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 matrix (ECM), cells are arrested in the G0/G1 phase. In this report, we demonstrate the evidence that TNF-alpha induced DNA synthesis of primary mouse hepatocytes in vitro by activating two distinct pathways. TNF-alpha induced drastic spreading of hepatocytes on hydrophobic plastic, while the adhesion was not influenced. The effect was time and dose 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 kinetic analysis and morphologic observation suggest that the effect was different from epidermal growth factor- or hepatocyte growth factor-induced transient hepatocyte spreading. TNF-alpha-induced hepatocyte spreading was blocked by cytochalasin D, Arg-Gly-Asp peptides, cycloheximide, or anti-integrin beta1 Ab. Results of competitive PCR for ECM proteins demonstrated that TNF-alpha increased the expression of laminin alpha3 and gamma1 chains in hepatocytes. These data suggested that TNF-alpha induced cell anchorage for hepatocytes by up-regulating ECM production. More importantly, TNF-alpha, but neither epidermal growth factor nor hepatocyte growth factor, induced DNA synthesis following the spreading in primary hepatocytes on hydrophobic plastic, while mere cell spreading on collagen did not induce DNA synthesis. The DNA synthesis was blocked by the inhibition of either cell spreading or DNA polymerase, demonstrating that TNF-alpha induced DNA synthesis in primary hepatocytes by activating two distinct pathways, i.e., forming the scaffold and inducing growth signals. Taken together, TNF-alpha bifunctionally regulates the proliferation of primary hepatocytes, serving as both an ECM inducer and a growth factor.
Watanabe
Y
Y
Department of Biomolecular Engineering, Tokyo Institute of Technology, Yokohama, Japan. ywatanab@bio.titech.ac.jp
Osaki
H
H
Akaike
T
T
D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D004261:DNA Replication / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D006133:Growth Substances / Q000494:pharmacology*; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D007797:Laminin / Q000235:genetics / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000187:drug effects*; D051379:Mice; D008813:Mice, Inbred ICR; D011137:Polystyrenes; D012333:RNA, Messenger / Q000096:biosynthesis; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects
D006133:Growth Substances; D007797:Laminin; D011137:Polystyrenes; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D004815:Epidermal Growth Factor; D017228:Hepatocyte Growth Factor