Establishment and characterization of porcine cytolytic cell lines and clones

M.C.M. de Bruin, E.M.A. van Rooij, J.L.M. Voermans, Y.E. de Visser, A.T.J. Bianchi, T.G. Kimman

    Research output: Contribution to journalArticleAcademicpeer-review

    23 Citations (Scopus)

    Abstract

    Although non-major-histocompatibility-complex-restricted cytolytic cells appear to significantly influence antiviral immunity in pigs, the phenotype and functional characteristics of these cells are not well defined. To allow a detailed analysis of these subsets, we established and characterized cell lines and clones of interleukin-2-activated (IL-2) cytolytic cells. Cell lines and clones were obtained from peripheral blood mononuclear cells of minipigs of the swine-leucocyte-antigen-complex (SLA) d/d haplotype. Cells were cultured in the presence of human recombinant IL-2 and cloned by double limiting dilution in the presence of gamma-irradiated L14 cells (a retrovirus immortalized B-lymphoblastoid cell line of the haplotype SLA(d/d)) or gamma- irradiated autologous peripheral blood mononuclear cells as feeder cells. Cytolytic cell lines and clones were characterized for their ability to kill different target cells and for their cell surface phenotype. All obtained clones expressed CD2 and CD8 and were negative for CD4. The following three subsets of cytolytic cells were identified: Subset 1) CD3- CD5- cells that killed K562 cells (a natural killer cell susceptible target cell line), as well as the pseudorabies virus (PRV)-infected or uninfected porcine kidney cells. These cells were considered to be typical natural killer cells Subset 2) CD3γ/δ+ CD5- T-cells that killed K562 cells and PRV virus-infected or uninfected porcine kidney cells, infected or uninfected L14 cells, and L14 cells constitutively expressing the PRV viral glycoprotein gB or gC. These cells were considered to be γ/δ T-cells with natural killer activity. Subset 3) CD3α/β+ CD5+ T-cells that killed L14 cells, PRV-infected L14 cells, and PRV gB- and gC-transfected L14 cells. These cells were possibly induced by the L14 feeder cells, used in the in vitro culture system. None of the cytolytic effector cells killed only MHC-matched vital infected cells. In conclusion, we describe a method to isolate, clone, and culture cytolytic cells from pigs. The clones could be cultured for 5 months, which allowed appropriate phenotypic and functional characterization of the various clones. Two of the subsets, CD3γ/δ T- and the natural killer cell subset may be involved in antiviral immunity in this species.
    Original languageEnglish
    Pages (from-to)337-347
    JournalVeterinary Immunology and Immunopathology
    Volume59
    Issue number3-4
    DOIs
    Publication statusPublished - 1997

    Fingerprint

    Swine
    Clone Cells
    cell lines
    clones
    Cell Line
    swine
    cells
    Suid Herpesvirus 1
    Suid herpesvirus 1
    natural killer cells
    Feeder Cells
    Natural Killer T-Cells
    K562 Cells
    T-lymphocytes
    Natural Killer Cells
    Haplotypes
    kidney cells
    mononuclear leukocytes
    Interleukin-2
    Antiviral Agents

    Cite this

    de Bruin, M. C. M., van Rooij, E. M. A., Voermans, J. L. M., de Visser, Y. E., Bianchi, A. T. J., & Kimman, T. G. (1997). Establishment and characterization of porcine cytolytic cell lines and clones. Veterinary Immunology and Immunopathology, 59(3-4), 337-347. https://doi.org/10.1016/S0165-2427(97)00085-8
    de Bruin, M.C.M. ; van Rooij, E.M.A. ; Voermans, J.L.M. ; de Visser, Y.E. ; Bianchi, A.T.J. ; Kimman, T.G. / Establishment and characterization of porcine cytolytic cell lines and clones. In: Veterinary Immunology and Immunopathology. 1997 ; Vol. 59, No. 3-4. pp. 337-347.
    @article{dbee1005299d49f58306e9dda6642b37,
    title = "Establishment and characterization of porcine cytolytic cell lines and clones",
    abstract = "Although non-major-histocompatibility-complex-restricted cytolytic cells appear to significantly influence antiviral immunity in pigs, the phenotype and functional characteristics of these cells are not well defined. To allow a detailed analysis of these subsets, we established and characterized cell lines and clones of interleukin-2-activated (IL-2) cytolytic cells. Cell lines and clones were obtained from peripheral blood mononuclear cells of minipigs of the swine-leucocyte-antigen-complex (SLA) d/d haplotype. Cells were cultured in the presence of human recombinant IL-2 and cloned by double limiting dilution in the presence of gamma-irradiated L14 cells (a retrovirus immortalized B-lymphoblastoid cell line of the haplotype SLA(d/d)) or gamma- irradiated autologous peripheral blood mononuclear cells as feeder cells. Cytolytic cell lines and clones were characterized for their ability to kill different target cells and for their cell surface phenotype. All obtained clones expressed CD2 and CD8 and were negative for CD4. The following three subsets of cytolytic cells were identified: Subset 1) CD3- CD5- cells that killed K562 cells (a natural killer cell susceptible target cell line), as well as the pseudorabies virus (PRV)-infected or uninfected porcine kidney cells. These cells were considered to be typical natural killer cells Subset 2) CD3γ/δ+ CD5- T-cells that killed K562 cells and PRV virus-infected or uninfected porcine kidney cells, infected or uninfected L14 cells, and L14 cells constitutively expressing the PRV viral glycoprotein gB or gC. These cells were considered to be γ/δ T-cells with natural killer activity. Subset 3) CD3α/β+ CD5+ T-cells that killed L14 cells, PRV-infected L14 cells, and PRV gB- and gC-transfected L14 cells. These cells were possibly induced by the L14 feeder cells, used in the in vitro culture system. None of the cytolytic effector cells killed only MHC-matched vital infected cells. In conclusion, we describe a method to isolate, clone, and culture cytolytic cells from pigs. The clones could be cultured for 5 months, which allowed appropriate phenotypic and functional characterization of the various clones. Two of the subsets, CD3γ/δ T- and the natural killer cell subset may be involved in antiviral immunity in this species.",
    author = "{de Bruin}, M.C.M. and {van Rooij}, E.M.A. and J.L.M. Voermans and {de Visser}, Y.E. and A.T.J. Bianchi and T.G. Kimman",
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    de Bruin, MCM, van Rooij, EMA, Voermans, JLM, de Visser, YE, Bianchi, ATJ & Kimman, TG 1997, 'Establishment and characterization of porcine cytolytic cell lines and clones' Veterinary Immunology and Immunopathology, vol. 59, no. 3-4, pp. 337-347. https://doi.org/10.1016/S0165-2427(97)00085-8

    Establishment and characterization of porcine cytolytic cell lines and clones. / de Bruin, M.C.M.; van Rooij, E.M.A.; Voermans, J.L.M.; de Visser, Y.E.; Bianchi, A.T.J.; Kimman, T.G.

    In: Veterinary Immunology and Immunopathology, Vol. 59, No. 3-4, 1997, p. 337-347.

    Research output: Contribution to journalArticleAcademicpeer-review

    TY - JOUR

    T1 - Establishment and characterization of porcine cytolytic cell lines and clones

    AU - de Bruin, M.C.M.

    AU - van Rooij, E.M.A.

    AU - Voermans, J.L.M.

    AU - de Visser, Y.E.

    AU - Bianchi, A.T.J.

    AU - Kimman, T.G.

    PY - 1997

    Y1 - 1997

    N2 - Although non-major-histocompatibility-complex-restricted cytolytic cells appear to significantly influence antiviral immunity in pigs, the phenotype and functional characteristics of these cells are not well defined. To allow a detailed analysis of these subsets, we established and characterized cell lines and clones of interleukin-2-activated (IL-2) cytolytic cells. Cell lines and clones were obtained from peripheral blood mononuclear cells of minipigs of the swine-leucocyte-antigen-complex (SLA) d/d haplotype. Cells were cultured in the presence of human recombinant IL-2 and cloned by double limiting dilution in the presence of gamma-irradiated L14 cells (a retrovirus immortalized B-lymphoblastoid cell line of the haplotype SLA(d/d)) or gamma- irradiated autologous peripheral blood mononuclear cells as feeder cells. Cytolytic cell lines and clones were characterized for their ability to kill different target cells and for their cell surface phenotype. All obtained clones expressed CD2 and CD8 and were negative for CD4. The following three subsets of cytolytic cells were identified: Subset 1) CD3- CD5- cells that killed K562 cells (a natural killer cell susceptible target cell line), as well as the pseudorabies virus (PRV)-infected or uninfected porcine kidney cells. These cells were considered to be typical natural killer cells Subset 2) CD3γ/δ+ CD5- T-cells that killed K562 cells and PRV virus-infected or uninfected porcine kidney cells, infected or uninfected L14 cells, and L14 cells constitutively expressing the PRV viral glycoprotein gB or gC. These cells were considered to be γ/δ T-cells with natural killer activity. Subset 3) CD3α/β+ CD5+ T-cells that killed L14 cells, PRV-infected L14 cells, and PRV gB- and gC-transfected L14 cells. These cells were possibly induced by the L14 feeder cells, used in the in vitro culture system. None of the cytolytic effector cells killed only MHC-matched vital infected cells. In conclusion, we describe a method to isolate, clone, and culture cytolytic cells from pigs. The clones could be cultured for 5 months, which allowed appropriate phenotypic and functional characterization of the various clones. Two of the subsets, CD3γ/δ T- and the natural killer cell subset may be involved in antiviral immunity in this species.

    AB - Although non-major-histocompatibility-complex-restricted cytolytic cells appear to significantly influence antiviral immunity in pigs, the phenotype and functional characteristics of these cells are not well defined. To allow a detailed analysis of these subsets, we established and characterized cell lines and clones of interleukin-2-activated (IL-2) cytolytic cells. Cell lines and clones were obtained from peripheral blood mononuclear cells of minipigs of the swine-leucocyte-antigen-complex (SLA) d/d haplotype. Cells were cultured in the presence of human recombinant IL-2 and cloned by double limiting dilution in the presence of gamma-irradiated L14 cells (a retrovirus immortalized B-lymphoblastoid cell line of the haplotype SLA(d/d)) or gamma- irradiated autologous peripheral blood mononuclear cells as feeder cells. Cytolytic cell lines and clones were characterized for their ability to kill different target cells and for their cell surface phenotype. All obtained clones expressed CD2 and CD8 and were negative for CD4. The following three subsets of cytolytic cells were identified: Subset 1) CD3- CD5- cells that killed K562 cells (a natural killer cell susceptible target cell line), as well as the pseudorabies virus (PRV)-infected or uninfected porcine kidney cells. These cells were considered to be typical natural killer cells Subset 2) CD3γ/δ+ CD5- T-cells that killed K562 cells and PRV virus-infected or uninfected porcine kidney cells, infected or uninfected L14 cells, and L14 cells constitutively expressing the PRV viral glycoprotein gB or gC. These cells were considered to be γ/δ T-cells with natural killer activity. Subset 3) CD3α/β+ CD5+ T-cells that killed L14 cells, PRV-infected L14 cells, and PRV gB- and gC-transfected L14 cells. These cells were possibly induced by the L14 feeder cells, used in the in vitro culture system. None of the cytolytic effector cells killed only MHC-matched vital infected cells. In conclusion, we describe a method to isolate, clone, and culture cytolytic cells from pigs. The clones could be cultured for 5 months, which allowed appropriate phenotypic and functional characterization of the various clones. Two of the subsets, CD3γ/δ T- and the natural killer cell subset may be involved in antiviral immunity in this species.

    U2 - 10.1016/S0165-2427(97)00085-8

    DO - 10.1016/S0165-2427(97)00085-8

    M3 - Article

    VL - 59

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    EP - 347

    JO - Veterinary Immunology and Immunopathology

    JF - Veterinary Immunology and Immunopathology

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    ER -