Notch signaling participates in liver fibrosis

Background and Aims: Humans have five Notch ligands (Jagged1, Jagged2, Delta like ligand (Dll)1, Dll3 and Dll4) and four receptors (Notch1, 2, 3 and 4). Mutations of Jagged1 or Notch2 lead to Alagille syndrome, a disease characterized by defective development of intralobular bile ducts. Alternatively, Notch signaling drives hepatic progenitor cells differentiating into cholangiocytes during acute and chronic hepatic regeneration. It is not known to date whether Notch family members a play role in liver fibrosis. Here we investigated effects of Notch ligands in HBV-infected patients and experimental models. Methods: Genechip analysis was used to investigate Notch associated gene expression in 7 healthy controls and 121 HBV infected patients with different fibrotic stages. Immunohistochemistry was performed to detect expression of Notch ligands/receptors in liver specimens from HBV patients. We examined effects of Notch ligands in primary rat hepatic stellate cells (HSC), CFSCs and primary hepatocytes. Finally, we investigated whether recombinant Notch ligands, rJagged1 and rDll4, influence liver fibrosis in mice treated with carbon tetrachloride (CCl4). Results: Microarray analyses identified nine Notch signaling associated genes, e.g. Jagged1, that were associated with fibrotic stages in HBV patients. Immunohistochemistry co-staining and confocal microscopy analyses revealed that Jagged1, Dll4 and Notch1 are expressed in activated HSC and myofibroblasts, the main collagen producing liver cells, in HBV patients. Dll4 immune score correlated with inflammatory grades and fibrotic stages in these patients. Recombinant Jagged-1 and Dll4 protein injection remarkably reduced serum ALT levels, hepatocyte apoptosis, inflammation and fibrosis in mice after four weeks of CCl4 treatment. In vitro, rJagged1 and rDll4 proteins significantly decreased TGF-β dependent mRNA expression of collagen I in HSCs. Consistent with animal model, rJagged1 and rDll4 incubation inhibited TNF-α and TGF-β induced hepatocyte apoptosis in vitro. Conclusions: Notch family proteins participate in liver fibrosis. rDll4 and rJagged-1 influence experimental liver fibrosis via interfering TGF-β effects in HSCs, inhibiting inflammatory cells and preventing hepatocyte apoptosis.