Comparative evaluation of antifibrotic activity of Salvadora persica extracts in CCl4-induced liver fibrosis in rats by reducing oxidative stress and inflammation via transcriptional modulation associated with TGF-β1/Smad signaling pathway

Plant Salvadora persica (SP) is well-known for its pharmacological activities, including its role in oral hygiene. In the present study, we investigated, the in vivo comparative hepatoprotective potential of SP root extracts against chronic liver injury fibrosis. In Wistar albino rats, liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl4) in olive oil (1:1 v/v) at 1 mL/kg, twice weekly for 9 weeks. Liver damage was evaluated by measuring serum liver biochemical parameters, antioxidant enzyme levels, and lipid peroxidation marker in liver homogenate. The mRNA expression level of inflammatory and profibrotic markers in liver tissue and histopathology of liver tissue by H&E staining and collagen deposition by Masson’s trichrome, and Van Gieson’s staining were determined. SP extracts, including aqueous (SP-AQ), ethanolic (SP-E), and petroleum ether (SP-PE), were analyzed by Gas Chromatography-Mass Spectrometry (GC-MS), followed by multivariate principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA). The findings demonstrated that SP extracts demonstrated significant hepatoprotective effects by improving rats’ body weight without significant increase in liver index, reducing liver biochemical markers, and reducing oxidative stress by increasing SOD and GPx levels and decreasing MDA level. In liver tissue, the mRNA expression levels of the inflammatory markers IL-1β, IL-6, and TNF-α and profibrotic markers TGF-β1, Smad2, Smad3, α-SMA, and Collagen1A1 were downregulated by modulating the genes expression levels related with the TGF-β1/Smad signaling pathway. Histopathology showed recovery from liver damage and reduced collagen deposition. Comparatively, antifibrotic activity was greater by SP-PE, modest by SP-E and lowest by SP-AQ extracts. Furthermore, phytochemical analysis of the SP extracts demonstrated considerable diversity in their phytoconstituent profiles due to difference in solvent polarity, with PCA and PLS-DA analyses confirming distinct clustering and discrimination among SP extracts. SP-PE extract, greater efficacy can be attributed to selection of a non-polar solvent and 4-propenyl-2,6-dimethoxyphenol, along with other dominant phytochemicals, which exerted synergistic biological effects. The SP extracts showed significant hepatoprotective activity with consistent biological effects in reducing oxidative stress, inflammation, and fibrosis in CCl4-induced liver fibrosis in rats. However, further studies of major individual compounds of SP-PE are warranted for clinical use in liver fibrosis.