Florfenicol impairs the immune responses to vaccination against foot-and-mouth disease in mice

Florfenicol is a new type of broad-spectrum antibacterial that has been used in veterinary clinics. It showed immunosuppressive activity on the immune responses to vaccination against foot-and-mouth disease virus (FMDV) serotype O in mice. In the present study, BALB/c mice were immunized subcutaneously with FMDV serotype O antigen on days 1 and 14. Beginning on day 21, mice were treated with a single daily oral dose of florfenicol (50, 100, and 200 mg/kg) for seven consecutive days. On day 28, blood samples were collected to analyze FMDV-specific immunoglobulin G (IgG), IgG1, and IgG2b antibodies, and splenocytes were harvested to assess lymphocyte proliferation, CD3(+) T- and CD19(+) B-lymphocyte subsets. The results presented here demonstrated that florfenicol not only significantly suppressed concanavalin A-, lipopolysaccharide-induced splenocyte proliferation but also decreased the percentage of CD19(+) B-cells in a dose-dependent manner and suppressed CD3(+) T-cell at high doses. Moreover, FMDV-specific IgG, IgG1, and IgG2b antibody levels in FMDV-immunized mice were reduced by florfenicol. These results suggested that florfenicol could suppress humoral and cellular immune responses to vaccination against foot-and-mouth disease (FMD) in mice.     

Comparison of immunological responses of plague vaccines F1+rV270 and EV76 in Chinese-origin rhesus macaque, Macaca mulatta

The subunit vaccine SV1 (20 μg F1 + 10 μg rV270) has been identified as the optimal formulation in mice, which provided a good protection against plague in mice, guinea pigs and rabbits. To compare SV1 and SV2 (200 μg F1 + 100 μg rV270) with live attenuated vaccine EV76, antibody responses, protective efficacy, cytokines (IFN‐γ, TNF‐α, IL‐2, IL‐4, IL‐10 and IL‐12) production, CD4/CD8 ratio and CD69⁺ T‐cell activation marker were determined in sera of the immunized Chinese‐origin rhesus macaques, Macaca mulatta. The immunized animals with SV1 or SV2 developed higher anti‐rV270 IgG titre, while those immunized with EV76 elicited a negligible IgG to V antigen, indicating that subunit vaccine (SV) had an advantage over EV76 in terms of the indispensable role of anti‐V antibody against Yersinia pestis. There was no significant antibody titre difference between SV1 and SV2, suggesting that the immune response may have been saturated at the dose level of SV1. There were no statistical changes for CD4/CD8 ratios, IL‐4 and CD69 levels between the three‐vaccine immunized groups. However, a significant higher level of IL‐12 was observed in the EV76 immunized animals, indicating that EV76 had an advantage over SV in respect of cellular immunity. Complete protection was observed for the immunized animals with SV and EV76, revealing that SV has a similar protective efficacy with EV76 against 6 × 10⁶ CFU of Y. pestis challenge by subcutaneous route in Chinese‐origin rhesus macaques.     

Adjuvant activity of monophosphoryl lipid A for nasal and oral immunization with soluble or liposome-associated antigen

The effectiveness of monophosphoryl lipid A (MPL) as a mucosal adjuvant was investigated following oral or intranasal (i.n.) administration of an aqueous adjuvant formulation of MPL (MPL-AF) added to soluble antigen or liposomal antigen or incorporated into liposomal antigen membranes. Groups of BALB/c female mice were immunized with 50 to 100 microg of free or liposomal Streptococcus mutans crude glucosyltransferase (C-GTF) with or without MPL-AF added to the vaccine or incorporated into the liposomal membrane. Plasma, saliva, vaginal wash, and fecal extract samples were collected biweekly following immunization and assessed for antigen-specific antibody activity by enzyme-linked immunosorbent assay (ELISA). Mice immunized by the i.n. route had higher levels of salivary, plasma, and vaginal immunoglobulin A (IgA) anti-C-GTF responses and higher levels of plasma IgG anti-C-GTF than the orally immunized groups. A second administration of the vaccine 14 weeks after the initial immunization resulted in an anamnestic response to C-GTF resulting in 10- and 100-fold increases in saliva and plasma IgA and plasma IgG, respectively (in the i.n. immunized groups). Mice receiving a second i.n. immunization with liposomal antigen and MPL-AF had higher salivary IgA anti-C-GTF responses than mice immunized with antigen plus MPL-AF or liposomal antigen (P < 0.05). Plasma IgG anti-C-GTF activity was highest in mice immunized by the i.n. route with antigen formulations containing MPL-AF (P < 0.05). These results demonstrate the effectiveness of MPL-AF as an adjuvant for potentiating mucosal and systemic immune responses to liposomal C-GTF following i.n. immunization.     

A clinically applicable adjuvant for an atherosclerosis vaccine in mice

Vaccination with MHC‐II‐restricted peptides from Apolipoprotein B (ApoB) with complete and incomplete Freund's adjuvant (CFA/IFA) is known to protect mice from atherosclerosis. This vaccination induces antigen‐specific IgG1 and IgG2c antibody responses and a robust CD4 T cell response in lymph nodes. However, CFA/IFA cannot be used in humans. To find a clinically applicable adjuvant, we tested the effect of vaccinating Apoe‐deficient mice with ApoB peptide P6 (TGAYSNASSTESASY). In a broad screening experiment, Addavax, a squalene‐based oil‐in‐water adjuvant similar to MF59, was the only adjuvant that showed similar efficacy as CFA/IFA. This was confirmed in a confirmation experiment for both the aortic arch and whole aorta analyzed by en face analysis after atherosclerotic lesion staining. Mechanistically, restimulated peritoneal cells from mice immunized with P6 in Addavax released significant amounts of IL‐10. Unlike P6 in CFA/IFA, vaccination with P6 in Addavax did not induce any detectable IgG1 or IgG2c antibodies to P6. These data suggest that squalene‐based adjuvants such as MF59 are good candidate adjuvants for developing a clinically effective atherosclerosis vaccine.     

Immune responses to vaccines involving a combined antigen–nanoparticle mixture and nanoparticle-encapsulated antigen formulation

Many physicochemical characteristics significantly influence the adjuvant effect of micro/nanoparticles; one critical factor is the kinetics of antigen exposure to the immune system by particle-adjuvanted vaccines. Here, we investigated how various antigen–nanoparticle formulations impacted antigen exposure to the immune system and the resultant antigen-specific immune responses. We formulated antigen with poly(lactic-co-glycolic acid) (PLGA) nanoparticles by encapsulating antigen within nanoparticles or by simply mixing soluble antigen with the nanoparticles. Our results indicated that the combined formulation (composed of antigen encapsulated in nanoparticles and antigen mixed with nanoparticles) induced more powerful antigen-specific immune responses than each single-component formulation. Mice immunized with the combined vaccine formulation displayed enhanced induction of antigen-specific IgG antibodies with high avidity, increased cytokine secretion by splenocytes, and improved generation of memory T cell. Enhanced immune responses elicited by the combined vaccine formulation might be attributed to the antigen-depot effect at the injection site, effective provision of both adequate initial antigen exposure and long-term antigen persistence, and efficient induction of dendritic cell (DC) activation and follicular helper T cell differentiation in draining lymph nodes. Understanding the effect of antigen–nanoparticle formulations on the resultant immune responses might have significant implications for rational vaccine design.     

CpG-ODN和氢氧化铝复合佐剂对流感病毒裂解疫苗免疫效果的影响

目的 研究CpG-ODN和氢氧化铝复合佐剂对流感病毒裂解疫苗体液免疫和细胞免疫效果的影响,为今后研制新佐剂流感疫苗和解决流感疫苗产能不足的问题提供依据.方法 以不同剂量的2009 H1N1流感病毒裂解疫苗为抗原,分别以CpG-ODN、氢氧化铝以及CpG-ODN和氢氧化铝复合佐剂为疫苗佐剂免疫BALB/c小鼠,通过ELISA、血凝抑制试验和假病毒中和试验等方法评价体液免疫效果,通过ELISPOT、胞内细胞因子染色和体内CTL杀伤等方法评价细胞免疫效果.结果 与无佐剂对照组相比,CpG-ODN或氢氧化铝单独使用能够在一定程度上增强体液免疫,2针免疫后不同抗原剂量组中抗原特异性IgG抗体滴度、血凝抑制抗体滴度和中和抗体滴度分别提高3～6倍、2～4倍和4～8倍.CpG-ODN和氢氧化铝复合佐剂具有更强的佐剂效应,2针免疫后不同抗原剂量组中抗原特异性IgG抗体滴度、血凝抑制抗体滴度和中和抗体滴度分别提高23～ 57倍、9～20倍和16～64倍.根据体液免疫结果,复合佐剂能够使流感病毒裂解疫苗的抗原用量降低至少16倍.此外,复合佐剂能够显著增强流感病毒裂解疫苗的细胞免疫应答,不但能够促进抗原特异性CD4+T细胞的IFN-γ分泌,而且能够促进抗原特异性CD8+T细胞的CTL杀伤活性.结论 CpG-ODN和氢氧化铝复合佐剂能够增强流感病毒裂解疫苗的体液免疫和细胞免疫应答并显著降低抗原用量.     

Rapeseed Oil and Ginseng Saponins Work Synergistically To Enhance Th1 and Th2 Immune Responses Induced by the Foot-and-Mouth Disease Vaccine

Previous investigations demonstrated that saponins isolated from the root of Panax ginseng C. A. Meyer (i.e., ginseng root saponin [GS-R]) had adjuvant activity. In the present study, the combined effects of rapeseed oil (RO) and GS-R on the immune responses elicited by foot-and-mouth disease (FMD) vaccine were investigated by measuring FMD virus (FMDV)-specific antibody levels, cytokine levels, lymphocyte proliferation, and long-lived IgG-secreting plasma cells from bone marrow in a mouse model. The results indicated that RO in combination with GS-R significantly enhanced serum IgG and isotype concentrations, gamma interferon (IFN-γ) and interleukin 5 (IL-5) levels, splenocyte proliferative responses to stimulations with concanavalin A (ConA), lipopolysaccharide (LPS), and FMDV antigen, and the numbers of IgG-secreting plasma cells in the bone marrow, suggesting that RO/GS-R enhanced both Th1 and Th2 immune responses. In addition, no significant difference was found between RO/GS-R and the commercial adjuvant oil ISA 206 in the promotion of FMD vaccine-induced immune responses. Considering the vegetable origin of RO and GS-R and the potent adjuvant activity, RO/GS-R should be studied further for the development of veterinary vaccines, especially for use in food animals in order to promote food safety.     

Artificially designed hepatitis B virus core particles composed of multiple epitopes of type A and O foot-and-mouth disease virus as a bivalent vaccine candidate

Recently, many countries, including China, have experienced a series of type A and O foot-and-mouth disease virus (FMDV) epidemics, causing serious economic losses. Although concerns about the safety of inactivated FMD vaccines have been raised, the development of a safe and effective subunit vaccine is necessary. We constructed two chimeric virus-like particles (VLPs; rHBc/AO and rHBc/AOT VLPs) displaying tandem repeats of B cell epitopes (VP1 residue 134-161 and 200-213) derived from type A and O FMDV and one T cell epitope (3 A residue 21-35) using the truncated hepatitis B virus core (HBc) carrier. Our results indicate that the chimeric HBc can self-assemble into VLPs with these FMDV epitopes displayed on the surface. Immunization with the chimeric VLPs induced specific IgG and neutralization antibodies against type A and O FMDV in mice. Compared with the commercial type A/O FMDV bivalent inactivated vaccine, rHBc/AO and rHBc/AOT VLPs significantly stimulated the production of Th1 type cytokines (IFN-γ and IL-2), whereas Th2 cytokine production (IL-4 and IL-10) was decreased. Compared with rHBc/AO, rHBc/AOT induced increased Th2 cytokine and specific IgG production. These results demonstrate that the VLPs constructed in the current study induced both humoral and cellular immune responses and may represent potential bivalent VLP vaccines targeting both FMDV type A and O strains.     

Hesperetin as an adjuvant augments protective anti-tumour immunity responses in B16F10 melanoma by stimulating cytotoxic CD8+ T cells

Hesperetin (HES) is a dihydroflavone with the molecular formula of C16H14O6. It has been reported that Hesperetin has antioxidant and anticancer effects. Recent studies showed that it can also regulate immune responses. To assess its potential function as a vaccine adjuvant, we formulated HES with inactivated B16F10 melanoma cells and determined whether it would enhance the activation of antigen-presenting cells by experiments in vivo and in vitro. We found that HES activated the PI3K-Akt signalling pathway in antigen-presenting cells (APCs), enhanced cytotoxic T lymphocyte (CTL) responses and deactivated tolerogenic T cells. We also observed that inactivated B16F10 cells in combination with HES vaccine inhibited the growth of mice tumours, resulting in improved overall survival compared to the effects of inactivated B16F10 cell vaccine. To verify that CD8⁺T cells play a key role in inhibiting the development of melanoma, we transferred the sorted CD8⁺T cells from immunized mice to B16F10 challenged models and found that the survival rate of tumour-bearing mice was significantly prolonged. Taken together, these results suggest that hesperetin can be used as a potential adjuvant to improve tumour immune responses and antigen immunogenicity.     

A DNA vaccine encoding the FMDV capsid precursor polypeptide P1 and the enhancing effect of bovine herpesvirus 1 VP22 protein as molecular adjuvant

DNA vaccines containing the capsid precursor polypeptide P1 gene of foot-and-mouth disease virus (FMDV) alone or combined with the VP22 gene of bovine herpesvirus 1 (BVP22) as molecular adjuvant were constructed and used for immunization of BALB/c mice. The latter were challenged with FMDV and their humoral as well as cell-mediated immune responses and virus clearance capacity were assayed. Both DNA vaccines elicited specific immune responses, however, the DNA vaccine with the BVP22 adjuvant showed stronger responses and more efficient virus clearance. A?stronger Th1 response was indicated by the IgG2a/IgG1 ratio. These results indicate that (i) a?DNA vaccine based on FMDV P1 can stimulate significant immune responses and virus clearance and (ii) BVP22 is a?potentially useful molecular adjuvant for such a?vaccine. Keywords: DNA vaccine; foot-and-mouth disease virus; bovine herpesvirus 1.     

Vitamin C supplementation improved the efficacy of foot-and-mouth disease vaccine

Vitamin C (VC) is an essential micronutrient and plays important roles in various biological processes including immune responses. Vaccination can stimulate immune responses effectively for prevention of virual infection. Adequate VC intake is required for the immune system to function efficiently. In this study, we evaluated the biological function of VC on immune response to inactivated foot-and-mouth disease (FMD) vaccine. The C57BL/6 mice were injected intraperitoneally with different dose of VC daily following immunization with inactivated FMD vaccine. The results showed high dose of VC supplementation significantly increased IgG level. Similarly, high dose of VC supplementation enhanced Th2 and Th17 cellular immune responses and also increased MHCII expression on dendritic cells. Taken together, these data indicate that high dose of VC supplementation can modulate Th2 and Th17 cellular immune responses to inactivated FMD vaccine and manifest the necessity for effective vaccination.     

Plasmids encoding foot-and-mouth disease virus VP1 epitopes elicited immune responses in mice and swine and protected swine against viral infection

VP1 is a capsid protein of foot-and-mouth disease virus (FMDV) and contains epitopes of the virus. Plasmids encoding two VP1 epitopes (amino acid residues 141-160 and 200-213) and a host-self immunoglobulin molecule were constructed to produce a new type of FMD DNA vaccine. Two plasmids, namely, pCEIM and pCEIS, containing mouse immunoglobulin (IgG) or swine IgG were subjected to immunogenicity testing in mice and swine, respectively. In mice administrated pCEIM in the abdomen using a genegun, both FMDV-specific T-cell proliferation and neutralizing antibodies were detected. In swine immunized with pCEIS at the back of the ear, immune responses were achieved after the second administration. Swine showed a T-cell proliferative response with a stimulation index (SI) of up to 8.1 and a neutralizing antibody response that was able to protect suckling mice from 10(2) LD(50) (lethal dose 50) FMDV challenge. To compare the immunogenicity of the DNA-based vaccine candidate, versus the protein-based vaccine candidates, a second group of swine was immunized with the protein F1-scIgG, which was encoded by the plasmid pCEIS. Injection with F1-scIgG elicited a T-cell proliferative response of SI < 1.7 and a neutralizing antibody response that protected suckling mice from up to 10(5) LD(50) FMDV challenge. In the challenge test, three of three swine immunized with pCEIS were fully protected from FMDV challenge.     

Robust immune response elicited by a novel and unique Mycobacterium tuberculosis protein using an optimized DNA/protein heterologous prime/boost protocol

An efficacious tuberculosis (TB) vaccine will probably need to induce both CD4 and CD8 T‐cell responses specific to a protective Mycobacterium tuberculosis antigen(s). To achieve this broad cellular immune response we tested a heterologous DNA/protein combination vaccine strategy. We used a purified recombinant protein preparation of a unique M. tuberculosis antigen (rMT1721) found in the urine of TB patients, an optimized plasmid DNA expressing this protein (DNA‐MT1721), and a Toll‐like receptor 4 agonist adjuvant. We found that priming mice with DNA‐MT1721 and subsequently boosting with rMT1721 elicited high titres of specific IgG1 and IgG2a antibodies as well as high magnitude and polyfunctional CD4⁺ T‐cell responses. However, no detectable CD8⁺ T‐cell response was observed using this regimen of immunization. In contrast, both CD4⁺ and CD8⁺ T‐cell responses were detected after a prime/boost vaccination regimen using rMT1721 as the priming antigen and DNA‐MT1721 as the boosting immunogen. These findings support the exploration of heterologous DNA/protein immunization strategies in vaccine development against TB and possibly other infectious diseases.     

Naloxone and alum synergistically augment adjuvant activities of each other in a mouse vaccine model of Salmonella typhimurium infection

Alum is the most commonly used adjuvant for human vaccination but is a poor inducer of cell mediated immunity and T helper 1 (Th1) responses. We have previously shown that naloxone (NLX), which is a general opioid antagonist, acts as an effective adjuvant in enhancing vaccine-induced cellular immunity and Th1 immune responses. Here, we tested the efficacy of an alum-NLX mixture, as a new adjuvant, in the induction of humoral and cellular immunity in response to endotoxin-removed lysate (ERL) of Salmonella typhimurium (S. typhimurium) as a model vaccine. BALB/c mice were divided into five vaccination groups. Mice in the experimental groups received either the ERL vaccine alone or in combination with the adjuvant alum, NLX or the alum-NLX mixture. Mice in the negative control group received phosphate-buffered saline. All mice were immunized on days 0 and 7. Two weeks after the last immunization, immune responses to S. typhimurium were assessed. Our results indicate that including the alum-NLX mixture as an adjuvant during vaccination increased the ability of the ERL vaccine to enhance lymphocyte proliferation, shifted the immune response toward a Th1 profile and increased S. typhimurium-specific IgG, IgG2a and the ratio of IgG2a to IgG1. This resulted in improved protective immunity against S. typhimurium. In conclusion, administering an alum-NLX mixture adjuvant in combination with the ERL vaccine enhances both humoral and cellular immunity, and shifts the immune response to a Th1 pattern.     

Enhancement of serological immune responses to foot-and-mouth disease vaccine by a supplement made of extract of cochinchina momordica seeds

Foot-and-mouth disease (FMD) is a highly contagious disease affecting cloven-hoofed animals. Vaccination against FMD is a routine practice in many countries where the disease is endemic. This study was designed first to investigate the extract of the seeds of Momordica cochinchinensis (Lour.) Spreng. (ECMS) for its adjuvant effect on vaccination of inactivated FMDV antigens in a guinea pig model and then to evaluate the supplement of ECMS in oil-emulsified FMD vaccines for its immunopotentiation in pigs. The results indicated that ECMS and oil emulsion act synergistically as adjuvants to promote the production of FMDV- and VP1-specific immunoglobulin G (IgG) and subclasses in guinea pigs. A supplement of ECMS in a commercial FMD vaccine significantly enhanced FMDV-specific indirect hemagglutination assay titers as well as VP1-specific IgG and subclasses in pigs. Therefore, ECMS could be an alternative approach to improving swine FMD vaccination when the vaccine is poor to induce an effective immune response.     

Enhanced immune response to foot-and-mouth disease vaccine by oral administration of ginseng stem-leaf saponins

Vaccination is an important approach to the control of foot-and-mouth disease (FMD). This study evaluated the effect of oral administration of ginseng stem–leaf saponins (GSLS) on the immune response to FMD vaccine and the gut mucosal immunity in mice. In experiment 1, mice were orally administered GSLS or not treated as a control. The animals were then immunized twice with FMD vaccine. Blood was sampled weekly within five weeks after the boost immunization for measurement of serum IgG and the isotypes. In experiment 2, mice were orally administrated GSLS or not treated as a control. After that, splenocytes were prepared from sacrificed mice for lymphocyte proliferation assay and intestinal tissues were sampled for immunohistochemistry and histological examination. The results showed that oral administration of GSLS significantly enhanced serum IgG and the isotype responses to FMD vaccine as well as the number of intestinal intraepithelial lymphocytes (IELs) and immunoglobulin A (IgA)+?cells. Therefore, GSLS may be a potent oral adjuvant and deserve further study to improve vaccination in susceptible animals.     

Efficacy of modified levamisole adjuvant on inactivated virus vaccine

To improve efficacy, especially for the cell-mediated response to inactivated viral vaccines, a modified levamisole (LMS) adjuvant formulation, designated LMS+, was evaluated for its efficacy in mice and chickens, using Newcastle Disease Virus (NDV) as a model pathogen. Compared with oil adjuvant, the killed NDV in LMS+ induced a significantly higher helper T cell type 1 response, as shown by higher levels of interleukin-2, interferon-gamma, T cell proliferation, and delayed-type hypersensitivity responses, without sacrificing the level of IgG production in mice. In addition, vaccine in LMS+ formulation increased the expression of MHC and costimulatory molecules as well as the number of CD11c+ dendritic cells, suggesting that the better response to the LMS+ formulation occurs partly via the maturation of dendritic cells and activation of MHC-antigen presentation and costimulation. Furthermore, this formulation provides 100% protection in chickens after challenge with a lethal dose of virulent NDV strain F48E9 at 1000 ELD50 (50% egg lethal dose). These results demonstrated that modified LMS+ adjuvant could be used to improve both humoral and cell-mediated responses for inactivated viral vaccines and its development as an effective inactivated viral vaccine is warranted.     

IL-17作为分子佐剂增强HIV DNA疫苗细胞免疫应答的研究

目的 为了进一步增强HIV DNA疫苗的免疫反应,本研究将IL-17作为HIV DNA疫苗的分子佐剂免疫小鼠,旨在探讨IL-17对HIVDNA疫苗诱导体液和细胞免疫应答的影响.方法 将小鼠IL-17构建到真核表达载体,与HW-1膜蛋白DNA疫苗pGX-Env联合免疫BALB/c小鼠;分别在第0、2周进行免疫,在第4周检测T淋巴细胞增殖指数、抗-Env IgG、细胞因子表达水平和体内细胞毒性T淋巴细胞杀伤作用(CTL)等免疫学指标.结果 IL-17能够增强HIV DNA疫苗的特定免疫反应.与单注射疫苗组相比,IL-17作为佐剂组的T细胞增殖、抗体水平和CD4~+T细胞分泌IFN-γ、IL-4和IL-17的表达均无明显增强,但对CD8~+T细胞分泌IFN-γ的表达和体内CTL的效果影响明显(P<0.05).结论 IL-17作为分子佐剂不足以影响Th细胞分化,然而却能够增强特异性CD8~+T细胞中IFN-γ的表达,尤其是增强体内CTL反应.此结果为增强艾滋病DNA疫苗CD8~+T细胞活性和用于艾滋病的治疗提供了一个新的思路.     

Granulocyte‐macrophage colony‐stimulating factor,a potent adjuvant for polarization to Th‐17 pattern: an experience on HIV‐1 vaccine model

Cytokines are mediators for polarization of immune response in vaccines. Studies show that co‐immunization of DNA vaccines with granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) can increase immune responses. Here, experimental mice were immunized with HIV‐1_{tat/pol/gag/env} DNA vaccine with GM‐CSF and boosted with recombinant vaccine. Lymphocyte proliferation with Brdu and CTL activity, IL‐4, IFN‐γ, IL‐17 cytokines, total antibody, and IgG1 and IgG2a isotypes were assessed with ELISA. Results show that GM‐CSF as adjuvant in DNA immunization significantly increased lymphocyte proliferation and IFN‐γ cytokines, but CTL response was tiny increased. Also GM‐CSF as adjuvant decreased IL‐4 cytokine vs mere vaccine group. IL‐17 in the group that immunized with mixture of DNA vaccine/GM‐CSF was significantly increased vs DNA vaccine group. Result of total antibody shows that GM‐CSF increased antibody response in which both IgG1 and IgG2a increased. Overall, results confirmed the beneficial effect of GM‐CSF as adjuvant to increase vaccine immunogenicity. The hallmark result of this study was to increase IL‐17 cytokine with DNA vaccine/GM‐CSF immunized group. This study for the first time provides the evidence of the potency of GM‐CSF in the induction of IL‐17 in response to a vaccine, which is important for control of infection such as HIV‐1.     

Immune responses induced by heterologous boosting of recombinant bacillus Calmette-Guerin with Ag85B-ESAT6 fusion protein in levamisole-based adjuvant

In the present study, we evaluated the effectiveness of a levamisole-based adjuvant (ADL) to enhance the ability of the Ag85B-ESAT6 fusion protein to boost immune responses after primary vaccination with recombinant bacillus Calmette-Guerin (rBCG) in Balb/c mice. The results were compared with that of the control adjuvant formulation of dimethyl dioctadecylammonium bromide (DDA) and monophosphoryl lipid A (MPL), which has previously been shown to induce T-helper type 1 (Th1)-biased responses. Enzyme-linked immunospot (ELISPOT) assay with Ag85B and ESAT6 derived peptides corresponding to CD4+ and CD8+ T cell restricted epitopes and cell surface immunostaining indicated that Ag85B-ESAT6/ADL predominantly triggered activation of CD4+ T cells. Functional CD8+ T cells with interferon (IFN)-γ production or cytotoxicity were undetectable all vaccinated mice. The ADL adjuvant modified T-helper (Th) subtypes by up-regulating multiple signature cytokines. Furthermore, profiles of the immunoglobulin G (IgG) subtypes indicated ADL enhanced the secretion of Th1-associated IgG2a antibodies and decreased the yield of Th2-associated IgG1 subtype. These observations suggest that the ADL adjuvant formulated with a protein booster may induce Th1-biased cellular and humoral immune responses to primary vaccination with a live attenuated bacterial TB vaccine.