Clonal growth of carp (Cyprinus carpio) T cells in vitro: Long-term proliferation of Th2-like cells

Carp kidney leukocytes co-cultured with a supporting cell layer resulted in proliferation of polyclonal CD4+ αβT cells as described previously. These bulk-cultured T cells expressed transcripts for both T helper 1 cells (Th1) master regulator (T-bet) and T helper 2 cells (Th2) master regulator (GATA-3). To identify the Th subsets in bulk-cultured T cells, single cells were picked up from the bulk culture, proliferated, and characterized. The majority of the clones displayed characteristics consistent with CD4+ αβT cell identity. These clones expressed both TCRα and TCRβ, but could not produce a TCRγδ heterodimer since they typically only expressed either TCRγ or TCRδ. These clones also expressed the TCR co-receptor genes CD4-1 or CD4-2, whereas they did not express CD8α or CD8β. In addition, GATA-3 was expressed whereas T-bet was not. Among these clones, one clone (KoThL5) continued to proliferate on the supporting cells and was successively transferred for more than 10 months and 90–100 passages. To characterize the KoThL5 cells by their cytokine production profile, they were stimulated with PHA and investigated by real-time RT-PCR. mRNA expression of Th2-related cytokine (IL-4/13B) was only enhanced in KoThL5 cells whereas both Th1-related cytokine (IFNγ) and Th2-related cytokines (IL-4/13A andIL-4/13B) were significantly enhanced in bulk-cultured T cells. Taken together, KoThL5 cells share some features with mammalian Th2 cells. s the first study to describe in vitro cultures of teleost cell with Th2-like features. The KoThL5 cell line has considerable potential for addressing questions concerning the properties of teleost Th2 cells.