Sustained antigen delivery improves germinal center reaction and increases antibody responses in neonatal mice.

Neonates and young infants are known to have limited responses to pediatric vaccines due to reduced germinal center formation. Extended vaccine antigen dosing was previously shown to expand germinal center formation and improve humoral responses in adult mice. We report that sustained antigen delivery through sequential dosing overcomes neonatal limitations to form germinal center reactions and improves humoral immunity. Thus, vaccine strategies that extend the release of vaccine antigens may reduce the number of doses, and time needed, to achieve protective immunity in neonates and young infants.

State of pneumococcal vaccine immunity.

Like the other invasive encapsulated bacteria, Streptococcus pneumoniae is also covered with a polysaccharide structure. Infants and elderly are most vulnerable to the invasive and noninvasive diseases caused by S. pneumoniae. Although antibodies against polysaccharide capsule are efficient in eliminating S. pneumoniae, the T cell independent nature of the immune response against polysaccharide vaccines renders them weakly antigenic. The introduction of protein conjugated capsular polysaccharide vaccines helped overcome the weak immunogenicity of pneumococcal polysaccharides and decreased the incidence of pneumococcal diseases, especially in pediatric population. Conjugate vaccines elicit T cell dependent response which involve the interaction of specialized CD4+ T cells, called follicular helper T cells (Tfh) with germinal center B cells in secondary lymphoid organs. Despite their improved immunogenicity, conjugate vaccines still need to be administered three to four times in infants during the first 15 month of their life because they mount poor Tfh response. Recent studies revealed fundamental differences in the generation of Tfh cells between neonates and adults. As the portfolio of pneumococcal conjugate vaccines continues to increase, better understanding of the mechanisms of antibody development in different age groups will help in the development of pneumococcal vaccines tailored for different ages.

IL6 suppresses vaccine responses in neonates by enhancing IL2 activity on T follicular helper cells.

The inability of neonates to develop CD4+FoxP3-CXCR5hiPD-1hi T follicular helper (TFH) cells contributes to their weak vaccine responses. In previous studies, we measured diminished IgG responses when IL-6 was co-injected with a pneumococcal conjugate vaccine (PCV) in neonatal mice. This is in sharp contrast to adults, where IL-6 improves vaccine responses by downregulating the expression of IL-2Rß on TFH cells and protecting them from the inhibitory effect of IL-2. In this study, we found that splenic IL-6 levels rapidly increased in both adult and neonatal mice following immunization, but the increase in neonatal mice was significantly more than that of adult mice. Moreover, immunized neonatal TFH cells expressed significantly more IL-2 as well as its receptors, IL-2Rα and IL-2Rß, than the adult cells. Remarkably, IL-6 co-injection with PCV vaccine further increased the production of IL-2 and the expression of its receptors by neonatal TFH cells, whereas excess IL-6 had totally opposite effect in immunized adult mice. Underscoring the role of IL-6 in activating the IL-2 mediated suppression of vaccine responses, immunization of IL-6 knock-out neonates led to improved antibody responses accompanied by expanded TFH cells as well as lower levels of IL-2 and IL-2 receptors on TFH cells. Moreover, CpG containing PCV improved TFH response in neonates by suppressing the expression of IL-2 receptors on TFH cells and inhibiting IL-2 activity. These findings unveil age-specific differences in IL-6 mediated vaccine responses and highlight the need to consider age-related immunobiological attributes in designing vaccines.

B cell receptor-induced IL-10 production from neonatal mouse CD19+CD43- cells depends on STAT5-mediated IL-6 secretion.

Newborns are unable to reach the adult-level humoral immune response partly due to the potent immunoregulatory role of IL-10. Increased IL-10 production by neonatal B cells has been attributed to the larger population of IL-10-producting CD43+ B-1 cells in neonates. Here, we show that neonatal mouse CD43- non-B-1 cells also produce substantial amounts of IL-10 following B cell antigen receptor (BCR) activation. In neonatal mouse CD43- non-B-1 cells, BCR engagement activated STAT5 under the control of phosphorylated forms of signaling molecules Syk, Btk, PKC, FAK, and Rac1. Neonatal STAT5 activation led to IL-6 production, which in turn was responsible for IL-10 production in an autocrine/paracrine fashion through the activation of STAT3. In addition to the increased IL-6 production in response to BCR stimulation, elevated expression of IL-6Rα expression in neonatal B cells rendered them highly susceptible to IL-6-mediated STAT3 phosphorylation and IL-10 production. Finally, IL-10 secreted from neonatal mouse CD43- non-B-1 cells was sufficient to inhibit TNF-α secretion by macrophages. Our results unveil a distinct mechanism of IL-6-dependent IL-10 production in BCR-stimulated neonatal CD19+CD43- B cells.

Circulating CD138 enhances disease progression by augmenting autoreactive antibody production in a mouse model of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a progressive autoimmune disease characterized by high levels of antibodies directed against nuclear antigens. Elevated serum CD138, a heparan sulfate-bearing proteoglycan, correlates with increased disease activity in patients with SLE, but the contribution of CD138 to lupus disease is not known. Corroborating patient data, we detected an increase in serum CD138 in MRL/MpJ-Faslpr/J (MRL/Lpr) mice (a model for SLE disease) parallel to disease activity. Although T-cell receptor ß (TCRß)+CD138+ T cells typically expand in MRL/Lpr mice as the disease progresses, surprisingly, TCRß+CD138- cells were the primary source of circulating CD138, as the transfer of TCRß+CD138- cells, but not TCRß+CD138+ cells, to young MRL/Lpr mice resulted in higher serum CD138 in the recipients. We found that trypsin was able to cleave CD138 from TCRß+CD138+ cells, and that trypsin was highly expressed in TCRß+CD138- cells. Moreover, trypsin inhibitors, the "defined trypsin inhibitor" and leupeptin, increased CD138 expression on TCRß+CD138- cells, suggesting a contribution of cleaved CD138 to the increase in blood CD138 levels. Furthermore, soluble CD138 was able to bind "a proliferation-inducing ligand" (APRIL) and enhance APRIL-mediated plasma cell generation and autoreactive antibody production through the phosphorylation of extracellular signal-regulated kinase in B cells. The APRIL receptor "transmembrane activator, calcium modulator, and cyclophilin ligand interactor" was involved in the enhancement of APRIL activity by CD138, as the synergistic effect of APRIL and CD138 was ablated in transmembrane activator, calcium modulator, and cyclophilin ligand interactor-deficient B cells. These findings indicate a regulatory role for soluble CD138 in B-cell differentiation and autoreactive antibody production in SLE disease.

Neonatal mice resist Plasmodium yoelii infection until exposed to para-aminobenzoic acid containing diet after weaning.

We developed a newborn (NB) mouse Plasmodium yoelii NL infection model to study malaria in early age. Surprisingly, the onset of parasitemia in P. yoelii challenged NB mice was delayed compared to adults and coincided with the weaning date when weanlings switched from maternal milk to normal chow diet. Also, compared to adult mice, parasitemia resolved much later (48 days vs 20 days post challenge) and the peak parasitemia was twice as high in weanlings. Concurrently, weanlings' germinal center reaction was delayed and diminished compared to adult mice. Maternal milk is deficient in para-aminobenzoic acid (PABA), which is required for de novo folate synthesis by Plasmodium. Suggesting a possible role for the protection afforded by PABA-deficient maternal milk, mice fed with a PABA-deficient diet after the weaning continued to control parasitemia. Despite the reduced parasitemia, these mice developed robust T follicular helper (Tfh) responses and were protected from a second P. yoelii challenge. The NB malaria model provides mechanistic insight into the human infant malaria manifestations where a diet solely based on breast-feeding reduces the incidence of severe malaria in infants. NB mice experiments also support further studies to investigate dietary PABA restriction in the management of severe malaria in infants.

Memory CD73+IgM+ B cells protect against Plasmodium yoelii infection and express Granzyme B.

To better understand anti-malaria protective immune responses, we examined the cellular mechanisms that govern protective immunity in a murine Plasmodium yoelii 17X NL (PyNL) re-infection model. Initially, we confirmed that immune B cells generated during a primary PyNL infection were largely responsible for protection from a second PyNL infection. Using the previously identified memory B cell markers CD80, PD-L2, and CD73, we found an increase in the frequency of CD80-PD-L2-CD73+ B cells up to 55 days after a primary PyNL infection and at 4-6 days following a second PyNL infection. Moreover, injection of enriched immune CD19+CD73+ B cells into nonimmune mice were significantly more protective against a PyNL infection than CD73- B cells. Interestingly, a substantial fraction of these CD73+ B cells also expressed IgM and granzyme B, a biomolecule that has been increasingly associated with protective responses against malaria.

Disease Stage-Specific Pathogenicity of CD138 (Syndecan 1)-Expressing T Cells in Systemic Lupus Erythematosus.

CD138 (syndecan 1), a member of the heparan-sulfate proteoglycan family, regulates diverse biological responses by interacting with chemokines, cytokines, growth factors, and adhesion molecules. Expression of CD138 has been detected on T cells from both healthy and sick mice mimicking systemic lupus erythematosus (SLE) disease. However, the characteristics and the role of CD138+ T cells in SLE pathogenesis remain largely unknown. We analyzed the lupus-prone MRL/Lpr mice and the control MRL/MpJ strain as well as the common laboratory strains Balb/c, and C57BL/6 for CD138-expression and found that only the MRL/Lpr strain harbored TCRß+CD138+ cells in various organs. The frequency of TCRß+CD138+ cells progressively expanded in MRL/Lpr mice with age and correlated with disease severity. Majority of the TCRß+CD138+ cells were CD4 and CD8 double-negative and 20% were CD4. At least a portion of TCRß+CD138+ cells originated from CD4+ cells because substantial number of CD4+TCRß+CD138- cells expressed CD138 after in vitro cultivation. Compared to TCRß+CD138- cells, TCRß+CD138+ cells exhibited central memory (Tcm) phenotype with reduced ability to proliferate and produce the cytokines IFNγ and IL-17. When co-cultured with B cells, the ability of TCRß+CD138+ cells to promote plasma cell formation and autoreactive antibody production was dependent on the presence of autoantigen, CD4 co-receptor expression and cell-to-cell contact. Surprisingly, adoptively transferred TCRß+CD138+ T cells slowed down disease progression in young recipient MRL/Lpr mice but had the opposite effect when DNA was co-administered with TCRß+CD138+ T cells or when TCRß+CD138+ cells were transferred to older MRL/Lpr mice with established disease. Thus, CD138-expressing T cells with Tcm phenotype enhance disease progression in SLE by rapidly activating autoreactive B cells when self-antigens are exposed to the immune system.

TACI Contributes to Plasmodium yoelii Host Resistance by Controlling T Follicular Helper Cell Response and Germinal Center Formation.

The delay in parasite-specific B cell development leaves people in malaria endemic areas vulnerable to repeated Plasmodium infections. Here, we investigated the role of transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), a molecule involved in the generation of antigen-specific antibody secreting cells, in host response to non-lethal Plasmodium yoelii infection. We found that TACI deficiency not only resulted in higher peak parasitemia levels in P. yoelii challenged mice, but also led to a delay in parasite clearance and anti-P. yoelii Merozoite Surface Protein 1(C-terminal 19-kDa fragment [rMSP-119]) protein and anti-rMSP-119 and anti-P. yoelii IgG antibody development. There was also a delay in the generation of splenic high affinity antibody secreting cells that recognize rMSP-119 protein as compared to wild-type mice. Interestingly, coinciding with the delay in parasite clearance there was a delay in the resolution of T follicular helper (TFH) cell and germinal center (GC) B cell responses in TACI -/- mice. The persistence of TFH and GC B cells is likely a result of enhanced interaction between TFH and GC B cells because inducible costimulator ligand (ICOSL) expression was significantly higher on TACI -/- GC B cells than wild-type cells. The difference in the kinetics of GC reaction appeared to also impact the emergence of plasma cells (PC) because there was a delay in the generation of TACI -/- mice PC. Nevertheless, following the recovery from P. yoelii infection, TACI -/- and wild-type mice were both protected from a rechallenge infection. Establishment of protective B cell response was responsible for the resolution of parasitemia because B cells purified from recovered TACI -/- or wild-type mice were equally protective when introduced to naïve wild-type mice prior to P. yoelii challenge. Thus, despite the increased susceptibility of TACI -/- mice to P. yoelii infection and a delay in the development of protective antibody levels, TACI -/- mice are able to clear the infection and resist rechallenge infection.

TACI-Deficient Macrophages Protect Mice Against Metaflammation and Obesity-Induced Dysregulation of Glucose Homeostasis.

Transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) is a receptor for the TNF superfamily cytokines, B cell-activating factor (BAFF), and A proliferation-inducing ligand (APRIL). Here, we demonstrate that TACI-deficient mice subjected to high-fat diet (HFD) are protected from weight gain and dysregulated glucose homeostasis. Resistance to HFD-induced metabolic changes in TACI-deficient mice does not involve TACI-mediated adipogenesis. Instead, accumulation of M2 macrophages (Mϕs), eosinophils, and type 2 innate lymphoid cells in visceral adipose tissue (VAT) is implicated in the protection from obesity-induced assaults. In support of this hypothesis, adoptively transferred TACI-deficient peritoneal or adipose tissue Mϕs, but not B cells, can improve glucose metabolism in the obese host. Interestingly, the transferred TACI-deficient Mϕs not only home to host VAT but also trigger the accumulation of host M2 Mϕs and eosinophils in VAT. The increase in host M2 Mϕs in VAT is likely a result of eosinophil recruitment in response to eotaxin-2 produced by TACI-deficient Mϕs. Insulin signaling experiments revealed that IL-10 secreted by TACI-deficient Mϕs is responsible for maintaining adipocyte insulin sensitivity. Thus, the adoptive transfer experiments offer a model where TACI-deficient Mϕs accumulate in VAT and protect against metaflammation and obesity-associated dysregulation of glucose metabolism.

Delayed onset of autoreactive antibody production and M2-skewed macrophages contribute to improved survival of TACI deficient MRL-Fas/Lpr mouse.

Anti-B cell activating factor belonging to TNF-family (BAFF) antibody therapy is indicated for the treatment of patients with active systemic lupus erythematosus (SLE). We hypothesized that the BAFF receptor, transmembrane activator and calcium-modulator and cyclophilin interactor (TACI) may be responsible for the generation of antibody secreting plasma cells in SLE. To test this hypothesis, we generated TACI deficient MRL-Fas/Lpr (LPR-TACI-/-) mouse. TACI deficiency resulted in improved survival of MRL-Fas/Lpr mice and delayed production of anti-dsDNA and anti-SAM/RNP antibodies. There was also a delay in the onset of proteinuria and the accumulation of IgG and inflammatory macrophages (Mϕs) in the glomeruli of young LPR-TACI-/- mice compared to wild-type mice. Underscoring the role of TACI in influencing Mϕ phenotype, the transfer of Mϕs from 12-week-old LPR-TACI-/- mice to age-matched sick wild-type animals led to a decrease in proteinuria and improvement in kidney pathology. The fact that, in LPR-TACI-/- mouse a more pronounced delay was in IgM and IgG3 autoreactive antibody isotypes and the kinetics of follicular helper T (Tfh) cell-development was comparable between the littermates suggest a role for TACI in T cell-independent autoantibody production in MRL-Fas/Lpr mouse prior to the onset of T cell-dependent antibody production.

IL-6 Impairs Vaccine Responses in Neonatal Mice.

The inability of infants to mount proper follicular helper T (TFH) cell response renders this age group susceptible to infectious diseases. Initial instruction of T cells by antigen presenting cells and subsequent differentiation into TFH cells are controlled by T cell receptor signal strength, co-stimulatory molecules and cytokines such as IL-6 and IL-21. In immunized adults, IL-6 promotes TFH development by increasing the expression of CXCR5 and the TFH master transcription factor, B cell lymphoma 6. Underscoring the importance of IL-6 in TFH generation, we found improved antibody responses accompanied by increased TFH cells and decreased follicular regulatory helper T (TFR) cells, a Foxp3 expressing inhibitory CD4+ T cell occupying the germinal center (GC), when a tetanus toxoid conjugated pneumococcal polysaccharide type 14 vaccine was injected in adult mice together with IL-6. Paradoxically, in neonates IL-6 containing PPS14-TT vaccine suppressed the already impaired TFH development and antibody responses in addition to increasing TFR cell population. Supporting the diminished TFH development, we detected lower frequency of phospho-STAT-3+ TFH in immunized neonatal T cells after IL-6 stimulation than adult cells. Moreover, IL-6 induced more phospho-STAT-3+ TFR in neonatal cells than adult cells. We also measured lower expression of IL-6R on TFH cells and higher expression on TFR cells in neonatal cells than adult cells, a possible explanation for the difference in IL-6 induced signaling in different age groups. Supporting the flow cytometry findings, microscopic examination revealed the localization of Treg cells in the splenic interfollicular niches of immunized adult mice compared to splenic follicles in neonatal mice. In addition to the limitations in the formation of IL-21 producing TFH cells, neonatal mice GC B cells also expressed lower levels of IL-21R in comparison to the adult mice cells. These findings point to diminished IL-6 activity on neonatal TFH cells as an underlying mechanism of the increased TFR: TFH ratio in immunized neonatal mice.

The Role of BAFF System Molecules in Host Response to Pathogens.

The two ligands B cell-activating factor of the tumor necrosis factor family (BAFF) and a proliferation-inducing ligand (APRIL) and the three receptors BAFF receptor (BAFF-R), transmembrane activator and calcium-modulating cyclophilin ligand interactor (TACI), and B cell maturation antigen (BCMA) are members of the "BAFF system molecules." BAFF system molecules are primarily involved in B cell homeostasis. The relevance of BAFF system molecules in host responses to microbial assaults has been investigated in clinical studies and in mice deficient for each of these molecules. Many microbial products modulate the expression of these molecules. Data from clinical studies suggest a correlation between increased expression levels of BAFF system molecules and elevated B cell responses. Depending on the pathogen, heightened B cell responses may strengthen the host response or promote susceptibility. Whereas pathogen-mediated increases in the expression levels of the ligands and/or the receptors appear to promote microbial clearance, certain pathogens have evolved to ablate B cell responses by suppressing the expression of TACI and/or BAFF-R on B cells. Other than its well-established role in B cell responses, the TACI-mediated activation of macrophages is also implicated in resistance to intracellular pathogens. An improved understanding of the role that BAFF system molecules play in infection may assist in devising novel strategies for vaccine development.

MRL Strains Have a BAFFR Mutation without Functional Consequence.

It has been shown that B cell activating factor receptor (BAFFR) is critical for B cell development and survival. In this study, we sought to evaluate the expression and function of BAFFR across multiple stains of mice that vary in their potential to develop systemic autoimmune disease. The inability of a commercial antibody to bind to BAFFR in the autoimmune prone mouse strains, MRL and MRL/Lpr led to the discovery of a mutation in TNFRSF13C gene (encoding BAFFR) that resulted in a Pro44Ser substitution in the N-terminus near the BAFF binding site in these strains. To define the biological consequences of mutant BAFFR, we compared the expression and activity of BAFFR in MRL and MRL/Lpr mice to BALB/c, which express the consensus version of TNFRSF13C. B cells from MRL and MRL/Lpr mice expressed mutant BAFFR on surface and were capable of responding to BAFF as exhibited by BAFF-mediated reduction in apoptosis and NF-κB2 activation. Signaling through MAPK ERK1/2 was not significantly induced by BAFF in MRL/Lpr mice; however, MAPK ERK1/2 signaling was intact in MRL mice. The inability of MRL/Lpr B cells to significantly activate ERK1/2 in response to BAFF was due to the high basal activity of the signaling pathway in these cells. In fact, basal activity of ERK1/2 in B cells correlated with the degree of autoimmune susceptibility exhibited by each strain. In addition, aged MRL/Lpr mice with severe autoimmune disease had high BAFF levels, low surface BAFFR, and high basal NF-κB2 activation, a pattern which is attributed to the high frequency of antibody secreting cells. We conclude that P44S BAFFR mutation does not hinder BAFFR function or enhance B cell activity in MRL/Lpr and MRL mice and that other susceptibility loci on the MRL background contributed to the hyperactivity of these cells.

Distinct Mechanisms Underlie Boosted Polysaccharide-Specific IgG Responses Following Secondary Challenge with Intact Gram-Negative versus Gram-Positive Extracellular Bacteria.

Priming of mice with intact, heat-killed cells of Gram-negative Neisseria meningitidis, capsular serogroup C (MenC) or Gram-positive group B Streptococcus, capsular type III (GBS-III) bacteria resulted in augmented serum polysaccharide (PS)-specific IgG titers following booster immunization. Induction of memory required CD4(+) T cells during primary immunization. We determined whether PS-specific memory for IgG production was contained within the B cell and/or T cell populations, and whether augmented IgG responses following booster immunization were also dependent on CD4(+) T cells. Adoptive transfer of purified B cells from MenC- or GBS-III-primed, but not naive mice resulted in augmented PS-specific IgG responses following booster immunization. Similar responses were observed when cotransferred CD4(+) T cells were from primed or naive mice. Similarly, primary immunization with unencapsulated MenC or GBS-III, to potentially prime CD4(+) T cells, failed to enhance PS-specific IgG responses following booster immunization with their encapsulated isogenic partners. Furthermore, in contrast to GBS-III, depletion of CD4(+) T cells during secondary immunization with MenC or another Gram-negative bacteria, Acinetobacter baumannii, did not inhibit augmented PS-specific IgG booster responses of mice primed with heat-killed cells. Also, in contrast with GBS-III, booster immunization of MenC-primed mice with isolated MenC-PS, a TI Ag, or a conjugate of MenC-PS and tetanus toxoid elicited an augmented PS-specific IgG response similar to booster immunization with intact MenC. These data demonstrate that memory for augmented PS-specific IgG booster responses to Gram-negative and Gram-positive bacteria is contained solely within the B cell compartment, with a differential requirement for CD4(+) T cells for augmented IgG responses following booster immunization.

TACI deficiency leads to alternatively activated macrophage phenotype and susceptibility to Leishmania infection.

The TNF family member, transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), is a key molecule for plasma cell maintenance and is required in infections where protection depends on antibody response. Here, we report that compared with WT mouse, TACI KO Μϕs expressed lower levels of Toll-like receptors (TLRs), CD14, myeloid differentiation primary response protein 88, and adaptor protein Toll/IL-1 receptor domain-containing adapter-inducing IFN-ß and responded poorly to TLR agonists. Analysis of Μϕ phenotype revealed that, in the absence of TACI, Μϕs adapt the alternatively activated (M2) phenotype. Steady-state expression levels for M2 markers IL-4Rα, CD206, CCL22, IL-10, Arg1, IL1RN, and FIZZ1 were significantly higher in TACI KO Μϕ than in WT cells. Confirming their M2 phenotype, TACI-KO Mϕs were unable to control Leishmania major infection in vitro, and intradermal inoculation of Leishmania resulted in a more severe manifestation of disease than in the resistant C57BL/6 strain. Transfer of WT Μϕs to TACI KO mice was sufficient to significantly reduce disease severity. TACI is likely to influence Mϕ phenotype by mediating B cell-activating factor belonging to the TNF family (BAFF) and a proliferation inducing ligand (APRIL) signals because both these ligands down-regulated M2 markers in WT but not in TACI-deficient Μϕs. Moreover, treatment of Μϕs with BAFF or APRIL enhanced the clearance of Leishmania from cells only when TACI is expressed. These findings may have implications for understanding the shortcomings of host response in newborns where TACI expression is reduced and in combined variable immunodeficiency patients where TACI signaling is ablated.

BAFF receptor and TACI in B-1b cell maintenance and antibacterial responses.

Although evidence of the protective immunity conferred by B-1b cells (CD19(+) B220(+) IgM(hi) Mac1(+) CD5(-)) has been established, the mechanisms governing the maintenance and activation of B-1b cells following pathogen encounter remain unclear. B cell-activating factor (BAFF) and a proliferation-inducing ligand (APRIL) mediate their function in mature B cells through the BAFF receptor (BAFFR) and transmembrane activator and CAML interactor (TACI). BAFFR-deficient mice have lower numbers of B-1b cells, and this reduction is directly proportional to BAFFR levels. The generation of B-1b cells is also dependent on the strength of B cell receptor (BCR) signaling. Mice with impaired BCR signaling, such as X-linked immunodeficient (xid) mice, have B-1b cell deficiency, indicating that both BCR- and BAFFR-mediated signaling are critical for B-1b cell homeostasis. Borrelia hermsii induces expansion and persistence of B-1b cells in xid mice, and these B-1b cells provide a heightened protective response. Toll-like receptor (TLR)-mediated stimulation of xid B cells results in a significant increase in TACI expression and restoration of TACI-mediated functions. The activation of TLR signaling by B. hermsii and BCR/TLR costimulation-mediated upregulation of BAFFR and TACI on B-1b cells suggests that B-1b cell maintenance and function following bacterial exposure may depend on BAFFR- and TACI-mediated signaling. In fact, the loss of both BAFFR and TACI results in a greater impairment in anti-B. hermsii responses compared to deficiency of BAFFR or TACI alone.

Impaired B cell receptor signaling is responsible for reduced TACI expression and function in X-linked immunodeficient mice.

Immune response to T cell independent type 2 (TI-2) Ags, such as bacterial polysaccharides, is severely impaired in X-linked immunodeficient (XID) mice. In this study, we investigated the involvement of a proliferation-inducing ligand (APRIL) or BAFF and their receptors in the unresponsiveness of XID mouse to TI-2 Ags. We discovered that whereas serum BAFF levels were increased, the expression of the APRIL and BAFF receptor transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI) was severely reduced in XID B cells. Moreover, B cells from XID mouse were unable to secrete Igs in response to APRIL or BAFF. In correlation with reduced TACI expression and impaired TACI function, APRIL or BAFF did not activate the classical NF-κB pathway in XID cells. Also correlating with the unaltered expression of BAFF receptor, BAFF stimulation induced the activation of the alternative NF-κB pathway in XID cells. Moreover, activation of MAPK pathway was ablated in APRIL-stimulated XID cells. Prestimulation of XID B cells with the TLR9 agonist, CpG led to a significant increase in TACI expression and restored TACI-mediated functions. CpG prestimulation also restored TACI-mediated signaling in APRIL- or BAFF-stimulated XID B cells. Finally, immunization of XID mouse with the prototype TI-2 Ag NP-Ficoll induced IgG and IgM Abs when CpG was given with NP-Ficoll. Collectively, these results suggest that reduced TACI expression is responsible for the unresponsiveness of XID mouse to TI-2 Ags and BCR activation controls TACI expression.

A host-restricted viral vector for antigen-specific immunization against Lyme disease pathogen.

Newcastle disease virus (NDV) is an avian virus that is attenuated in primates and is a potential vaccine vector for human use. We evaluated NDV as a vector for expressing selected antigens of the Lyme disease pathogen Borrelia burgdorferi. A series of recombinant NDVs were generated that expressed intracellular or extracellular forms of two B. burgdorferi antigens: namely, the basic membrane protein A (BmpA) and the outer surface protein C (OspC). Expression of the intracellular and extracellular forms of these antigens was confirmed in cultured chicken cells. C3H or Balb/C mice that were immunized intranasally with the NDV vectors mounted vigorous serum antibody responses against the NDV vector, but failed to mount a robust response against either the intracellular or extracellular forms of BmpA or OspC. By contrast, a single immunization of hamsters with the NDV vectors via the intranasal, intramuscular, or intraperitoneal route resulted in rapid and rigorous antibody responses against the intracellular or extracellular forms of BmpA and OspC. When groups of hamsters were separately inoculated with various NDV vectors and challenged with B. burgdorferi (10(8)cells/animal), immunization with vector expressing either intracellular or extracellular BmpA was associated with a significant reduction of the pathogen load in the joints. Taken together, our studies highlighted the importance of NDV as vaccine vector that can be used for simple yet effective immunization of hosts against bacterial infections including Lyme disease.