TGF-β downstream Smad3 and MAPK signaling are mutually antagonistic in liver progenitor cells

Abstract

Introduction: Liver progenitor cells (LPCs) are a subpopulation of cells which are always found in chronic injuried liver, and are contribute to liver fibrosis and liver cancer initiation under the circumstance of chronic inflammation. However, the precise role of LPCs in responding to chronic inflammatory signals are remain elusive. Method: We observed the effects of TGF-β, one of the most commonly detected cytokine in chronic hepatitis, in LPCs, by using CCK-8, F-actin staining, western blot, and transwell analyses in liver progenitoc cell lines (WB-F344 and LE/6). Western blot and luciferase reporter analyses were utilized to measure the activation of Smad and MAPK signaling in LPCs. We then overexpressed the expression of Smad3 in LPCs by adenovirus carrying wild type Smad3, mutated Smad3 at specific serine phosphorylation site respectively, and observed their downstream effects. Result: In LPCs, TGF-β showed limited cytostatic and epithelial-mesenchymal transition (EMT) effects, and TGF-β activated both Smad and MAPK (Erk, JNK and p38 MAPK) signaling. TGFβ-Smad signaling was the inducer of cytostasis and EMT, whereas TGFβ-MAPK signaling showed opposite effects. Moreover, the activity of TGF-β downstream Smad and MAPK signaling were mutually restricted. Mechansitically, TGF-β activated Smad signaling through serine phosphorylation of both C-terminal and linker region of Smad2 and Smad3 in LPCs, and TGFβ-MAPK signaling inhibited the phosphorylation of Smad3 at C-terminal, but it reinforced the phosphorylation of Smad3 at the linker region. Phosphorylation of Smad3 at C-terminal contributed to TGF-β induced cytostasis and EMT effects, whereas the phosphorylation of Smad3 at the linker region inhibited these effects in LPCs. Conclusion: TGF-β downstream Smad and MAPK signaling were mutually antagonistic in LPCs through phosphorylation of Smad3 at different serine sites, and this antagonism may help LPCs to survive in the circumstance of chronic inflammation. Additionally, imbalanced antagonisitc Smad3 and MAPK signaling may contribute to the malignant transformation of LPCs.