Cell-dependent B-cell Responses Research Articles

Percutaneous sensitization is limited by in situ inhibition of cutaneous dendritic cell migration through skin-resident regulatory T cells

Percutaneous sensitization is associated with various allergic diseases, including asthma and food allergies. However, the immunologic mechanisms underlying how the skin regulates percutaneous sensitization are still unclear. We aimed to investigate whether and how CD4+Foxp3+ regulatory T (Treg) cells residing in the skin regulate percutaneous sensitization in the skin. Selective reduction of numbers of cutaneous Treg cells was achieved by means of intradermal injection of diphtheria toxin into the ear skin of Foxp3DTR mice, in which Treg cells specifically express the diphtheria toxin receptor fused with green fluorescent protein. Thirty percent to 40% of cutaneous Treg cells were capable of IL-10 production in both mice and human subjects. Selective reduction of cutaneous Treg cells at the sensitization site promoted migration of antigen-bearing dendritic cells (DCs) to the draining lymph nodes (dLNs). Accordingly, sensitization through the skin with reduced numbers of Treg cells led to enhanced antigen-specific immune responses in the dLNs, including both effector T-cell differentiation and T cell-dependent B-cell responses, such as the development of germinal center B cells expressing IgG1 and IgE. Furthermore, antigen-bearing cutaneous DC migration was enhanced in mice with IL-10 deficiency restricted to the cutaneous Treg cell compartment, suggesting an important role of cutaneous IL-10+ Treg cells in limiting percutaneous sensitization. Treg cells with a skin-homing phenotype in skin dLNs expressed high levels of IL-10, suggesting that they contribute to renewal and maintenance of the cutaneous IL-10+ Treg cell population. Skin-resident Treg cells limit percutaneous sensitization by suppressing antigen-bearing DC migration through in situ IL-10 production.

The transcriptional coactivator Bob1 promotes the development of follicular T helper cells via Bcl6.

Follicular T helper (Tfh) cells are key regulators of the germinal center reaction and long‐term humoral immunity. Tfh cell differentiation requires the sustained expression of the transcriptional repressor Bcl6; however, its regulation in CD4+ T cells is incompletely understood. Here, we report that the transcriptional coactivator Bob1, encoded by the Pou2af1 gene, promotes Bcl6 expression and Tfh cell development. We found that Bob1 together with the octamer transcription factors Oct1/Oct2 can directly bind to and transactivate the Bcl6 and Btla promoters. Mixed bone marrow chimeras revealed that Bob1 is required for the expression of normal levels of Bcl6 and BTLA, thereby controlling the pool size and composition of the Tfh compartment in a T cell‐intrinsic manner. Our data indicate that T cell‐expressed Bob1 is directly involved in Tfh cell differentiation and required for mounting normal T cell‐dependent B‐cell responses.

Survival analysis and microarray profiling identify Cd40 as a candidate for the Salmonella susceptibility locus, Ity5.

The outcome of infection with Salmonella Typhimurium in mouse models of human typhoid fever is dependent upon a coordinated complex immune response. A panel of recombinant congenic strains (RCS) derived from reciprocal backcross of A/J and C57BL/6J mice was screened for their susceptibility to Salmonella infection and two susceptibility loci, Ity4 (Immunity to Typhimurium locus 4) and Ity5, were identified. We validated Ity5 in a genetic environment free of the impact of Ity4 using a cross between A/J and 129S6. Using a time-series analysis of genome-wide transcription during infection, comparing A/J with AcB60 mice having a C57BL/6J-derived Ity5 interval, we have identified the differential expression of the positional candidate gene Cd40, Cd40-associated signaling pathways, and the differential expression of numerous genes expressed in neutrophils. CD40 is known to coordinate T cell-dependent B-cell responses and myeloid cell activation. In fact, CD40 signaling is altered in A/J mice as seen by impaired IgM upregulation during infection, decreased Ig class switching, neutropenia, reduced granulocyte recruitment in response to infection and inflammation, and decreased ERK1/2 activity. These results suggest that altered CD40 signaling and granulocyte recruitment in response to infection are responsible for the Ity5-associated Salmonella susceptibility of A/J mice.

The effect of aging on the frequency, phenotype and cytokine production of human blood CD4 + CXCR5 + T follicular helper cells: comparison of aged and young subjects.

BackgroundT cell-dependent B-cell responses decline with age, indicating declined cognate helper activity of aged CD4 + T cells for B cells. However, the mechanisms remain unclear. T follicular helper (Tfh) cells, a novel T helper subset, play an essential role in helping B cells differentiation into long-lived plasma cells in germinal center (GC) or short-lived plasma cells. In the present study, we proposed that there might existe changes of proportion, phenotype or cytokine production of blood Tfh cells in healthy elderly individuals compared with healthy young individuals.ResultsThe results showed that frequencies of aged blood CXCR5 + CD4 + Tfh cells increased compared with young subjects. Both aged and young blood CXCR5 + CD4 + Tfh cells constitutively expressed CD45RO, CCR7 and CD28, and few of these cells expressed CD69 or HLA-DR, which indicated that they were resting memory cells. There was no significant difference of IL-21 frequency production by aged blood CXCR5 + CD4 + Tfh determined by FACS compared with young individuals, however, aged PBMCs produced significantly higher levels of IL-21 evaluated by ELISA. Furthermore, there were no significant differences of percentages of IFN-γ, IL-4, IL-17 or IL-22 production by aged Tfh cells compared with their counterparts of young individuals respectively. However, frequencies of IL-17+ cells within aged CD4 + CXCR5-T cells were markedly lower than in the young individuals. Furthermore we observed different frequencies of IFN-γ, IL-17, IL-4 or IL-22 production by Tfh or by CD4 + CXCR5- cells in aged and young subjects respectively.ConclusionsOur data demonstrated that the frequencies of blood memory CXCR5 + CD4 + Tfh cells increased in the elderly population. There were similar frequencies of Th characterized cytokine production such as IL-21, IFN-γ, IL-4, IL-17 or IL-22 in aged and young Tfh cells. However, aged PBMCs produced a significantly higher amount of IL-21 compare to young subjects.

CD8 T cells induce T-bet–dependent migration toward CXCR3 ligands by differentiated B cells produced during responses to alum-protein vaccines

Antibody-forming cells (AFCs) expressing the chemokine receptor CXCR3 are recruited to sites of inflammation where they help clear pathogens but may participate in autoimmune diseases. Here we identify a mechanism that induces CXCR3 expression by AFC and germinal center (GC) B cells. This happens when CD8 T cells are recruited into CD4 T cell-dependent B-cell responses. Ovalbumin-specific CD4 T cells (OTII) were transferred alone or with ovalbumin-specific CD8 T cells (OTI) and the response to subcutaneous alum-precipitated ovalbumin was followed in the draining lymph nodes. OTII cells alone induce T helper 2-associated class switching to IgG1, but few AFC or GC B cells express CXCR3. By contrast, OTI-derived IFN-γ induces most responding GC B cells and AFCs to express high levels of CXCR3, and diverse switching to IgG2a, IgG2b, with some IgG1. Up-regulation of CXCR3 by GC B cells and AFCs and their migration toward its ligand CXCL10 are shown to depend on B cells' intrinsic T-bet, a transcription factor downstream of the IFN-γR signaling. This model clarifies how precursors of long-lived AFCs and memory B cells acquire CXCR3 that causes their migration to inflammatory foci.

Increased production of soluble inducible costimulator in patients with diffuse cutaneous systemic sclerosis

Inducible costimulator (ICOS) is crucial for T cell proliferation, production of various cytokines, and T cell-dependent B-cell responses. To determine the serum soluble ICOS (sICOS) level and its association with clinical parameters in patients with systemic sclerosis (SSc), serum sICOS level was examined by enzyme-linked immunosorbent assay in 38 patients with SSc and 24 healthy individuals. The expression of ICOS and ICOS ligand in skin was examined immunohistochemically. There was no significant difference in serum sICOS level between patients with SSc and healthy individuals. Patients with diffuse cutaneous SSc had higher levels of sICOS than those with limited cutaneous SSc (P < 0.05) or healthy individuals (P < 0.05). Serum sICOS level correlated positively with the severity of skin sclerosis. Patients with SSc and elevated sICOS level more often had interstitial lung disease and decreased vital capacity than those with normal sICOS level. The serum sICOS level was significantly greater in patients with early phase SSc than those with late phase SSc. ICOS and ICOS ligand immunostaining were observed on infiltrating dermal mononuclear cells in lesional skin tissue. These results suggest that the serum level of sICOS is increased in patients with diffuse cutaneous SSc and correlates with the severity and activity of skin sclerosis and interstitial lung disease. ICOS may contribute to the development of SSc. In addition, measurement of serum sICOS level in patients with early SSc may offer an important means for further evaluation of SSc disease severity.

Signal inhibition by the dual-specific phosphatase 4 impairs T cell-dependent B-cell responses with age

T cell-dependent B-cell responses decline with age, suggesting defective CD4 T-cell function. CD4 memory T cells from individuals older than 65 y displayed increased and sustained transcription of the dual-specific phosphatase 4 (DUSP4) that shortened expression of CD40-ligand (CD40L) and inducible T-cell costimulator (ICOS) (both P < 0.001) and decreased production of IL-4, IL-17A, and IL-21 (all P < 0.001) after in vitro activation. In vivo after influenza vaccination, activated CD4 T cells from elderly individuals had increased DUSP4 transcription (P = 0.002), which inversely correlated with the expression of CD40L (r = 0.65, P = 0.002), ICOS (r = 0.57, P = 0.008), and IL-4 (r = 0.66, P = 0.001). In CD4 KO mice reconstituted with DUSP4 OT-II T cells, DUSP4 had a negative effect on the expansion of antigen-specific B cells (P = 0.003) and the production of ova-specific antibodies (P = 0.03) after immunization. Silencing of DUSP4 in memory CD4 T cells improved CD40L (P < 0.001), IL-4 (P = 0.007), and IL-21 (P = 0.04) expression significantly more in the elderly than young adults. Consequently, the ability of CD4 memory T cells to support B-cell differentiation that was impaired in the elderly (P = 0.004) was restored. Our data suggest that increased DUSP4 expression in activated T cells in the elderly in part accounts for defective adaptive immune responses.

CD27 deficiency is associated with combined immunodeficiency and persistent symptomatic EBV viremia

CD27 is a lymphocyte costimulatory molecule that regulates T-cell, natural killer (NK) cell, B-cell, and plasma cell function, survival, and differentiation. On the basis of its function and expression pattern, we considered CD27 a candidate gene in patients with hypogammaglobulinemia. We sought to describe the clinical and immunologic phenotypes of patients with genetic CD27 deficiency. A molecular and extended immunologic analysis was performed on 2 patients lacking CD27 expression. We identified 2 brothers with a homozygous mutation in CD27 leading to absence of CD27 expression. Both patients had persistent symptomatic EBV viremia. The index patient was hypogammaglobulinemic, and immunoglobulin replacement therapy was initiated. His brother had aplastic anemia in the course of his EBV infection and died from fulminant gram-positive bacterial sepsis. Immunologically, lack of CD27 expression was associated with impaired T cell-dependent B-cell responses and T-cell dysfunction. Our findings identify a role for CD27 in human subjects and suggest that this deficiency can explain particular cases of persistent symptomatic EBV viremia with hypogammaglobulinemia and impaired T cell-dependent antibody generation.

Potential of Nasopharynx-associated Lymphoid Tissue for Vaccine Responses in the Airways

Nasopharynx-associated lymphoid tissue (NALT), constituting Waldeyer's ring in humans, is a unique inductive site for B-cell responses and plasma cell generation. This makes the nasal route of vaccine administration interesting for induction of mucosal and systemic antibodies. The unpaired nasopharyngeal tonsil (adenoids) and the paired palatine tonsils are prominent NALT structures, functionally similar to the paired rodent NALT structures located dorsal to the cartilaginous soft palate. Human NALT is more strategically located, however, because its elements are exposed to both airborne and alimentary antigens and have antigen-retaining crypts. It also shows similarities with lymph nodes and participates both in systemic- and secretory-type mucosal immunity. Primary follicles occur at 16 weeks of gestation, which is similar to Peyer's patches but different from rodent NALT whose organogenesis begins at birth. The formation of germinal centers reflecting B-cell activation does not take place until shortly after birth, and terminal differentiation of plasma cell can be seen about 2 weeks postnatally. Germinal centers arise in T cell-dependent B-cell responses and are associated with somatic hypermutation of Ig V-region genes. Downstream switching to various Ig isotypes also takes place, with or without concurrent expression of the J-chain gene. The J chain is a crucial part of dimeric IgA and pentameric IgM, making these Ig polymers able to interact with the epithelial polymeric Ig receptor. This interaction is central in the formation of secretory IgA and secretory IgM. Accumulating evidence suggests a major role for NALT in antibody immunity of the respiratory tract and associated glands.

Mechanisms of B cell autoimmunity in SLE

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that is known to be associated with polyclonal B-cell hyperreactivity. The underlying causes of the diffuse B-cell over-reactivity are unclear, but potential candidates include (a) intrinsic hyper-reactivity leading to polyclonal B-cell activation with disturbed activation thresholds and ineffective negative selection; (b) lack of immunoregulatory functions; (c) secondary effects of an overactive inflammatory environment, such as overactive germinal center and ectopic follicular activity; and/or (d) disturbed cytokine production by non-B immune cells. These mechanisms are not mutually exclusive and may operate to varying extents and at varying times in SLE. Phenotypic and molecular studies as well as the results of recent clinical trials have begun to provide new insights to address these possibilities. Of importance, new information has made it possible to distinguish between the contribution played by abnormalities in central checkpoints that could lead to a pre-immune repertoire enriched in autoreactive B cells, on the one hand, and the possibility that autoimmunity arises in the periphery from somatic hypermutation and abnormal selection during T cell-dependent B-cell responses on the other. There is an intriguing possibility that apoptotic material bound to the surface of follicular dendritic cells positively selects autoreactive B cells that arise from non-autoreactive B-cell precursors as a result of somatic hypermutation and thereby promotes the peripheral emergence of autoimmunity.

TLR7 regulates dendritic cell-dependent B-cell responses through BlyS in immune thrombocytopenic purpura

Immune thrombocytopenic purpura (ITP) is an autoimmune disorder in which anti-platelet antibodies induce platelet destruction owing to an imbalanced immune response. Several studies have established that B cells play an important role in the production of anti-platelet antibodies and that dendritic cells (DC) are professional antigen-presenting cells of the immune system that may lead to the development of autoantibody through B cells. We aimed at investigating the role of B cells and DC in the pathogenesis of ITP through B-lymphocyte stimulator (BlyS) and toll-like receptor 7 (TLR7) signals. Twenty-two patients with ITP and 20 healthy controls were enrolled in this study. Serum BlyS, its mRNA and TLR7 mRNA were measured using ELISA kits or RT-PCR, and CD14(+) or CD19(+) monocytes were investigated for the pathogenesis of ITP with in vitro culture systems. We demonstrated that serum BlyS levels in patients with ITP were significantly higher than those in healthy controls and that there was a positive correlation between serum BlyS levels and glycoprotein-specific antibody levels in patients with ITP. We found that TLR7 regulates dendritic cell-dependent B-cell responses through BlyS in patients with ITP. Dendritic cells stimulated with R848 (TLR7 ligand) are able to produce vast amounts of BlyS, which is crucial for B-cell survival, proliferation and differentiation, and increase anti-platelet antibodies production in in vitro coculture systems. These findings provide new insights into the pathogenesis of ITP by which TLR7 regulates DC-dependent B-cell responses through BlyS.

Costimulatory blockade of CD154-CD40 in combination with T-cell lymphodepletion results in prevention of allogeneic sensitization

AbstractSensitization is a critical unresolved challenge in transplantation. We show for the first time that blockade of CD154 alone or combined with T-cell depletion prevents sensitization. Allogeneic skin grafts were rejected by recipients treated with anti-αβ T-cell receptor (TCR), anti-CD154, anti-OX40L, or anti–inducible costimulatory pathway (ICOS) mAb alone with a kinetic similar to untreated recipients. However, the production of anti–donor MHC antibody was prevented in mice treated with anti-CD154 mAb only, suggesting a specific role for the CD154-CD40 pathway in B-cell activation. The impairment of T cell–dependent B-cell responses by blocking CD154 occurs through inhibiting activation of T and B cells and secretion of IFN-γ and IL-10. Combined treatment with both anti-CD154 and anti–αβ TCR abrogated antidonor antibody production and resulted in prolonged skin graft survival, suggesting the induction of both T- and B-cell tolerance with prevention of allogeneic sensitization. In addition, we show that the tolerance induced by combined treatment was nondeletional. Moreover, these sensitization-preventive strategies promote bone marrow engraftment in recipients previously exposed to donor alloantigen. These findings may be clinically relevant to prevent allosensitization with minimal toxicity and point to humoral immunity as playing a dominant role in alloreactivity in sensitized recipients.

Abnormal T cell–dependent B-cell responses in SCID mice receiving allogeneic bone marrow in utero

In allogeneic hematopoietic stem cell transplant recipients, restoration of humoral immunity is delayed and can remain impaired for years. In many severe combined immune deficiency (SCID) patients given haploidentical bone marrow (BM), lesions in humoral immunity are exacerbated by poor engraftment of donor B cells. The nature of these defects is important to understand as they render patients susceptible to infection. Previous work in mice suggested that in utero transplantation (IUT) of allogeneic BM might offer several advantages for the correction of primary immune deficiencies. In SCID mice given fully allogeneic BM in utero, the lymphoid compartment was restored with minimal evidence of graft-versus-host disease (GVHD). The present report examines B-cell reconstitution and function in mice that have received allogeneic IUT. Results are compared with those of adult mice given total body irradiation (TBI) followed by transplantation with allogeneic BM. In addition to enumerating the various B-cell subsets present in BM, spleen, and peritoneal cavity (PC), B-cell competence was assessed by challenging mice with T cell-independent (TI) and T cell-dependent (TD) antigens. The results demonstrated that all B-cell subsets in the BM and periphery were restored in allogeneic IUT and TBI mice, as were antibody responses after TI challenge. Upon immunization with TD antigens, however, IUT and TBI mice exhibited suboptimal responses as measured by the capacity to isotype switch and generate germinal center (GC) B cells. Thus, although allogeneic BM transplantation results in complete recovery of the B-cell compartment, certain elements of the humoral response remain defective.

Visualizing immune responses in vivo

Helper T cell dependent B-cell responses develop in the complex microenvironments of secondary lymphoid organs. New strategies for visualizing antigen-responsive lymphocytes offer direct insight into how differentiation proceeds in vivo.

Adoptive transfer of immunity to hepatitis B virus after T cell-depleted allogeneic bone marrow transplantation

Recipients of allogeneic bone marrow transplantation are pancytopenic for several weeks and immunosuppressed for many months as a result of myeloablative therapy required to eliminate the basic disease and to prevent allograft rejection. After bone marrow transplantation, these patients remain profoundly immunosuppressed by the chemotherapy and immunotherapy used as prophylaxis against graft-vs.-host disease, treatment of established disease or both. These patients are usually dependent on multiple blood products and are therefore at risk for hepatitis B virus infection, which may run a fulminant course. Active immunization against hepatitis B virus in the immediate pre-bone marrow transplantation and post-bone marrow transplantation periods was found to be ineffective, probably because of the absence of T cell-dependent B-cell responses, which persists for approximately 1 yr after bone marrow transplantation. We studied adoptive transfer of immunity to hepatitis B virus through bone marrow transplantation in two populations of patients. The first group (A) consisted of 12 pairs of BMT donors and recipients, in which all bone marrow donors were positive for antibodies to HBc and HBs as a result of previously acquired hepatitis B virus infection and resolution; all recipients were negative to antibodies to HBc and HBs. The second group (B) consisted of eight pairs of donors and recipients in which all the donors were actively immunized against hepatitis B virus before bone marrow transplantation; all recipients were negative for all hepatitis B virus markers. All bone marrow transplantation recipients were monitored for antibodies to hepatitis B virus antigens.(ABSTRACT TRUNCATED AT 250 WORDS)

Adoptive transfer of immunity to hepatitis B virus in mice by bone marrow transplantation from immune donors

Recipients of allogeneic bone marrow transplantation are immunosuppressed as a result of their primary disease and by myeloablative therapy. Such patients are dependent on multiple blood products and are at risk for hepatitis B virus infection. Active immunization against hepatitis B in the immediate pre- and post-transplant periods is ineffective, pre- sumably because of decreased T cell–dependent B-cell responses. This study was designed to evaluate, in a mouse model system, the transfer of immunity against hepatitis B to bone marrow transplant recipients through immunization of bone marrow donors against hepatitis B before transplantation. Bone marrow donor BALB/c mice were immunized with a recombinant hepatitis B vaccine. Seroconversion to HBs antibody occurred within 4 wk of primary immunization, and antibody levels in treated donor mice rose above 300 mIU/ml after a single booster injection. Bone marrow recipient mice, conditioned by sublethal irradiation, were injected intravenously with bone marrow cells obtained from syngeneic HBs antibody–positive immune donors. Antibody was detected in 10% of bone marrow recipients within 30 days of transplantation and in 56% 1 mo after a booster injection that led to a secondary rise in HBs antibody. Adoptive transfer of immunity to hepatitis B also occurred after transplantation of T cell–depleted bone marrow cells from hepatitis B–immune donors, albeit at a lower HBs antibody level. These results indicate that immunity to hepatitis B can be transferred in mice by bone marrow transplantation from hepatitis B–immune donors to immunosuppressed recipients. Furthermore, such bone marrow transplant recipients were able to respond to a booster immunization, suggesting secondary immune response in bone marrow recipients after primary response in donors. These data support the rationale for active vaccination of potential human bone marrow donors against hepatitis B, followed by booster vaccination to bone marrow recipients. (HEPATOLOGY 1993;17:955–959.)

Adoptive transfer of immunity to hepatitis B virus in mice by bone marrow transplantation from immune donors

Recipients of allogeneic bone marrow transplantation are immunosuppressed as a result of their primary disease and by myeloablative therapy. Such patients are dependent on multiple blood products and are at risk for hepatitis B virus infection. Active immunization against hepatitis B in the immediate pre- and post-transplant periods is ineffective, presumably because of decreased T cell-dependent B-cell responses. This study was designed to evaluate, in a mouse model system, the transfer of immunity against hepatitis B to bone marrow transplant recipients through immunization of bone marrow donors against hepatitis B before transplantation. Bone marrow donor BALB/c mice were immunized with a recombinant hepatitis B vaccine. Seroconversion to HBs antibody occurred within 4 wk of primary immunization, and antibody levels in treated donor mice rose above 300 mIU/ml after a single booster injection. Bone marrow recipient mice, conditioned by sublethal irradiation, were injected intravenously with bone marrow cells obtained from syngeneic HBs antibody-positive immune donors. Antibody was detected in 10% of bone marrow recipients within 30 days of transplantation and in 56% 1 mo after a booster injection that led to a secondary rise in HBs antibody. Adoptive transfer of immunity to hepatitis B also occurred after transplantation of T cell-depleted bone marrow cells from hepatitis B-immune donors, albeit at a lower HBs antibody level. These results indicate that immunity to hepatitis B can be transferred in mice by bone marrow transplantation from hepatitis B-immune donors to immunosuppressed recipients.(ABSTRACT TRUNCATED AT 250 WORDS)