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Pilot Study on Interferon-γ-producing T Cell Subsets after the Protective Vaccination with Radiation-attenuated Cercaria of Schistosoma japonicum in the Miniature Pig Model.

Abdel-Hafeez EH, Watanabe K, Kamei K, Kikuchi M, Chen H, Daniel B, Yu C, Hirayama K - Trop Med Health (2014)

Bottom Line: To characterize the protective immune response of the pig model vaccinated by attenuated cercaria, flow cytometric analysis of the reactive T cell subsets was performed.CD4+ αβ-T cells as well as CD4+/CD8α(mid) double positive and/or CD8α(high) αβ-T cells were the major IFN-γ-producing CD3+ T cells.On the contrary, γδ T cells did not produce intracellular IFN-γ.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunogenetics, Institute of Tropical Medicine (NEKKEN), Nagasaki University , 1-14-2 Sakamoto, Nagasaki 852-8523, Japan ; Department of Parasitology, Faculty of Medicine, Minia University , Minia, 61519, Egypt.

ABSTRACT
CLAWN miniature pig has been shown to serve as a suitable host for the experimental infection of Schistosoma japonicum. In this study, we found that radiation-attenuated cercaria (RAC) vaccine gave CLAWN miniature pigs protective immunity against subsequent challenge infection with S. japonicum cercaria. To characterize the protective immune response of the pig model vaccinated by attenuated cercaria, flow cytometric analysis of the reactive T cell subsets was performed. The intracellular interferon (IFN)-γ and the cell surface markers revealed the peripheral blood CD3+ T-lymphocytes produced significant amounts of IFN-γ during the immunization period and after the challenge infection. CD4+ αβ-T cells as well as CD4+/CD8α(mid) double positive and/or CD8α(high) αβ-T cells were the major IFN-γ-producing CD3+ T cells. On the contrary, γδ T cells did not produce intracellular IFN-γ. Our results suggested that RAC-vaccinated miniature pigs showed effective protective immunity through the activation of αβ T cells bearing antigen specific T-cell receptors but not through the activation of γδ T cells.

No MeSH data available.


Related in: MedlinePlus

Flow cytometric analysis of the cellular source of IFN-γ in RAC-immunized miniature pig PBLPeripheral blood of the immunized pigs was collected at the time of scarification. PBMC were then stimulated with SWA for 3 days. The samples were cultured with PMA, ionomycin and breferdin A for 4 h. Cells were stained with CD3, CD16, γδ TCR, CD4, CD8α and IFN-γ antibodies. Lymphocyte-gated cells were analyzed. IFN-γ expression was examined in (a) CD3, γδ TCR+ (b) CD16+ (c) CD4+ and/or CD8α cells.
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Figure 3: Flow cytometric analysis of the cellular source of IFN-γ in RAC-immunized miniature pig PBLPeripheral blood of the immunized pigs was collected at the time of scarification. PBMC were then stimulated with SWA for 3 days. The samples were cultured with PMA, ionomycin and breferdin A for 4 h. Cells were stained with CD3, CD16, γδ TCR, CD4, CD8α and IFN-γ antibodies. Lymphocyte-gated cells were analyzed. IFN-γ expression was examined in (a) CD3, γδ TCR+ (b) CD16+ (c) CD4+ and/or CD8α cells.

Mentions: Our observation of cytokine response suggested that RAC immunization in miniature pigs elicits an IFN-γ-mediated immune response, similar to the findings reported for S. mansoni infection in mice [22, 23]. Therefore, we examined the cellular source of IFN-γ, as few papers have reported the source of IFN-γ during S. japonicum infection in pigs. In order to examine the cellular source of IFN-γ, immunized miniature pig PBMC were cultured in the presence of SWA and then stimulated with PMA and ionomycin for 4 h. The PBMC were then stained for intracellular IFN-γ and analyzed using flow cytometry (Fig. 3). Based on forward and side scatter, IFN-γ-positive cells were lymphocytes (data not shown). Among the lymphocytes observed, almost all of the IFN-γ-positive cells were also positive for CD3 and negative for γδ TCR (Fig. 3a). Thus, we suggest that the conventional T cells expressing αβ TCR are the most likely major source of IFN-γ in the immunized miniature pigs. We also examined the natural killer (NK) cells, which are strong producers of IFN-γ [24], using CD16 as a marker of NK cells among other lymphocytes. CD16 is a low affinity receptor for IgG and is expressed on monocytes and a population of NK cells [25]. Additional specific markers of porcine NK cells have yet to be clearly established [18].


Pilot Study on Interferon-γ-producing T Cell Subsets after the Protective Vaccination with Radiation-attenuated Cercaria of Schistosoma japonicum in the Miniature Pig Model.

Abdel-Hafeez EH, Watanabe K, Kamei K, Kikuchi M, Chen H, Daniel B, Yu C, Hirayama K - Trop Med Health (2014)

Flow cytometric analysis of the cellular source of IFN-γ in RAC-immunized miniature pig PBLPeripheral blood of the immunized pigs was collected at the time of scarification. PBMC were then stimulated with SWA for 3 days. The samples were cultured with PMA, ionomycin and breferdin A for 4 h. Cells were stained with CD3, CD16, γδ TCR, CD4, CD8α and IFN-γ antibodies. Lymphocyte-gated cells were analyzed. IFN-γ expression was examined in (a) CD3, γδ TCR+ (b) CD16+ (c) CD4+ and/or CD8α cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
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Figure 3: Flow cytometric analysis of the cellular source of IFN-γ in RAC-immunized miniature pig PBLPeripheral blood of the immunized pigs was collected at the time of scarification. PBMC were then stimulated with SWA for 3 days. The samples were cultured with PMA, ionomycin and breferdin A for 4 h. Cells were stained with CD3, CD16, γδ TCR, CD4, CD8α and IFN-γ antibodies. Lymphocyte-gated cells were analyzed. IFN-γ expression was examined in (a) CD3, γδ TCR+ (b) CD16+ (c) CD4+ and/or CD8α cells.
Mentions: Our observation of cytokine response suggested that RAC immunization in miniature pigs elicits an IFN-γ-mediated immune response, similar to the findings reported for S. mansoni infection in mice [22, 23]. Therefore, we examined the cellular source of IFN-γ, as few papers have reported the source of IFN-γ during S. japonicum infection in pigs. In order to examine the cellular source of IFN-γ, immunized miniature pig PBMC were cultured in the presence of SWA and then stimulated with PMA and ionomycin for 4 h. The PBMC were then stained for intracellular IFN-γ and analyzed using flow cytometry (Fig. 3). Based on forward and side scatter, IFN-γ-positive cells were lymphocytes (data not shown). Among the lymphocytes observed, almost all of the IFN-γ-positive cells were also positive for CD3 and negative for γδ TCR (Fig. 3a). Thus, we suggest that the conventional T cells expressing αβ TCR are the most likely major source of IFN-γ in the immunized miniature pigs. We also examined the natural killer (NK) cells, which are strong producers of IFN-γ [24], using CD16 as a marker of NK cells among other lymphocytes. CD16 is a low affinity receptor for IgG and is expressed on monocytes and a population of NK cells [25]. Additional specific markers of porcine NK cells have yet to be clearly established [18].

Bottom Line: To characterize the protective immune response of the pig model vaccinated by attenuated cercaria, flow cytometric analysis of the reactive T cell subsets was performed.CD4+ αβ-T cells as well as CD4+/CD8α(mid) double positive and/or CD8α(high) αβ-T cells were the major IFN-γ-producing CD3+ T cells.On the contrary, γδ T cells did not produce intracellular IFN-γ.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunogenetics, Institute of Tropical Medicine (NEKKEN), Nagasaki University , 1-14-2 Sakamoto, Nagasaki 852-8523, Japan ; Department of Parasitology, Faculty of Medicine, Minia University , Minia, 61519, Egypt.

ABSTRACT
CLAWN miniature pig has been shown to serve as a suitable host for the experimental infection of Schistosoma japonicum. In this study, we found that radiation-attenuated cercaria (RAC) vaccine gave CLAWN miniature pigs protective immunity against subsequent challenge infection with S. japonicum cercaria. To characterize the protective immune response of the pig model vaccinated by attenuated cercaria, flow cytometric analysis of the reactive T cell subsets was performed. The intracellular interferon (IFN)-γ and the cell surface markers revealed the peripheral blood CD3+ T-lymphocytes produced significant amounts of IFN-γ during the immunization period and after the challenge infection. CD4+ αβ-T cells as well as CD4+/CD8α(mid) double positive and/or CD8α(high) αβ-T cells were the major IFN-γ-producing CD3+ T cells. On the contrary, γδ T cells did not produce intracellular IFN-γ. Our results suggested that RAC-vaccinated miniature pigs showed effective protective immunity through the activation of αβ T cells bearing antigen specific T-cell receptors but not through the activation of γδ T cells.

No MeSH data available.


Related in: MedlinePlus