Journal: International Journal of Molecular Sciences

Article Title: Pirfenidone Suppresses Liver Fibrosis Through Inhibition of TGF-β-Associated Lipid Metabolic Remodeling in Hepatic Stellate Cells

doi: 10.3390/ijms27094061

Figure Lengend Snippet: TGF-β signaling enhances intracellular lipid accumulation in HSCs, which is suppressed by pirfenidone. ( A ) BODIPY 493/503 staining images of LX2 and HSC-T6 cells after serum starvation followed by TGF-β1 or TGF-β2 (10 ng/mL) treatment for 48 h. ( B ) Quantification of mean BODIPY fluorescence intensity in LX2 and HSC-T6 cells. ( C ) Oil Red O staining images of LX2 and HSC-T6 cells treated with TGF-β1 or TGF-β2 (10 ng/mL, 48 h) in the presence or absence of PFD (250 μg/mL) after serum starvation. The red square indicates the region shown at higher magnification in the upper left corner of the same subfigure. ( D ) Quantification of Oil Red O-stained lipid droplets per cell in LX2 and HSC-T6 cells. Quantification was performed from randomly selected fields under blinded conditions. Data are presented as the mean ± SD from three independent biological replicates. Significance was determined by one-way ANOVA with Tukey’s HSD post hoc test. Two-way ANOVA with Šídák’s post hoc test was used for comparisons between PFD-treated and untreated groups. *, p < 0.05; **, p < 0.01; ns indicates not significant ( p ≥ 0.05).

Article Snippet: Pirfenidone (PFD) was purchased from TargetMol (Cat# T2386, Boston, MA, USA).

Techniques: Staining, Fluorescence, Immunofluorescence

Journal: International Journal of Molecular Sciences

Article Title: Pirfenidone Suppresses Liver Fibrosis Through Inhibition of TGF-β-Associated Lipid Metabolic Remodeling in Hepatic Stellate Cells

doi: 10.3390/ijms27094061

Figure Lengend Snippet: TGF-β signaling enhances OA-associated intracellular lipid accumulation in HSCs, which is inhibited by pirfenidone. ( A ) BODIPY 493/503 staining images of LX2 and HSC-T6 cells under serum starvation and pretreated with TGF-β1 or TGF-β2(10 ng/mL) for 24 h and then incubated with 200 μM OA for the final 4 h. ( B ) Quantification of OA-associated intracellular lipid accumulation in LX2 and HSC-T6 cells. ( C ) Oil Red O staining images of cells under serum starvation ± PFD (250 μg/mL) for 24 h. Groups: control; OA (200 μM, final 4 h); TGF-β1 (10 ng/mL, 24 h) + OA (200 μM, final 4 h). The red square indicates the region shown at higher magnification in the upper left corner of the same subfigure ( D ) Quantification of Oil Red O-stained lipid droplets per cell in LX2 and HSC-T6 cells. Quantification was performed from randomly selected fields under blinded conditions. Data are presented as the mean ± SD from three independent biological replicates. Significance was determined by one-way ANOVA with Tukey’s HSD post hoc test. Two-way ANOVA with Šídák’s post hoc test was used for comparisons between PFD-treated and untreated groups. *, p < 0.05; **, p < 0.01; ns indicates not significant ( p ≥ 0.05).

Article Snippet: Pirfenidone (PFD) was purchased from TargetMol (Cat# T2386, Boston, MA, USA).

Techniques: Staining, Incubation, Control

Journal: International Journal of Molecular Sciences

Article Title: Pirfenidone Suppresses Liver Fibrosis Through Inhibition of TGF-β-Associated Lipid Metabolic Remodeling in Hepatic Stellate Cells

doi: 10.3390/ijms27094061

Figure Lengend Snippet: Low-dose OA activates AKT/ERK/p70 S6K signaling in HSCs, and this response is attenuated by pirfenidone. ( A , B ) Western blot detection of phospho-AKT, AKT, phospho-ERK, ERK and phospho-p70 S6K, p70 S6K in LX2 cells treated with OA or PA (25 μM) for 30 or 60 min, with 5% and 10% FBS as positive controls. ( C ) Western blot detection of phospho-AKT, AKT, phospho-ERK, ERK and phospho-p70 S6K, p70 S6K in LX2 and HSC-T6 cells treated with OA (25 μM) in the presence or absence of PFD (250 μg/mL) for 30 min.

Article Snippet: Pirfenidone (PFD) was purchased from TargetMol (Cat# T2386, Boston, MA, USA).

Techniques: Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Pirfenidone Suppresses Liver Fibrosis Through Inhibition of TGF-β-Associated Lipid Metabolic Remodeling in Hepatic Stellate Cells

doi: 10.3390/ijms27094061

Figure Lengend Snippet: Pirfenidone suppresses SREBP1/FASN signaling in activated HSC-rich regions in vivo. ( A , B ) Immunohistochemical staining of SREBP1 and FASN in liver sections from control, CCl 4 , and CCl 4 + PFD (400 mg/kg/day) groups. ( C , D ) Quantification of SREBP1 and FASN positive staining. ( E ) Multiplex immunofluorescence images of liver sections stained for α-SMA, SREBP1, and FASN. Quantification was performed from randomly selected fields under blinded conditions. Data are presented as the mean ± SD. Significant changes are indicated as follows: **, p < 0.01, compared to the control group (unpaired Student’s t -test).

Article Snippet: Pirfenidone (PFD) was purchased from TargetMol (Cat# T2386, Boston, MA, USA).

Techniques: In Vivo, Immunohistochemical staining, Staining, Control, Multiplex Assay, Immunofluorescence, Immunohistochemistry

Journal: International Journal of Molecular Sciences

Article Title: Pirfenidone Suppresses Liver Fibrosis Through Inhibition of TGF-β-Associated Lipid Metabolic Remodeling in Hepatic Stellate Cells

doi: 10.3390/ijms27094061

Figure Lengend Snippet: Pirfenidone attenuates CCl 4 -induced liver fibrosis and HSC activation in vivo. ( A ) H&E staining of liver sections from control, CCl 4 , and CCl 4 + PFD (400 mg/kg/day) groups. ( B ) Immunohistochemical staining of α-SMA in liver sections from the indicated groups. ( C ) Immunohistochemical staining of COL1A1 in liver sections from the indicated groups. ( D , E ) Quantification of α-SMA and COL1A1 positive staining. Quantification was performed from randomly selected fields under blinded conditions. Data are presented as the mean ± SD. Significance was determined by one-way ANOVA with Tukey’s HSD post hoc test. ** p < 0.01; ns indicates not significant ( p ≥ 0.05).

Article Snippet: Pirfenidone (PFD) was purchased from TargetMol (Cat# T2386, Boston, MA, USA).

Techniques: Activation Assay, In Vivo, Staining, Control, Immunohistochemical staining, Immunohistochemistry, H&E Stain