The anti-fibrosis efficacy of Ganoderma lucidum (GL) was investigated on thioacetamide (TAA)-induced liver fibrosis. Experimental fibrosis was induced by (200 mg/kg TAA, i.p.) twice weekly for 6 weeks in male Sprague-Dawley rats. GL was administered to TAA rats, either as (250/500 mg/kg, i.p) daily for further 3 weeks. Repeated administration of TAA caused liver fibrosis evidenced by significant elevation of hepatic TGF-β1 accompanied by an increase in the hydroxyproline content, oxidative stress levels and liver biomarkers. Contrarily, GL treatment significantly reduced ALT, AST, total bilirubin, MDA, NO concentrations and restored SOD activity. H & E and Masson trichrome staining confirmed that GL suppressed liver fibrosis, inflammation and attenuated the histopathological alterations. GL also elicited a statistical decrease in TGF-β1 level and hydroxyproline content with a concomitant decline in NFk-B and α-SMA immunoexpression in the TAA treated rats. Furthermore, GL downregulated TNF-α and IL-1β levels in TAA-treated rats compared to the control group. Conclusion: GL possesses a pronounced protective activity against TAA-induced fibrosis in rats; It significantly inhibits oxidative stress, inflammation and diminishes fibrosis by: inhibiting NFk-B activation, reducing TGF-β mediated by PI3K-AKT pathway thus restoring the liver function, the effect was in a dose dependent manner.