TNF-ALPHA BIFUNCTIONALLY INDUCES PROLIFERATION IN PRIMARY HEPATOCYTES- ROLE OF CELL ANCHORAGE AND SPREADING

In the absence of a growth factor or an appropriate extracellular matr ix (ECM), cells are arrested in the G(0)/G(1) phase, In this report, w e demonstrate the evidence that TNF-alpha induced DNA synthesis of pri mary mouse hepatocytes in vitro by activating two distinct pathways, T NF-alpha induced drastic spreading of hepatocytes on hydrophobic plast ic, while the adhesion was not influenced, The effect was time and dos e dependent, The cell spreading was accompanied by the phosphorylation of paxillin, indicating the stimulation of focal adhesion molecules, TNF-alpha-induced spreading of hepatocytes was not transient, and kine tic analysis and morphologic observation suggest that the effect was d ifferent from epidermal growth factor- or hepatocyte growth factor-ind uced transient hepatocyte spreading, TNF-alpha-induced hepatocyte spre ading was blocked by cytochalasin D, Arg-Gly-Asp peptides, cycloheximi de, or anti-integrin beta(1) Ab, Results of competitive PCR for ECM pr oteins demonstrated that TNF-alpha increased the expression of laminin alpha(3) and gamma(1) chains in hepatocytes, These data suggested tha t TNF-alpha induced cell anchorage for hepatocytes by up-regulating EC M production. More importantly, TNF-alpha, but neither epidermal growt h factor nor hepatocyte growth factor, induced DNA synthesis following the spreading in primary hepatocytes on hydrophobic plastic, while me re cell spreading on collagen did not induce DNA synthesis, The DNA sy nthesis was blocked by the inhibition of either cell spreading or DNA polymerase, demonstrating that TNF-alpha induced DNA synthesis in prim ary hepatocytes by activating two distinct pathways, i.e., forming the scaffold and inducing growth signals, Taken together, TNF-alpha bifun ctionally regulates the proliferation of primary hepatocytes, serving as both an ECM inducer and a growth factor.