In vivo oral administration effects of various oligodeoxynucleotides containing synthetic immunostimulatory motifs in the immune response to pseudorabies attenuated virus vaccine in newborn piglets.

Numerous studies have demonstrated that oligonucleotides containing CpG motifs (CpG ODN) are efficient immunoadjuvants to various antigens administered by parenteral routes to mice. Recently, it has been found that CpG ODNs also is a promising mucosal adjuvant in mice. To date, there have been no studies to screen the optimal CpG sequence and modified ODN backbone to piglets in vivo, when delivered by oral route. We have previously demonstrated that human-specific CpG ODN is a potent adjuvant to pseudorabies live attenuated virus (PRV) vaccine when administered subcutaneously (SC) or ocularly in piglets. In this study, we screened and evaluated the optimal CpG sequences (porcine-specific, human-specific, mouse-specific ODN) and optimal backbone (SOS-backbone consisting of a nuclease-resistant phosphorothioate guanosines at the 5' and the 3'-end and with a phosphodiester (O) in the center and phosphorothioate (S) backbone (S-backbone)) to PRV vaccine delivered orally in piglets. The proliferation of peripheral blood mononuclear cells (PBMCs), IFN-gamma and IL-4 in serum, and the titre of IgG, IgG2/IgG1 isotype in serum and IgA in intestinal washings and feces to PRV vaccine were tested at different time-points. The results suggested that, CpG ODNs augmented systemic (IgG in serum, T-cell proliferation) and mucosal (IgA in intestinal washings and feces) immune responses against antigen. CpG ODNs stimulated both T-helper type1 (Th1) (IgG2) and Th2 (IgA) responses when delivered orally. With the same backbone, the porcine-specific ODN-induced responses were comparable with human-specific ODNs, but stronger than mouse-specific CpG ODNs. SOS-backbone induced a stronger IFN-gamma and proliferative responses than S-backbone, while antibody responses induced by SOS-backbones were slightly less or similar with S-backbone. The in vivo data demonstrate for the first time that porcine-specific and human-specific ODNs both are optimal sequences for mucosal system in piglets.

Vaccination with porcine reproductive and respiratory syndrome killed virus vaccine and immunostimulatory oligodeoxynucleotides induces specific immunity in piglets.

A large number of studies demonstrated the immunostimulatory effects of CpG oligonucleotides (ODN), particularly in mice. In present study, the objective was to investigate the immunoadjuvant effects of CpG ODN to porcine reproductive and respiratory syndrome (PRRS) killed virus vaccine (PRRSV) and its protective effects against PRRS virus in piglets. The PRRSV-specific antibodies titres and serum IgG1/IgG2 titres, the proliferation of lymphocytes, PRRSV-specific IFN-gamma, the expression of major histocompatibility complex class II (MHCII) of peripheral blood mononuclear cells (PBMCs) and post-challenge clinical protection were examined to identify the immune responses of the piglets. The results were found that the above-mentioned immune responses of the piglets inoculated with CpG ODN plus PRRSV were significantly stronger than those indued by PRRSV or PBS control groups. All these data suggested that CpG ODN could be employed as effective immunoadjuvant to raise the humoral and cellular responses of the piglets to PRRSV.

In vivo immunostimulatory effects of CpG ODN in newborn piglets.

The in vivo immunoadjuvant effects of CpG oligodeoxynucleotides (CpG ODN) have been studied extensively in mice and relatively fewer studies have been done in other species. But so far, the innate immunostimulatory effects of CpG ODN have been demonstrated just in mouse, monkey, sheep and chicken in some reports. The purpose of this study is to determine the potential effects of CpG ODN in newborn piglets. The proportion of CD4(+), CD8(+) T lymphocytes subpopulations and the major histocompability complex (MHC-II) antigen expression of peripheral blood mononuclear cells (PBMCs) and IFN-gamma in serum were tested at various time-points. The results suggested that, the CD4(+)/CD8(+) ratio decreased over time in piglets inoculated with phosphate buffer saline (PBS) alone, however, it was stable in CpG ODN-inoculated piglets; the use of CpG ODN can prevent effectively the reduction of the proportion of CD4(+) T lymphocytes. The MHC-II antigen expression and IFN-gamma level of CpG ODN-injected piglets were significantly higher than those of PBS-injected piglets. The ODN-induced responses were stronger in animals injected with CpG ODN formulated in 30% emulsigen than in PBS. The innate immunostimulatory activity of CpG ODN appeared to be in dose-dependent manner. These in vivo data demonstrate for the first time that CpG ODN can stimulate innate immune system in newborn piglets.

Vaccination with Newcastle disease vaccine and CpG oligodeoxynucleotides induces specific immunity and protection against Newcastle disease virus in SPF chicken.

Oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODN) have been proven to be immunoprotective in mouse models. However, little work has been conducted on in vivo immune responses in chicken with CpG ODN. The objective of this study was to investigate the immunoadjuvant effects of CpG ODN to Newcastle disease (ND) vaccine and its protective effects against ND virus in SPF chicken. In this report, the titre of serum IgG to ND vaccine and the proliferation of lymphocytes were monitored in SPF chickens. The results demonstrated that the above-mentioned immune responses were significantly stronger in chickens that received CpG ODN than in the birds that received only ND vaccine. Furthermore, ND vaccine plus CpG ODN protected SPF chicken from challenge with an otherwise lethal dose of ND virus. These data suggest that CpG ODN holds considerable promise as an adjuvant for future vaccines against ND virus.

In vivo effects of oligodeoxynucleotides containing synthetic immunostimulatory motifs in the immune response to swine streptococcic septicemia vaccine in weaned piglets.

The diverse immunostimulatory effects of CpG oligodeoxynucleotides (CpG ODN) have been demonstrated extensively in mice and human. Although the immunoadjuvant effects of CpG ODN in pigs were also studied in several reports, until now, little work has been carried out with regard to their effects on the adaptive immune system of newly weaned piglets. In this study, swine streptococcic septicemia killed vaccine (SSSK vaccine) was used as antigen, we assessed the in vivo immunostimulatory effects of different CpG motifs in newly weaned piglets. The proportion of CD4(+), CD8(+) T lymphocytes subpopulations and proliferation of peripheral blood mononuclear cells (PBMCs), IFN-gamma and IL-6 in serum, and the titre of IgG and IgG2/IgG1 isotype to SSSK vaccine in serum were tested at different time-points. The results suggested that, the CD4(+)/CD8(+) ratio decreased significantly in weaned piglets inoculated with phosphate buffer saline (PBS) alone, however, it was stable in CpG ODN-coinoculated newly weaned piglets. IFN-gamma and IL-6 levels, the titres of specific antibodies IgG, IgG2 and proliferative responses of CpG ODN-coinjected piglets were all significantly higher than those of SSSK vaccine alone or PBS or GpC ODN-coinjected piglets. The porcine-specific ODN-induced responses were stronger in animals injected with human-specific or mouse-specific CpG ODN. These in vivo data demonstrate for the first time that CpG ODN can stimulate adaptive immune system in weaned piglets.

The efficacy of CpG oligodinucleotides, in combination with conventional adjuvants, as immunological adjuvants to swine streptococcic septicemia vaccine in piglets in vivo.

Oligodinucleotides containing CpG motifs (CpG ODN) are strong adjuvants for immune responses, particularly in mice, the immunostimulatory effects of CpG in combination with aluminum hydroxide (alum) or Emulsigen (Em) were investigated in cattle, rabbits or mice, but not piglets. In this report, using the swine streptococcus as model bacteria, the efficacy of CpG ODN as an adjuvant for piglets was assessed alone and in combination with alum (CpG/alum) or Em (CpG/Em). The CpG/alum or CpG/Em combination elicited greater immune responses to swine streptococcic septicemia killed vaccine (SSSK vaccine) compared with CpG alone, or alum or Em. A GpC/alum or GpC/Em combination did not have the same effects as CpG/alum or CpG/Em suggesting that the adjuvanticity was related to the CpG motifs. In addition, we also found that the 10% Em in combination with CpG ODN had similar immunological effects as 30% Em combination. Our results demonstrate that the addition of CpG ODN to alum or to Em significantly improves the efficiency of the adjuvants in piglets.

CpG oligodinucleotides induce strong humoral and cellular responses to swine streptococcic septicemia vaccine in piglets in vivo.

Oligodinucleotides containing CpG motifs (CpG ODN) are strong adjuvants for immune responses, particularly in mice, but data on humoral and cellular immune responses in piglets are scarce. In this report, using the swine streptococcus as model bacteria, CpG ODN was used as immunoadjuvants to enhance the immune responses of the piglets to swine streptococcic septicemia killed vaccine (SSSK vaccine). The titre of specific antibodies to SSSK vaccine, the proliferation of lymphocytes, SSSK-specific interferon-gamma(IFN-gamma) and IL-6, the expression of major histocompatibility complex class II (MHC-II) and CD14 of peripheral blood mononuclear cells (PBMCs) were examined to identify the immune responses of the piglets. The results were found that the above-mentioned immune responses of the piglets with CpG ODN were significantly stronger than standard immunization protocols. All these data suggested that immunostimulatory CpG ODN was promising immune enhancers for vaccination against SSSK vaccine.

Co-administration of porcine-specific CpG oligodeoxynucleotide enhances the immune responses to pseudorabies attenuated virus vaccine in newborn piglets in vivo.

Oligonucleotides containing CpG motifs (CpG ODN) are strong adjuvants for humoral and cellular immune responses in mice, but data on immune responses in piglets are scarce. In this report, porcine-specific CpG ODN were used as immunoadjuvants to enhance the immune responses of the newborn piglets to Pseudorabies attenuated virus (PRV) vaccine. The titres of specific antibodies and serum IgG1/IgG2 ratio to PRV vaccine, the proliferation of peripheral blood mononuclear cells (PBMCs), IL-4 and interferon-gamma(IFN-gamma) in piglets serum were examined to identify the immune response of the newborn piglets. The results showed that piglets immunized with PRV vaccine and CpG ODN presented high titers of PRV-specific antibodies and IgG2 isotype, a Th1-dominated (IFN-gamma) cytokine profile, together with inducing higher proliferation of PBMCs. All these data indicate that CpG ODN are potential effective adjuvants for the PRV vaccine in newborn piglets.

Effects of CpG ODN on CD4+ and CD8+ T subpopulations in the immune response to porcine reproductive and respiratory syndrome killed virus vaccine.

CpG ODN is a noval immunostimulatory reagent In this research, the effects of immunostimulatory CpG oligodeoxynucleotides (CpG ODN) on CD4+ and CD8+ T lymphocytes subpopulations in the newborn piglets blood were tested at different time with porcine reproductive and respiratory syndrome killed virus vaccine (PRRS vaccine) with or without CpG ODN. The results suggested that, the CD4+/CD8+ ratio decreased with age in piglets inoculated with vaccine alone or GpC ODN with vaccine or phosphate buffer saline (PBS), however, it was stable in piglets co-inoculated with CpG ODN and PRRS vaccine (p>0.05), the use of CpG ODN can prevent effectively the reduction of the proportion of CD4+ T lymphocytes. High titers of PRRS specific antibody can also be tested in the newborn piglets serum immunized PRRS vaccine and CpG ODN (p<0.05).