Macrophage-derived complement component C4 can restore humoral immunity in C4-deficient mice

Mice with a disrupted C4 locus (C4(-/-)) have an impaired immune response to thymus-dependent Ags. To test the role of bone marrow-derived C4 in humoral immunity, we reconstituted deficient animals with wild-type bone marrow or an enriched fraction of bone marrow-derived macrophages. C4 chimeras were immunized with 4-hydroxy-3-nitrophenyl(5) conjugated to keyhole limpet hemocyanin (NP(5)- KLH) or infected with HSV-1, and the Ab response was evaluated. Wild-type bone marrow rescued the humoral immune response to both Ags, i.e., the soluble Ag and HSV-1, demonstrating that local C4 production is sufficient for humoral responses. Although the C4 chimeric animals lacked detectable C4 in their sera, C4 mRNA was identified in splenic sections by in situ hybridization, and C4 protein deposits were identified in the germinal center areas of splenic follicles by immunofluorescence staining. Macrophages derived from bone marrow produced sufficient C4 protein to restore the humoral response to NP(5)-KLH in C4-deficient animals when administered along with Ag. Cell-sorting experiments, followed by C4-specific RT-PCR, identified splenic macrophages (CD11b(+), CD11c(-)) as a cellular source for C4 synthesis within the spleen.     

Defective vaccine-induced immunity to Schistosoma mansoni in P strain mice. I. Analysis of antibody responses

Inbred P4 strain mice have previously been shown to be uniquely defective in their resistance to challenge infection induced by irradiated cercariae of Schistosoma mansoni. To assess whether the low levels of resistance developed by vaccinated P mice could be due to a defective antibody response, we compared the anti-schistosomulum antibody responses in vaccinated P animals with those occurring in vaccinated C57BL/6J (B6) mice, a strain that consistently develops high levels of resistance to challenge infection. Our results indicate that vaccinated P mice develop levels of total anti-schistosomulum antibodies that are significantly lower than those occurring in B6 mice for at least 15 wk after immunization, with the exception of the fifth week, at which time the responses are indistinguishable. Further analysis revealed that the defect in P strain antibody response occurs specifically in the IgM isotype and that specific IgM levels in P mice are less than one-half the levels in B6 mice at every time point examined. In contrast, no differences in total IgM immunoglobulins were evident when sera from normal (nonvaccinated) P and B6 mice were compared. P mouse anti-schistosomulum IgG antibody responses reached the same levels as those observed in B6 mice by 5 wk after vaccination. However, a much faster decay in IgG antibody levels occurred after this time point in P animals. No differences were observed when the levels of anti-schistosomulum antibodies occurring in each of the major IgG isotypes (IgG1, IgG2a, IgG2b, IgG3) were compared in sera from P and B6 mice vaccinated 4 wk previously. Similarly, vaccinated P and B6 mice were found to mount indistinguishable IgG anamnestic responses after challenge infection. Finally, no differences between vaccinated P and B6 mice were observed when immediate (30 min) skin test and mast cell degranulation responses to a soluble schistosome antigenic preparation were compared. The above findings suggest that P strain mice have a specific defect in their ability to mount IgM antibody responses after immunization with irradiated cercariae. The possible contribution of this defect in IgM response to the decreased resistance of vaccinated P mice to challenge infection is discussed.     

Defective vascular development in connexin 45-deficient mice

In order to reveal the biological function(s) of the gap-junction protein connexin 45 (Cx45), we generated Cx45-deficient mice with targeted replacement of the Cx45-coding region with the lacZ reporter gene. Heterozygous Cx45(+/)(-) mice showed strong expression of the reporter gene in vascular and visceral smooth muscle cells. Cx45-deficient embryos exhibited striking abnormalities in vascular development and died between embryonic day (E) 9.5 and 10.5. Differentiation and positioning of endothelial cells appeared to be normal, but subsequent development of blood vessels revealed impaired formation of vascular trees in the yolk sac, impaired allantoic mesenchymal ingrowth and capillary formation in the labyrinthine part of the placenta, and arrest of arterial growth, including a failure to develop a smooth muscle layer surrounding the major arteries of the embryo proper. As a consequence, the hearts of most Cx45-deficient embryos were dilated. The abnormal development of the vasculature in the yolk sac of Cx45(-)(/)(-) embryos could be caused by defective TGFbeta signalling, as the amount of TGF beta1 protein in the epithelial layer of the yolk sac was largely decreased in the E9.5 Cx45(-)(/)(-) embryo, compared with the wild-type embryo. The defective vascular development was accompanied by massive apoptosis, which began in some embryos at E8.5 and was abundant in virtually all tissues of the embryos at E9.5. We conclude that in Cx45(-)(/)(-) embryos, vasculogenesis was normal, but subsequent transformation into mature vessels was interrupted. Development of different types of vessels was impaired to a varying extent, which possibly reflects the complementation by other connexin(s).     

Defective osteoblast function in ICAP-1-deficient mice

The integrin receptor family plays important roles in cell-to-cell and cell-to-extracellular matrix interactions through the recruitment of accessory molecules. One of them, the integrin cytoplasmic domain-associated protein-1 (ICAP-1; also known as ITGB1BP1), specifically interacts with the cytoplasmic domain of the beta1 integrin subunit and negatively regulates its function in vitro. To address the role of ICAP-1 in vivo, we ablated the Icap-1 gene in mice. We report an unexpected role of ICAP-1 in osteoblast function during bone development. Icap-1-deficient mice suffer from reduced osteoblast proliferation and delayed bone mineralization, resulting in the retarded formation of bone sutures. In vitro studies reveal that primary and immortalized Icap-1-null osteoblasts display enhanced adhesion and spreading on extracellular matrix substrates, probably owing to an increase in beta1 integrin activation. Finally, we provide evidence that ICAP-1 promotes differentiation of osteoprogenitors by supporting their condensation through modulating the integrin high affinity state.     

Role of t lymphocytes in the humoral immune response. II. T cell-mediated regulation of antibody avidity.

The effect of activated T lymphocytes (ATC) on the avidity distribution of PFC in the secondary response was studied in normal mice. The total PFC response was not significantly changed for either direct or indirect PFC by administration of ATC before secondary antigen challenge. However, marked suppression occurred of indirect PFC that secreted high avidity antibody; no suppression was seen of high avidity direct PFC. At the same time, significant stimulation was seen of relative and absolute frequencies of indirect PFC that secreted middle and low avidity antibody. These effects were dependent on Thy 1-bearing, nylon nonadherent cells which demonstrated carrier specificity. In further characterization of these effects, it was found that increasing the number of ATC transferred produced progressive loss of high avidity PFC and compensatory increase in lower avidity PFC. Moreover, in these experiments, suppression of the high avidity response was inducible with the administration of ATC 5 weeks before to 3 days after the secondary immunization. Thus, it is likely that the avidity-modifying effects are dependent on T lymphocytes which influence the late stages of B lymphocyte maturation.     

Chronic spinal cord injury impairs primary antibody responses but spares existing humoral immunity in mice

Spinal cord injury (SCI) results in immune depression. To better understand how injury inhibits humoral immunity, the effects of chronic thoracic SCI on B cell development and immune responses to thymus-independent type 2 and thymus-dependent Ags were determined. Mice received complete crush injury or control laminectomy at either thoracic level 3, which disrupts descending autonomic control of the spleen, or at thoracic level 9, which conserves most splenic sympathetic activity. Although mature B cell numbers were only mildly reduced, bone marrow B cell production was transiently but profoundly depressed immediately after injury. Despite the return of normal B cell production 4 wk after SCI, mice receiving thoracic level 3 injury showed a significant reduction in their ability to mount primary thymus-independent type 2 or thymus-dependent immune responses. The latter were marked by decreases in germinal center B cells as well as class-switched high-affinity Ab-secreting cells. Importantly, injury did not affect affinity maturation per se, pre-existing B cell memory, or secondary humoral immune responses. Taken together, these findings show that chronic high thoracic SCI impairs the ability to mount optimal Ab responses to new antigenic challenges, but spares previously established humoral immunity.     

IgM-mediated, T cell-independent suppression of humoral immunity.

The immunological unreactive state occurring in (T,G)-A-L nonresponder mice after secondary antigen challenge was investigated. Syngeneic IgM anti-(T,G)-A-L antibody-containing plasma, transferred at the time of the time of primary challenge, induced persistent suppression of autologous specific antibody production. Removal of plasma IgM with goat anti-mu antisera removed the ability of the plasma to supress. The induction and maintenance of the suppressed state were not different in thymectomized or sham-thymectomized animals. Primed animals subjected to graft-vs-host reaction (GVHR) at the time of secondary challenge switched over to IgG production. Animals suppressed by passive antibody transfer reacted to GVHR, at the time of secondary challenge, with specific IgM but not IgG antibody production. Transfused normal spleen cells partially abrogated suppression only when (suppressed) hosts had been lethally irradiated. Spleen cells from antigen-plus-antibody suppressed donors, upon transfer to previously normal, syngeneic hosts, were less immunocompetent than spleen cells from untreated donors. These data are consistent with a model of IgM mediated, T cell-independent persistent suppression of humoral immunity.     

Ineffective humoral immunity in the elderly

As individuals age, dysfunction of the immune system leads to an increased incidence of infectious disease. Due to the complex network of cellular interactions and the multi-factorial process of aging, numerous impairments in humoral immunity have been reported. Advances in technology have allowed scientists to begin to identify the molecular mechanisms behind the age-associated decline.     

Regulation of the immune response by antibody. II. The effects of different classes of passive guinea pig antibody on humoral and cell-mediated immunity in rats

The ability of different classes of passively administered guinea pig antibody (γ1, γ2, and IgM) to regulate humoral and cell-mediated immunity to flagellin, polymerized flagellin (POL), and sheep red blood cells (SRBC) was investigated in rats. It was found that at high concentrations, all classes of antibody suppressed the primary antibody responses and usually enhanced the delayed-type hypersensitivity induced by the three antigens. With flagellin and SRBC, the different classes of passive antibody varied in their suppressing and enhancing properties, being in the order: γ2 > γ1 = IgM. At low concentrations, γ1 and IgM enhanced the primary antibody response and suppressed the delayed hypersensitivity induced by flagellin. Such an effect was not observed with either POL or SRBC. Priming for a secondary antibody response was less readily suppressed by all classes of passive antibody. The removal of macrophage cytophilic antibody from γ2 converted this antibody to a preparation (γ2 absorbed) which had effects on humoral and cell-mediated immunity approaching that of γ1 antibody.     

Inhibition of humoral immunity in vivo by monoclonal antibody to L3T4: studies with soluble antigens in intact mice

Monoclonal antibody (MAb) to the mouse "helper" T cell antigen L3T4 inhibits the T cell response to class II major histocompatibility antigens on antigen-presenting cells in vitro and in thymectomized mice. To examine the effect of MAb to L3T4 on humoral immunity in euthymic mice, we treated BALB/c mice with 1 mg of anti-L3T4 i.p. at the time of immunization with either bovine serum albumin (BSA) or chicken egg ovalbumin (OA) in complete Freund's adjuvant. Administration of MAb to L3T4 selectively depleted greater than 90% of L3T4+ cells from the blood, spleen, and lymph nodes, but it had little effect on thymocytes. Mice treated with anti-L3T4 were unable to generate an IgG response to either BSA or OA. Treatment with anti-L3T4 also prevented the antigen-specific IgM response to these antigens, although it did not prevent nonspecific stimulation of IgM anti-BSA and anti-OA antibodies induced by adjuvant in the absence of antigen. Humoral immunity was inhibited even when treatment was delayed until 48 hr after immunization. These findings indicate that T cell help for humoral immunity can be abrogated in intact mice by MAb to L3T4.     

Opiate activity of a bone marrow humoral factor which stimulates antibody production

The radioceptor method was used to demonstrate that humoral factor of the bone marrow, a stimulant of antibody production (SAP), contains substances that competitively remove 3H-morphine and 3H-met-enkephalin from opiate receptors of the rat brain, and 3H-met-enkephalin from specific binding sites on human lymphocytes. Comparison of the magnitudes EC50 SAP obtained during the removal of labeled opiates allows a suggestion to be made that opiates contained by the preparation under study have a greater capability to interact with delta-type opiate receptors rather than with those of the mu-type.