CYCLOSPORINE-A INDUCES APOPTOSIS IN RAT HEPATOCYTES IN CULTURE

Cyclosporin 4 (CsA) at concentrations up to 1 mu M induced apoptosis i n a dose-dependent manner in cultured rat hepatocytes for 48 h in the presence of insulin and epidermal growth factor (EGF). The effect of C sA was evidenced by the DNA fragmentation pattern constituted by fragm ents of multiples of 180-200 base pairs, which is a characteristic of programmed cell death. The metabolic activity did not change significa ntly in the presence of 0.1 mu M CsA and diminished to 49% of control in the presence of 1 mu M CsA. Changes in the metabolic activity were correlated with a decrease in both [methyl-H-3]thymidine uptake and DN A content, which reflects a decrease in the cell number. The treatment of cells with CsA (1 mu M) decreased the metabolic activity/DNA conte nt ratio by 24% with respect to dimethyl sulphoxide (DMSO) control, wh ich also suggests, under these conditions, that the necrosis achieved is at most only 24%. In addition, the changes observed (apoptotic proc ess, arrest of the cell cycle and apparition of a necrotic process) we re correlated with an increase in the high-affinity guanosine triphosp hatase (GTPase) enzymes.