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Ikaros is a broad transcription factor pointed as a critical regulator of lymphocyte development. Recent reports have emphasized that distinct isoforms of Ikaros control the dichotomy of the hematopoietic system into lymphoid and myeloid lineages. In addition, expression of dominant-negative isoforms of Ikaros is linked to abnormal hematopoiesis, which could culminate in hematological disorders due to loss of function of the protein. B-1 cells are an intriguing subtype of B-lymphocytes that preserves some myeloid characteristics. These cells are able to differentiate into phagocytes (B-1CDP - B-1 cell derived phagocytes) in vitro and in vivo. During such process, reprogramming of gene expression occurs: lymphoid genes are turned off, while expression of myeloid genes is increased. This study aims to investigate whether Ikaros could be related to the control of B-1 cell plasticity. Interestingly, Ikaros expression by B-1CDP cells was found to be relatively low, and the protein is abnormally localized in the cytoplasm. Moreover, the isoforms expressed by B-1 cells are different from those expressed by other lymphocytes, with expression of active isoforms being almost absent in B-1CDP. Based on these findings, Ikaros could be an important factor driving the differentiation and proliferation of B-1 cells., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

B-1 lymphocytes are present in large numbers in the mouse peritoneal cavity, as are macrophages, and are responsible for natural IgM production. These lymphocytes migrate to inflammatory foci and are also involved in innate immunity. It was also demonstrated that B-1 cells are able to differentiated into phagocytes (B-1CDP), which is characterized by expression of F4/80 and increased phagocytic activity. B-1 cell responses to antigens and adjuvants are poorly characterized. It has been shown that Propionibacterium acnes suspensions induce immunomodulatory effects in both macrophages and B-2 lymphocytes. We recently demonstrated that this bacterium has the ability to increase B-1 cell populations both in vitro and in vivo. P. acnes induces B-1CDP differentiation, increases the expression of TLR2, TLR4 and TLR9 and augments the expression of CD80, CD86 and CD40 in B-1 and B-1CDP cells. Because P. acnes has been shown to modulate TLR expression, in this study, we investigated the role of TLR2 and TLR4 in B-1 cell population, including B-1CDP differentiation and phagocytic activity in vitro and in vivo. Interestingly, we have demonstrated that TLR2 signaling could be involved in the increase in the B-1 cell population induced by P. acnes. Furthermore, the early differentiation of B-1CDP is also dependent of TLR2. It was also observed that TLR signals also interfere in the phagocytic ability of B-1 cells and their phagocytes. According to these data, it is clear that P. acnes promotes an important adjuvant effect in B-1 cells by inducing them to differentiate into B-1CDP cells and modulates their phagocytic functions both in vivo and in vitro. Moreover, most of these effects are mediated primarily via TLR2. These data reinforce the findings that such bacterial suspensions have powerful adjuvant properties. The responses of B-1 cells to exogenous stimulation indicate that these cells are important to the innate immune response., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

The role of B-1 cells in the hyperproliferative hematologic disease has been described. Several reports bring evidences that B-1 cells are the main cell population in the chronic lymphatic leukemia. It is also described that these cells have an important involvement in the lupus erythematous systemic. The murine model used to investigate both disease models is NZB/NZW. Data from literature point that mutation in micro-RNA 15a and 16 are the responsible for the B-1 hyperplasia in these mice. Interestingly, it was demonstrated that NZB/NZW B-1 cells are radioresistant, contrariwise to observe in other mouse lineage derived B-1 cells and B-2 cells. However, some reports bring evidences that a small percentage of B-1 cells in healthy mice are also able to survive to irradiation. Herein, we aim to investigate the malignant potential of ionizing-radiation resistant B-1 cells in vitro. Our main goal is to establish a model that mimics the neoplastic transformation originate to a damage exposure of DNA, and not only related to intrinsic mutations. Data shown here demonstrated that radiation-resistant B-1 cells were able to survive long periods in culture. Further, these cells show proliferation index increase in relation to non-irradiated B-1 cells. In addition, radiation resistant B-1 cells showed hyperploid, morphologic alterations, increased induction of apoptosis after anti-IgM stimulation. Based on these results, we could suggest that radiation resistant B-1 cells showed some modifications in that could be related to induction of malignant potential., (Copyright © 2016. Published by Elsevier GmbH.)

Ikaros, a zinc finger transcription factor, is an important regulator of the hematopoietic system. Several studies have suggested the role of Ikaros in the development, maturation, activation and differentiation of lymphocytes. To elucidate this mechanism, it is important to understand how this transcription factor works in the dichotomy of the hematopoietic system, a topic that remains uncertain. Herein, we investigated the role of Ikaros in the control of the lymphomyeloid phenotype of B-1 lymphocytes. We found that Ikaros, as well as its target genes, are expressed in B-1 cells,. Moreover, Ikaros positively regulates the expression of Flt3, Gfi and Il7r, while it down-regulates PU.1. During the induction of differentiation of B-1 cells toward phagocytes, Ikaros transcription was reduced. Taken together, these data pointed to the relevance of Ikaros in the maintenance of the promiscuous gene profile of B-1 cells. It could be suggested that Ikaros functions as a guardian of B-1 lymphoid pattern, and that its absence directs the differentiation of B-1 cells into phagocytes., (Copyright © 2015 Elsevier GmbH. All rights reserved.)

The Wnt/β-catenin signaling pathway has been shown to play an important role in controlling the proliferation, survival and differentiation of hematopoietic cells. Several Wnt/β-catenin signaling components influence hematopoietic cells fate. B-1 cells are self-renewing and spontaneously express both myeloid and lymphoid restricted transcription factors. B-1 lymphocytes play a major role in autoimmunity and are related to CD5(+) B-cell lymphomas and leukemias, such as CLL (chronic lymphocytic leukemia). Herein, we demonstrate that Wnt/β-catenin pathway is important to B-1 cell survival in vitro. The loss of Wnt signals by quercetin treatment induces a reduction in the proliferation and survival of B-1 cells. Furthermore, the quercetin treatment diminishes IL-6 production by peritoneal cells, a cytokine important to the maintenance of B-1 cells in vitro. Importantly, the IL-6 addition to B-1 cell culture prevents cells from apoptosis, even in the presence of quercetin. These data suggest that a deregulation in β-catenin signals could result in alterations in B-1 cell proliferation and differentiation. The correlation between Wnt/β-catenin and IL-6 could point out a mechanism for the expansion of B-1 cells in autoimmune disease and neoplasia., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

Macrophages respond to endogenous and non-self stimuli acquiring the M1 or M2 phenotypes, corresponding to classical or alternative activation, respectively. The role of B-1 cells in the regulation of macrophage polarization through the secretion of interleukin (IL)-10 has been demonstrated. However, the influence of B-1 cells on macrophage phenotype induction by an immunogen that suppress their ability to secrete IL-10 has not been explored. Here, we studied the peritoneal macrophage pattern induced by liposomes comprised of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (Chol) carrying ovalbumin (OVA) (Lp DPPC/OVA), and the involvement of B-1 cells in macrophage polarization. Peritoneal cells from BALB/c, B-1 cells-deficient BALB/xid and C57BL/6 mice immunized with Lp DPPC/OVA and OVA in soluble form (PBS/OVA) were analyzed and stimulated or not in vitro with lipopolysaccharide (LPS). Peritoneal macrophages from BALB/c and C57BL/6 mice immunized with Lp DPPC/OVA showed an M2-like phenotype as evidenced by their high arginase activity without LPS stimulation. Upon stimulation, these macrophages were reprogrammable toward the M1 phenotype with the upregulation of nitric oxide (NO) and a decrease in IL-10 secretion. In addition, high IFN-γ levels were detected in the culture supernatant of peritoneal cells from BALB/c and C57BL/6 mice immunized with Lp DPPC/OVA. Nevertheless, still high levels of arginase activity and undetectable levels of IL-12 were found, indicating that the switch to a classical activation state was not complete. In the peritoneal cells from liposomes-immunized BALB/xid mice, levels of arginase activity, NO, and IL-6 were below those from wild type animals, but the last two products were restored upon adoptive transfer of B-1 cells, together with an increase in IFN-γ secretion. Summarizing, we have demonstrated that Lp DPPC/OVA induce an M2-like pattern in peritoneal macrophages reprogrammable to M1 phenotype after LPS stimulation, with the involvement of B-1 cells., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

B-1 cells can be differentiated from B-2 cells because they are predominantly located in the peritoneal and pleural cavities and have distinct phenotypic patterns and activation properties. The role of both cell populations in cancer progression is still controversial. Previous studies have indicated that direct contact between B-1 cells and B16 melanoma tumor cells (B16) increases the metastatic potential of the tumor cells. However, cellular changes that are induced in B-1 cells during the interaction between these two cell types have not been evaluated. In the present study, it is hypothesized that B-1 cells are modified after their interaction with tumor cells, leading to both increased cell viability and rate of proliferation. Additionally, soluble factors that were secreted by B16 cells were sufficient to augment B-1 cell viability and to modify the production of IL-10 by B-1 cells. Impressively, after direct or indirect contact with the B16 cells, B-1 cells became resistant to radiation-induced cell death. Thus, future studies that assess the importance of concomitant immunity and other conventional therapies in cancer treatment are needed., (Copyright © 2012 Elsevier GmbH. All rights reserved.)

The existence of murine peritoneal osteoclast precursors has been already described. Also, recent reports evidenced an interplay between B lymphocytes and osteoclasts development. B-1 cells comprise a B-lymphocyte subset that resides mostly in pleural and peritoneal cavities. It has been demonstrated that B-1 cells can differentiate into mononuclear phagocytes and form multinucleated giant cells. Based on these findings, we investigated the role of B-1 lymphocytes in bone resorption and osteoclastogenesis. In vivo experimental periodontitis induced in B-1 deficient Xid mice demonstrated that bone resorption is impaired in these animals. However, reconstitution of Xid mice with B-1 cells increased bone resorption to near Balb/c values. B-1 cell derived phagocytes express the receptor activator of nuclear factor-κB (RANK) and the macrophage colony-stimulating factor receptor (M-CSFR). When cultured with RANK-ligand (RANKL) and M-CSF, B-1 cells became tartrate resistant acid phosphatase (TRAP) positive multinucleated cells, a typical osteoclast phenotype. Lacunae formation was observed when cells were cultivated onto a calcium phosphate analog, indicating functional differentiation of B1 cells into osteoclast-like cells. The dynamics of their IgM expression showed that this lymphoid marker was downregulated along the differentiation of B-1 lymphocytes into osteoclasts. Our results unveiled the first evidence that B-1 cells have a role in osteoclastogenesis and bone resorption and offer new insights in the relationship between bone and lymphoid cells., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Sepsis syndrome is caused by inappropriate immune activation due to bacteria and bacterial components released during infection. This syndrome is the leading cause of death in intensive care units. Specialized B-lymphocytes located in the peritoneal and pleural cavities are known as B-1 cells. These cells produce IgM and IL-10, both of which are potent regulators of cell-mediated immunity. It has been suggested that B-1 cells modulate the systemic inflammatory response in sepsis. In this study, we conducted in vitro and in vivo experiments in order to investigate a putative role of B-1 cells in a murine model of LPS-induced sepsis. Macrophages and B-1 cells were studied in monocultures and in co-cultures. The B-1 cells produced the anti-inflammatory cytokine IL-10 in response to LPS. In the B-1 cell-macrophage co-cultures, production of proinflammatory mediators (TNF-α, IL-6 and nitrite) was lower than in the macrophage monocultures, whereas that of IL-10 was higher in the co-cultures. Co-culture of B-1 IL-10(-/-) cells and macrophages did not reduce the production of the proinflammatory mediators (TNF-α, IL-6 and nitrite). After LPS injection, the mortality rate was higher among Balb/Xid mice, which are B-1 cell deficient, than among wild-type mice (65.0% vs. 0.0%). The Balb/Xid mice also presented a proinflammatory profile of TNF-α, IL-6 and nitrite, as well as lower levels of IL-10. In the early phase of LPS stimulation, B-1 cells modulate the macrophage inflammatory response, and the main molecular pathway of that modulation is based on IL-10-mediated intracellular signaling., (Copyright © 2010 Elsevier GmbH. All rights reserved.)

Wound healing is a complex phenomenon whose mechanisms are not fully understood. Although inflammatory cells are recruited to the site of the lesion there are no reports concerning the participation of B lymphocytes in tissue repair. As demonstrated in our laboratory, B-1 cells migrate to a non-specific inflammatory focus and differentiate into a phagocyte. It has been reported that BALB/Xid mice are deficient in B-1 cells. Using this model, here we report that BALB/Xid mice have an increased inflammatory response, a delayed wound-healing process, a prominent neutrophilic infiltration of the lesion, and an increased neovascularization of the lesion as compared with BALB/c and BALB/Xid reconstituted with B-1 cells. The infiltration of B-1 cells into the wound was demonstrated. As B-1 cells secret and use interleukin 10 (IL-10) as an autocrine growth factor, the possible participation of this interleukin in the kinetics of wound healing was investigated. Results show that C57/BL6 IL-10 KO mice had an increased inflammatory response when compared with C57/BL6 and C57/BL6 IL-10 KO mice reconstituted with B-1 cells, thus suggesting that the observed effects of B-1 cells in the healing process is mediated by this interleukin. However, the mechanisms by which IL-10 influence these phenomena remain to be uncovered., (2009 Elsevier GmbH. All rights reserved.)

A mononuclear phagocyte derived from B1b cells (B1CDP) has been described. As these cells migrate from the peritoneal cavity to non-specific inflammatory lesion sites and are highly phagocytic via Fc and mannose receptors, their microbicidal ability of these cells was investigated using the Coxiella burnetii cell infection model in vitro. In this report, the pattern of infection and C. burnetii phase II survival in B1CDP phagosomes was compared with the pattern of infection of peritoneal macrophages from Xid mice (PMphi) and bone marrow derived macrophages (BMMphi). Infection was assessed by determining the large parasitophorous vacuole formation, the relative focus forming units and the quantification of DAPI (4',6-diamino-2-phenylindole) fluorescence images acquired by confocal microscopy. When compared to macrophages, B1CDP are more permissive to the bacterial infection and less effective to kill them. Further, results suggest that IL-10 secreted by B1 cells are involved in their susceptibility to infection by C. burnetti, since B1CDP from IL-10 KO mice are more competent to control C. burnetii infection than cells from wild type mice. These data contribute further to characterize B1CDP as a novel mononuclear phagocyte.

Protective immunity in paracoccidioidomycosis is mainly mediated by cellular immunity. The role of B cells in this disease, in particular B-1 cells, is poorly understood. The aim of this study was to characterize the participation of B-1 cells in resistance or susceptibility of BALB/c and BALB/Xid mice to P. brasiliensis (Pb) pulmonary infection. BALB/Xid, which lacks B-1 cells, exhibited higher resistance to infection when compared with BALB/c mice. However, adoptive transfer of B-1 cells to BALB/Xid mice drastically increased the susceptibility of these animals to Pb infection. The fungal burden in BALB/c and B-1-reconstituted BALB/Xid was significantly higher as compared to BALB/Xid strain. Compact, well-organized granulomas were observed in the lungs of BALB/Xid mice, whereas large lesions with necrotic center with a plethora of fungi developed in BALB/c mice. It was also shown that B-1 cells impair phagocytosis of Pb by macrophages in vitro via secretion of IL-10, which was increased upon stimulation with a purified Pb antigen, gp43. Finally, in vivo blockade of IL-10 led to a better control of infection by the highly susceptible B10.A mouse. These findings suggest that B-1 cells play a major role in resistance/susceptibility to Pb infection in murine models, most likely via production of IL-10.

Macrophages constitute one of the primary cellular mechanisms that impairs parasite invasion of host tissues. The phagocytic and microbicidal properties of these cells can be modulated by specific membrane receptors involved in cell-microorganism interactions. Gp43, the main antigen secreted by Paracoccidiodes brasiliensis (Pb), the causative agent of Paracoccidioidomycosis, is a high mannose glycoprotein. The role played by gp43 in the pathogenesis of the disease is not completely known. Here, we describe the influence of this molecule on the interaction between peritoneal murine macrophages and Pb. Phagocytosis of Pb, live or heat-killed, by adherent peritoneal cells from both, B10.A (susceptible) and A/Sn (resistant) mice, was evaluated. Addition of different concentrations of gp43 to the culture medium inhibited, in a dose-dependent pattern, phagocytosis of live or heat-killed Pb by peritoneal macrophages from both B10.A and A/Sn mice. Gp43 also inhibits phagocytosis of zymosan particles but did not interfere with the uptake of opsonized sheep red blood cells. It was also shown that both gp43 and heat-killed Pb have an inhibitory effect on the release of NO by zymosan stimulated macrophages. Finally, we demonstrated that gp43 inhibits the fungicidal ability of macrophages from both lineages. Based on these data, it is suggested that gp43 can be considered one of the evasion mechanisms for the installation of primary infection in susceptible hosts.