ABSTRACT
<p><b>OBJECTIVE</b>To investigate whether Notch signaling is activated in hepatic stellate cells (HSCs), and to determine whether manipulation of the Notch signaling pathway can effect the activation of HSCs.</p><p><b>METHODS</b>The expression of Notch signaling components in unactivated or TGF-b1-activated HSC-T6 cells was detected by Taqman Probe-based gene expression analysis. Differential expression of Notch3 and Jagged1 was detected by immunofluorescence analysis. Notch3-mediated expression of the myofibroblastic markers, a-SMA and collagen I, was detected in HSC-T6 cells transfected with pcDNA3.1-N3ICD or Notch3 siRNA by Western blotting.</p><p><b>RESULTS</b>Notch signaling components were expressed in both unactivated and activated HSC-T6 cells, but the TGF-b1-treated cells showed significantly higher expression levels of Jagged1 (3.9-fold, F = 2543.482), Notch3 (4.2-fold, F = 287.982), and HES1 (3.2-fold, F = 1719.851). Transfection-mediated over-expression of Notch3 led to significantly increased expression of a-SMA (6.8-fold, t = 13.157) and collagen I (5.5-fold, t = 9.810) (both P less than 0.01). Transient knock-down of Notch3 expression by siRNA decreased expression of the myofibroblastic markers (a-SMA by approximately 90%, t = 19.863 and collagen I by 84%, t = 10.376; both, P less than 0.01). Moreover, knock-down of Notch3 antagonized the TGF-b1-induced expression of a-SMA and collagen I.</p><p><b>CONCLUSION</b>Notch signaling may participate in liver fibrogenesis by regulating HSC activation. Selective interruption of Notch3 may represent a new anti-fibrotic strategy to treat liver fibrosis.</p>
Subject(s)
Animals , Rats , Calcium-Binding Proteins , Genetics , Metabolism , Cell Line , Hepatic Stellate Cells , Metabolism , Intercellular Signaling Peptides and Proteins , Genetics , Metabolism , Jagged-1 Protein , Membrane Proteins , Genetics , Metabolism , RNA, Messenger , Genetics , RNA, Small Interfering , Receptor, Notch3 , Receptors, Notch , Genetics , Metabolism , Serrate-Jagged Proteins , Signal TransductionABSTRACT
<p><b>OBJECTIVE</b>To establish a mouse model for human chronic HBV infection, and to investigate the role of PD-1/PD-L1 signaling pathway in antiviral immunity.</p><p><b>METHODS</b>A mouse model was established by hydrodynamic injection of the plasmid pAAV/HBV1.2-GFP into the tail vein of C57BL/6 mice, HBV markers were assayed at different time points after injection. After intraperitoneal injection of anti-PD-L1 monoclonal antibody, the serum ALT, and HBV DNA in the serum, liver and kidney were assayed.</p><p><b>RESULTS</b>The chronic HBV infection mouse model were established successfully, serum HBsAg and high load of HBV DNA were detectable 90 days after plasmid injection. After blocking of the PD-1/PD-L1 pathway, the serum ALT level of mice were significantly increased (P < 0.01), and the HBV DNA load in serum (P < 0.01), liver (P < 0.05) and kidney (P < 0.05) were decreased significantly.</p><p><b>CONCLUSION</b>Blocking the PD-1/PD-L1 signaling pathway can enhance antiviral response in mice with chronic HBV infection.</p>
Subject(s)
Animals , Male , Mice , Antigens, Surface , Metabolism , Apoptosis Regulatory Proteins , Metabolism , B7-1 Antigen , Metabolism , B7-H1 Antigen , DNA, Viral , Disease Models, Animal , Hepatitis B virus , Metabolism , Hepatitis B, Chronic , Allergy and Immunology , Metabolism , Virology , Membrane Glycoproteins , Metabolism , Mice, Inbred C57BL , Peptides , Metabolism , Programmed Cell Death 1 Receptor , Signal TransductionABSTRACT
Objective To investigate whether functional Wnt-?-catenin signaling is present in activated hepatic stellate cells (HSC),and the effect of blocking this signaling on activation of HSC. Methods?-catenin expression in HSC was examined by immunocytochemistry.Wnt signalings in HSC-T6 were assessed using a T cell factor (TCF)-dependent luciferase reporter gene (pTOP- FLASH) assay.Wnt signalings in HSC-T6 were blocked by transfecting with a dominant negative TCF (dnTCF) expression plasmid,then the expression of alpha smooth muscle actin (?-SMA) and collagen typeⅠwere examined by Western blot.Results?-catenin staining was positive in the nuclei of HSC-T6.Luciferase activity in the cells transfected with pTOPFLASH was significantly higher than that in the cells transfected with pFOPFLASH (P