Details

Autor(en) / Beteiligte

• Stuge, Tor Brynjar

Titel

Functional and phenotypic characterization of channel catfish (Ictalurus punctatus) cytotoxic cells generated in mixed leukocyte cultures

Ort / Verlag

ProQuest Dissertations & Theses

Erscheinungsjahr

1998

Quelle

ProQuest Dissertations & Theses A&I

Beschreibungen/Notizen

• Cytotoxic studies employing catfish lymphoid cell lines as allogeneic targets have demonstrated that a subpopulation of catfish nonimmune PBLs nonspecifically kill cells expressing nonself alloantigens. In vitro mixed leukocyte cultures of catfish PBLs stimulated with various irradiated lymphoid cell lines exhibit vigorous cellular proliferation and greatly enhanced nonspecific cytotoxicity ($\sim$100 fold) towards allogeneic but not autologous cell line targets. The heterogeneous nature of these MLC generated cells and the lack of appropriate surface markers precluded definitive assessment of the cell type(s) responsible for the observed cytotoxic responses. In attempts to identify catfish cytotoxic effector cells, freshly isolated and MLC stimulated PBLs from both immune and nonimmune catfish were cloned by limiting dilution in the presence of irradiated allogeneic stimulators and culture supernatant from a catfish T cell line (as a presumed source of growth promoting factors). A number of clonal cell lines were successfully generated using this strategy. Cytotoxic and RT-PCR characterizations of these clonal cells identified at least three different types of channel catfish cytotoxic effector cells: (1) TCR $\alpha/\beta\sp-$ nonspecific cytotoxic (NK-like) cells; (2) TCR $\alpha/\beta\sp-$ cytotoxic cells demonstrating significant allospecificity; and (3) TCR $\alpha/\beta\sp+$ and IgM$\sp-$ allospecific cytotoxic T cells (CTL). The use of alloantisera specifically blocked target lysis by catfish TCR $\alpha/\beta\sp+$ CTL clones, but not NK-like clones, indicating that target recognition by catfish CTL is dependent upon the presence of alloantigen. These results, coupled with the ability to generate nonspecific cytotoxic activity in autologous MLC, indicate that activation and target recognition of catfish NK-like cells may involve non-MHC-like molecules. Further characterization of the catfish NK-like clones revealed that one of these clones may express a putative Fc receptor due to the presence of polyclonal IgM on their surface. Finally, a monoclonal antibody developed against catfish NK-like cells was found to react specifically with the catfish NK-like clones and also with a subpopulation of catfish PBLs (${\sim}$10-20%) which exhibits nonspecific cytotoxicity toward allogeneic targets. The development of cytotoxic cell lines and specific monoclonal antibodies will likely serve as unique tools for understanding both the function and evolution of cytotoxic cells.