| Summary: | 博士 === 國立陽明大學 === 傳統醫藥研究所 === 101 === During prolonged liver injury, activation of hepatic stellate cells (HSCs) is crucially involved in the pathogenesis of liver fibrosis. Excessive oxidative stress, cytokines release due to hepatic inflammation and apoptotic bodies from hepatocytes are implicated in the activation of HSCs and hepatic fibrogenesis. Strategies of antioxidation, anti-inflammation and hepato-protection have been proposed to inhibit the activation of HSCs and attenuate hepatic fibrosis. Armepavine (Arm), an active component of Nelumbo nucifera Semen (Nn), has been reported to exert antioxidative, anti-inflammatory and hepatoprotective effects. This study investigated the in vitro and in vivo anti-fibrotic effects of an armepavine-enriched extract of Nn (Nelumbo nucifera Semen) and Arm.
A hepatic stellate cell line of rat origin (HSC-T6), which could be activated by TNF-α, TGF-β1 or LPS was used as a cellular model, and we used extract of Nn (0-80 μg/ml) and Arm (0-10 μM) as treatments. The extract of Nn (0-80 M) inhibited TNF-α-stimulated α-SMA secretion, intracellular ROS production, and suppressed IκBα phosphorylation and NF-κB transcriptional activity induced by TNF-α. Nn (0-80 M) also inhibited mRNA expressions of fibrosis-related genes including α-SMA. Moreover, the extract of Nn (0-80 μg/ml) significantly reduced TGF-β1-induced collagen deposition, Smad 2/3 phosphorylation and α-SMA secretion.
Arm was further shown to reduce TNF-α-induced expression of α-SMA and procollagen type I, intracellular ROS production, collagen deposition, NF-κB transactivation performed by the p-NF-κB-luc luciferase assay, MAPK phosphorylations including p38, ERK1/2 and JNK, translocation of transcription factors-NF-κB, JunD and C/EBPβ in HSC-T6 cells. Furthermore, Arm (10 µM) could significantly inhibit TNF-α stimulated protein in HSCs expression of angiopoietin-1 from 337 ± 24% to 242 ± 19% in HSCs. The in vitro study suggested Arm inhibited TNF-α-stimulated HSC activation MAPK phosphorylations and multiple transcription factors-NF-κB, JunD and C/EBPβ. Arm (10 µM) also significantly inhibited LPS-induced α-SMA secretion (from 174 ± 37% to 98 ± 4%) in HSCs.
We induced hepatic fibrosis in rats by thioacetamide (TAA) administration and bile duct ligation (BDL), to investigate the in vivo effects of the extract of Nn and Arm on hepatic fibrosis. Both the extract of Nn 0.5 g/kg and Arm (3 and 10 mg/kg) significantly reduced the fibrosis scores of livers, levels of plasma AST and ALT activities and hepatic collagen contents. Arm (3 and 10 mg/kg) significantly reduced mRNA expression levels of IL-6, TGF-β1, TIMP-1, col1α2, iNOS, and ICAM-1 genes, but up-regulated metallothionein gene in TAA and BDL rats. Furthermore, immuno-staining results showed that α-SMA- and NF-κB- positive cells (activated HSCs) were decreased in the fibrotic livers of TAA and BDL rats treated by Arm.
In conclusion, results from this study suggested that the extract of Nn and Arm exerted both in vitro and in vivo antifibrotic effects in rats, possibly through anti-NF-κB, JunD, and C/EBPβ activation pathways. The data of this study might support drug development and clinical application of Arm and Nn.
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