Conventional type 1 dendritic cells are essential for the development of primary biliary cholangitis.

BACKGROUND & AIMS: Primary biliary cholangitis (PBC) is a progressive-cholestatic autoimmune liver disease. Dendritic cells (DC) are professional antigen-presenting cells and their prominent presence around damaged bile ducts of PBC patients are documented. cDC1 is a rare subset of DC known for its cross-presentation abilities and interleukin 12 production. Our aim was to assess the role of cDC1 in the pathogenesis of PBC. METHODS: We utilized an inducible murine model of PBC and took advantage of the DC reporter mice Zbtb46gfp and the Batf3-/- mice that specifically lack the cDC1 subset. cDC1 cells were sorted from blood of PBC patients and healthy individuals and subjected to Bulk-MARS-seq transcriptome analysis. RESULTS: Histopathology assessment demonstrated peri-portal inflammation in wild type (WT) mice, whereas only minor abnormalities were observed in Batf3-/- mice. Flow cytometry analysis revealed a two-fold reduction in hepatic CD8/CD4 T cells ratio in Batf3-/- mice, suggesting reduced intrahepatic CD8 T cells expansion. Histological evidence of portal fibrosis was detected only in the WT but not in Batf3-/- mice. This finding was supported by decreased expression levels of pro-fibrotic genes in the livers of Batf3-/- mice. Transcriptome analysis of human cDC1, revealed 78 differentially expressed genes between PBC patients and controls. Genes related to antigen presentation, TNF and IFN signalling and mitochondrial dysfunction were significantly increased in cDC1 isolated from PBC patients. CONCLUSION: Our data illustrated the contribution the cDC1 subset in the pathogenesis of PBC and provides a novel direction for immune based cell-specific targeted therapeutic approach in PBC.

The Critical Role of Chemokine (C-C Motif) Receptor 2-Positive Monocytes in Autoimmune Cholangitis.

The therapy of primary biliary cholangitis (PBC) has lagged behind other autoimmune diseases despite significant improvements in our understanding of both immunological and molecular events that lead to loss of tolerance to the E2 component of pyruvate dehydrogenase, the immunodominant autoepitope of PBC. It is well known that Ly6Chi monocytes are innate immune cells infiltrating inflammatory sites that are dependent on the expression of C-C motif chemokine receptor 2 (CCR2) for emigration from bone marrow. Importantly, humans with PBC have a circulating monocyte pro-inflammatory phenotype with macrophage accumulation in portal tracts. We have taken advantage of an inducible chemical xenobiotic model of PBC and recapitulated the massive infiltration of monocytes to portal areas. To determine the clinical significance, we immunized both CCR2-deficient mice and controls with 2OA-BSA and noted that CCR2 deficiency is protective for the development of autoimmune cholangitis. Importantly, because of the therapeutic potential, we focused on inhibiting monocyte infiltration through the use of cenicriviroc (CVC), a dual chemokine receptor CCR2/CCR5 antagonist shown to be safe in human trials. Importantly, treatment with CVC resulted in amelioration of all aspects of disease severity including serum total bile acids, histological severity score, and fibrosis stage. In conclusion, our results indicate a major role for Ly6Chi monocytes and for CCR2 in PBC pathogenesis and suggest that inhibition of this axis by CVC should be explored in humans through the use of clinical trials.

Talin1 is required for integrin-dependent B lymphocyte homing to lymph nodes and the bone marrow but not for follicular B-cell maturation in the spleen.

Talin1 is a key integrin coactivator. We investigated the roles of this cytoskeletal adaptor and its target integrins in B-cell lymphogenesis, differentiation, migration, and function. Using CD19 Cre-mediated depletion of talin1 selectively in B cells, we found that talin1 was not required for B-cell generation in the bone marrow or for the entry of immature B cells to the white pulp of the spleen. Loss of talin1 also did not affect B-cell maturation into follicular B cells but compromised differentiation of marginal zone B cells. Nevertheless, serum IgM and IgG levels remained normal. Ex vivo analysis of talin1-deficient spleen B cells indicated a necessary role for talin1 in LFA-1 and VLA-4 activation stimulated by canonical agonists, but not in B-cell chemotaxis. Consequently, talin1 null B splenocytes could not enter lymph nodes nor return to the bone marrow. Talin1 deficiency in B cells was also impaired in the humoral response to a T cell-dependent antigen. Collectively, these results indicate that talin1 is not required for follicular B-cell maturation in the spleen or homeostatic humoral immunity but is critical for integrin-dependent B lymphocyte emigration to lymph nodes and optimal immunity against T-dependent antigens.

c-Met and its ligand hepatocyte growth factor/scatter factor regulate mature B cell survival in a pathway induced by CD74.

The signals regulating the survival of mature splenic B cells have become a major focus in recent studies of B cell immunology. Durable B cell persistence in the periphery is dependent on survival signals that are transduced by cell surface receptors. In this study, we describe a novel biological mechanism involved in mature B cell homeostasis, the hepatocyte growth factor/scatter factor (HGF)/c-Met pathway. We demonstrate that c-Met activation by HGF leads to a survival cascade, whereas its blockade results in induction of mature B cell death. Our results emphasize a unique and critical function for c-Met signaling in the previously described macrophage migration inhibitory factor/CD74-induced survival pathway. Macrophage migration inhibitory factor recruits c-Met to the CD74/CD44 complex and thereby enables the induction of a signaling cascade within the cell. This signal results in HGF secretion, which stimulates the survival of the mature B cell population in an autocrine manner. Thus, the CD74-HGF/c-Met axis defines a novel physiologic survival pathway in mature B cells, resulting in the control of the humoral immune response.

Mad3 negatively regulates B cell differentiation in the spleen by inducing Id2 expression.

Immature B cells migrate to the spleen where they differentiate into mature cells. This final maturation step is crucial to enable B cells to become responsive to antigens and to participate in the immune response. Previously, we showed that Id2 acts as a negative regulator of the differentiation of immature B cells occurring in the spleen. Id2 expression has been found to depend on Myc-Max-Mad transcriptional complexes in mammary epithelial cells. Nearly all studies to date have shown that Mad proteins inhibit proliferation, presumably by antagonizing the function of Myc proteins. In the current study, we followed the Mad family members during peripheral B cell differentiation. We show that Mad3 actively regulates B cell differentiation. Our results demonstrate that high expression levels of Mad3 in immature B cells induce Id2 expression, which inhibits transcription of genes essential for B cell differentiation. During their differentiation to mature cells, B cells reduce their Mad3 expression, enabling the maturation process to occur.

Macrophage migration inhibitory factor induces B cell survival by activation of a CD74-CD44 receptor complex.

Macrophage migration inhibitory factor (MIF) is an upstream activator of innate immunity that regulates subsequent adaptive responses. It was previously shown that in macrophages, MIF binds to a complex of CD74 and CD44, resulting in initiation of a signaling pathway. In the current study, we investigated the role of MIF in B cell survival. We show that in B lymphocytes, MIF initiates a signaling cascade that involves Syk and Akt, leading to NF-kappaB activation, proliferation, and survival in a CD74- and CD44-dependent manner. Thus, MIF regulates the adaptive immune response by maintaining the mature B cell population.

CD74 induces TAp63 expression leading to B-cell survival.

Most mature follicular B cells circulate within the periphery in a quiescent state, without actively contributing to an acute immune response. Lasting B-cell persistence in the periphery is dependent on survival signals that are transduced by cell surface receptors. We recently demonstrated that cell surface CD74 controls mature B-cell survival. Stimulation of cell surface CD74 leads to NF-kappaB activation, which enables entry of the stimulated B cells into the S phase, induction of DNA synthesis, and cell division, and augments the expression of survival genes. In the present study, we investigated CD74 target genes to determine the identities of the molecules whose expression is modulated by CD74, thereby regulating B-cell survival. We report that CD74 activates the p65 member of the NF-kappaB family, which in turn up-regulates the expression of p53-related TAp63 proteins. TAp63 then binds and transactivates the Bcl-2gene and induces the production of Bcl-2 protein, thereby providing the cells with increased survival capacity. Thus, the CD74/NF-kappaB/TAp63 axis defines a novel antiapoptotic pathway in mature B cells, resulting in the shaping of both the B-cell repertoire and the immune response.

IL-8 secreted in a macrophage migration-inhibitory factor- and CD74-dependent manner regulates B cell chronic lymphocytic leukemia survival.

Chronic lymphocytic leukemia (CLL) is a malignant disease of small mature lymphocytes. Previous studies have shown that CLL B lymphocytes express relatively large amounts of CD74 mRNA relative to normal B cells. In the present study, we analyzed the molecular mechanism regulated by CD74 in B-CLL cells. The results presented here show that activation of cell-surface CD74, expressed at high levels from an early stage of the disease by its natural ligand, macrophage migration-inhibition factor (MIF), initiates a signaling cascade that contributes to tumor progression. This pathway induces NF-kappaB activation, resulting in the secretion of IL-8 which, in turn, promotes cell survival. Inhibition of this pathway leads to decreased cell survival. These findings could form the basis of unique therapeutic strategies aimed at blocking the CD74-induced, IL-8- dependent survival pathway.

Cell-surface CD74 initiates a signaling cascade leading to cell proliferation and survival.

CD74 is an integral membrane protein that was thought to function mainly as an MHC class II chaperone. However, CD74 was recently shown to have a role as an accessory-signaling molecule. Our studies demonstrated that CD74 regulates B-cell differentiation by inducing a pathway leading to the activation of transcription mediated by the NF-kappaB p65/RelA homodimer and its coactivator, TAF(II)105. Here, we show that CD74 stimulation with anti-CD74 antibody leads to an induction of a signaling cascade resulting in NF-kappaB activation, entry of the stimulated cells into the S phase, elevation of DNA synthesis, cell division, and augmented expression of BCL-X(L). These studies therefore demonstrate that surface CD74 functions as a survival receptor.