The Different Calcium Requirements of the Mitogenic Effects Elicited in Primary Neonatal Rat Hepatocytes by the Diterpene Phorbol Esters 12-O-Tetradecanoylphorbol-13-Acetate and Sapintoxin A

Abstract

A single exposure to a wide range of concentrations (10−13−10−5 mol/L) of the powerful, complete tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulated 4-day-old primary neonatal rat hepatocytes to synthesize new DNA and to divide within 24 hours independently on the actual (either high [1.8 mmol/L] or low [0.01 mmol/L]) concentration of calcium (Ca2+) in the surrounding HiWoBa2000 synthetic growth medium. Conversely, the exposure to the same range of doses of sapintoxin A (SAP A; a fluorescent phorbol ester activating the Ca2+/phospholipid-dependent protein kinase [PK-C]) without acting by itself either as a complete or as a second-stage tumour promoter) enhanced the proliferative activity of primary hepatocytes only on condition that a high level of Ca2+ was present in the growth medium. On the other hand, studies on the kinetics of the inhibition of new DNA synthesis showed that the induction of the G0/G1 and G1/S transitions by TPA in hepatocytes incubated in the Ca2+-devoid HiWoBa20000 medium still required cellular Ca2+-, CaM-, and PK-C-dependent metabolic events. Hence, the diverse conditioning of the mitogenic activity of tumour promoting and non-promoting phorbol esters by the level of extracellular Ca2+ suggests as a quite likely event the involvement in the process of differently Ca2+-dependent isoenzymes of their receptor PK-C.