Immunization Response Research Articles

Heterosubtypic protective immunity against widely divergent influenza subtypes induced by fusion protein 4sM2 in BALB/c mice

BackgroundRegular reformulation of currently available vaccines is necessary due to the unpredictable variability of influenza viruses. Therefore, vaccine based on a highly conserved antigen with capability of induction of effective immune responses could be a potential solution. Influenza matrix protein-2 (M2) is highly conserved across influenza subtypes and a promising candidate for a broadly protective influenza vaccine. For the enhancement of broad protection, four tandem copies of consensus M2 gene containing extracellular (ED) and cytoplasmic (CD) without the trans-membrane domain (TM) reconstituted from H1N1, H5N1 and H9N2 influenza viruses were linked and named as 4sM2. The construct was effectively expressed in Escherichia coli, purified and proteins were used to immunize BALB/c mice. Humoral and cell-mediated immune responses were investigated following administration.ResultsMice were intramuscularly immunized with 4sM2 protein 2 times at 2 weeks interval. Two weeks after the last immunization, first humoral and cell mediated immune response specific to sM2 protein were evaluated and the mice were challenged with a lethal dose (10MLD50) of divergent subtypes A/EM/Korea/W149/06(H5N1), A/PR/8/34(H1N1), A/Aquatic bird/Korea/W81/2005(H5N2), A/Aquatic bird/Korea/W44/2005(H7N3), and A/Chicken/Korea/116/2004(H9N2) viruses. The efficacy of 4sM2 was evaluated by determining survival rates, body weights and residual lung viral titers. Our studies demonstrate that the survival of mice immunized with 4sM2 was significantly higher (80–100% survival) than that of unimmunized mice (0% survival). We also examined the long lasting protection against heterosubtype H5N2 virus and found that mice vaccinated with 4sM2 displayed 80% of protection even after 6 months of final vaccination.ConclusionTaken together, these results suggest that prokaryotic expressed multimeric sM2 protein achieved cross protection against lethal infection of divergent influenza subtypes which are lasting for the long time.

Lentiviral vectors containing mouse Csf1r control elements direct macrophage-restricted expression in multiple species of birds and mammals

The development of macrophages requires signaling through the lineage-restricted receptor Csf1r. Macrophage-restricted expression of transgenic reporters based upon Csf1r requires the highly conserved Fms-intronic regulatory element (FIRE). We have created a lentiviral construct containing mouse FIRE and promoter. The lentivirus is capable of directing macrophage-restricted reporter gene expression in mouse, rat, human, pig, cow, sheep, and even chicken. Rat bone marrow cells transduced with the lentivirus were capable of differentiating into macrophages expressing the reporter gene in vitro. Macrophage-restricted expression may be desirable for immunization or immune response modulation, and for gene therapy for lysosomal storage diseases and some immunodeficiencies. The small size of the Csf1r transcription control elements will allow the insertion of large “cargo” for applications in gene therapy and vaccine delivery.

The relationship between concentration of specific antibody at birth and subsequent response to primary immunization

Background and aimsTrans-placentally acquired antibodies can protect infants from infection in the first months of life. However, high concentrations of antibody at birth may impact the infant's own immune response to primary immunization. We examine the relationship between concentration of specific antibody to Bordetella pertussis, Haemophilus influenzae type b (Hib), tetanus toxoid and pneumococcal antigens at birth and following primary immunization. MethodsHealthy mother–infant pairs were recruited from a UK maternity unit. Peripheral blood samples were obtained at birth and 4 weeks after primary immunization. Specific antibody concentrations were determined using enzyme-linked immunosorbent assays. Pertussis antibody concentrations >50IU/ml, Tetanus antibody levels >0.1IU/ml and Hib antibody levels >0.15mg/l were regarded as protective. ResultsFollowing primary immunization, 35/36 (97%) infants had specific antibody concentrations associated with protection against Hib, 32/36 (89%) against pertussis and 36/36 (100%) against tetanus. Concentrations of all specific antibodies were significantly higher than at birth (p<0.0001), except anti-tetanus toxoid, p=0.41. However, there was an inverse correlation between infant antibody concentration at birth and fold-increase in antibody concentration post-immunization for tetanus: rs −0.86 (95%CI −0.93 to −0.74), p<0.0001; pneumococcus: rs −0.82 (95% CI −0.91 to −0.67), p<0.0001; pertussis: rs −0.77 (95% CI −0.89 to −0.58), p<0.0001 and Hib: rs −0.66 (95%CI −0.82 to −0.42), p<0.0001. The highest concentrations of specific IgG at birth were associated with lower concentrations post-immunization for tetanus (p=0.009) and pneumococcus (p=0.03). This association was not observed for Hib (p=0.88) or pertussis (p=0.14). ConclusionHigher antibody concentration at birth appeared to inhibit the response to infant immunization for tetanus and pneumococcus; the effect was less marked for Hib and pertussis. However, the majority of infants achieved high antibody levels post-immunization. This supports maternal immunization, as high levels of maternally derived antibody at birth may not inhibit infants’ immunization responses in a clinically relevant manner.

Neutral and Acidic Oligosaccharides Supplementation Does Not Increase the Vaccine Antibody Response in Preterm Infants in a Randomized Clinical Trial

BackgroundIn preterm infants, a decreased immunological response and lower serological effectiveness are observed after immunizations due to ineffectiveness of both humoral and cellular immune mechanisms.ObjectiveTo determine the effect of 80% neutral oligosaccharides [small-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides (scGOS/lcFOS)] in combination with 20% pectin-derived acidic oligosaccharides (pAOS) on antibody concentrations after DTaP-IPV-Hib immunization in preterm infants.DesignIn this randomized clinical trial, preterm infants with gestational age <32 weeks and/or birth weight <1500 g received enteral supplementation with scGOS/lcFOS/pAOS or placebo (maltodextrin) between days 3 and 30 of life. Blood samples were collected at 5 and 12 months of age.ResultsIn total, 113 infants were included. Baseline and nutritional characteristics were not different in both groups. Geometric mean titers were not different after prebiotic supplementation at 5 months, Ptx (37/44 EU/ml), FHA (78/96 EU/ml), Prn (78/80 EU/ml), Diphtheria (0.40/0.57 IU/ml), Tetanus (0.74/0.99 IU/ml) and Hib (0.35/0.63 µg/ml), and at 12 months Ptx (55/66 EU/ml), FHA (122/119 EU/ml), Prn (116/106 Eu/ml), Diphtheria (0.88/1.11 IU/ml), Tetanus (1.64/1.79 IU/ml) and Hib (2.91/2.55 µg/ml).ConclusionsEnteral supplementation of neutral (scGOS/lcFOS) and acidic oligosaccharides (pAOS) does not improve the immunization response in preterm infants.Trial RegistrationControlled-Trials.com ISRCTN16211826 ISRCTN16211826

Monitoring the evolution of antigen-specific antibodies in circulation and their corresponding B cell clones in longitudinal immunization case studies through the use of NG-XMT (P4186)

Abstract Previous studies aimed at determining the affinity maturation pathways of monoclonal antibodies during an immunization have relied on comparing germ-line with mature forms of antibodies present in either transgenic B cell receptor mouse models or by cloning circulating human B cells present in peripheral blood (PB) of pre and post-immunized individuals. These approaches have been insightful, but they do not identify in confidence the true B cell clone(s) producing the antigen-specific antibodies found in the sera of immunized animals or humans. We recently developed a technology, NG-XMTTM, encompassing tandem mass spectrometry and next generation sequencing1,2, which for the first time enables the 1) identification of the anatomical location of the B cell clone(s) corresponding to the antigen-specific serum antibodies 2) investigation of clonal diversity of antigen-specific B cell clones residing in primary and secondary lymphoid organs and circulating in PB and 3) analysis of the evolving affinity mature serum antibodies, in two longitudinal immunization studies of rabbits and humans. These results not only advance our understanding of the affinity maturation process and the dynamics of memory B cells and plasma cell development that occurs during an immunization response, they also allow us to use this information to generate useful recombinant antibodies that can be utilized in passive immunization strategies. 1Nat Biotechnol. 2012,30:447; 2Nat Biotechnol. 2012,30:1039

Immune suppression in cynomolgus monkeys by XPro9523

The CTLA4-Ig fusion proteins abatacept and belatacept are clinically proven immunosuppressants used for rheumatoid arthritis and renal transplant, respectively. Given that both biologics are typically administered chronically by infusion, a need exists for a next-generation CTLA4-Ig with more convenient dosing. We used structure-based protein engineering to optimize the affinity of existing CTLA4-Ig therapeutics for the ligands CD80 and CD86, and for the neonatal Fc receptor, FcRn. From a rationally designed library, we identified four substitutions that enhanced binding to human CD80 and CD86. Coupled with two IgG1 Fc substitutions that enhanced binding to human FcRn, these changes comprise the novel CTLA4-Ig fusion protein, XPro9523. Compared with abatacept, XPro9523 demonstrated 5.9-fold, 23-fold, and 12-fold increased binding to CD80, CD86, and FcRn, respectively; compared with belatacept, CD80, CD86, and FcRn binding increased 1.5-fold, 7.7-fold, and 11-fold, respectively. XPro9523 and belatacept suppressed human T cell proliferation and IL-2 production more potently than abatacept. XPro9523 also suppressed inflammation in the mouse collagen-induced arthritis model. In cynomolgus monkeys, XPro9523 saturated CD80 and CD86 more effectively than abatacept and belatacept, potently inhibited IgM and IgG immunization responses, and demonstrated longer half-life. Pharmacokinetic modeling of its increased potency and persistence suggests that, in humans, XPro9523 may demonstrate superior efficacy and dosing convenience compared with abatacept and belatacept.

Ptpn22 promotes immunization responses to Influenza A (P4043)

Abstract A variant allele of PTPN22 is a potent susceptibility factor for autoimmune diseases, including Rheumatoid Arthritis and Type I Diabetes. Ptpn22 is required in myeloid cells for selective promotion of Toll-Like Receptor signaling leading to Type I IFN production. Since Type I IFN are essential for anti-viral immunity and for efficient responses to viral immunization, we investigated the role of PTPN22 in response to vaccination. We tested the ability of Ptpn22 knockout mice to mount a protective immune response to vaccination against Influenza A virus. We found that Ptpn22 is required for efficient generation of both influenza-specific cytotoxic T lymphocyte and neutralizing humoral responses after vaccination with inactivated influenza. Further, upon challenge with live influenza, vaccinated Ptpn22 KO animals exhibit increased mortality, suggesting that defects in antibody and CTL responses associated with Ptpn22 deficiency result in decreased protection engendered by inactivated influenza vaccination. These findings indicate a requirement for Ptpn22 in protective immune responses to influenza vaccine. Moreover, they suggest that further studies of vaccine responsiveness should be mounted in human carriers of the disease-associated PTPN22 variant, since such carriers exhibit diminished Toll-Like Receptor-induced Type I IFN production by myeloid cells.

Rapid Improvement of Canine Cognitive Dysfunction with Immunotherapy designed for Alzheimer's Disease

Immunotherapy against amyloid-β(Aβ) may improve rodent cognitive function by reducing amyloid neuropathology and is being validated in clinical trials with positive preliminary results. However, for a complete understanding of the direct and long-term immunization responses in the aged patient, and also to avoid significant side effects, several key aspects remain to be clarified. Thus, to investigate brain Aβ clearance and Th2 responses in the elderly, and the reverse inflammatory events not found in the immunized rodent, better Alzheimer's disease (AD) models are required. In the aged familiar canine with a Cognitive Dysfunction Syndrome (CDS) we describe the rapid effectiveness and the full safety profile of a new active vaccine candidate for human AD prevention and treatment. In these aged animals, besidesa weak immune system, the antibody response activated a coordinated central and peripheral Aβ clearance, that rapidly improved their cognitive function in absence of any side effects. Our results also confirm the interest to use familiar dogs to develop innovative and reliable therapies for AD.

CCR5 Antagonism Impacts Vaccination Response and Immune Profile in HIV-1 Infection

Maraviroc (MVC) is the first licensed antiretroviral therapeutic agent to target a host cell surface molecule, and successful HIV-1 entry blockade by this C-C chemokine receptor type 5 (CCR5)-antagonist potentiates immunomodulation. We hypothesized that MVC intensification impacts immunization responses, T-cell phenotype, function and delayed type hypersensitivity (DTH) in HIV-1(+) subjects. A 24-wk, double-blinded, placebo-controlled study of the addition of MVC to suppressive antiretroviral therapy in HIV-1(+) persons was performed. Subjects received DTH tests, intramuscular tetanus, meningococcal and oral cholera immunizations. Antibody titers, T-cell function and phenotype were assessed. Of 157 patients referred, 47 were randomized 1:1; MVC:placebo. MVC enhanced meningococcal neo-immunization, blunted cholera response and expedited lymphoproliferation to tetanus boost, without affecting recall humoral response. Anti-HIV-1 group-specific antigen (Gag) and tetanus toxoid (TTox) function improved significantly, HIV-1-associated CD8 T-cell skewing normalized, and the percentage of late-stage and major histocompatibility complex (MHC) class II expressing CD4 T-cells increased. Activated CD4(+) CD38(+) human leukocyte antigen (HLA)-DR(+) T-cells declined, and costimulation shifted to coinhibition. DTH was unchanged. Maraviroc intensification, through antagonism of the cell surface molecule CCR5, favorably influences immune profiles of HIV-1(+) patients, supporting its immunomodulatory use in HIV-1 infection and potentially in other immunologically relevant settings.

A case of hepatitis a vaccination failure in an HIV-positive man who had sex with men in Israel

Hepatitis A virus (HAV) vaccination is recommended for men who have sex with men (MSM) and other susceptible populations, who are at increased risk for HAV infection, such as HIV-positive persons. Vaccines failures are uncommon, and in HIV-positive individuals whose CD4 count is ≥ 500 cells/mm(2), seroconversion is achieved in 73-94% of vaccinees following the second dose. Data were retrieved from the patient's file at the sexually transmitted disease clinic and the AIDS clinic describing this rare case of vaccine failure. A 35-year-old, HIV-positive MSM was vaccinated against HAV on 2007, while his CD4 count was 551 cells/mm(2). Two years later, he was hospitalized due to acute HAV. The patient's serum drawn two months prior to the onset of acute HAV was retrospectively tested and showed no response to the vaccine. The source of the HAV infection was not identified. The patient's partner who was HIV-negative and had been vaccinated simultaneously with the same batch developed protective antibodies. In conclusion, HIV-positive patients and their providers should be informed about HAV vaccine failure, and post-immunization serologies to hepatitis should be considered to evaluate immunization response. Alternative approaches to develop immunity are needed for non-responders.

A long-lasting dendritic cell DNA vaccination system using lysinylated amphiphiles with mannose-mimicking head-groups

Dendritic cells (DCs) pulsed/transduced with tumor-associated or viral antigens have shown promise in combating cancer and infectious diseases. Despite significant progresses, development of a biologically safe DC-based genetic immunization (DNA vaccination) system capable of providing truly long-lasting protective immunity remains a significant scientific challenge. Here we show that immunization with autologous DCs pre-transfected with electrostatic complexes (lipoplexes) of a plasmid DNA encoding melanoma tumor associated antigen and liposomes of two lysinylated cationic amphiphiles with mannose-mimicking quinic and shikimic acid head-groups provides long-lasting (300 days post tumor challenge) protective immunity with significant memory response (more than six months after the second tumor challenge) in more than 80% immunized mice. The presently described non-viral ex vivo DC-transfection system may be exploited in inducing long-lasting immune response in DC-based genetic immunization.

Demasking of epithelial cell adhesion molecule (EpCAM) on circulating epithelial tumor cells by Tween®20 treatment in breast cancer patients

The epithelial cell adhesion molecule (EpCAM) embedded in the plasma membrane of circulating epithelial tumor cells (CETC) is used for detection and enrichment of circulating tumor cells in peripheral blood and as a target for anti-epithelial antibodies elicited during immune response in anti-tumor immunization. Although an efficient immune response against EpCAM can be generated, the clinical application of such approaches has not been successful so far and the detection of circulating epithelial cells is highly variable. One reason for these discrepancies may be that not all circulating tumor cells are equally accessible for the specific antibody. A possible reason might be masking of EpCAM by glycoproteins or membrane lipoproteins preventing antibody binding. We have tested the application of detergents as demasking agents known to be successful in demasking red blood cell epitopes and determined how and in which way they affect integral membrane proteins and membrane lipids. The results showed that the polysorbate Tween®20, a non-ionic detergent like organic solvent is able to demask EpCAM on CETC and makes it better accessible to its specific antibody retaining at the same time full cell viability. The data presented in this study suggest that EpCAM is present on part of circulating tumor cells in a masked form and that it is possible to demask EpCAM on CETC of breast cancer patients using Tween®20 treatment. But further studies are needed to elucidate the mechanism of demasking.

Female genital tract immunity: distinct immunological challenges for vaccine development

The population explosion and unintended pregnancies, sexually transmitted diseases including human immunodeficiency virus, and cervical cancer, are major challenges to health worldwide. Their prevention might be achieved through vaccination-based approaches to activate specific immunity against pathogen- or fertility-associated antigens in the female genital tract (FGT). This article aims to review methodologies for enhancing adaptive immunity in the FGT to maximize the response to vaccination. Most components of the adaptive and innate mucosal immune system are present in the FGT and several features are common with the nasopharynx/bronchial and gastrointestinal tracts. In contrast to other mucosal sites, the FGT has minimal local lymphoid tissue. Other sites primarily produce IgA and IgM while in the FGT, especially the vaginocervix, IgG is the predominant immunoglobulin secreted. In rodents, data exist to substantiate a common mucosal immune system interconnecting the nasal/bronchial, gastrointestinal, and female genital tracts. The intranasal route seems the most efficacious to induce an immunity in the FGT especially when combined with a systemic or parenteral route. In humans, for induction of secretory IgA and IgG antibodies in the FGT, immunization by the nasal or the vaginal route is effective. In vaginal immunization, a strong and consistent antibody response is best achieved following vaccination during the follicular phase of the menstrual cycle. Antibodies administered systemically percolate into the FGT and can provide immunoprotection against target molecules or cells. Thus, as well as active immunization using selected routes, the passive immunization approach may provide a viable alternative to vaccinology for future development.

NCI, NHLBI/PBMTC First International Conference on Late Effects After Pediatric Hematopoietic Cell Transplantation: Persistent Immune Deficiency in Pediatric Transplant Survivors

Defective immune reconstitution is a major barrier to successful hematopoietic cell transplantation (HCT), and has important implications in the pediatric population. There are many factors that affect immune recovery, including stem cell source and graft-versus-host disease (GVHD). Complete assessment of immune recovery, including T and B lymphocyte evaluation, innate immunity, and response to neoantigens, may provide insight as to infection risk and optimal time for immunizations. The increasing use of cord blood grafts requires additional study regarding early reconstitution and impact upon survival. Immunization schedules may require modification based upon stem cell source and immune reconstitution, and this is of particular importance as many children have been incompletely immunized, or not at all, before school entry. Additional studies are needed in children post-HCT to evaluate the impact of differing stem cell sources upon immune reconstitution, infectious risks, and immunization responses.

Effect of rituximab on human in vivo antibody immune responses

B-lymphocyte depletion with rituximab has been shown to benefit patients with various autoimmune diseases. We have previously demonstrated that this benefit is also apparent in patients with newly diagnosed type 1 diabetes. The effect of rituximab on invivo antibody responses, particularly during the period of B-lymphocyte depletion, is incompletely determined. This study was designed to assess this knowledge void. In patients with recent-onset type 1 diabetes treated with rituximab (n= 46) or placebo (n= 29), antibody responses to neoantigen phiX174 during B-lymphocyte depletion and with hepatitis A(as a second neoantigen) and tetanus/diphtheria (as recall antigens) after B-lymphocyte recovery were studied. Anti- tetanus, diphtheria, mumps, measles, and rubella titers were measured before and after treatment by means of ELISA. Antibody titers and percentage IgM versus percentage IgG to phiX174 were measured by means of phage neutralization. B-lymphocyte subsets were determined by means of flow cytometry. No change occurred in preexisting antibody titers. Tetanus/diphtheria and hepatitis Aimmunization responses were protective in the rituximab-treated subjects, although significantly blunted compared with those seen in the controls subjects, when immunized at the time of B-lymphocyte recovery. Anti-phiX174 responses were severely reduced during the period of B-lymphocyte depletion, but with B-lymphocyte recovery, anti-phiX174 responses were within the normal range. During the time of B-lymphocyte depletion, rituximab recipients had a decreased antibody response to neoantigens and significantly lower titers after recall immunization with diphtheria and tetanus toxoid. With recovery, immune responses return toward normal. Immunization during the time of B-lymphocyte depletion, although ineffective, does not preclude a subsequent response to the antigen.

Does neonatal circumcision alter behavioral pain response during subsequent routine immunizations?

Pain experienced during infancy may have long-term effects on future pain response. Study objectives were to determine whether neonatal circumcision altered behavioral pain response during subsequent immunizations compared to uncircumcised infants and whether pretreatment of circumcision pain affected subsequent immunization response. Using data from our previously reported randomized clinical trial, 97 male infants presented for immunizations at 4 and 6-months of age, 82 were circumcised. Seventy-five infants received analgesia prior to circumcision; seven infants did not receive preemptive analgesia.

Chitosan-DNA nanoparticles enhancing the immune response of Helicobacter pylori Lpp20 DNA vaccine following the mucosal immunization

Objective To investigate the immune response of mucosal immunization of new chitosan(CS) nanoparticles coating DNA vaccine. Methods The chitosan nanoparticles containing plasmid DNA encoding H. pylori lipoprotein Lpp20 gene were prepared using a complex coacervation method and then its speciality were analyzed. We then administered the naked plasmid DNA and chitosan-DNA nanoparticles to 6-week-old female BALB/c mice by intranasal or oral mucosal routes to observe the humoral and cellular immune responses. Results Naked plasmid pcDNA3.1 ( + )/Lpp20 and chitosan-pcDNA3. 1 ( + )/Lpp20 nanoparticles both induced effective immune response in mice through mucosal vaccination. Specific IgG and sIgA antibodies of chitosan-pcDNA3. 1 ( + )/Lpp20 nanoparticles groups were higher than that of naked pcDNA3.1 ( + )/Lpp20 group. The concentration of cytokines IFN-γ and IL-4 in cultural supernatant of T lymphocytes from chitosan-pcDNA3. 1 ( + )/Lpp20 nanoparticles immunized mice increased greatly than that of control groups. After stimulated by corresponding antigen, the stimulation index of intranasal or oral delivery of chitosan-pcDNA3. 1 ( + )/Lpp20 nanoparticles group was significantly higher than that of pcDNA3.1( + )/Lpp20 group, CS group and PBS control group. Moreover, systemic and mucosal immune responses in mice induced by intranasal immunization were stronger than that of oral immunization. Conclusion Chitosan nanoparticles enhanced the immune response of pcDNA3.1 ( + )/Lpp20 DNA vaccine by intranasal or oral administration in BALB/c mice. Compared to oral administration, intranasal delivery of chitosan-pcDNA3.1 ( + )/Lpp20 DNA nanoparticles could induce stronger cellular and humoral immune responses in BALB/c mice. Key words: Chitosan nanoparticles; Helicobacter pylori; Lpp20; DNA vaccine; Immunogenicity

Feedback regulation of proliferation vs. differentiation rates explains the dependence of CD4 T-cell expansion on precursor number

The mechanisms regulating clonal expansion and contraction of T cells in response to immunization remain to be identified. A recent study established that there was a log-linear relation between CD4 T-cell precursor number (PN) and factor of expansion (FE), with a slope of ∼-0.5 over a range of 3-30,000 precursors per mouse. The results suggested inhibition of precursor expansion either by competition for specific antigen-presenting cells or by the action of other antigen-specific cells in the same microenvironment as the most likely explanation. Several molecular mechanisms potentially accounting for such inhibition were examined and rejected. Here we adopt a previously proposed concept, "feedback-regulated balance of growth and differentiation," and show that it can explain the observed findings. We assume that the most differentiated effectors (or memory cells) limit the growth of less differentiated effectors, locally, by increasing the rate of differentiation of the latter cells in a dose-dependent manner. Consequently, expansion is blocked and reversed after a delay that depends on initial PN, accounting for the dependence of the peak of the response on that number. We present a parsimonious mathematical model capable of reproducing immunization response kinetics. Model definition is achieved in part by requiring consistency with available BrdU-labeling and carboxyfluorescein diacetate succinimidyl ester (CFSE)-dilution data. The calibrated model correctly predicts FE as a function of PN. We conclude that feedback-regulated balance of growth and differentiation, although awaiting definite experimental characterization of the hypothetical cells and molecules involved in regulation, can explain the kinetics of CD4 T-cell responses to antigenic stimulation.

Intranasal Vaccination with the Recombinant Listeria monocytogenes ΔactA prfA*Mutant Elicits Robust Systemic and Pulmonary Cellular Responses and Secretory Mucosal IgA

We previously showed that recombinant (r) Listeria monocytogenes carrying ΔactA and a selected prfA* mutation (r-Listeria ΔactA prfA*) secreted >100-fold more immunogen in broth culture than wild-type r-Listeria or r-Listeria ΔactA and elicited much greater cellular and humoral immune responses than r-Listeria ΔactA after intravenous vaccination of mice. Here, we conducted comparative studies evaluating vaccine-elicited immune responses in systemic and mucosal sites after intranasal, intravenous, intraperitoneal, or subcutaneous immunization of mice with r-Listeria ΔactA prfA* vaccine candidates. Intranasal vaccination of mice with r-Listeria ΔactA prfA* vaccine candidates elicited a robust gamma interferon-positive (IFN-γ(+)) cellular response in systemic sites, although intravenous or intraperitoneal immunization was more efficient. Surprisingly, intranasal vaccination elicited an appreciable pulmonary IFN-γ(+) cellular response that was nonstatistically higher than the magnitude induced by the intravenous route but was significantly greater than that elicited by subcutaneous immunization. Furthermore, although intranasal r-Listeria ΔactA prfA* delivery induced poor systemic IgG responses, intranasal vaccination elicited appreciable secretory immunogen-specific IgA titers that were similar to or higher in mucosal fluid than those induced by subcutaneous and intravenous immunizations. Thus, intranasal vaccination with r-Listeria ΔactA prfA* appears to be a useful approach for eliciting robust systemic and pulmonary cellular responses and measurable secretory mucosal IgA titers.

Parents' attitudes to neonatal research involving venepuncture

The objective of the study was to explore parental experiences of being offered participation in a previous neonatal research study involving venepuncture. The method employed was a questionnaire-based exploration of parents' attitudes in those approached to participate in a study of term and preterm immunization responses (Preterm Immunisation Study [PREMIS]). We explored experience of the initial approach, knowledge of study, venepuncture and views on research ‘in general’. In all, 59% of families responded. Highest response rates were for those participating in PREMIS (87% term/69% preterm) and lowest in decliners (34% and 35%). Responding parents participating in PREMIS were well informed, positive about research and did not find the venepuncture problematic. Sixty percent of responding parents who declined PREMIS attributed their declining to the need for venepuncture. In conclusion, parents participating or declining a neonatal study involving venepuncture are different, but participating parents were well informed and seemed able to judge that participation was right for them such that in consenting families venepuncture itself is not problematic.