Immunization Of Neonatal Mice Research Articles

Activation of regulatory dendritic cells by Mertk coincides with a temporal wave of apoptosis in neonatal lungs.

Multiple mechanisms restrain inflammation in neonates, most likely to prevent tissue damage caused by overly robust immune responses against newly encountered pathogens. Here, we identify a population of pulmonary dendritic cells (DCs) that express intermediate levels of CD103 (CD103int) and appear in the lungs and lung-draining lymph nodes of mice between birth and 2 weeks of age. CD103int DCs express XCR1 and CD205 and require expression of the transcription factor BATF3 for development, suggesting that they belong to the cDC1 lineage. In addition, CD103int DCs express CCR7 constitutively and spontaneously migrate to the lung-draining lymph node, where they promote stromal cell maturation and lymph node expansion. CD103int DCs mature independently of microbial exposure and TRIF- or MyD88-dependent signaling and are transcriptionally related to efferocytic and tolerogenic DCs as well as mature, regulatory DCs. Correlating with this, CD103int DCs show limited ability to stimulate proliferation and IFN-γ production by CD8+ T cells. Moreover, CD103int DCs acquire apoptotic cells efficiently, in a process that is dependent on the expression of the TAM receptor, Mertk, which drives their homeostatic maturation. The appearance of CD103int DCs coincides with a temporal wave of apoptosis in developing lungs and explains, in part, dampened pulmonary immunity in neonatal mice. Together, these data suggest a mechanism by which DCs sense apoptotic cells at sites of noninflammatory tissue remodeling, such as tumors or the developing lungs, and limit local T cell responses.

Development of a TLR7/8 agonist adjuvant formulation to overcome early life hyporesponsiveness to DTaP vaccination

Infection is the most common cause of mortality early in life, yet the broad potential of immunization is not fully realized in this vulnerable population. Most vaccines are administered during infancy and childhood, but in some cases the full benefit of vaccination is not realized in-part. New adjuvants are cardinal to further optimize current immunization approaches for early life. However, only a few classes of adjuvants are presently incorporated in vaccines approved for human use. Recent advances in the discovery and delivery of Toll-like receptor (TLR) agonist adjuvants have provided a new toolbox for vaccinologists. Prominent among these candidate adjuvants are synthetic small molecule TLR7/8 agonists. The development of an effective infant Bordetella pertussis vaccine is urgently required because of the resurgence of pertussis in many countries, contemporaneous to the switch from whole cell to acellular vaccines. In this context, TLR7/8 adjuvant based vaccine formulation strategies may be a promising tool to enhance and accelerate early life immunity by acellular B. pertussis vaccines. In the present study, we optimized (a) the formulation delivery system, (b) structure, and (c) immunologic activity of novel small molecule imidazoquinoline TLR7/8 adjuvants towards human infant leukocytes, including dendritic cells. Upon immunization of neonatal mice, this TLR7/8 adjuvant overcame neonatal hyporesponsiveness to acellular pertussis vaccination by driving a T helper (Th)1/Th17 biased T cell- and IgG2c-skewed humoral response to a licensed acellular vaccine (DTaP). This potent immunization strategy may represent a new paradigm for effective immunization against pertussis and other pathogens in early life.

A comparative study of adjuvants effects on neonatal plasma cell survival niche in bone marrow and persistence of humoral immune responses.

The neonatal immune system is distinct from the immune system of older individuals rendering neonates vulnerable to infections and poor responders to vaccination. Adjuvants can be used as tools to enhance immune responses to co-administered antigens. Antibody (Ab) persistence is mediated by long-lived plasma cells that reside in specialized survival niches in the bone marrow, and transient Ab responses in early life have been associated with decreased survival of plasma cells, possibly due to lack of survival factors. Various cells can secrete these factors and which cells are the main producers is still up for debate, especially in early life where this has not been fully addressed. The receptor BCMA and its ligand APRIL have been shown to be important in the maintenance of plasma cells and Abs. Herein, we assessed age-dependent maturation of a broad range of bone marrow accessory cells and their expression of the survival factors APRIL and IL-6. Furthermore, we performed a comparative analysis of the potential of 5 different adjuvants; LT-K63, mmCT, MF59, IC31 and alum, to enhance expression of survival factors and BCMA following immunization of neonatal mice with tetanus toxoid (TT) vaccine. We found that APRIL expression was reduced in the bone marrow of young mice whereas IL-6 expression was higher. Eosinophils, macrophages, megakaryocytes, monocytes and lymphocytes were important secretors of survival factors in early life but undefined cells also constituted a large fraction of secretors. Immunization and adjuvants enhanced APRIL expression but decreased IL-6 expression in bone marrow cells early after immunization. Furthermore, neonatal immunization with adjuvants enhanced the proportion of plasmablasts and plasma cells that expressed BCMA both in spleen and bone marrow. Enhanced BCMA expression correlated with enhanced vaccine-specific humoral responses, even though the effect of alum on BCMA was less pronounced than those of the other adjuvants at later time points. We propose that low APRIL expression in bone marrow as well as low BCMA expression of plasmablasts/plasma cells in early life together cause transient Ab responses and could represent targets to be triggered by vaccine adjuvants to induce persistent humoral immune responses in this age group.

Adenovector 26 encoded prefusion conformation stabilized RSV-F protein induces long-lasting Th1-biased immunity in neonatal mice

While RSV is a major cause of respiratory morbidity in infants, vaccine development is hindered by the immaturity and Th2-bias of the infant immune system and the legacy of enhanced respiratory disease (ERD) after RSV infection following immunization with formalin inactivated (FI)-RSV vaccine in earlier clinical trials. Preclinical studies have demonstrated that an adenoviral vector-based RSV F vaccine candidate (Ad26.RSV.FA2) induces Th1-biased protective immune responses, without signs of ERD upon subsequent RSV challenge. We here developed an Ad26 vector encoding the RSV F protein stabilized in its prefusion conformation (Ad26.RSV.preF). In adult mice, Ad26.RSV.preF induced superior, Th1-biased IgG2a-dominated humoral responses as compared to Ad26.RSV.FA2, while maintaining the strong Th1-biased cellular responses. Similar to adult mice, Ad26.RSV.preF induced robust and durable humoral immunity in neonatal mice, again characterized by IgG2a-dominated RSV F-binding antibodies, and high and stable virus-neutralizing titers. In addition, vaccine-elicited cellular immune responses were durable and characterized by IFN-γ-producing CD4+ and CD8+ T cells, with a profound Th1 bias. In contrast, immunization of neonatal mice with FI-RSV resulted in IgG1 RSV F-binding antibodies associated with a Th2 phenotype, no detectable virus-neutralizing antibodies, and a Th2-biased cellular response. These results are supportive for the clinical development of Ad26.RSV.preF for use in infants.

Adjuvant pretreatment with alum protects neonatal mice in sepsis through myeloid cell activation.

The high mortality in neonatal sepsis has been related to both quantitative and qualitative differences in host protective immunity. Pretreatment strategies to prevent sepsis have received inadequate consideration, especially in the premature neonate, where outcomes from sepsis are so dismal. Aluminium salts-based adjuvants (alum) are used currently in many paediatric vaccines, but their use as an innate immune stimulant alone has not been well studied. We asked whether pretreatment with alum adjuvant alone could improve outcome and host innate immunity in neonatal mice given polymicrobial sepsis. Subcutaneous alum pretreatment improves survival to polymicrobial sepsis in both wild-type and T and B cell-deficient neonatal mice, but not in caspase-1/11 null mice. Moreover, alum increases peritoneal macrophage and neutrophil phagocytosis, and decreases bacterial colonization in the peritoneum. Bone marrow-derived neutrophils from alum-pretreated neonates produce more neutrophil extracellular traps (NETs) and exhibit increased expression of neutrophil elastase (NE) after in-vitro stimulation with phorbol esters. In addition, alum pretreatment increases bone marrow and splenic haematopoietic stem cell expansion following sepsis. Pretreatment of neonatal mice with an alum-based adjuvant can stimulate multiple innate immune cell functions and improve survival. These novel findings suggest a therapeutic pathway for the use of existing alum-based adjuvants for preventing sepsis in premature infants.

Immunization of Newborn Mice Accelerates the Architectural Maturation of Lymph Nodes, But AID-Dependent IgG Responses Are Still Delayed Compared to the Adult.

Lymph nodes (LNs) have evolved to maximize antigen (Ag) collection and presentation as well as lymphocyte proliferation and differentiation—processes that are spatially regulated by stromal cell subsets, including fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs). Here, we showed that naïve neonatal mice have poorly organized LNs with few B and T cells and undetectable FDCs, whereas adult LNs have numerous B cells and large FDC networks. Interestingly, immunization on the day of birth accelerated B cell accumulation and T cell recruitment into follicles as well as FDC maturation and FRC organization in neonatal LNs. However, compared to adults, the formation of germinal centers was both delayed and reduced following immunization of neonatal mice. Although immunized neonates poorly expressed activation-induced cytidine deaminase (AID), they were able to produce Ag-specific IgGs, but with lower titers than adults. Interestingly, the Ag-specific IgM response in neonates was similar to that in adults. These results suggest that despite an accelerated structural maturation of LNs in neonates following vaccination, the B cell response is still delayed and reduced in its ability to isotype switch most likely due to poor AID expression. Of note, naïve pups born to Ag-immunized mothers had high titers of Ag-specific IgGs from day 0 (at birth). These transferred antibodies confirm a mother-derived coverage to neonates for Ags to which mothers (and most likely neonates) are exposed, thus protecting the neonates while they produce their own antibodies. Finally, the type of Ag used in this study and the results obtained also indicate that T cell help would be operating at this stage of life. Thus, neonatal immune system might not be intrinsically immature but rather evolutionary adapted to cope with Ags at birth.

Prostaglandin E2 production during neonatal respiratory infection with mouse adenovirus type 1

Neonatal mice are more susceptible than adults to mouse adenovirus type 1 (MAV1) respiratory infection. In adult mice, MAV-1 respiratory infection induces production of prostaglandin E2 (PGE2), a lipid mediator that exerts suppressive effects on a variety of host immune functions. We tested the hypothesis that exaggerated PGE2 production in neonatal mice contributes to increased susceptibility to MAV-1. PGE2 concentrations were lower in lungs of uninfected neonatal mice than in adults. PGE2 production was induced by both MAV-1 and a nonspecific stimulus to a greater degree in neonatal mice than in adults, but only in adults was PGE2 induced in a virus-specific manner. Lung viral loads were equivalent in PGE2-deficient neonatal mice and wild type controls, as was virus-induced expression of IFN-γ, IL-17A, and CCL5 in the lungs. PGE2 deficiency had minimal effect on production of virus-specific IgG or establishment of protective immunity in neonatal mice. Collectively, our data indicate that lung PGE2 production is exaggerated early in life, but this effect does not mediate increased susceptibility to MAV-1 infection.

Susceptibility to Acute Mouse Adenovirus Type 1 Respiratory Infection and Establishment of Protective Immunity in Neonatal Mice

There is an incomplete understanding of the differences between neonatal immune responses that contribute to the increased susceptibility of neonates to some viral infections. We tested the hypothesis that neonates are more susceptible than adults to mouse adenovirus type 1 (MAV-1) respiratory infection and are impaired in the ability to generate a protective immune response against a second infection. Following intranasal infection, lung viral loads were greater in neonates than in adults during the acute phase but the virus was cleared from the lungs of neonates as efficiently as it was from adult lungs. Lung gamma interferon (IFN-γ) responses were blunted and delayed in neonates, and lung viral loads were higher in adult IFN-γ(-/-) mice than in IFN-γ(+/+) controls. However, administration of recombinant IFN-γ to neonates had no effect on lung viral loads. Recruitment of inflammatory cells to the airways was impaired in neonates. CD4 and CD8 T cell responses were similar in the lungs of neonates and adults, although a transient increase in regulatory T cells occurred only in the lungs of infected neonates. Infection of neonates led to protection against reinfection later in life that was associated with increased effector memory CD8 T cells in the lungs. We conclude that neonates are more susceptible than adults to acute MAV-1 respiratory infection but are capable of generating protective immune responses.

Immunization of neonatal mice with LAMP/p55 HIV gag DNA elicits robust immune responses that last to adulthood

Successful T cell priming in early postnatal life that can generate effective long-lasting responses until adulthood is critical in HIV vaccination strategies because it prevents early sexual initiation and breastfeeding transmission of HIV. A chimeric DNA vaccine encoding p55 HIV gag associated with lysosome-associated membrane protein 1 ( LAMP-1; which drives the antigen to the MIIC compartment), has been used to enhance cellular and humoral antigen-specific responses in adult mice and macaques. Herein, we investigated LAMP-1/gag vaccine immunogenicity in the neonatal period in mice and its ability to generate long-lasting effects. Neonatal vaccination with chimeric LAMP/gag generated stronger Gag-specific immune responses, as measured by the breadth of the Gag peptide-specific IFN-γ, proliferative responsiveness, cytokine production and antibody production, all of which revealed activation of CD4+ T cells as well as the generation of a more robust CTL response compared to gag vaccine alone. To induce long-lived T and B cell memory responses, it was necessary to immunize neonates with the chimeric LAMP/gag DNA vaccine. The LAMP/gag DNA vaccine strategy could be particularly useful for generating an anti-HIV immune response in the early postnatal period capable of inducing long-term immunological memory.

IC31®, a Two‐Component Novel Adjuvant Mixed with a Conjugate Vaccine Enhances Protective Immunity against Pneumococcal Disease in Neonatal Mice

IC31 is a novel adjuvant which combines the immunostimulatory effects of an 11-mer antibacterial peptide (KLKL(5)KLK) and a synthetic oligodeoxynucleotide (ODN1a) which is a Toll-like receptor 9 agonist without containing cytosine phosphate guanine (CpG) motifs. The effects of IC31 on neonatal immune response to vaccination have not been reported. Neonatal mice were immunized once or twice with a Streptococcus pneumoniae serotype 1 polysaccharide conjugate containing Tetanus Toxoid (Pnc1-TT) carrier protein, with or without IC31 or CpG-ODN. IC31 significantly enhanced IgG1, IgG2a and IgG2b antibodies (Ab) to the serotype 1 polysaccharide. One dose of Pnc1-TT and low dose IC31 elicited high Ab levels that protected the neonatal mice completely from bacteraemia and significantly reduced lung infection following i.n. challenge with serotype 1 pneumococcal strain. One-sixth of an adult murine dose of IC31 was sufficient and optimal for induction of protective immunity in neonatal mice. Two doses of Pnc1-TT with or without adjuvants protected the neonatal mice completely, but more rapid Ab response was observed when IC31 was given with the Pnc1-TT. IC31 is a promising new adjuvant for neonatal vaccinations, rapidly enhancing protective humoral responses when combined with Pnc1-TT.

Immunization of neonatal mice induces protective immunity against Hericobacter pylori

Immunization of neonatal mice induces protective immunity against Hericobacter pylori

Induction of TH1 and TH2 immunity in neonatal mice.

The neonatal period has been thought of as a window in ontogeny, during which the developing immune system is particularly susceptible to tolerization. In the present study, the classic system for induction of neonatal tolerance to protein antigens was reexamined in mice. The presumably tolerogenic protocol was found to trigger a vigorous T helper cell type 2 (TH2) immune response. Thus, neonatal "tolerization" induces immune deviation, not tolerance in the immunological sense. Neonates are not immune privileged but generate TH2 or TH1 responses, depending on the mode of immunization.