Serum IgG Production Research Articles

340 Anti-CD20 attenuates lung humoral and cellular immune responses: implications for gene therapy in Cystic Fibrosis

OBJECTIVES/GOALS: Re-administration of inhaled gene therapies has the potential to overcome low correction efficiencies and limiting immune responses observed in previous trials of gene therapy for Cystic Fibrosis. We therefore tested the hypothesis that pre-treatment with a B-cell depleting α ± CD20 antibody would permit vector re-administration. METHODS/STUDY POPULATION: We first selected Adenoviral (Ad) vectors to study initially due to their well-known ability to elicit potent immune responses. Mice were dosed with a depleting α ± CD20 antibody or isotype control 2 days prior to delivery of Ad-Luc. 4 weeks later, mice were euthanized to assess the development of anti-vector immune responses. Flow cytometry and single-cell RNA sequencing were used to evaluate the development of lung-resident memory cells. Serum and airway antibody responses were assessed by ELISA. After 4 weeks, mice were dosed with Ad-LacZ and euthanized 3 days later to assess efficiency of second-round gene transfer by β-galactosidase activity assay. Similar methods were used in a pilot experiment with Adeno-associated virus vector (AAV), but with euthanasia 3 weeks after secondary gene transfer. RESULTS/ANTICIPATED RESULTS: Delivery of Ad vectors leads to the development of lung-resident memory B and T-cells. The depletion of B-cells prior to first-round vector delivery attenuated airway T-cell infiltration and serum IgG production, abrogated mucosal IgG and IgA production, and completely rescued secondary gene transfer. Genetically modified mouse models suggest secreted antibodies are critical in prevention of vector redosing. AAV vectors were found to be less immunogenic than Ad vectors, with only partial reduction of second-round gene transfer. However, anti-CD20 provided no benefit for AAV redelivery. DISCUSSION/SIGNIFICANCE: Mucosal humoral immunity is critical in preventing re-administration of Adenoviral vectors. Impairment of B-cell responses by α ± CD20 treatment prior to vector delivery allows re-administration and may help overcome low efficiencies of CF gene therapy. AAV vectors may be less susceptible to neutralization by pre-existing immunity.

Choice of adjuvant and antigen composition alters the immunogenic profile of a SARS-CoV-2 subunit vaccine

Introduction: Since their introduction, adjuvanted recombinant subunit vaccines against COVID-19 have played a pivotal role in protecting global populations. Optimizing the immune response’s quality, amplitude, and durability to these vaccines depends on the appropriate adjuvant choice and dose in combination with the selected antigen.Methods: Here, we employed a preclinical mouse model to study the adaptive humoral and cellular immune responses to a SARS-CoV-2 receptor binding domain (RBD) antigen formulated with one of four different immune agonists [GLA, 3M-052, CpG-1826 (CpG), and dmLT], in combination with one of two different immune-stimulating formulations, a stabilized squalene emulsion (SE) or aluminum hydroxide (Alum). Using a weighted desirability index, we established an immunogenicity ranking for each adjuvant in combination with the RBD antigen.Results: We found that formulations of the RBD with Alum in combination with either 3M-052 or CpG led to at least a 2-log increase in serum IgG production and a 1.3- to 2.2-log increase in the number of bone marrow-derived antibody-secreting cells compared to the RBD formulated with Alum without an additional agonist. In contrast, the RBD formulated with SE in combination with 3M-052 or CpG did not elicit an IgG response greater than the unadjuvanted control. Additionally, RBD formulated with 3M-052 or CpG on Alum generated a 0.8- or 1.6-log lower splenocyte IL-5 response (a pro-Th2 marker), respectively, than Alum without an additional agonist. When formulated with 3M-052-Alum, a bivalent vaccine containing the original lineage (Wuhan-Hu-1) and the Delta variant (B.1.617.2) RBD antigens led to a more than 2-log increase in neutralizing antibodies against an Omicron variant (B.1.1.529) pseudovirus in vaccinated animals compared to animals that received the monovalent RBD antigen.Discussion: Our results suggest that optimal immune responses to subunit antigens may be achieved through an orthogonal approach that applies adjuvant formulation, antigen combination, and advances in rational vaccine development techniques.

Recombinant hemagglutinin protein and DNA-RNA-combined nucleic acid vaccines harbored by yeast elicit protective immunity against H9N2 avian influenza infection

A safe, convenience, and effective vaccine for controlling avian influenza virus infection is crucial in scale poultry production. Yeasts are considered useful vaccine vehicles for the delivery of antigens, which has been used to protect human and animal health. We report here the development of H9N2 strain hemagglutinin (HA)-based recombinant protein vaccines (rH9HA) and DNA-RNA-combined vaccine (rH9-DNA-RNA) in Saccharomyces cerevisiae for the first time. The immunogenicity assay indicated that both rH9HA and rH9-DNA-RNA could induce robust production of serum IgG, mucosal sIgA, and cellular immune responses. The reshape and diversification of gut microbiota and an enriched Lactobacillus, Debaryomyces were observed after oral immunization with rH9HA or rH9-DNA-RNA yeast vaccine, which might contribute to modulate the intestinal mucosal immunity and antiviral process. Oral immunized birds with either rH9HA or rH9-DNA-RNA were effectively protected from H9N2 virus challenge. Our findings suggested that yeast-derived H9N2 HA-based recombinant protein vaccines and DNA-RNA-combined nucleic acid vaccines are feasible and efficacious, opening up a new avenue for rapid and cost-effective production of avian influenza vaccines to achieve good protection effect.

Intestinal lysozyme releases Nod2 ligand(s) to promote the intestinal mucosal adjuvant activity of cholera toxin.

Commensal bacteria boost serum IgG production in response to oral immunization with antigen and cholera toxin (CT) in a manner that depends on Nod2 (nucleotide-binding oligomerization domain-containing protein 2). In this study, we examined the role of intestinal lysozyme (Lyz1) in adjuvant activity of CT. We found that Lyz1 released Nod2 ligand(s) from bacteria. Lyz1 deficiency reduced the level of circulating Nod2 ligand in mice. Lyz1 deficiency also reduced the production of IgG and T-cellspecific cytokines after oral immunization in mice. Supplementing Lyz1-deficient mice with MDP restored IgG production. Furthermore, overexpression of Lyz1 in intestinal epithelium boosted the antigen-specific IgG response induced by CT. Collectively, our results indicate that Lyz1 plays an important role in mediating the immune regulatory effect of commensal bacteria through the release of Nod2 ligand(s).

A probiotic treatment increases the immune response induced by the nasal delivery of spore-adsorbed TTFC

BackgroundSpore-forming bacteria of the Bacillus genus are widely used probiotics known to exert their beneficial effects also through the stimulation of the host immune response. The oral delivery of B. toyonensis spores has been shown to improve the immune response to a parenterally administered viral antigen in mice, suggesting that probiotics may increase the efficiency of systemic vaccines. We used the C fragment of the tetanus toxin (TTFC) as a model antigen to evaluate whether a treatment with B. toyonensis spores affected the immune response to a mucosal antigen.ResultsPurified TTFC was given to mice by the nasal route either as a free protein or adsorbed to B. subtilis spores, a mucosal vaccine delivery system proved effective with several antigens, including TTFC. Spore adsorption was extremely efficient and TTFC was shown to be exposed on the spore surface. Spore-adsorbed TTFC was more efficient than the free antigen in inducing an immune response and the probiotic treatment improved the response, increasing the production of TTFC-specific secretory immunoglobin A (sIgA) and causing a faster production of serum IgG. The analysis of the induced cytokines indicated that also the cellular immune response was increased by the probiotic treatment. A 16S RNA-based analysis of the gut microbial composition did not show dramatic differences due to the probiotic treatment. However, the abundance of members of the Ruminiclostridium 6 genus was found to correlate with the increased immune response of animals immunized with the spore-adsorbed antigen and treated with the probiotic.ConclusionOur results indicate that B. toyonensis spores significantly contribute to the humoral and cellular responses elicited by a mucosal immunization with spore-adsorbed TTFC, pointing to the probiotic treatment as an alternative to the use of adjuvants for mucosal vaccinations.

Immunomodulatory effects in children in conditions of the exposure to strontium due to intake with drinking water

The study of the immune regulation in children living under conditions of exogenous strontium exposure revealed a decrease in phagocytosis, production of serum IgG and IgM, the expression level of CD95+ T- cell receptors, as well as the elevation of specific IgG to strontium and cytokine tumor necrosis factor and the change in the balance of immune osteometabolism regulators RANKL and osteoprotegerin.

Variability of immunoregulatory and genetic markers in conditions of the combined effects of industrial environmental factors

The study of the immune status markers and characteristics of genetic polymorphism in potash mining workers under the impact of combined industrial factors was performed. We showed the inhibition ofphagocytosis and production of serum IgG and IgM, the deficiency of mediators of the immune and endothelial regulation of interleukin-17, VEGF, GAD1 and GAS6. The revealed changes are associated with the negative genetic fixation of key polymorphisms responsible as well for the detoxification of the first and second phase of the neuro-immune regulation, the state of the vascular endothelium and apoptosis, as both due to heterozygous and due to the homozygous variant genotypes of genes of cytochrome (CYP1A1), Sulfotransferase (SULT1A1), toll-like receptor (TLR4), tumor necrosis factor (TNF), endothelial growth factor (VEGFA)

Early IgG Response to Foot and Mouth Disease Vaccine Formulated with a Vegetable Oil Adjuvant.

The present study evaluated soybean oil (SO) containing vitamin E (VE) and ginseng saponins (GS) (SO-VE-GS) for their adjuvant effect on foot-and-mouth disease (FMD) vaccine. Since mineral oil ISA 206 is a common adjuvant used in the FMD vaccine, it was used as a control adjuvant in this study. VE and GS were found to have a synergistic adjuvant effect. When mice were immunized with the FMD vaccine emulsified in SO with VE and GS, significantly higher serum IgG, IgG1, and IgG2a were found than VE and GS used alone. SO-VE-GS and ISA 206 behaved differently in adjuvant activities. When mice were immunized with the FMD vaccine adjuvanted with SO-VE-GS, significantly higher and earlier production of serum IgG was found than that adjuvanted with ISA 206. Although both adjuvants significantly increased the number of bone marrow plasma cells, a stimulation index of lymphocytes (SI) as well as the production of IL-4 and IL-6, SO-VE-GS promoted significantly higher SI and the ratio of CD4+/CD8+ T cells with production of increased IFN-γ and decreased TGF-β1 as compared with the ISA 206 group. The data suggested that SO-VE-GS activated Th1/Th2 immune responses. Transcriptome analysis of splenocytes showed that differentially expressed genes (DEGs), immune-related gene ontology (GO) terms, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were significantly enriched in the SO-VE-GS group. Therefore, the potent adjuvant effect of SO-VE-GS on the FMD vaccine may be attributed to the immune-related gene profile expressed in lymphocytes. Due to its plant origin and due to being much cheaper than imported mineral oil ISA 206, SO-VE-GS deserves further study in relation to vaccines used in food animals.

Sphingomyelin Synthase 1 Enhances BCR Signaling to Promote Lupus-Like Autoimmune Response

Sphingomyelin synthase 1 (SMS1) has been reported to participate in hepatitis and athereosclerosis. However, its role in autoimmune response is not clear. In this study, we investigate the possible involvement of SMS1 in B-cell activation and lupus-like autoimmunity. SMS1 knockout lupus-like animal model and SLE patient samples were utilized. We found that SMS1 deficiency suppressed B-cell activation in culture, which was restored by exogenous SM supplementation. Accordingly, the BCR-mediated early signal transduction including the colocalization of BCR with F-actin or pY/pBtk, and the phosphorylation of intracellular Fyn and Syk were impaired in SMS1 knockout B cells. Furthermore, SMS1 knockout mice showed reduced production and deposition of autoantibodies, accompanied by less severe kidney pathological changes after pristane induction. SMS1 deficiency also displayed lower autoantibody titers and 24h urine protein excretion in bm12-induced lupus, which were associated with reduced B-cell activation. Adoptively transferred wide-type B cells partially recovered B-cell activation and autoantibody production in SMS1 deficient bm12-induced lupus mice. Moreover, the SMS1 mRNA level in B cells of SLE patients was increased and positively correlated with the serum IgG production and globulin titers. These data suggest that SMS1 is involved in lupus-like autoimmunity via regulating BCR signal transduction and B cell activation. Funding Statement: This work was supported by grants from National Natural Science Foundation of China (no. 31270965 and 81771754 to Lingli Dong) and Integrated Innovative Team for Major Human Diseases Program of Tongji Medical College, HUST to Lingli Dong and Xiangping Yang. Declaration of Interests: The authors declare that there is no conflict of interest in this study. Ethics Approval Statement: This study was approved by the ethic committee of Tongji Hospital. Informed consents were obtained from all participants.

Immunoprotective response induced by recombinant glycoprotein D in the BALB/c respiratory mouse model of Equid alphaherpesvirus 1 infection

Equid alphaherpesvirus 1 (EHV-1) infection causes abortion, respiratory disease, perinatal deaths and neurological disorders in horses. The natural infection and available vaccines provide only partial and short-lived protection against reinfections. In the present study, we analyzed the ability of purified baculovirus-expressed glycoprotein D (gD) administered by different routes to induce protective immunity in BALB/c mice after challenge with the EHV-1 AR8 strain. Clinical signs varied among the different groups of mice immunized by parenteral routes, and, although gD induced a specific serum IgG response, it did not prevent the virus from reaching the lungs. Intranasally immunized mice showed no clinical signs, and virus isolation from lungs, histological lesions and antigen detection by immunohistochemistry were negative. In addition, by this route, gD did not stimulate the production of serum IgG and IgA. However, a specific IgA response in the respiratory tract was confirmed in intranasally immunized mice. Thus, we conclude that the mucosal immune response could reduce the initial viral attachment and prevent the virus from reaching the lungs. Our findings provide additional data to further study new immunization strategies in the natural host.

A bivalent vaccine derived from attenuated Salmonella expressing O-antigen polysaccharide provides protection against avian pathogenic Escherichia coli O1 and O2 infection

Avian pathogenic Escherichia coli (APEC), a leading cause of avian airsacculitis and colibacillosis, is responsible for significant economic loss in the poultry industry. APEC serogroups O1, O2, and O78 are predominantly associated with disease. Lipopolysaccharide (LPS) O-antigen has been shown to be a potent antigen for inducing specific protective immune responses. Therefore, we sought to develop a multivalent polysaccharide vaccine to prevent most APEC infections. We previously reported the stable expression of plasmid pSS27 encoding the APEC O1 O-antigen gene cluster (10.8 kb) in attenuated Salmonella enterica serovar Typhimurium S740 provided excellent protection against APEC O1 challenge. In this study, the plasmid pSS28 harboring the APEC O2 O-antigen polysaccharide gene cluster (15.5 kb) was constructed. Biosynthesis of pSS28-encoded APEC O2 O-antigen in Salmonella vaccine strain S740 was validated by Western blot. The recombinant Salmonella vaccine strain S740 (pSS28) elicited homologous protection against virulent wild-type APEC O2 challenge in a chicken model. Furthermore, through equal-volume mixing the two monovalent vaccine strains S740 (pSS27) and S740 (pSS28), a bivalent vaccine candidate against both APEC O1 and O2 was developed. Immunization of chickens with the bivalent vaccine elicited production of serum IgG and mucosal sIgA antibodies against the LPS of both APEC O1 and O2. Moreover, antibodies induced by the bivalent vaccine promoted opsonization, provoked complement-mediated bactericidal activity, and elicited protection against lethal challenge with both virulent APEC O1 and O2 strains. These results demonstrate that the bivalent vaccine comprised of S740 (pSS27) and S740 (pSS28) is a promising vaccine candidate against APEC O1 and O2 infection.

Consumption of Diet Containing Free Amino Acids Exacerbates Colitis in Mice.

Dietary proteins can influence the maturation of the immune system, particularly the gut-associated lymphoid tissue, when consumed from weaning to adulthood. Moreover, replacement of dietary proteins by amino acids at weaning has been shown to impair the generation of regulatory T cells in the gut as well as immune activities such as protective response to infection, induction of oral and nasal tolerance as well as allergic responses. Polymeric and elemental diets are used in the clinical practice, but the specific role of intact proteins and free amino acids during the intestinal inflammation are not known. It is plausible that these two dietary nitrogen sources would yield distinct immunological outcomes since proteins are recognized by the immune system as antigens and amino acids do not bind to antigen-recognition receptors but instead to intracellular receptors such as mammalian target of rapamycin (mTOR). In this study, our aim was to evaluate the effects of consumption of an amino acid-containing diet (AA diet) versus a control protein-containing diet in adult mice at steady state and during colitis development. We showed that consumption of a AA diet by adult mature mice lead to various immunological changes including decrease in the production of serum IgG as well as increase in the levels of IL-6, IL-17A, TGF-β, and IL-10 in the small and large intestines. It also led to changes in the intestinal morphology, to increase in intestinal permeability, in the number of total and activated CD4+ T cells in the small intestine as well as in the frequency of proliferating cells in the colon. Moreover, consumption of AA diet during and prior to development of dextran sodium sulfate-induced colitis exacerbated gut inflammation. Administration of rapamycin during AA diet consumption prevented colitis exacerbation suggesting that mTOR activation was involved in the effects triggered by the AA diet. Therefore, our study suggests that different outcomes can result from the use of diets containing either intact proteins or free amino acids such as elemental, semielemental, and polymeric diets during intestinal inflammation. These results may contribute to the design of nutritional therapeutic intervention for inflammatory bowel diseases.

Immunization of mice with soluble lysate of interferon gamma expressing Plasmodium berghei ANKA\xa0induces high IFN-\u03b3 production

BackgroundEfforts in search of lasting malaria vaccine have led to the development of transgenic rodent malaria parasites. As a result, wild type Plasmodium berghei ANKA (WTPbA) has recently been transformed to express mouse interferon gamma (mIFN-γ). The immunomodulatory effect of this transgenic parasite on WTPbA infection has been demonstrated. However, the protective immune responses after repeated immunization with soluble lysate of this parasite has not been investigated.MethodsSoluble lysate of transgenic PbA (TPbA) was prepared and concentration of IFN-γ in lysate determined by ELISA. Four groups of 20 BALB/c mice each (two treatment groups and two control groups) were setup. Treatment Groups 1 and 2 were primed (at day 0) with lysate of TPbA containing 75 pg/ml IFN-γ and live TPbA parasites respectively. Infection in Group 2 mice was cured with Coartem™ at 450 mg/kg for 3 days. At day 14 post-priming, both groups were boosted twice at day 14 and day 28 with lysate of TPbA containing 75 pg/ml IFN-γ and 35 pg/ml IFN-γ respectively. Blood and spleen samples were collected at day 0, day 14, day 21 and day 28 for preparation of serum and cell cultures respectively. Serum IgG and cytokines (TNF-α and IFN-γ) levels in culture supernatant were measred by ELISA.Survivorship and parasitemia were daily monitored for 21 days. Data were statistically analyzed using ANOVA student’s t test. A p value of <0.05 was considered significant.ResultsAt day 28 post-priming, IFN-γ production in Group 1 was tenfold higher than in RBC control group (p = 0.070) There was significant difference in IFN-γ production among the groups at day 28 (p < 0.0001). TNF-α production in Group 1 mice increased fourfold in Group 2 mice from day 14 to day 28 post-immunization (p = 0.0005). There was no significant effect on serum IgG production. Mice in treatment groups survived 5 to 4 days longer compared to non-immunized group.ConclusionThe study has demonstrated that, repeated immunization with soluble lysate of TPbA induces Th 1 response leading to increased IFN-γ and TNF-γ production.

Polyethyleneimine is a potent systemic adjuvant for glycoprotein antigens.

Polyethyleneimine (PEI) is an organic polycation used extensively as a gene and DNA vaccine delivery reagent. Although the DNA targeting activity of PEI is well documented, its immune activating activity is not. We recently reported that PEI has robust mucosal adjuvanticity when administered intranasally with glycoprotein antigens. Here, we show that PEI has strong immune activating activity after systemic delivery. PEI administered subcutaneously with viral glycoprotein (HIV-1 gp140) enhanced antigen-specific serum IgG production in the context of mixed Th1/Th2-type immunity. PEI elicited higher titers of both antigen binding and neutralizing antibodies than alum in mice and rabbits and induced an increased proportion of antibodies reactive with native antigen. In an intraperitoneal model, PEI recruited neutrophils followed by monocytes to the site of administration and enhanced antigen uptake by antigen-presenting cells. The Th bias was modulated by PEI activation of the Nlrp3 inflammasome; however its global adjuvanticity was unchanged in Nlrp3-deficient mice. When coformulated with CpG oligodeoxynucleotides, PEI adjuvant potency was synergistically increased and biased toward a Th1-type immune profile. Taken together, these data support the use of PEI as a versatile systemic adjuvant platform with particular utility for induction of secondary structure-reactive antibodies against glycoprotein antigens.

Non-recombinant display of the B subunit of the heat labile toxin of Escherichia coli on wild type and mutant spores of Bacillus subtilis

BackgroundMucosal infections are a major global health problem and it is generally accepted that mucosal vaccination strategies, able to block infection at their entry site, would be preferable with respect to other prevention approaches. However, there are still relatively few mucosal vaccines available, mainly because of the lack of efficient delivery systems and of mucosal adjuvants. Recombinant bacterial spores displaying a heterologous antigen have been shown to induce protective immune responses and, therefore, proposed as a mucosal delivery system. A non-recombinant approach has been recently developed and tested to display antigens and enzymes.ResultsWe report that the binding subunit of the heat-labile toxin (LTB) of Escherichia coli efficiently adsorbed on the surface of Bacillus subtilis spores. When nasally administered to groups of mice, spore-adsorbed LTB was able to induce a specific immune response with the production of serum IgG, fecal sIgA and of IFN-γ in spleen and mesenteric lymph nodes (MLN) of the immunized animals. Dot blotting experiments showed that the non-recombinant approach was more efficient than the recombinant system in displaying LTB and that the efficiency of display could be further increased by using mutant spores with an altered surface. In addition, immunofluorescence microscopy experiments showed that only when displayed on the spore surface by the non-recombinant approach LTB was found in its native, pentameric form.ConclusionOur results indicate that non-recombinant spores displaying LTB pentamers can be administered by the nasal route to induce a Th1-biased, specific immune response. Mutant spores with an altered coat are more efficient than wild type spores in adsorbing the antigen, allowing the use of a reduced number of spores in immunization procedures. Efficiency of display, ability to display the native form of the antigen and to induce a specific immune response propose this non-recombinant delivery system as a powerful mucosal vaccine delivery approach.

Atypical and classical memory B cells produce Plasmodium falciparum neutralizing antibodies

Antibodies can protect from Plasmodium falciparum (Pf) infection and clinical malaria disease. However, in the absence of constant reexposure, serum immunoglobulin (Ig) levels rapidly decline and full protection from clinical symptoms is lost, suggesting that B cell memory is functionally impaired. We show at the single cell level that natural Pf infection induces the development of classical memory B cells (CM) and atypical memory B cells (AtM) that produce broadly neutralizing antibodies against blood stage Pf parasites. CM and AtM contribute to anti-Pf serum IgG production, but only AtM show signs of active antibody secretion. AtM and CM were also different in their IgG gene repertoire, suggesting that they develop from different precursors. The findings provide direct evidence that natural Pf infection leads to the development of protective memory B cell antibody responses and suggest that constant immune activation rather than impaired memory function leads to the accumulation of AtM in malaria. Understanding the memory B cell response to natural Pf infection may be key to the development of a malaria vaccine that induces long-lived protection.

Evaluation of immune response to artificial infections of Haemonchus contortus in Gulf Coast Native compared with Suffolk lambs

The Gulf Coast Native (Native) breed of sheep among many others is identified as being relatively resistant to Haemonchus contortus, an abomasal nematode parasite of small ruminants. Understanding the mode of immune response that helps these breeds of sheep control infection could help design and implement appropriate control programs. In this experiment, the components of the immune response during the early infection period in resistant Native lambs were evaluated and compared with susceptible Suffolk breed of sheep. Groups (n=5) of six month old Native and Suffolk lambs were given infective larvae as one time (single) or trickle experimental infections. Fecal, blood, and serum samples were collected on days 0, 2, 7, 14 and 21 post-infection. Abomasal mucosa and regional lymph node samples were collected at the time of necropsy on days 14 and 21. There was no significant difference in number of worms recovered at necropsy but the ratio of adult versus larvae was significantly greater in single infected Suffolk than Native lambs. Native lambs had significantly greater numbers of mast cells and eosinophils in the abomasal mucosa and serum IgG production was significantly greater compared to Suffolk lambs. Native lambs also showed a trend of increased level of serum IgA and IgE compared to Suffolk lambs.

Comparative studies of local antibody and cellular immune responses to influenza infection and vaccination with live attenuated reassortant influenza vaccine (LAIV) utilizing a mouse nasal-associated lymphoid tissue (NALT) separation method

The first and most significant barrier against influenza infection is the mucosal-associated lymphoid tissue of the upper airways and rodent nasopharyngeal-associated lymphoid tissue (NALT) is considered equivalent to the lymphoid tissue of human Valdryer's ring. This study is the first attempt to analyze and compare local and systemic cellular and antibody immune responses in NALT and spleen in a mouse model of experimental influenza infection and intranasal vaccination with LAIV (live attenuated reassortant influenza vaccine). It was shown that the vaccine strain completely inherited the ability to induce high-grade local antibody responses (secretory IgA + IgG + IgM), local cellular lymphoproliferative activity, CD4 +, CD8 + and CD19 + lymphocyte and cytokine production responses from the virulent parental strain but it had less capacity to stimulate production of serum IgG, accumulation of CD8 + cells and IFN-γ production in the spleen. Primary non-complicated influenza infection and primary vaccination were accompanied by a short-term (24 h) increase in the levels of lymphocyte apoptosis in both NALT and spleen. However, experimental data indicated that vaccination with LAIV and uncomplicated forms of influenza infection did not influence immune system apoptosis following a secondary immune response.

Recombinant Mycobacterium bovis BCG vector system expressing SIV Gag protein stably and persistently induces antigen-specific humoral immune response concomitant with IFNγ response, even at three years after immunization

A vaccine against HIV-1 infection absolutely needs the ability to effectively elicit virus-specific immunity over a long term; nevertheless, there have been few studies indicating that the immunoinductivity of such a candidate vaccine has been researched for several years running. In a previous report, we demonstrated that recombinant BCG (rBCG) expressing the full-length gag gene of simian immunodeficiency virus (SIV) (rBCG-SIVgag) induced Gag-specific delayed-type hypersensitivity, T cell proliferation, gamma interferon (IFNγ), and serum IgG responses in guinea pigs immunized intradermally (i.d.) with 0.1 mg for the 1-year period of study. Especially, the production of long-lasting Gag-specific serum IgG in the vaccinated animals perhaps reflects the persistent antigenic stimulation by rBCG-SIVgag. How long, we questioned, will such immune responses to Gag engendered by the rBCG-SIVgag vaccination persist without booster immunizations? To learn this, we examined Gag-specific IgG production in sera and Gag-specific IFNγ mRNA expression in peripheral blood mononuclear cells (PBMC) in guinea pigs vaccinated with rBCG-SIVgag i.d. (0.1 mg) or orally (80 mg × 2) during a 3-year period. As a result, Gag-specific serum IgG was highly generated for 3 years at similar levels between the i.d. and the orally immunized guinea pigs (IgG2 > IgG1). The enhancement of IFNγ mRNA expression by in vitro restimulation with Gag antigen was also detected in PBMC from the two immunization groups throughout the 3-year observation period. In guinea pigs immunized i.d. with rBCG-SIVgag, a high level of Gag-specific IFNγ response was observed at 1 year after vaccination, whereas the response has waned gradually. The current study indicates that i.d. and oral inoculations of rBCG-SIVgag elicit stable, strong, Gag-specific serum IgG production while exhibiting the different kinetics of Gag-specific IFNγ responses between i.d. and oral vaccination routes. This suggests that the rBCG vector system expressing an appropriate size of foreign antigen gene should be suited for the induction of the antigen-specific humoral immunity concomitant with an IFNγ response.

Human immunoglobulin production in immunodeficient mice: enhancement by immunosuppression of host and in vitro activation of human mononuclear cells.

The affect of host and donor related factors on successful engraftment of human cells into mice was examined to minimize the variability that has been observed in successful development of human-mouse chimera for the study of human disease and immune physiology and regulation. Human immunoglobulin production in severe combined immunodeficiency (SCID) mice engrafted with human peripheral blood mononuclear cells (PBMC) was augmented by immunosuppressing recipient mice and activating donor PBMC. Immunosuppression of recipient mice with 3 Gy of gamma-irradiation induced a 10-fold increase in human IgG in the sera of engrafted SCID mice. Variation in production of human IgG in recipient mice correlated with preinjection phenotype and activation status of injected PBMC. Mice injected with PBMC with a low CD4/CD8 ratio (less than 0.5) produced no detectable circulating human immunoglobulin. When the CD4/CD8 ratio was greater than 1.5, human IgG was detected in sera of PBMC-recipient SCID mice. Serum IgG increased 10-fold following in vitro activation of donor PBMC with anti-CD3, IL-2 and Staphylococcus aureus. Successful engraftment and serum IgG production was evidenced by an increase in the recovery of activated human IgG+ cells in the spleens of mice with maximal IgG production. Optimization of functional engraftment required modification of both the host (SCID mice) and the donor cells.