LONG-TERM PRIMARY CULTURE OF EPITHELIAL-CELLS FROM RAINBOW-TROUT (ONCORHYNCHUS-MYKISS) LIVER

Long-term primary cultures of epithelial cells from rainbow trout (Onc orhynchus mykiss) liver have been established. Nearly homogenous (>97% ) populations of hepatocytes were placed into primary culture and rema ined viable and proliferative for at least 70 d. In addition to hepato cytes, proliferative biliary cells persisted in the cultures for at le ast 30 d. Finally, a third type of epithelial cell, which we have term ed a ''spindle cell,'' consistently appeared and proliferated to confl uence in these cultures. The confluent cultures of spindle cells were successfully subcultured and passaged. The initial behavior, growth, a nd optimization of serum and media requirements for these cells is des cribed. All three cell types proliferated as measured by thymidine inc orporation, autoradiography, proliferating cellular nuclear antigen an alysis, and propidium iodine staining. Further efforts to characterize the cells included western blotting and immunohistochemical staining with antibodies to cytokeratins previously reported in fish liver. Fro m these data, it appears that all three cell populations are epithelia l in nature. Furthermore, significant changes in actin organization, o ften indicative of transformation or pluripotent cells, were observed with increased time in primary culture.