that the loss of thymic factors during senescence may be a contributing factor to the decreased cellular immune function. This study used Haar's in vitro model to investigate the bone marrow-thymus axis in aged mice. Erythroid-depleted bone-marrow cells from 3-month- and 24-month-old CBA (Thy 1.2) mice were placed in the upper half of a blind-well chamber with thymus supernatant in the lower half. Experimental cells were treated with thymus supernatant for 1 hr prior to migration. This study confirmed that pre-thymic stem cells in aged bone marrow are deficient in their ability to migrate to the thymus supernatant. It also revealed that treatment of the old bone marrow with thymus supernatant, made from neonatal thymus cultures, could dramatically improve the thymus migrating ability of the aged bone-marrow stem cells.     

Different chemokine expression in lethal and non-lethal murine West Nile virus infection

West Nile (WN) virus is a mosquito-borne flavivirus that can cause lethal encephalitis in humans and horses. The WN virus endemic in New York City (NY) in 1999 caused large-scale mortality of wild birds that was not evident in endemic areas in other parts of the world, and the pathogenesis of the WN virus strain isolated in NY (NY strain) appears to differ from that of previously isolated strains. However, the pathogenesis of NY strain infection remains unclear. This study examined CC (RANTES/CCL5, MIP-1 alpha/CCL3, MIP-1 beta/CCL4) and CXC (IP-10/CXCL10, B lymphocyte chemoattractant (BLC/CXCL13), and B cell- and monocyte-activating chemokine (BMAC/CXCL14)) chemokine expression during lethal NY strain and non-lethal Eg101 strain infection in mice. We found that the mRNA of the CC chemokines, RANTES, MIP-1 alpha, MIP-1 beta, and IP-10 was highly up-regulated in the brain of NY strain-infected mice. By contrast, BLC mRNA was not detected in either group of mice, and BMAC mRNA was highly up-regulated in late stage of infection with the non-lethal Eg101 strain relative to levels in NY strain-infected mice.     

Transplantation of retrovirally transduced bone marrow prevents autoimmune disease in aged mice by peripheral tolerance mechanisms

Transplantation of bone marrow (BM) engineered to express self-antigen has been shown to protect 100% of young mice from myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), with thymic clonal deletion as a tolerance mechanism. Here, we asked whether aged mice can also be tolerised following transplantation with self-antigen-engineered BM and whether castration-induced thymus regrowth can enhance this outcomes. Then, 50% of aged mice were protected from EAE regardless of castration-induced thymus regrowth. EAE-free and diseased mice demonstrated MOG-specific lymphocyte proliferation and antibody production regardless of castration-induced thymus regrowth, consistent with lack of intrathymic deletion of self-antigen-reactive T cells. Although low chimerism levels ( < 4%) were observed, EAE-free mice showed significantly higher chimerism levels in lymphocytes in peripheral lymphoid organs compared with thymus. CD4⁺CD25⁺ regulatory T cells were elevated in lymph nodes of EAE-free mice. We conclude that transplantation of self-antigen expressing BM protects 50% of aged mice and castration-induced thymic regrowth had no effect on outcomes. Peripheral tolerance mechanisms are implicated since protection is associated with higher chimerism levels in peripheral T and B lymphocytes and with elevated regulatory T cells.     

Transgene-mediated hyper-expression of IL-5 inhibits autoimmune disease but increases the risk of B cell chronic lymphocytic leukemia in a model of murine lupus

IL-5 preferentially activates B1 cells to produce natural antibodies cross-reactive to self antigens. To determine the role of IL-5 in antibody-mediated autoimmune disease, we generated systemic lupus erythematosus (SLE)-prone (NZB x NZW)F1 mice congenic for IL-5 transgene (TG-F1). The transgene unexpectedly reduced the incidence of lupus nephritis. Anti-DNA antibodies in sera and those produced by splenic B cells in vitro were markedly decreased in TG-F1 mice, while total polyclonal Ig levels were comparable to those in IL-5 transgene-negative (NZB x NZW)F1 (non-TG-F1) littermates. Flow cytometry-sorted splenic B1 cells showed a significant reduction of anti-DNA antibody synthesis in response to IL-5, while proliferative responses to IL-5 did not significantly differ between TG-F1 and non-TG-F1 mice. As TG-F1 mice aged, frequencies of peripheral B1 cells progressively increased, and the mice frequently developed B cell chronic lymphocytic leukemia (B-CLL). Our results suggest that dysregulated, continuous high expression of IL-5 in SLE-prone mice may directly or indirectly mediate a skewed signaling of proliferation/differentiation of self-antigen-activated B1 cells, leading to suppression of autoimmune disease, but instead to aberrant expansion of B1 cells, giving rise to B-CLL. Thus, this model may provide a clue to the pathogenesis of both SLE and B-CLL.     

Transplantation of retrovirally transduced bone marrow prevents autoimmune disease in aged mice by peripheral tolerance mechanisms

Transplantation of bone marrow (BM) engineered to express self-antigen has been shown to protect 100% of young mice from myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), with thymic clonal deletion as a tolerance mechanism. Here, we asked whether aged mice can also be tolerised following transplantation with self-antigen-engineered BM and whether castration-induced thymus regrowth can enhance this outcomes. Then, 50% of aged mice were protected from EAE regardless of castration-induced thymus regrowth. EAE-free and diseased mice demonstrated MOG-specific lymphocyte proliferation and antibody production regardless of castration-induced thymus regrowth, consistent with lack of intrathymic deletion of self-antigen-reactive T cells. Although low chimerism levels (?相似文献