Defensins act as potent adjuvants that promote cellular and humoral immune responses in mice to a lymphoma idiotype and carrier antigens

Defensins released by neutrophils are able to kill a broad spectrum of microbes. They also induce leukocyte migration in vitro and elicit inflammatory leukocyte responses at s.c. injection sites in mice. In vitro experiments showed that human defensins enhanced concanavalin A-stimulated murine spleen cell proliferation and IFN-gamma production. This led us to examine the effects of human defensins on specific immune responses in vivo. BALB/c mice were immunized with 50 microg of keyhole limpet hemocyanin (KLH) adsorbed to aluminum hydroxide and administered with defensins in aqueous solution. Intraperitoneal administration of defensins significantly increased the production of KLH-specific IgG1, IgG2a and IgG2b antibodies 14 days after immunization. In vitro splenic KLH-specific proliferative responses were higher in mice treated with KLH and defensins than in those treated with KLH alone. Increased IFN-gamma and, to a lesser extent, IL-4 production were also detected in the supernatants of ex vivoKLH-activated spleen cells from mice treated with defensins. Finally, defensins significantly enhanced the antibody response to a syngeneic tumor antigen, lymphoma Ig idiotype and also augmented resistance to tumor challenge. These results indicate that defensins act as potent immune adjuvants by inducing the production of lymphokines, which promote T cell-dependent cellular immunity and antigen-specific Ig production. Thus, defensins appear to function as neutrophil-derived signals that promote adaptive immune responses.     

Compartmentalized multicellular crosstalk in lymph nodes coordinates the generation of potent cellular and humoral immune responses

Distributed throughout the body, lymph nodes (LNs) constitute an important crossroad where resident and migratory immune cells interact to initiate antigen-specific immune responses supported by a dynamic 3-dimensional network of stromal cells, that is, endothelial cells and fibroblastic reticular cells (FRCs). LNs are organized into four major subanatomically separated compartments: the subcapsular sinus (SSC), the paracortex, the cortex, and the medulla. Each compartment is underpinned by particular FRC subsets that physically support LN architecture and delineate functional immune niches by appropriately providing environmental cues, nutrients, and survival factors to the immune cell subsets they interact with. In this review, we discuss how FRCs drive the structural and functional organization of each compartment to give rise to prosperous interactions and coordinate immune cell activities. We also discuss how reciprocal communication makes FRCs and immune cells perfect compatible partners for the generation of potent cellular and humoral immune responses.     

The coiled-coil N-terminal domain of the heparin-binding haemagglutinin is required for the humoral and cellular immune responses in mice

Heparin-binding haemagglutinin (HBHA) is a 28-kDa mycobacterial adhesin, composed of three functional domains. Previous work has shown that the C-terminal methylated domain is important for adherence, and it is involved in protective T cell immunity in mouse models. However, the role of the coiled-coil N-terminal domain of HBHA in its overall immunogenic capacity remains elusive. Herein, a comparison of the antibody and cellular immune responses after subcutaneous and intranasal immunization of mice with HBHA (native and recombinant) revealed that the methylation pattern is important but not essential for this property. Subcutaneous immunization of mice with a truncated protein, rHBHADeltaC, which lacks the C-terminal methylated domain, was sufficient to trigger humoral and cellular immune responses to HBHA in mice. Altogether we provide evidence that the coiled-coil N-terminal domain is required for HBHA immunogenicity in vivo.     

Adjuvant effects of orally administered saponins on humoral and cellular immune responses in mice

We have described adjuvant effects of orally administered Quillaja saponins on the immune responses of mice fed inactivated rabies antigen (AG). The in vivo lymphocyte proliferation in mice fed antigen + saponin (AG + SAP) was significantly greater than that in mice fed antigen (AG) alone. Further, the mitogen-induced cell proliferative responses in animals primed with AG + SAP was markedly increased compared with those in the AG group. These changes in clonal expansion were associated with an enhanced helper T cell (Th) and B cell co-operation. The in vivo cell proliferation and in vitro mitogen-induced responses of mice fed AG + SAP correlated with enhanced antibody synthesis. In mice fed saponin alone, there were significant increases in clonal expansion and lymphocyte function. Our present data indicate that the immunocompetence in animals fed AG + SAP was indeed evoked by saponins. Cytotoxic T lymphocyte activity in mice fed SAP or AG + SAP was detected 7 days after booster, in contrast to 21 days in mice fed AG alone. The natural killer cell activity in mice fed SAP alone was greatly enhanced and persisted for an extended period of time.     

土茯苓对细胞免疫和体液免疫的影响

土茯苓水提取物在抗原致敏后及攻击后给药均明显地抑制了picryl chloride(PC)所致的小鼠接触性皮炎(PC-DTH)和绵羊红细胞(SRBC)所致的足蹠反应(SRBC-DTH),其中攻击后给药时作用较强.土茯苓还明显地抑制了二甲苯所致的耳壳及蛋清所致的小鼠足蹠炎症反应。此外,土茯苓对小鼠抗SR-BC 抗体形成的细胞数(IgM-及IgG-PFC 数)无明显影响,但其溶血空斑明显地较对照组为大,同时,血清溶血素水平未见降低,而呈增加趋势。以上结果表明,土茯苓对体液免疫反应无抑制作用,但可选择性地抑制细胞免疫反应,后者主要系影响致敏T 淋巴细胞释放淋巴因子以后的炎症过程。     

Nature of immune lymph node factors stimulating humoral and cellular immune responses

Institute of Immunology, Ministry of Health of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. D. Ado.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 107, No. 4, pp. 460–462, April, 1989.     

Regulatory effects of osteoprotegerin on cellular and humoral immune responses

The interaction between receptor activator of nuclear factor-kappaB ligand (RANKL) and RANK has been reported to regulate immunity in addition to bone metabolism. The aim of this study was to determine if osteoprotegerin (OPG), an inhibitor of the RANKL-RANK interaction and possibly a new drug against osteoporosis, would adversely affect immunity. OPG was used to treat mice developing different models of cellular and humoral immune responses and also in vitro in T and B cell assays. In mice, OPG does not affect cell-mediated reactions such as contact hypersensitivity to the hapten oxazolone and liver damage, granuloma formation, and infectious load induced by mycobacterial infection. However, OPG increases humoral reactions such as the production of IgM, IgG, and IgE against the T cell dependent antigen keyhole limpet hemocyanin and the production of IgM against the T cell independent antigen Pneumovax. In vitro, OPG modestly co-stimulates T cells but does not affect the proliferation of B cells. OPG has modest immunoregulatory effects that seem to be confined to the humoral response to specific antigens.     

Immunization with a synthetic multiepitope antigen induces humoral and cellular immune responses to hepatitis C virus in mice

The immunogenicity of a synthetic multiepitope PCX3 antigen, which contains triple tandem repeats of five conserved epitopes from hepatitis C virus (HCV) polyprotein, was studied in BALB/c mice given three intraperitoneal injections of antigen with Freund's adjuvant. Both a strong antibody response and specific cytotoxic T lymphocytes were induced. The specific anti-PCX3 IgG was able to bind HCV particles from hepatitis C patient sera by incubation overnight. In particular, in transgenic mice with chimeric human livers, anti-PCX3 antibody was able to lower the viral load in two of five mice and to eliminate HCV infection in three of five mice by 2 wk after inoculation with HCV-positive serum from patients. These results indicated that the synthetic multiepitope PCX3 antigen elicits a potent humoral and cellular immune response against HCV.     

Independence of measles-specific humoral and cellular immune responses to vaccination

With a larger, independent cohort and more sophisticated measures, we sought to confirm our work that indicated independence of humoral and cellular immunity following measles vaccination. We recruited an age-stratified random cohort of 764 healthy subjects from all socioeconomic strata, all with medical-record documentation of 2 age-appropriate doses of measles-containing vaccine. We quantified measles-specific neutralizing antibody levels and assayed the interferon-γ (IFN-γ) enzyme-linked immunosorbent spot assay (ELISPOT) response to measles virus. We also measured secreted cytokines from the peripheral blood mononuclear cells (PBMCs) in response to measles virus by performing enzyme-linked immunosorbent assays as secondary measures of cellular immune status. The median antibody level and median IFN-γ ELISPOT response were 844 mIU/mL (interquartile range [IQR]: 418-1,752) and 36 (IQR: 13.00-69.00) spot-forming cells (per 2 × 10(5) PBMCs), respectively. We observed only a very weak and negative correlation (Spearman's r(s) or ρ of -0.090 [95% confidence interval: -0.162--0.018]). We observed a similar lack of quantitatively important correlations between the neutralizing antibody level and any of the secondary measures. Our data confirm the independence of humoral and cellular immune responses after the second dose of measles vaccination. As researchers pursue novel measles vaccine and measles vaccine delivery systems, they must not infer that humoral responses predict cellular responses.     

Interrelations of humoral and cellular immune responses in experimental canine gastritis

Experimental canine gastritis is an animal model with features similar to the gastritis of human pernicious anaemia. In this study eight mongrel dogs immunized with autologous or homologous gastric juice in Freund' scomplete adjuvant developed gastritis whereas four mongrel dogs immunized with the adjuvant alone did not. Cellular immunity to gastric juice antigens demonstrated by skin testing and peripheral leucocyte migration inhibition coincided with the appearance of gastritis, whereas parietal cell antibodies appeared some 2 weeks later. The work demonstrates the combined humoral and cellular immune response of the dogs to gastric juice, and suggests that the cellular response may be of primary importance in the development of gastritis.     

Impairment of cellular but not humoral immune responses in chronic pulmonary and disseminated paracoccidioidomycosis in mice.

Humoral and cellular immune responses were measured during the progression of chronic pulmonary and disseminated paracoccidioidomycosis in mice. The chronic disease was established by pulmonary infection of mice with different doses of the yeast form of Paracoccidioides brasiliensis isolate GAP. Levels of antibodies to P. brasiliensis, detected in serum by immunodiffusion and enzyme-linked immunosorbent assay, directly correlated with the size of the infectious challenge. Significant delayed-type hypersensitivity (DTH) responses to antigen were largely restricted to week 1 after pulmonary infection with intranasally administered high doses (5.0 x 10(6) or 1.1 x 10(7) CFU per inoculum). In vitro lymphoproliferative responses of peripheral blood lymphocytes (PBL) to P. brasiliensis antigens were significant only at 2 weeks after infection with intranasally administered 1.1 x 10(7) CFU. Responses of PBL to concanavalin A were depressed (50% of control response) as early as 8 weeks and reached a nadir at 10 to 18 weeks after infection. Infected mice made antibodies to sheep erythrocytes (SRBC) (10(9) intravenously [i.v.]) normally at all times tested after infection. In contrast, infected mice sensitized to SRBC (10(6) i.v.) had significantly depressed DTH responses to SRBC at 9 and 20 weeks postinfection compared with noninfected mice. These results indicated that in this model, normal humoral responses developed to homologous and heterologous antigens. In contrast, the T cellular immune responses were depressed with progression and chronicity of the disease. Thus, this model closely mimics the immunological findings in human paracoccidioidomycosis.     

Co-delivery of LIGHT expression plasmid enhances humoral and cellular immune responses to HIV-1 Nef in mice

The immunogenicity and efficacy of a DNA vaccine can be greatly enhanced when a gene adjuvant is used. LIGHT, a member of TNF superfamily, can function as a costimulatory molecule for human naïve T cells to proliferate and can be a potential gene adjuvant. In the current study, the eukaryotic expression plasmid pcDNA-nef was constructed by inserting a full-length nef gene into pcDNA3.1(+), and an in vitro transfection experiment suggested that the nef gene could be expressed successfully in mammalian cells. BALB/c mice were immunized with HIV-1 nef DNA vaccine plasmids alone or in combination with LIGHT expression plasmids, and the specific humoral and cellular immune responses were measured. The data showed that HIV-1 nef DNA vaccine plasmids could induce anti-Nef antibodies, Nef-specific lymphocyte proliferation and CTL activity, whereas stronger specific immune responses were induced in mice when co-immunizing with HIV-1 nef DNA vaccine plasmids and LIGHT expression plasmids, suggesting that the eukaryotic expression vector encoding HIV-1 nef is capable of inducing specific immune responses towards HIV-1 Nef and that LIGHT could be considered as a gene adjuvant for HIV-1 DNA vaccination.     

Comparison of specificities of humoral and cellular immune responses to haptens

In this study we induced humoral and cellular immunity to three isomers of aminosulfanilic and aminobenzoic acids and compared their respective specificities. These studies were facilitated by using a new method for preferentially inducing a cellular immune response to haptens: the haptens were covalently coupled to mycobacteria and injected into animals in incomplete Freund's adjuvant. Humoral antibody was iduced by coupling the same isomers to bovine serum albumin and injecting them into animals in complete Freund's adjuvant. Both groups of animals were examined for hapten-specific cellular immunity (migration inhibition factor, delayed skin tests) and for antibody response (passive hemolysin test). The results show that although both cell-mediated and humoral responses can discriminate the para, meta and ortho isomeric forms of the haptens used, the antibody response demonstrated much less cross-reactivity. The relevance of this finding to different requirements for B and T cells in antigenic recognition is discussed.     

Characterization of humoral and cellular immune responses elicited by meningococcal carriage

In order to study the immune response elicited by asymptomatic carriage of Neisseria meningitidis, samples of serum, peripheral blood mononuclear cells (PBMCs), and saliva were collected from a cohort of more than 200 undergraduate students in Nottingham, United Kingdom, who were subject to high rates of acquisition and carriage of meningococci. Serum immunoglobulin G levels were elevated following increases in the rate of carriage, and these responses were specific for the colonizing strains. In order to investigate T-cell responses, PBMCs from 15 individuals were stimulated with a whole-cell lysate of the H44/76 meningococcal strain (B:15:P1.7,16), stained to detect cell surface markers and intracellular cytokines, and examined by flow cytometry. The cells were analyzed for expression of CD69 (to indicate activation), gamma interferon (IFN-gamma) (a representative T-helper 1 subset [Th1]-associated cytokine), and interleukin-5 (IL-5) (a Th2-associated cytokine). Following a brief meningococcal stimulation, the numbers of CD69(+) IFN-gamma(+) CD56/16(+) NK cells were much higher than cytokine-positive CD4(+) events. Both IFN-gamma(+) and IL-5(+) events were detected among the CD69(+) CD4(+) population, leading to the conclusion that an unbiased T-helper subset response was elicited by meningococcal carriage.     

Combination of protein and viral vaccines induces potent cellular and humoral immune responses and enhanced protection from murine malaria challenge

The search for an efficacious vaccine against malaria is ongoing, and it is now widely believed that to confer protection a vaccine must induce very strong cellular and humoral immunity concurrently. We studied the immune response in mice immunized with the recombinant viral vaccines fowlpox strain FP9 and modified virus Ankara (MVA), a protein vaccine (CV-1866), or a combination of the two; all vaccines express parts of the same preerythrocytic malaria antigen, the Plasmodium berghei circumsporozoite protein (CSP). Mice were then challenged with P. berghei sporozoites to determine the protective efficacies of different vaccine regimens. Two immunizations with the protein vaccine CV-1866, based on the hepatitis B core antigen particle, induced strong humoral immunity to the repeat region of CSP that was weakly protective against sporozoite challenge. Prime-boost with the viral vector vaccines, FP9 followed by MVA, induced strong T-cell immunity to the CD8⁺ epitope Pb9 and partially protected animals from challenge. Physically mixing CV-1866 with FP9 or MVA and then immunizing with the resultant combinations in a prime-boost regimen induced both cellular and humoral immunity and afforded substantially higher levels of protection (combination, 90%) than either vaccine alone (CV-1866, 12%; FP9/MVA, 37%). For diseases such as malaria in which different potent immune responses are required to protect against different stages, using combinations of partially effective vaccines may offer a more rapid route to achieving deployable levels of efficacy than individual vaccine strategies.     

Normal development of the thymus-dependent limb of humoral immune responses in mice

The capacity of mice to produce antibody-forming cells (AFC) to sheep erythrocytes and to low doses of two haptenated proteins increases logarithmically with age in the postnatal period. This increase is prevented by thymectomy. These findings, together with those on the ability of suckling mice to make high numbers of AFC to high doses of at least one of the antigens used, indicate that T cells are the limiting cell type in neonatal humoral immune responses. An analysis of variance of the number of AFC in individual spleens as a measure for T cells showed that the data are in agreement with random generation and subsequent proliferation of these cells during ontogenesis. Different antigen reactivities carried by these cells seem to arise at different times during fetal life.     

Immunostimulatory RNA is a potent inducer of antigen-specific cytotoxic and humoral immune response in vivo

Single-stranded RNA stimulates immune cells and induces the secretion of pro-inflammatory cytokines and type I IFN. As adjuvant RNA can induce a T(h)2 type of humoral response, however, its potency in the induction of cytotoxic T cells in vivo has not been analyzed. Here we show that immunization with the antigen ovalbumin (OVA) and the adjuvant phosphodiester RNA complexed to the cationic lipid N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methylsulfate (DOTAP) induced a Toll-like receptor-7-dependent cytotoxic T cell and humoral response. Staining with SIINFEKL-K(b) tetramers demonstrated the induction of antigen-specific T cells that were functional in in vivo cytotoxic T cell assays against SIINFEKL-loaded target cells. In infection experiments with OVA-secreting Listeria monocytogenes, the cytotoxic T cell response strongly reduced the bacterial load in liver and spleen. The RNA-driven humoral response was characterized by OVA-specific antibodies of the IgG1 isotype whereas CpG-DNA induced antigen-specific antibodies of the IgG2a (BALB/c) or IgG2c (C57BL/6) isotype. Furthermore, stimulation with RNA did not induce splenomegaly, a common feature of CpG-DNA-driven immune activation in mice. Taken together, our data confirm that RNA can be used as a safe adjuvant and induces a strong antibody response of the IgG1 isotype. Additionally, we demonstrate that RNA induces an antigen-specific immunity characterized by a potent cytotoxic T cell response to infection.