IL-1 is an effective adjuvant for mucosal and systemic immune responses when coadministered with protein immunogens

Mucosal immunization with soluble protein Ag alone may induce Ag-specific tolerance, whereas mucosal immunization with Ag in the presence of a mucosal adjuvant may induce Ag-specific systemic and mucosal humoral and cell-mediated immune responses. The most widely used and studied mucosal adjuvant is cholera toxin (CT). Although the mechanism of adjuvanticity of CT is not completely understood, it is known that CT induces mucosal epithelial cells to produce the proinflammatory cytokines IL-1, IL-6, and IL-8 and up-regulates macrophage production of IL-1 and the costimulatory molecule B7.2. Because IL-1 may duplicate many of the activities of CT, we evaluated IL-1alpha and IL-1beta for their ability to serve as mucosal adjuvants when intranasally administered with soluble protein Ags. IL-1alpha and IL-1beta were as effective as CT for the induction of Ag-specific serum IgG, vaginal IgG and IgA, systemic delayed-type hypersensitivity, and lymphocyte proliferative responses when intranasally administered with soluble protein Ag. Our results indicate that IL-1alpha and IL-1beta may be useful as mucosal vaccine adjuvants. Such an adjuvant may be useful, and possibly required, for vaccine-mediated protection against pathogens that infect via the mucosal surfaces of the host such as HIV.     

Assessing immune competence in pigs by immunization with tetanus toxoid

Immune competence can be tested by challenging organisms with a set of infectious agents. However, disease control requirements impose restrictions on the infliction of infections upon domestic pigs. Alternatively, vaccinations induce detectable immune responses that reflect immune competence. Here, we tested this approach with tetanus toxoid (TT) in young domestic pigs. To optimize the vaccination protocol, we immunized the pigs with a commercial TT vaccine at the age of 21 or 35 days. Booster immunizations were performed either 14 or 21 days later. TT-specific antibodies in plasma as well as lymphoproliferative responses were determined both 7 and 14 days after booster immunization using ELISA and lymphocyte transformation tests, respectively. In addition, general IgG and IgM plasma concentrations and mitogen-induced proliferation were measured. The highest TT-specific antibody responses were detected when blood samples were collected 1 week after a booster immunization conducted 21 days after primary immunization. The pigs’ age at primary immunization did not have a significant influence on TT-specific antibody responses. Similarly, the TT-specific proliferative responses were highest when blood samples were collected 1 week after booster immunization, while age and time of primary and booster immunization were irrelevant in our setup. While general IgG and IgM plasma levels were highly age dependent, there were no significant age effects for TT-specific immune responses. In addition, mitogen-induced proliferation was independent of immunization as well as blood sampling protocols. In summary, our model of TT vaccination provides an interesting approach for the assessment of immune competence in young pigs. The detected vaccination effects were not biased by age, even though our data were acquired from immune systems that were under development during our tests.     

Mucosal and systemic immune responses to a human immunodeficiency virus type 1 epitope induced upon vaginal infection with a recombinant influenza A virus

The humoral and cellular immune responses in the genital mucosa likely play an important role in the prevention of sexually transmitted infections, including infection with human immunodeficiency virus type 1 (HIV-1). Here we show that vaginal infection of progesterone-treated BALB/c mice with a recombinant influenza virus bearing the immunodominant P18IIIB cytotoxic T-lymphocyte (CTL) epitope of the gp160 envelope protein from an HIV-1 IIIB isolate (P18IIIB; RIQRGPGRAFVTIGK) can induce a specific immune response in regional mucosal lymph nodes, as well as in a systemic site (the spleen). A single inoculation of mice with the recombinant influenza virus induced long-lasting (at least 5 months) antigen-specific CTL memory detectable as a rapid recall of effector CTLs upon vaginal infection with recombinant vaccinia virus expressing HIV-1 IIIB envelope gene products. Long-term antigen-specific CTL memory was also induced and maintained in distant mucosal tissues when mice were intranasally immunized with the recombinant influenza virus. These results indicate that mucosal immunization and, in particular, local vaginal immunization with recombinant influenza virus can provide strong, durable immune responses in the female genital tract of mice.     

Enhanced immune response to pneumococcal infection in malnourished mice nasally treated with heat-killed Lactobacillus casei

The present study analyzed whether nasal administration of viable and non-viable Lactobacillus casei CRL 431 to immunocompromised mice was capable of increasing resistance against Streptococcus pneumoniae. Weaned mice were malnourished after consuming a PFD for 21 days. Malnourished mice were fed a BCD for 7 days or BCD for 7 days with viable or non-viable L. casei nasal treatments on day 6 and day 7 (BCD+LcV and BCD+LcN, respectively). The MNC group received PFD whereas the WNC mice consumed BCD. MNC mice showed greater lung colonization, more severe lung injuries, impaired leukocyte recruitment and reduced antibodies and cytokine production when compared with WNC mice. Administration of L. casei increased the resistance of malnourished mice to the infection. Both BCD+LcV and BCD+LcN treatments prevented the dissemination of the pathogen to the blood and induced its lung clearance. BCD+LcV or BCD+LcN groups showed improved production of TNF-α and activity of phagocytes in the respiratory tract, an effect that was not observed in the BCD control group. In addition, IL-4 and IL-10 were significantly increased in BCD+LcV and BCD+LcN groups, which correlated with the increase in the levels of specific respiratory IgA. The nasal treatments with L. casei were also effective at stimulating the production of specific IgG at both the systemic and the respiratory levels. The comparative study between the viable and the non-viable bacteria demonstrated that viability would be an important factor to achieve maximum protective effects. However, the results from this study suggest that heat-killed lactic acid bacteria are also effective in the immunomodulation of the systemic and respiratory immune system.     

Specific immune responses after booster immunization with tetanus toxoid in man: Study of kinetics,family segregation,and linkage to HLA of in vitro lymphocyte proliferative responses and serum-antibody responses

Kinetics and family transmission of antigen-specific in vitro cell-mediated responses were investigated in 68, and serum-antibody responses to tetanus toxoid (TT) in 73 individuals from a total of 12 families. Proliferative responses to highly purified TT monomer were studied in 6- to 7-day lymphocyte cultures. The effect of booster immunization was detectable 7 (D7) and 30 (D30), but not 120 days (D120) later. The sex of donors was not found to have any influence. A significant influence of the time interval since the last immunization was found for the responses at D7 and D30. Data were correspondingly adjusted for segregation and linkage analyses. Several transmission hypotheses for the data obtained at D7 and D30 were evaluated by likelihood ratio tests. Observations at D30 were compatible with the hypothesis of a control by a dominant genetic determinant for high responses closely linked to the major histocompatibility complex region. No such evidence could be found for D7. After booster immunization, mean antibody levels determined on D7, D30 (peak of response), and D120 were found to be higher than those prior to immunization (D0). The sex of the donors was found to have no influence on antibody responses. The time interval since the last immunization and the age of donors both had a slight influence, and data were correspondingly adjusted for segregation and linkage analyses, which showed no evidence of genetic control of the antibody responses or of linkage to HLA.     

The clearance of tetanus toxoid/anti-tetanus toxoid immune complexes from the circulation of humans. Complement- and erythrocyte complement receptor 1-dependent mechanisms

The role of complement and its receptor on erythrocytes (CR1) in the physiologic elimination of large immune complexes from the circulation of humans was assessed. Large radiolabeled soluble tetanus toxoid- anti-tetanus toxoid complexes were injected i.v. into three normal individuals and three patients with SLE. These complexes were prepared in antibody excess and were 45S in size, fixed C and bound to E CR1 in vitro. The percentage of complexes bound in vitro was directly proportional to CR1 number/E in four normal subjects and three SLE patients. After i.v. injection into normal subjects, complexes were cleared rapidly, with a monoexponential rate constant (10.3 to 11% complexes cleared/min). In the SLE patients, clearance was best explained by two phases: the first occurred within the first minute indicating immediate trapping of a fraction of the complexes (19.5 to 25.3% of injected complexes trapped), the second was monoexponential and was similar to the normal range. A large fraction of complexes bound within the first minute to E in vivo; the percentage of binding was variable, ranging from 16.3% to 71.5% and was related to E CR1 number. In a second study complexes were injected that had been attached to autologous E by opsonization with C in vitro. Their elimination was similarly monoexponential, except in one SLE patient in whom there was significant initial trapping (30.9%). A fraction of these complexes were released from E within the first minute, the percentage release being greatest in the patient with the lowest CR1 number (81.4%). E bearing immune complexes remained in the circulation and were not transiently sequestered in the liver or spleen. This is the first study of the clearance of soluble immune complexes in vivo in humans and shows that C and CR1 on E participate in immune complex clearance reactions, and that abnormal clearance can be detected in the form of rapid removal of immune complexes from the circulation.     

DNA-plasmids of HIV-1 induce systemic and mucosal immune responses

DNA-based immunization has been shown to induce protective immunity against several microbial pathogens including HIV-1. Several routes of DNA vaccination have been exploited. However, the properties of the immune responses seem to differ with the different routes used for DNA delivery, ultimately affecting the outcome of experimental challenge. We measured the primary immune response following one vaccination. This report presents differences associated with three different DNA delivery routes: intramuscular injection, intranasal application, and gene-gun based immunization. Induction of systemic humoral immune responses was achieved most efficiently by either intranasal or gene-gun mediated immunization, followed by intramuscular injection. Mucosal IgA was reproducibly induced by intranasal instillation of the DNA, and found in lung washings, faeces, and vaginal washings. Cytotoxic T cells were not induced by a single immunization, but were observed after three immunizations using intramuscular injections.     

Network connectivity is shown to change in C57BL/6 mice during a continuing immune response subsequent to tetanus toxoid hyperimmunization

We have already demonstrated (Stojanovic et al., 2009) a connection between tetanus toxoid (TTd) hyperimmunization and the induction of anti-phospholipid syndrome (APS) in BALB/c mice. Here we show that C57BL/6 mice subjected to an identical procedure do not exhibit any like pathology attributable to anti-phospholipid antibodies; we explain that this absence results from idiotypic connectivity. Six groups of C57BL/6 mice were hyperimmunized with TTd in aluminum hydroxide or glycerol, with or without pretreatments. Pretreated mice had been injected with polyclonal or nonspecific immune stimulators, such as complete Freund's adjuvant (CFA) or glycerol. The epitope specificity of induced antibodies was tested by indirect ELISA using a tetanus toxoid immunogen and these autoantigens: phospholipids, gangliosides, laminin. Idiotypic connectivity was tested by competitive ELISA and gauged from the degree to which the interaction of idiotypic/anti-idiotypic complementary antibodies was inhibited in the presence of immunized sera antibodies. Higher idiotypic connectivity was noted amongst pretreated mice. There was a positive correlation between higher connectivity and autoantibody levels that acted to favor the participation of natural autoantibodies in the inhibitory process. We conclude that idiotypic connectivity plays a protective role in immunization-induced autoimmunity.     

Intranasal immunization with inactivated influenza virus enhances immune responses to coadministered simian-human immunodeficiency virus-like particle antigens

Intranasal immunization with inactivated influenza virus vaccine can provide protective immunity, whereas many other antigens are less effective when used for mucosal immunization. To determine whether influenza virus could enhance immune responses to an antigen coadministered to a mucosal surface, we studied the intranasal immunization of mice with a mixture of simian-human immunodeficiency virus (SHIV) virus-like particles (VLPs) and inactivated influenza virus. Compared to mice immunized with SHIV VLPs alone, mice coimmunized with SHIV VLPs and inactivated influenza virus showed significant increases in serum immunoglobulin G (IgG) and mucosal IgA antibodies specific to the human immunodeficiency virus envelope protein, neutralizing activities, numbers of gamma interferon- and interleukin 4-secreting lymphocytes, and cytotoxic-T-lymphocyte activities. The levels of enhancement of immune response by coimmunization with inactivated influenza virus were equivalent to those induced by inclusion of immunostimulatory CpG oligodeoxynucleotides (CpG DNA). We also observed that SHIV VLPs bind to influenza virus virions, forming mixed aggregates. These results indicate that inactivated influenza virus can play a role as a mucosal adjuvant to coadministered antigens.     

Spermatocyte chromosome alterations in mice inoculated with antirabies vaccine and tetanus toxoid separately and in combination

The genotoxic potential of antirabies vaccine (ARV) and tetanus toxoid (TT) inoculated separately and in combination was evaluated by spermatocyte chromosome analysis in mice. Adult males were vaccinated with a dose of 1 ml/kg ARV once daily for 7 days, or with a dose of 1 ml/kg TT once only, or with both (TT being administered along with the last dose of ARV). Spermatocytes were analyzed over 1-6 post-vaccination weeks for numerical and structural changes as well as for univalent formation. Controls were age matched non-vaccinated males. Significantly high incidences of structural aberrations from wk 4 onwards and at wk 6 following inoculation of ARV and TT, respectively, suggested susceptibility of stem cells to the vaccines. Combined vaccination (ARV plus TT) showed an additive effect. Occurrence of univalents involving autosomes as well as sex chromosomes was very common in all the vaccinated groups, the sex chromosomes being prevalently involved. Structural aberrations showed a positive correlation with time, while univalent formation showed a negative correlation. It appears that factors for clastogenicity and univalent formation are different for both the vaccines.