Glia maturation factor-γ is involved in S1P-induced marginal zone B-cell chemotaxis and optimal IgM production to type II T-independent antigen.

Marginal zone B cells (MZBs) represent a unique B-cell sub-population that rapidly differentiate into IgM-secreting plasma cells in response to T-independent (T-I) antigen. Sphingosine 1-phosphate (S1P) promotes MZB localization to the marginal zone. However, intracellular molecules involved in MZB localization and migration remain largely unknown. Here, we show that MZBs lacking the glia maturation factor-γ (GMFG) are impaired in chemotaxis toward S1P under both in vitro and in vivo conditions, suggesting that GMFG is an effector downstream of S1P receptors. GMFG undergoes serine phosphorylation upon S1P stimulation and is required for S1P-induced desensitization of S1P receptor 1 (S1PR1). Compared with wild-type mice, Gmfg-/- mice produce elevated levels of 4-hydroxy-3-nitrophenyl-acetyl (NP)-specific IgM against a T-I type II antigen, NP-Ficoll, accompanied by dysregulated MZB localization. These results identify GMFG as a regulator of S1P-induced MZB chemotaxis and reveal a role for MZB localization in the marginal zone for optimal IgM production against a T-I antigen.

CTLA-4 expression by B-1a B cells is essential for immune tolerance.

CTLA-4 is an important regulator of T-cell function. Here, we report that expression of this immune-regulator in mouse B-1a cells has a critical function in maintaining self-tolerance by regulating these early-developing B cells that express a repertoire enriched for auto-reactivity. Selective deletion of CTLA-4 from B cells results in mice that spontaneously develop autoantibodies, T follicular helper (Tfh) cells and germinal centers (GCs) in the spleen, and autoimmune pathology later in life. This impaired immune homeostasis results from B-1a cell dysfunction upon loss of CTLA-4. Therefore, CTLA-4-deficient B-1a cells up-regulate epigenetic and transcriptional activation programs and show increased self-replenishment. These activated cells further internalize surface IgM, differentiate into antigen-presenting cells and, when reconstituted in normal IgH-allotype congenic recipient mice, induce GCs and Tfh cells expressing a highly selected repertoire. These findings show that CTLA-4 regulation of B-1a cells is a crucial immune-regulatory mechanism.

Distinct roles of BCNP1 in B-cell development and activation.

B-cell novel protein 1 (BCNP1) has recently been identified as a new B-cell receptor (BCR) signaling molecule but its physiological function remains unknown. Here, we demonstrate that mice deficient in BCNP1 exhibit impaired B-cell maturation and a reduction of B-1a cells. BCNP1-deficient spleen B cells show enhanced survival, proliferation and Ca2+ influx in response to BCR cross-linking as compared with wild-type spleen B cells. Consistently, mutant B cells show elevated phosphorylation of SYK, B-cell linker protein (BLNK) and PLCγ2 upon BCR cross-linking. In vivo, BCNP1-deficient mice exhibit enhanced humoral immune responses to T-independent and T-dependent antigens. Moreover, aged mutant mice contain elevated levels of serum IgM and IgG3 antibodies and exhibit polyclonal and monoclonal B-cell expansion in lymphoid organs. These results reveal distinct roles for BCNP1 in B-cell development, activation and homeostasis.

MZB1 promotes the secretion of J-chain-containing dimeric IgA and is critical for the suppression of gut inflammation.

IgA is the most abundantly produced antibody in the body and plays a crucial role in gut homeostasis and mucosal immunity. IgA forms a dimer that covalently associates with the joining (J) chain, which is essential for IgA transport into the mucosa. Here, we demonstrate that the marginal zone B and B-1 cell-specific protein (MZB1) interacts with IgA through the α-heavy-chain tailpiece dependent on the penultimate cysteine residue and prevents the intracellular degradation of α-light-chain complexes. Moreover, MZB1 promotes J-chain binding to IgA and the secretion of dimeric IgA. MZB1-deficient mice are impaired in secreting large amounts of IgA into the gut in response to acute inflammation and develop severe colitis. Oral administration of a monoclonal IgA significantly ameliorated the colitis, accompanied by normalization of the gut microbiota composition. The present study identifies a molecular chaperone that promotes J-chain binding to IgA and reveals an important mechanism that controls the quantity, quality, and function of IgA.

Role of the IgM Fc Receptor in Immunity and Tolerance.

Immunoglobulin (Ig) M is the first antibody isotype to appear during evolution, ontogeny and immune responses. IgM not only serves as the first line of host defense against infections but also plays an important role in immune regulation and immunological tolerance. For many years, IgM is thought to function by binding to antigen and activating complement system. With the discovery of the IgM Fc receptor (FcµR), it is now clear that IgM can also elicit its function through FcµR. In this review, we will describe the molecular characteristics of FcµR, its role in B cell development, maturation and activation, humoral immune responses, host defense, and immunological tolerance. We will also discuss the functional relationship between IgM-complement and IgM-FcµR pathways in regulating immunity and tolerance. Finally, we will discuss the potential involvement of FcµR in human diseases.

The B cell novel protein 1 (BCNP1) regulates BCR signaling and B cell apoptosis.

The BCR plays a central role in B cell development, survival, activation, and differentiation. We have identified the B cell novel protein 1 (BCNP1) as a new regulator of BCR signaling. BCNP1 contains a pleckstrin homology domain, three proline-rich motifs, and a potential SH2 binding site, and is predominantly expressed by B cells. We found that BCNP1 overexpression in WEHI231 immature B cells potentiated α-IgM-induced apoptosis. Conversely, BCNP1-deficient WEHI231 cells, generated by CRISPR-Cas9-mediated genome editing, exhibited reduced apoptosis after BCR crosslinking. Biochemical analyses revealed that BCNP1 physically interacted with the B cell linker protein (BLNK), Grb2, and PLCγ2. Moreover, absence of BCNP1 resulted in accelerated dephosphorylation of BLNK, reduced phosphorylation of SYK and PLCγ2, and decreased Ca2+ influx after BCR crosslinking. These results demonstrate that BCNP1 promotes BCR signaling by modulating the phosphorylation of BLNK, SYK, and PLCγ2.

EAF2 mediates germinal centre B-cell apoptosis to suppress excessive immune responses and prevent autoimmunity.

Regulated apoptosis of germinal centre (GC) B cells is critical for normal humoral immune responses. ELL-associated factor 2 (EAF2) regulates transcription elongation and has been shown to be an androgen-responsive potential tumour suppressor in prostate by inducing apoptosis. Here we show that EAF2 is selectively upregulated in GC B cells among various immune cell types and promotes apoptosis of GC B cells both in vitro and in vivo. EAF2 deficiency results in enlarged GCs and elevated antibody production during a T-dependent immune response. After immunization with type II collagen, mice lacking EAF2 produce high levels of collagen-specific autoantibodies and rapidly develop severe arthritis. Moreover, the mutant mice spontaneously produce anti-dsDNA, rheumatoid factor and anti-nuclear antibodies as they age. These results demonstrate that EAF2-mediated apoptosis in GC B cells limits excessive humoral immune responses and is important for maintaining self-tolerance.

Effects of Ciji Hua'ai Baosheng granule formula (CHBGF) on life time, pathology, peripheral blood cells of tumor chemotherapy model mouse with H22 hepatoma carcinoma cells.

BACKGROUND: Ciji Hua'ai Baosheng Granule Formula (CHBGF) is a traditional Chinese empirical formula that can help the tumor patients who have received chemotherapy antagonize the toxin and side-effects so as to improve and prolong the life. This study is to evaluate the effects of CHBGF on improving life quality in terms of survival time, pathology of tumor tissue and ameliorating peripheral blood cells in mouse chemotherapy model with subcutaneous transplanted tumor or ascitic tumor of H22 hepatoma carcinoma cells at an overall level. MATERIALS AND METHODS: 71 mice among the 92 Kunming mice were injected subcutaneously into the right anterior armpit with H22 hepatoma carcinoma cells, after 7 days, which had formed tumors and were used peritoneal injection of Cytoxan (CTX) (200mg/kg) to establish the mouse chemotherapy model with transplanted tumor, and then which were commensurately divided into 8 groups by random digits table. 21 mice were injected into peritoneal cavity to use CTX and the same method to establish the model. The groups for evaluating the effects on the survival time were the model, CHBGF and positive control group respectively with 7 mice in each group. The groups for evaluating the effects on anti-cancer were the model group, three treatment groups and positive control group with 10 mice in each group. The survival-time-observing groups were given intragastric administration of normal saline, CHBGF (64g/kg) once a day, and peritoneal injection of 5-Fluorouracil (25mg/kg) once every other day respectively. The survival time of each group was observed. The five anti-cancer-observing groups were given intragastric administration of normal saline, CHBGF (64g/kg, 32g/kg and 16g/kg) once a day, and peritoneal injection of 5-Fluorouracil (25mg/kg) once every other day respectively. After treatment for 21 days, the transplanted tumors were peeled off. Blood was collected through pricking eyeball and analyzed by hematology analyzer. And postchemotherapy transplanted tumor inhibition ratios were calculated. Pathological changes of tumor tissues and blood smears were observed with light microscope. RESULTS: The life prolonging rate of CHBGF (64g/kg) group with transplanted tumor is 20.14%, and their survival time was longer than that of the 5-Fluorouracil group (P<0.05). Life prolonging rate of CHBGF (64g/kg) group with ascitic tumor is 64.15%, the survival time was longer than that of the model group (P<0.01) and the 5-Fluorouracil group (P<0.05). The growth of the transplanted tumor in model group was faster than that in CHBGF (64g/kg) group and 5-Fluorouracil group (P<0.05). The tumor average weight of the positive drug and the CHBGF (64g/kg, 32g/kg) groups was lighter than that of the model group (P<0.05 or P<0.01). The inhibition ratios of CHBGF (64g/kg, 32g/kg and 16g/kg) groups are 31.15%, 21.31%, and 13.11% respectively. Under light microscope, in the positive drug and three CHBGF groups the pathological deteriorated severity of tumor tissue observed was milder than that in the model group, the distribution of WBC in CHBGF groups was more obvious than that of the model and 5-Fluorouracil groups. The WBC and PLT decrease in CHBGF (64g/kg, 32g/kg and 16g/kg) groups is less than the model and the 5-Fluorouracil group (P<0.05 or P<0.01), the number of RBC and HGB just in the CHBGF (64g/kg) group was more than that of the model group or the 5-Fluorouracil group (P<0.05). CONCLUSION: Ciji Hua'ai Baosheng Granule Formula can prolong the survival time of the mice chemotherapy model of both subcutaneous transplanted tumor and ascitic tumor of H22 hepatoma carcinoma cells, has some determinate inhibitory effects on the growth of subcutaneous transplanted tumor chemo-treated, and has the therapeutic effect on antagonizing decrease of WBC and PLT caused by chemotherapy.