EFFECTS OF EXTRACELLULAR CA2+ AND HCO3- ON EPIDERMAL GROWTH FACTOR-INDUCED DNA-SYNTHESIS IN CULTURED RAT HEPATOCYTES

Background/Aims: The elevation of cytosolic free calcium concentration ([Ca2+](i)) and intracellular pH mediate the growth factor-initiated proliferation of many cells, but it is not known if they trigger mitosis in resting hepatocytes. The maintenance of [Ca2+](i) and intracellular pH depends partly on extracellular calcium concentration ([Ca2+](e)) and extracellular bicarbonate concentration ([HCO3-](e)). Therefore, the effects of [Ca2+](e) and [HCO3-](e) on hepatocyte proliferation were examined. Methods: Epidermal growth factor induced proliferation in primary cultures of rat hepatocytes. [H-3]thymidine incorporation into DNA and nuclear labeling indices were measured. Results: Between 0.2 and 0.9 mmol/L of [Ca2+](e), the proliferative response to epidermal growth factor increased, and total hepatocellular Ca2+ content was increased. Increasing [HCO3-](e) also stimulated DNA synthesis in a concentration-dependent manner, maximal at 35 mmol/L, Using optimal [Ca2+](e) (0.9 mmol/L) and [HCO3-](e) (35 mmol/L), a synergistic stimulation of hepatocellular DNA synthesis was shown. Voltage-dependent Ca2+ channel blockers failed to inhibit hepatocyte proliferation when administered in concentrations that inhibit proliferation in other cell types. Conclusions: [Ca2+](e) and [HCO3-](e) are both essential for hepatocyte proliferation, and their effects are synergistic. The entry of extracellular Ca2+ is criticai for epidermal growth factor-induced DNA synthesis in hepatocytes, but this is not mediated by voltage-dependent Ca2+ channels.