Interleukin-1 family cytokines as mucosal vaccine adjuvants for induction of protective immunity against influenza virus

A safe and potent adjuvant is needed for development of mucosal vaccines against etiological agents, such as influenza virus, that enter the host at mucosal surfaces. Cytokines are potential adjuvants for mucosal vaccines because they can enhance primary and memory immune responses enough to protect against some infectious agents. For this study, we tested 26 interleukin (IL) cytokines as mucosal vaccine adjuvants and compared their abilities to induce antigen (Ag)-specific immune responses against influenza virus. In mice intranasally immunized with recombinant influenza virus hemagglutinin (rHA) plus one of the IL cytokines, IL-1 family cytokines (i.e., IL-1α, IL-1β, IL-18, and IL-33) were found to increase Ag-specific immunoglobulin G (IgG) in plasma and IgA in mucosal secretions compared to those after immunization with rHA alone. In addition, high levels of both Th1- and Th2-type cytokines were observed in mice immunized with rHA plus an IL-1 family cytokine. Furthermore, mice intranasally immunized with rHA plus an IL-1 family cytokine had significant protection against a lethal influenza virus infection. Interestingly, the adjuvant effects of IL-18 and IL-33 were significantly decreased in mast cell-deficient W/W(v) mice, indicating that mast cells have an important role in induction of Ag-specific mucosal immune responses induced by IL-1 family cytokines. In summary, our results demonstrate that IL-1 family cytokines are potential mucosal vaccine adjuvants and can induce Ag-specific immune responses for protection against pathogens like influenza virus.     

Comparison of Th1- and Th2-associated immune reactivities stimulated by single versus multiple vaccination of mice with irradiated Schistosoma mansoni cercariae.

Mice immunized against Schistosoma mansoni by a single percutaneous exposure to radiation-attenuated parasite larvae demonstrate partial resistance to challenge infection that has been shown to correlate with development of cell-mediated immunity, whereas mice hyperimmunized by multiple exposure to attenuated larvae produce antibodies capable of transferring partial protection to naive recipients. Measurement of Ag-specific lymphokine responses in these animals suggested that the difference in resistance mechanisms may be due to the differential induction of Th subset response by the two immunization protocols. Thus, upon Ag stimulation, singly immunized mice predominantly demonstrated responses associated with Th1 reactivity, including IL-2 and IFN-gamma production, whereas multiply immunized animals showed increased IL-5, IL-4, and IgG1 antibody production associated with enhanced Th2 response. These responses demonstrated some degree of organ compartmentalization, with splenocytes demonstrating higher Th1-related lymphokine production and cells from draining lymph nodes showing stronger proliferation and Th2 type reactivity. However, hyperimmunized mice also continued to demonstrate substantial Th1-associated immune reactivity. Moreover, in vivo Ag challenge elicited activated larvacidal macrophages in hyperimmunized animals. These observations indicate that protective cell-mediated mechanisms associated with induction of CD4+ Th1 cell reactivity predominate in singly vaccinated mice. Further vaccination stimulates Th2 responses, such as enhanced IgG1 production, that may also contribute to protective immunity.     

Dendritic cells (DC) activated by CpG DNA ex vivo are potent inducers of host resistance to an intracellular pathogen that is independent of IL-12 derived from the immunizing DC

We used the model of murine leishmaniasis to evaluate the signals enabling Ag-pulsed dendritic cells (DC) to prime a protective Th1 response in vivo. Bone marrow-derived DC (BMDC) that had been activated by TNF-alpha or CD40 ligation were not able to induce protection against leishmaniasis in susceptible BALB/c mice. In contrast, all mice vaccinated with a single dose of Leishmania major Ag-pulsed BMDC stimulated by prior in vitro exposure to CpG-containing oligodeoxynucleotides (ODN) were completely protected, had a dramatic reduction in parasite burden, and developed an Ag-specific Th1 response. Importantly, systemic administration of CpG ODN was not required. Protection mediated by ex vivo CpG ODN-activated and Ag-pulsed DC was solid, as documented by resistance to reinfection with a higher parasite dose, and long-lasting, as immunized mice were still protected against L. major challenge 16 wk after vaccination. A significantly increased level of protection could also be elicited in resistant C57BL/6 mice. Surprisingly, IL-12 expression by the immunizing BMDC was not required for induction of host resistance. In contrast, the availability of IL-12 derived from recipient cells was essential for the initial triggering of protective immunity by transferred BMDC. Together, these findings demonstrate that the type of stimulatory signal is critical for activating the potential of DC to induce a Th1 response in vivo that confers complete protection against an intracellular pathogen. Moreover, they show that the impact of activated DC on the initiation of a protective Th cell response in vivo may be independent of their ability to produce IL-12.     

The combination of DNA vectors expressing IL-12 + IL-18 elicits high protective immune response against cutaneous leishmaniasis after priming with DNA-p36/LACK and the cytokines,followed by a booster with a vaccinia virus recombinant expressing p36/LACK

Protocols of immunization based on the DNA prime/vaccinia virus (VV) boost regime with recombinants expressing relevant antigens have been shown to elicit protection against a variety of pathogens in animal model systems, and various phase I clinical trials have been initiated with this vaccination approach. We have previously shown that mice immunized with a DNA vector expressing p36/LACK of Leishmania infantum followed by a booster with VVp36/LACK induced significant protection against Leishmania major infection. To further improve this protocol of immunization, here we investigated whether the cytokines interleukin-12 (IL-12) and IL-18 could enhance protection against L. major infection in BALB/c mice. We found that priming with DNA vectors expressing p36/LACK and either IL-12 or IL-18, followed by a booster with a VV recombinant expressing the same L. infantum LACK antigen, elicit a higher cellular immune response than by using the same protocol in the absence of the cytokines. The cytokine IL-12 triggered a higher number of IFN-gamma-secreting cells specific for p36 protein than IL-18. When immunized animals were challenged with promastigotes, the highest protection against L. major infection was observed in animals primed with DNAp36 + DNA IL-12 + DNA IL-18 and boosted with VVp36. This protection correlated with a Th1 type of immune response. Our findings revealed that in prime/booster protocols, co-expressing IL-12 and IL-18 during priming is an efficient approach to protect against leishmaniasis. This combined prime/booster immunization regime could have wide use in fighting against parasitic and other infectious diseases.     

GM-CSF transgene-based adjuvant allows the establishment of protective mucosal immunity following vaccination with inactivated Chlamydia trachomatis

Cellular and humoral immune responses induced following murine Chlamydia trachomatis infection confer almost sterile protection against homologous reinfection. On the other hand, immunization with inactivated organism induces little protective immunity in this model system. The underlying mechanism(s) that determines such divergent outcome remains unclear, but elucidating the mechanism will probably be important for chlamydial vaccine development. One of the distinct differences between the two forms of immunization is that chlamydia replication in epithelial cells causes the secretion of a variety of proinflammatory cytokines and chemokines, such as GM-CSF, that may mobilize and mature dendritic cells and thereby enhance the induction of protective immunity. Using a murine model of C. trachomatis mouse pneumonitis lung infection and intrapulmonary adenoviral GM-CSF transfection, we demonstrate that the expression of GM-CSF in the airway compartment significantly enhanced systemic Th1 cellular and local IgA immune responses following immunization with inactivated organisms. Importantly, immunized mice had significantly reduced growth of chlamydia and exhibited less severe pulmonary inflammation following challenge infection. The site of GM-CSF transfection proved important, since mice immunized with inactivated organisms after GM-CSF gene transfer by the i.p. route exhibited little protection against pulmonary challenge, although i.p. immunization generated significant levels of systemic Th1 immune responses. The obvious difference between i.p. and intrapulmonary immunization was the absence of lung IgA responses following i.p. vaccination. In aggregate, the findings demonstrate that the local cytokine environment is critical to the induction of protective immunity following chlamydial vaccination and that GM-CSF may be a useful adjuvant for a chlamydial vaccine.     

Effect of adjuvants on immune response and protective immunity elicited by recombinant Hsp60 (GroEL) of Salmonella typhi against S. typhi infection

Heat shock proteins (Hsps) have been reported to be dominant antigens for the host immune response to various pathogens and thus, have great potential for use in vaccination. In the present study, we evaluated the immunogenicity and protective efficacy of GroEL of Salmonella enterica serovar Typhi against lethal infection by S. typhi Ty2 in mice with or without adjuvants. Anti GroEL–IgG titers were significantly higher in mice immunized with either GroEL-alone or in combination with alum/Complete Freund’s adjuvant (CFA) as compared to the control. Analysis of antibody isotypes suggested predominance of Th2 type immune response in GroEL + alum immunized animals as revealed by higher IgG1/IgG2a ratio. Whereas, immunization of animals with GroEL + CFA or GroEL-alone shifted the immune response toward Th1 phenotype. Mice immunized with GroEL with or without adjuvants, showed a significant increase in lymphocyte proliferation and cytokine levels. The animals immunized with GroEL + CFA or GroEL-alone showed higher IFN-γ and IL-2 levels than alum group, indicating Th1 response whereas IL-4 levels (Th2 response) were found to be highest in alum group as compared to other two immunized groups. Immunization of mice with GroEL-alone, GroEL + alum, and GroEL + CFA provided 70, 50 and 80% protection, respectively, against lethal challenge by S. typhi in mice. The differences in the percentage protection among various groups were attributed to the differences in the immune responses generated by respective immunizations. The present study shows that GroEL forms an ideal candidate molecule to develop a recombinant protein based vaccine against human typhoid.     

Modulation of IL-33/ST2 signaling as a potential new therapeutic target for cardiovascular diseases

IL-33 belongs to the IL-1 family of cytokines, which function as inducers of Th2 cytokine production by binding with ST2L and IL-1RAcP. This, in turn, activates various signaling pathways, including the mitogen-activated protein kinase (MAPK), the inhibitor of Kappa-B kinase (IKK) pathway, and the phospholipase D-sphingosine kinase pathway. IL-33 has demonstrated protective effects against various cardiovascular diseases (CVDs) by inducing Th2 cytokines and promoting alternative activating M2 polarization. However, the soluble decoy form of ST2 (sST2) mitigates the biological effects of IL-33, exacerbating CVDs. Furthermore, IL-33 also plays a significant role in the development of asthma, arthritis, atopic dermatitis, and anaphylaxis through the activation of Th2 cells and mast cells. In this review, we aim to demonstrate the protective role of IL-33 against CVDs from 2005 to the present and explore the potential of serum soluble ST2 (sST2) as a diagnostic biomarker for CVDs. Therefore, IL-33 holds promise as a potential therapeutic target for the treatment of CVDs.     

Immune response and cytokine production following immunization with experimental herpes simplex virus 1 (HSV-1) vaccines

Balb/c mice were immunized with the recombinant fusion protein gD1/313 (FpgD1/313 representing the ectodomain of HSV-1 gD), with the non-pathogenic ANGpath gE-del virus, with the plasmid pcDNA3.1-gD expressing full-length gD1 and with the recombinant immediate early (IE) HSV-1 protein ICP27. Specific antibodies against these antigens (as detected by ELISA) reached high titers with the exception of the DNA vaccine. High-grade protection against challenge with the virulent strain SC16 was found following immunization with the pcDNA3.1-gD plasmid and with the gE-del virus. Medium grade, but satisfactory protection developed after immunization with the FpgD1/313 and minimum grade protection was seen upon immunization with the IE/ICP27 polypeptide. A considerable response of peripheral blood cells (PBL) and splenocytes in the lymphocyte transformation test (LTT) was found in mice immunized with FpgD1/313, with the pcDNA3.1-gD plasmid and with the live ANGpathgE-del virus. For lymphocyte stimulation in vitro, the FpgD1/313 antigen was less effective than the purified gD1/313 polypeptide (cleaved off from the fusion protein); both proteins elicited higher proliferation at the 5 mug per 0.1 mL dose than at the 1 mug per 0.1 mL dose. The secretion of Th type 1 (TNF, IFN-gamma and IL-2) and Th type 2 (IL-4 and IL-6) cytokines was tested in the medium fluid of purified PBL and splenocyte cultures; their absolute values were expressed in relative indexes. The PBL from FpgD1/313 immunized mice showed increased secretion of both T(H)1 (TNF) as well as T(H)2 (IL-4) cytokines (7-10-fold, respectively). Splenocytes from FpgD1/313 immunized mice showed a significant (23-fold) increase in IL-4 production.     

Th1-Th17 Cells Mediate Protective Adaptive Immunity against Staphylococcus aureus and Candida albicans Infection in Mice

We sought to define protective mechanisms of immunity to Staphylococcus aureus and Candida albicans bloodstream infections in mice immunized with the recombinant N-terminus of Als3p (rAls3p-N) vaccine plus aluminum hydroxide (Al(OH₃) adjuvant, or adjuvant controls. Deficiency of IFN-γ but not IL-17A enhanced susceptibility of control mice to both infections. However, vaccine-induced protective immunity against both infections required CD4+ T-cell-derived IFN-γ and IL-17A, and functional phagocytic effectors. Vaccination primed Th1, Th17, and Th1/17 lymphocytes, which produced pro-inflammatory cytokines that enhanced phagocytic killing of both organisms. Vaccinated, infected mice had increased IFN-γ, IL-17, and KC, increased neutrophil influx, and decreased organism burden in tissues. In summary, rAls3p-N vaccination induced a Th1/Th17 response, resulting in recruitment and activation of phagocytes at sites of infection, and more effective clearance of S. aureus and C. albicans from tissues. Thus, vaccine-mediated adaptive immunity can protect against both infections by targeting microbes for destruction by innate effectors.     

Relative Contribution of Th1 and Th17 Cells in Adaptive Immunity to Bordetella pertussis: Towards the Rational Design of an Improved Acellular Pertussis Vaccine

Whooping cough caused by Bordetella pertussis is a re-emerging infectious disease despite the introduction of safer acellular pertussis vaccines (Pa). One explanation for this is that Pa are less protective than the more reactogenic whole cell pertussis vaccines (Pw) that they replaced. Although Pa induce potent antibody responses, and protection has been found to be associated with high concentrations of circulating IgG against vaccine antigens, it has not been firmly established that host protection induced with this vaccine is mediated solely by humoral immunity. The aim of this study was to examine the relative contribution of Th1 and Th17 cells in host immunity to infection with B. pertussis and in immunity induced by immunization with Pw and Pa and to use this information to help rationally design a more effective Pa. Our findings demonstrate that Th1 and Th17 both function in protective immunity induced by infection with B. pertussis or immunization with Pw. In contrast, a current licensed Pa, administered with alum as the adjuvant, induced Th2 and Th17 cells, but weak Th1 responses. We found that IL-1 signalling played a central role in protective immunity induced with alum-adsorbed Pa and this was associated with the induction of Th17 cells. Pa generated strong antibody and Th2 responses, but was fully protective in IL-4-defective mice, suggesting that Th2 cells were dispensable. In contrast, Pa failed to confer protective immunity in IL-17A-defective mice. Bacterial clearance mediated by Pa-induced Th17 cells was associated with cell recruitment to the lungs after challenge. Finally, protective immunity induced by an experimental Pa could be enhanced by substituting alum with a TLR agonist that induces Th1 cells. Our findings demonstrate that alum promotes protective immunity through IL-1β-induced IL-17A production, but also reveal that optimum protection against B. pertussis requires induction of Th1, but not Th2 cells.     

Evaluation of the C-Terminal Fragment of Entamoeba histolytica Gal/GalNAc Lectin Intermediate Subunit as a Vaccine Candidate against Amebic Liver Abscess

Background

Entamoeba histolytica is an intestinal protozoan parasite that causes amoebiasis, including amebic dysentery and liver abscesses. E. histolytica invades host tissues by adhering onto cells and phagocytosing them depending on the adaptation and expression of pathogenic factors, including Gal/GalNAc lectin. We have previously reported that E. histolytica possesses multiple CXXC sequence motifs, with the intermediate subunit of Gal/GalNAc lectin (i.e., Igl) as a key factor affecting the amoeba''s pathogenicity. The present work showed the effect of immunization with recombinant Igl on amebic liver abscess formation and the corresponding immunological properties.

Methodology/Principal Findings

A prokaryotic expression system was used to prepare the full-length Igl and the N-terminal, middle, and C-terminal fragments (C-Igl) of Igl. Vaccine efficacy was assessed by challenging hamsters with an intrahepatic injection of E. histolytica trophozoites. Hamsters intramuscularly immunized with full-length Igl and C-Igl were found to be 92% and 96% immune to liver abscess formation, respectively. Immune-response evaluation revealed that C-Igl can generate significant humoral immune responses, with high levels of antibodies in sera from immunized hamsters inhibiting 80% of trophozoites adherence to mammalian cells and inducing 80% more complement-mediated lysis of trophozoites compared with the control. C-Igl was further assessed for its cellular response by cytokine-gene qPCR analysis. The productions of IL-4 (8.4-fold) and IL-10 (2-fold) in the spleen cells of immunized hamsters were enhanced after in vitro stimulation. IL-4 expression was also supported by increased programmed cell death 1 ligand 1 gene.

Conclusions/Significance

Immunobiochemical characterization strongly suggests the potential of recombinant Igl, especially the C-terminal fragment, as a vaccine candidate against amoebiasis. Moreover, protection through Th2-cell participation enabled effective humoral immunity against amebic liver abscesses.     

Escherichia coli heat-labile enterotoxin promotes protective Th17 responses against infection by driving innate IL-1 and IL-23 production

Escherichia coli heat-labile enterotoxin (LT) is a powerful mucosal adjuvant; however, it is associated with toxic effects when delivered intranasally, and its mechanism of action is poorly understood. In this article, we demonstrate that LT acts as a highly effective adjuvant when administered parenterally, promoting Ag-specific IL-17, as well as IFN-γ, IL-4, and IL-10 production in response to coadministered Ags. We found that the adjuvant activity of LT was mediated in part by inducing dendritic cell (DC) activation; LT promoted CD80 and CD86 expression by DCs and enhanced IL-1α, IL-1β, and IL-23 production. An LT mutant, LTK63, that lacks enzyme activity was less effective than the wild-type toxin in promoting DC maturation and the development of Ag-specific Th17 cells. LT enhanced IL-23 and IL-1α production from DCs via activation of ERK MAPK and IL-1β secretion through activation of caspase-1 and the NLRP3 inflammasome. These cytokines played a major role in promoting Th17 responses by LT and LTK63. The induction of Th17 cells in vivo in response to LT and LTK63 as adjuvants was significantly reduced in IL-1RI-deficient mice. Finally, using a murine respiratory infection model, we demonstrated that LT can act as a highly effective adjuvant for a pertussis vaccine, promoting Ag-specific Th17 cells and protection against Bordetella pertussis challenge, which was significantly reduced in IL-17-defective mice. Our findings provide clear evidence that LT can promote protective immune responses in part through induction of innate IL-1 and, consequently, Th17 cells.     

DNA Vaccine Encoding the Chimeric Form of Schistosoma mansoni Sm-TSP2 and Sm29 Confers Partial Protection against Challenge Infection

Schistosomiasis is an important parasitic disease worldwide that affects more than 207 million people in 76 countries and causes approximately 250,000 deaths per year. The best long-term strategy to control schistosomiasis is through immunization combined with drug treatment. Due to the ability of DNA vaccines to generate humoral and cellular immune responses, such vaccines are considered a promising approach against schistosomiasis. Sm29 and tetraspanin-2 (Sm-TSP2) are two proteins that are located in the S. mansoni tegument of adult worms and schistosomula and induce high levels of protection through recombinant protein immunization. In this study, we transfected BHK-21 cells with plasmids encoding Sm29, Sm-TSP2 or a chimera containing both genes. Using RT-PCR analysis and western blot, we confirmed that the DNA vaccine constructs were transcribed and translated, respectively, in BHK-21 cells. After immunization of mice, we evaluated the reduction in worm burden. We observed worm burden reductions of 17-22%, 22%, 31-32% and 24-32% in animals immunized with the pUMVC3/Sm29, pUMVC3/SmTSP-2, pUMVC3/Chimera and pUMVC3/Sm29 + pUMVC3/SmTSP-2 plasmids, respectively. We evaluated the humoral response elicited by DNA vaccines, and animals immunized with pUMVC3/Sm29 and pUMVC3/Sm29 + pUMVC3/SmTSP-2 showed higher titers of anti-Sm29 antibodies. The cytokine profile produced by the spleen cells of immunized mice was then evaluated. We observed higher production of Th1 cytokines, such as TNF-α and IFN-γ, in vaccinated mice and no significant production of IL-4 and IL-5. The DNA vaccines tested in this study showed the ability to generate a protective immune response against schistosomiasis, probably through the production of Th1 cytokines. However, future strategies aiming to optimize the protective response induced by a chimeric DNA construct need to be developed.     

Partial Protective Effect of Intranasal Immunization with Recombinant Toxoplasma gondii Rhoptry Protein 17 against Toxoplasmosis in Mice

Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite that infects a variety of mammals, including humans. An effective vaccine for this parasite is therefore needed. In this study, RH strain T. gondii rhoptry protein 17 was expressed in bacteria as a fusion with glutathione S-transferase (GST) and the recombinant proteins (rTgROP17) were purified via GST-affinity chromatography. BALB/c mice were nasally immunised with rTgROP17, and induction of immune responses and protection against chronic and lethal T. gondii infections were investigated. The results revealed that mice immunised with rTgROP17 produced high levels of specific anti-rTgROP17 IgGs and a mixed IgG1/IgG2a response of IgG2a predominance. The systemic immune response was associated with increased production of Th1 (IFN-γand IL-2) and Th2 (IL-4) cytokines, and enhanced lymphoproliferation (stimulation index, SI) in the mice immunised with rTgROP17. Strong mucosal immune responses with increased secretion of TgROP17-specific secretory IgA (SIgA) in nasal, vaginal and intestinal washes were also observed in these mice. The vaccinated mice displayed apparent protection against chronic RH strain infection as evidenced by their lower liver and brain parasite burdens (59.17% and 49.08%, respectively) than those of the controls. The vaccinated mice also exhibited significant protection against lethal infection of the virulent RH strain (survival increased by 50%) compared to the controls. Our data demonstrate that rTgROP17 can trigger strong systemic and mucosal immune responses against T. gondii and that ROP17 is a promising candidate vaccine for toxoplasmosis.     

Pentavalent outer membrane vesicles of Vibrio cholerae induce adaptive immune response and protective efficacy in both adult and passive suckling mice models

Recently, we demonstrated oral immunizations with single serotype outer membrane vesicles of Vibrio cholerae induced serogroup specific protective immunity in the RITARD model. In our present study, we advanced our research by formulating multi-serotype outer membrane vesicles, mixing the OMVs of five virulent V. cholerae strains. Four doses of oral immunization with cholera pentavalent outer membrane vesicles (CPMVs) induced V. cholerae specific B and T cell responses. CPMVs-immunized mice generated long lasting serum IgG, IgA, IgM as well as mucosal sIgA and also elicited a higher percentage of CD4+ T cell distribution in spleen. Our study revealed that in vitro CPMVs-activated dendritic cells were secreting T cell polarizing cytokines, IL-12p40, IL-4, IL-6 and IL-1β. Moreover, purified splenic CD4+ T cells of immunized mice also secreted IL-4, IL-13 and IL-17 cytokines, indicating the initiation of Th2 and Th17 cell mediated immune responses. CPMVs immunized adult female mice and their offspring were significantly protected from heterologous challenge with wild type V. cholerae. CPMVs could be exploited for the development of a novel non-living vaccine against circulating cholera in near future.     

A multivalent combination of experimental antituberculosis DNA vaccines based on Ag85B and regions of difference antigens

Two candidate DNA vaccines based on the proteins CFP10 and CFP21 encoded by regions of difference (RDs) of Mycobacterium tuberculosis were evaluated individually and in multivalent combination with the immunodominant protein Ag85B for induction of protective immune responses against experimental tuberculosis. Experimental DNA vaccines induced substantial levels of cell-mediated immune responses as indicated by marked lymphocyte proliferation, significant release of the Th1 cytokines IFN-gamma and IL-12 (p40), and predominant cytotoxic T cell activity. High levels of antigen-specific IgG1 and IgG2a antibodies observed in the sera of immunized mice depicted strong humoral responses generated by DNA vaccine constructs. The multivalent combination of three DNA vaccine constructs induced maximal T cell and humoral immune responses. All the experimental vaccines imparted significant protection against challenge with M. tuberculosis H(37)Rv (in terms of colony-forming unit reduction in lungs and spleen) as compared to vector controls. The level of protection exhibited by multivalent DNA vaccine formulation was found to be equivalent to that of Mycobacterium bovis BCG observed both at 4 and 8 weeks post-challenge. These results show the protective potential of the multivalent DNA vaccine formulation used in this study.     

Protective immunity against Bordetella pertussis by a recombinant DNA vaccine and the effect of coinjection with a granulocyte-macrophage colony stimulating factor gene

A recombinant pertussis DNA vaccine was described here with its immunogenicity and the ability to induce protection against B. pertussis infection in mice. Three immunodominant antigen gene fragments of pertussis, pertussis toxin subunit 1 (pts1), fragments of pertactin (prn) and filamentous hemagglutinin (fha), were recombined as fragment pts1-prn-fha named ppf, and it was cloned to plasmid pVAX1 as pVAX1/ppf. Compared to those injected with pVAX1, the mice injected with pVAX1/ppf significantly elicited more antigen specific antibody anti-PTS1, anti-PRN, anti-FHA and cytokine IL-10, IFN-gamma. When pGM-CSF was coinjected with pVAX1/ppf, the mice showed significantly increases of the three antibodies and cytokine IL-10, IL-4, IFN-gamma and TNF-alpha compared to those injected with pVAX1 only. The mice in group pVAX1/ppf & pGM-CSF, in particular; induced much more anti-PTS1, IL-4 and TNF-alpha than those in group pVAX1/ppf. In the intracerebral mouse protection test, the mice immunized with pVAX1/ppf or pVAX1/ppf & pGM-CSF induced protection to a lethal dose of B. pertussis. The results indicate that recombinant DNA vaccine and pGM-CSF coinjection can induce protective immunity against B. pertussis, demonstrating a valuable method to prevent pertussis.     

Vaccination with Leishmania soluble antigen and immunostimulatory oligodeoxynucleotides induces specific immunity and protection against Leishmania donovani infection

In this report, we investigated the effect of ODN containing immunostimulatory CG motifs as adjuvant with soluble antigen (SA) from Leishmania donovani. BALB/c mice were vaccinated with the soluble antigen with or without CpG-ODN as adjuvant and then challenged with L. donovani metacyclic promastigotes. CpG-ODN alone resulted in partial protection against challenge with L. donovani. Immunization of mice with SA and CpG-ODN showed enhanced reduction in parasite load ( approximately 60%) when compared to SA ( approximately 40%) immunized mice. Immunization with SA by itself resulted in a mixed Th1/Th2 response whereas co-administration of SA with CpG-ODN resulted in a strong Th1 promoting isotype as they together promoted production of immunoglobulin G2a. Leishmania-specific Th1 cytokine response was induced by co-administering CpG-ODN and SA as they together promoted production of IFN-gamma and IL-12. In the present study, we demonstrate that immunostimulatory phosphorothioate-modified ODN are promising immune enhancers for vaccination against visceral leishmaniaisis.