Long-term humoral and cellular immunity induced by a single immunization with replication-defective adenovirus recombinant vector

This study examines the suitability of replication-defective adenovirus vectors for engineering recombinant vaccines. The immunological abilities and limitations of E1-deleted adenoviruses containing the lac Z gene (Ad-β-gal) were investigated by examining the humoral and cellular immune responses to the β-galactosidase protein. BALB/c mice (H-2^d) were given in a single injection of recombinant adenovirus. The cytotoxic T lymphocyte (CTL) response of spleen cells was evaluated. Recognized target cells were H-2^d-derived tumor cells transfected by the lac Z gene, or incubated with the 876–884 β-galactosidase peptide known to be restricted by the L^d molecule of the major histocompatibility complex. A long-lasting β-galactosidase-specific cytotoxic T cell response was obtained. By contrast, CTL from mice immunized with the L^d-restricted peptide were less specific for the endogenous epitope presented by the transfectants expressing β-galactosidase. Ad-β-gal-immunized mice were also protected against an intra-cerebral challenge with a recombinant vaccinia virus expressing the lac-Z gene. These results suggest that Ad-β-gal-induced CTL have protective abilities in vivo. The induction of β-galactosidase-specific T helper lymphocytes and humoral IgG responses were also examined. A proliferative response occurred only late after immunization and the primed T lymphocytes produced interleukin-2, but no interleukin-4. A humoral IgG response to the β-galactosidase protein was detected 15–30 days after a single immunization and remained stable for 6 months without boosting. Lastly, we followed the evolution of the immune response over the course of successive immunizations. The magnitude and kinetics of the cellular and humoral responses were similar to those obtained after a single immunization. Consistent with these observations, an adenovirus-specific neutralizing antibody response was detected as early as the second immunization. Thus, a single immunization with a replication-defective adenovirus recombinant vector induces long-lasting humoral and cellular immune responses specific to the transgene product.     

Assay of humoral immunity to mumps virus.

One hundred randomly chosen sera from blood donors from North London were assayed for antibodies to mumps virus by plaque reduction and microtitre neutralisation assay, haemagglutination inhibition and in-house ELISA. The assay reproducibility was determined, and there was reasonable agreement between antibody levels measured by the two neutralising methods. Neutralising antibody levels measured by either method were low but the strain used in the assay had a large effect on the antibody titres observed. Titres measured by neutralisation assay, HI assay and ELISA did not correlate well. Assessment of immunity to mumps virus remains problematical.     

Intranasal immunization with mumps virus DNA vaccine delivered by influenza virosomes elicits mucosal and systemic immunity

To improve the efficiency of liposome-mediated DNA transfer as a tool for gene therapy or vaccinology, we have further developed a new delivery system based on the modified immunopotentiating reconstituted influenza virus (IRIV). In this study, we engineered a plasmid DNA vector expressing the mumps virus hemagglutinin or the fusion protein. The administration of this DNA vaccine delivered by influenza virosomes, in combination with the mucosal adjuvant Escheriagen via the intranasal route, was efficient for inducing an immune response, both mucosally and systemically, in mice. The production of IgG2a mumps virus-specific antibodies and the secretion of interleukin 10 (IL-10) by antigen-specific T cells indicated that not only Th1 but also Th2 responses were induced by this DNA vaccine formulation. These results suggest that cationic virosomes in combination with Escheriagen may have great potential as an efficient delivery system for intranasal DNA immunization and provide an immune barrier at the mucosal sites.     

Heterogeneity of humoral immune abnormalities in children with Nijmegen breakage syndrome: an 8-year follow-up study in a single centre

During an 8-year period of observation, defects of immune responses were characterized and monitored in 40 of 50 Polish children with Nijmegen breakage syndrome referred to the Children's Memorial Health Institute in Warsaw. The following parameters were determined at diagnosis: (1) concentrations of serum IgM, IgG, IgA; (2) concentrations of IgG subclasses; and (3) lymphocyte subpopulations. In addition, naturally acquired specific antibodies against Streptococcus pneumoniae were determined in 20 patients with a history of recurrent respiratory infections. During follow-up, total serum immunoglobulins and IgG subclasses were monitored systematically in 17 patients who did not receive immunomodulatory therapy. Moreover, anti-HBs antibody response was measured after vaccination of 20 children against HBV. We found that the immune deficiency in NBS is profound, highly variable, with a tendency to progress over time. Systematic monitoring of the humoral response, despite good clinical condition, is essential for early medical intervention.     

Comparison of cell-mediated and humoral immunity in the dog lung after localized lung immunization

This study evaluated cell-mediated immunity (CMI) and antibody production serially in control and immunized lung lobes of beagle dogs. A fiberoptic bronchoscope was used to immunize selected airways in the left cardiac lung lobe with 10(10) sheep red blood cells (SRBC); the right cardiac lobe received saline as control. The immune responses produced by this localized lung immunization were evaluated in cells and fluids obtained from blood and serial bronchial washings of the immunized and control lung lobes from 5 to 21 days after immunization. The antigen-specific production of procoagulant activity (LPCA) and inhibition of migration of alveolar macrophages by SRBC antigen were used to measure CMI in lavage cells from immunized and control lung lobes. The level of specific IgM and IgG antibody in lavage fluid from the control and immunized lung lobes was evaluated with the enzyme-linked immunosorbent assay (ELISA). The results of this study showed a significantly greater percentage of neutrophils and lymphocytes in the lavage fluid from the immunized lung lobes than from the control lung lobes. The increased number of lymphocytes in the immunized lung lobes showed a positive correlation with increased antibody concentrations. In contrast to the antibody response, CMI as measured by LPCA and MIF assays was positive, with nearly equal responses in control and immunized lung lobes. Even though there were only a few lymphocytes in the control lung lobes, there were apparently enough specific immune lymphocytes present that produced mediators after antigen stimulation to result in positive CMI responses.     

分子佐剂C3d增强hCGβ基因免疫体液免疫效应

目的：该实验室前期工作已成功构建真核表达质粒pcDNA3-hCGβ、pcDNA3-hCGβ-C3d3且证明其能在真核表达系统中有效表达。通过动物DNA免疫，以期证实分子佐剂C3d增强免疫避孕疫苗免疫原性。方法：抽提与纯化pcDNA3、pcDNA3-hCGβ、pcDNA3-hCGβ-C3d3质粒，进行动物DNA免疫。免疫剂量分别为5、10、20pmol，共免疫2次，每次间隔3周。末次免疫后6周采血，用间接ELISA分析实验动物外周血抗hCGβ抗体效价。结果：C3d分子佐剂能明显提高抗hCGβ抗体滴度；且其作用呈剂量依赖性。结论：分子佐剂确实能增强hCGβ免疫原性；此结果将有助于免疫避孕疫苗的技术进步及实际应用。     

Specific cellular and humoral immunity in children with grass pollen asthma

Lymphocyte stimulation, as determined by incorporation of thymidine, to rye grass extract in twenty-three children with bronchial reactivity to rye grass and to house dust mite, did not differ significantly from four children with reactivity to house dust mite alone, or from nine children with asthma but without a bronchial response to these allergens. Sixteen children underwent hyposensitization with rye grass extract or treatment with placebo. There was no consistent effect of hyposensitization on the lymphocyte stimulation indices to rye grass. A decrease in lymphocyte responsiveness occurred to rye grass and to house dust mite after the grass pollen season but was not statistically significant. Analysis of changes in lymphocyte responsiveness to both house dust mite and rye grass of the children most highly sensitized to rye allergen, showed that the lymphocyte responsiveness to rye grass fell during the pollen season (P<0.05) but this effect was not seen with house dust mite. The study suggests that a decrease in lymphocyte responsiveness to rye grass allergen in children with large amounts of anti-rye IgE antibodies is antigen specific and may be seen following seasonal exposure.     

Tardive dyskinesia: a 3-year follow-up study

A prospective study of tardive dyskinesia was carried out to gain information regarding the natural history of the condition and to identify risk factors. Out of an original cohort of 182 psychiatric patients receiving maintenance antipsychotic drugs 99 were available for reassessment after 3 years. In this follow-up group the point prevalence of oro-facial dyskinesia increased from 39% to 47% over the 3-year period. Twenty-two patients developed the disorder, while remission occurred in 14 others. Risk factors predicting the presence of oro-facial dyskinesia at follow-up included being over 50 years of age and the presence of akathisia. There was no convincing association between the duration of antipsychotic drug treatment and the presence or severity of oro-facial dyskinesia. Patients receiving over 1000 mg chlorpromazine equivalents of antipsychotic drug per day were unlikely to have the condition. The amount of purposeless trunk and limb movement present proved to be a relatively stable phenomenon, showing only a slight increase with age and no change over the follow-up period. The implications of these findings are discussed, with particular consideration being given to the effects of loss of patients to follow-up.     

Induction of humoral immunity and pulmonary mast cells in mice and rats after immunization with aerosolized antigen.

Rats (BN X Wistar) and mice (CBA/Ca) were immunized by exposure in 10-day periods to an aerosol of ovalbumin (OA). In rats this immunization resulted in IgE antibodies detectable at very low levels in bronchial washings, whereas IgG, IgA and IgM antibodies were recorded both in serum and in bronchial washings. In mice, exposure to aerosolized antigen resulted in specific IgE and IgG antibodies in serum. The levels of IgM antibodies were low and no IgA antibodies could be recorded with the enzyme-linked immunosorbent assay (ELISA). Histological examination of lung tissue from immunized rats and mice revealed increased numbers of cells with characteristics of both immature and mature mast cells. In addition, in the rats these cells were more closely located to the bronchi in immunized than in control animals. In the latter animals the mast cells were located around the blood vessels. Immature mast cells were located in the bronchiole-associated lymphatic tissue (BALT) which showed a marked proliferation in immunized animals. The findings indicate that sensitization via the airways provides possibilities to develop a model in rodents for studies of IgE-mediated allergy in the lung.     

Transcriptional signatures associated with rubella virus-specific humoral immunity after a third dose of MMR vaccine in women of childbearing age

Multiple factors linked to host genetics/inherent biology play a role in interindividual variability in immune response outcomes after rubella vaccination. In order to identify these factors, we conducted a study of rubella-specific humoral immunity before (Baseline) and after (Day 28) a third dose of MMR-II vaccine in a cohort of 109 women of childbearing age. We performed mRNA-Seq profiling of PBMCs after rubella virus in vitro stimulation to delineate genes associated with post-vaccination rubella humoral immunity and to define genes mediating the association between prior immune response status (high or low antibody) and subsequent immune response outcome. Our study identified novel genes that mediated the association between prior immune response and neutralizing antibody titer after a third MMR vaccine dose. These genes included the following: CDC34; CSNK1D; APOBEC3F; RAD18; AAAS; SLC37A1; FAS; and JAK2. The encoded proteins are involved in innate antiviral response, IFN/cytokine signaling, B cell repertoire generation, the clonal selection of B lymphocytes in germinal centers, and somatic hypermutation/antibody affinity maturation to promote optimal antigen-specific B cell immune function. These data advance our understanding of how subjects’ prior immune status and/or genetic propensity to respond to rubella/MMR vaccination ultimately affects innate immunity and humoral immune outcomes after vaccination.     

Associations between cytokine/cytokine receptor single nucleotide polymorphisms and humoral immunity to measles, mumps and rubella in a Somali population

We genotyped a Somali population ( n  = 85; age ≤30 years) for 617 cytokine and cytokine receptor single nucleotide polymorphisms (SNPs) using Illumina GoldenGate genotyping to determine associations with measles, mumps and rubella immunity. Overall, 61 significant associations ( P  ≤ 0.01) were found between SNPs belonging to cytokine receptor genes regulating T helper (Th)1 ( IL12RB2 , IL2RA and B ) and Th2 ( IL4R and IL10RB ) immunity, and cytokine ( IL1B , TNFA , IL6 and IFNB1 ) and cytokine receptor ( IL1RA , IFNAR2 , IL18R1 , TNFRSF1A and B ) genes regulating innate immunity and variations in antibody levels to measles, mumps and/or rubella. SNPs within two major inflammatory cytokine genes, TNFA and interleukin ( IL ) 6 , showed associations with measles-specific antibodies. Specifically, the minor allele variant of rs1799964 ( TNFA −1211 C>T ) was associated with primarily seronegative values (median enzyme immunoassay index values ≤0.87; P  = 0.002; q  = 0.23) in response to measles disease and/or vaccination. A heterozygous variant CT for rs2069849 ( IL6 +4272C>T ; Phe201Phe) was also associated with seronegative values and a lower median level of antibody response to measles disease and/or vaccination ( P  = 0.004; q  = 0.36) or measles vaccination alone ( P  = 0.008). Several SNPs within the coding and regulatory regions of cytokine and cytokine receptor genes showed associations with mumps and rubella antibody levels but were less informative as strong linkage disequilibrium patterns and lower frequencies for minor alleles were observed among these SNPs. Our study identifies specific SNPs in innate immune response genes that may play a role in modulating antibody responses to measles vaccination and/or infection in Somali subjects.     

Generation of antibody-producing hybridomas following one single immunization with a targeted DNA vaccine

The standard protocol for generating antibody (Ab)-producing hybridomas is based on fusion of plasmacytoma cells with Ab-producing B cells harvested from immunized mice. To increase the yield of hybridomas, it is important to use immunization protocols that induce a high frequency of B cells producing specific Abs. Our laboratory has developed a vaccine format, denoted vaccibody that promotes the immune responses towards the delivered antigen. The vaccine format targets antigens in a bivalent form to surface receptors on antigen-presenting cells (APCs). Here, we used the fluorescent protein (FP) mCherry as antigen and targeted it to APCs by use of either the natural ligand CCL3/MIP-1α or single-chain variable fragment specific for major histocompatibility complex class II. The vaccine format was delivered to mouse muscle as DNA combined with electroporation. By this procedure, we developed two monoclonal Abs that can be utilized to detect the FC mCherry in various applications. The data suggest that the targeted DNA vaccine format can be utilized to enhance the number of Ab-producing hybridomas and thereby be a tool to improve the B cell hybridoma technology.     

The immune system in the elderly: I. Specific humoral immunity

Profound and complex changes in the immune response occur during the aging process. Immunosenescence is reflected by a sum of disregulations of the immune system and its interaction with other systems. Many of the changes would appear to implicate age-related deficiencies of the immune responses. The term immunosenescence designates therefore a sort of deterioration of the immune function which is believed to manifest itself in the increased susceptibility to cancer, autoimmune disease, and infectious disease. Evidence has been accumulating from several studies which suggest an association between immune function and individual longevity. However, there are observations, especially in very old healthy people, that several immune functions are unexpectedly well preserved and substantially comparable to those observed in young subjects. These findings raise the question of whether the alterations that can be observed in the immune parameters of the elderly are a cause or a result of underlying disease processes. Moreover, studies on centenarians revealed a remodeling of the immune system rather than a deterioration, suggesting that the changes observed during immunosenescence do not correspond to immunodeficiency. The underlying mechanisms of these events are however still unclear. The purpose of the present review is to assess the status of research on the immunobiology of aging. In this first section, we focus attention on the B cell biology of aging. In clinical practice, the changes in humoral immune responsiveness and antibody-mediated defense mechanisms could greatly influence the incidence and outcome of bacterial infections and autoimmune diseases as well as the response to vaccines.     

Cell-mediated and humoral immunity induced by a live Francisella tularensis vaccine.

Live attenuated Francisella tularensis vaccine induced long-lasting humoral and cell-mediated immune responses in all 13 subjects studied. Lymphocyte blast transformation reactivity to F. tularensis appeared 2 weeks after vaccination in most subjects and remained unchanged for up to 1.5 years. Similarly, in most recipients, antibodies against F. tularensis were detectable by both the enzyme-linked immunosorbent assay (ELISA) and the agglutination method from 2 weeks after vaccination, although diagnostically significant agglutination titers (greater than or equal to 80) were not detectable until 4 weeks after vaccination. Maximal agglutination titers of 80 to 2,560 appeared at 4 to 8 weeks, and in spite of decreasing tendency, titers as high as 320 were still present 1.5 years after vaccination. ELISA showed the simultaneous, but not parallel, appearance of different immunoglobulin classes, immunoglobulin M (IgM) reaching individual maximal values 1.8 months after vaccination on average, at the same time as agglutinating antibodies, 1 week earlier than IgA, and about 1 month earlier than IgG. All of these immunoglobulin classes persisted in significant amounts up to 1.5 years, with IgG generally dominant. Long-lasting IgA and IgM responses after vaccination, as also after infection, suggested that the serodiagnosis of tularemia generally requires two consecutive serum samples with a significant increase in the titer.     

A single subcutaneous or intranasal immunization with adenovirus-based SARS-CoV-2 vaccine induces robust humoral and cellular immune responses in mice

Optimal vaccines are needed for sustained suppression of SARS-CoV-2 and other novel coronaviruses. Here, we developed a recombinant type 5 adenovirus vector encoding the gene for the SARS-CoV-2 S1 subunit antigen (Ad5.SARS-CoV-2-S1) for COVID-19 immunization and evaluated its immunogenicity in mice. A single immunization with Ad5.SARS-CoV-2-S1 via S.C. injection or I.N delivery induced robust antibody and cellular immune responses. Vaccination elicited significant S1-specific IgG, IgG1, and IgG2a endpoint titers as early as 2 weeks, and the induced antibodies were long lasting. I.N. and S.C. administration of Ad5.SARS-CoV-2-S1 produced S1-specific GC B cells in cervical and axillary LNs, respectively. Moreover, I.N. and S.C. immunization evoked significantly greater antigen-specific T-cell responses compared to unimmunized control groups with indications that S.C. injection was more effective than I.N. delivery in eliciting cellular immune responses. Mice vaccinated by either route demonstrated significantly increased virus-specific neutralization antibodies on weeks 8 and 12 compared to control groups, as well as BM antibody forming cells (AFC), indicative of long-term immunity. Thus, this Ad5-vectored SARS-CoV-2 vaccine candidate showed promising immunogenicity following delivery to mice by S.C. and I.N. routes of administration, supporting the further development of Ad-based vaccines against COVID-19 and other infectious diseases for sustainable global immunization programs.     

Activation of cell-mediated immunity following immunization with pneumococcal conjugate or polysaccharide vaccine

The immunogenicity of pneumococcal polysaccharide (PS) vaccines can be improved by conjugating PS to a polypeptide carrier that alters the immune response from T-cell independent to T-cell dependent. In order to study the influence of PS or protein antigens as inducers of cell-mediated responses, 30 adults were immunized with a 23-valent pneumococcal PS vaccine (PS-group) or an 11-valent, tetanus and diphtheria mixed carrier conjugate vaccine with (adjuvant group) or without aluminium adjuvant (nonadjuvant group). Cell-mediated responses were analyzed on days 0, 14 and 28 after vaccination by measuring lymphocyte proliferation and production of interferon (IFN)-gamma (Th1 marker) or interleukin (IL)-4 and IL-5 (Th2 markers) cytokines after in vitro stimulation with the PS and protein components of the vaccines. Tetanus and diphtheria proteins were the main inducers of lymphocyte proliferative and cytokine responses. Conjugate vaccines induced increased proliferative responses to the tetanus or diphtheria protein, but not to the PS components. In the PS-group, a lymphocyte proliferative response to protein antigens was not observed. The number of antigen-specific and nonspecific IFN-gamma-secreting cells detected by ELISPOT tended to increase in all three groups in response to protein or to PS antigen. No major differences were detected in the number of IL-4-secreting cells measured 14 and 28 days after vaccination. The conjugate vaccine with adjuvant was associated with Th2 type of activation indicated by an enhanced IL-5 secretion in response to the tetanus and diphtheria protein antigens.