Perturbation of hepatocyte growth regulation is associated with a number of liver diseases such as fibrosis and cancer. These diseases are mediated by a network of growth factors and cytokines that regulate the induction of hepatocyte proliferation and apoptosis. In this study, we have investigated the role of signaling pathways activated by tumor necrosis factor alpha (TNF-alpha) and epidermal growth factor (EGF) in the regulation of apoptosis induced by transforming growth factor beta(1) (TGF-beta(1)), because this physiological factor is believed to regulate spontaneous apoptosis in the liver. We show that pretreatment with (10 ng/mL) EGF or (25 ng/mL) TNF-alpha can suppress TGF-beta(1)-induced apoptosis by 73% and 50%, respectively, in isolated rat hepatocytes. However, suppression of TGF-beta(1)-induced apoptosis by EGF and TNF-alpha occurs via different protein kinase signaling pathways. Using specific inhibitors, we show that suppression of apoptosis by EGF is dependent on activation of phosphoinositide 3-kinase (PI 3-kinase) and the extracellular signal regulated kinase (ERK) mitogen-activated protein (MAP) kinase pathways, but not p38 MAP kinase. In contrast, suppression of TGF-beta(1)-induced apoptosis by TNF-alpha does not require PI 3-kinase and protein kinase B (PKB or Akt)-mediated pathways, but is dependent on ERK and p38 MAP kinase activity. These data contribute to our understanding of the intracellular survival signals that play a role in normal liver homeostasis and in diverse pathological conditions.