• The Journal of Internal Korean Medicine
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• v.42 no.6
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• pp.1223-1236
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• 2021

Objectives: The aim of the current study was to investigate the effect of Sosiho-tang on thioacetamide (TAA)-induced liver fibrosis in mice and to elucidate its underlying mechanisms. Methods: The mice were divided into 4 groups: Normal mice (Normal), TAA-induced control mice (Control), TAA-induced and silymarin-treated (50 mg/kg) mice (Silymarin), and TAA-induced and Sosiho-tang treated (200 mg/kg) mice (SSHT). Liver fibrosis was induced via intraperitoneal injection of TAA three times a week for 8 weeks. Silymarin and Sosiho-tang were concomitantly administered for 8 weeks. Serum and liver tissues were then collected and the anti-oxidant and inflammatory protein levels in the liver tissues were evaluated using western blotting. Results: SSHT administration significantly reduced the levels of AST, ALT, ammonia, and MPO in the serum. SSHT also significantly down-regulated liver NADPH oxidase and regulated the Nrf2/Keap1 signaling pathway. SSHT treatment downregulated the liver NF-κB levels and suppressed inflammatory cytokines. SSHT treatment also decreased bile acid-related factors, such as CYP7A1 and NTCP, and fibrosis-related factors, such as α-SMA and Collagen I. Conclusions: Taken together, these data suggest that SSHT administration suppressed the progression of liver fibrosis by activating the Nrf2/Keap1 pathway and inhibiting NF-κB.

• Biomolecules & Therapeutics
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• v.31 no.4
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• pp.434-445
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• 2023

We investigated whether FTY-720 might have an effect on bleomycin-induced pulmonary fibrosis through inhibiting TGF-β1 pathway, and up-regulating autophagy. The pulmonary fibrosis was induced by bleomycin. FTY-720 (1 mg/kg) drug was intraperitoneally injected into mice. Histological changes and inflammatory factors were observed, and EMT and autophagy protein markers were studied by immunohistochemistry and immunofluorescence. The effects of bleomycin on MLE-12 cells were detected by MTT assay and flow cytometry, and the related molecular mechanisms were studied by Western Blot. FTY-720 considerably attenuated bleomycin-induced disorganization of alveolar tissue, extracellular collagen deposition, and α-SMA and E-cadherin levels in mice. The levels of IL-1β, TNF-α, and IL-6 cytokines were attenuated in bronchoalveolar lavage fluid, as well as protein content and leukocyte count. COL1A1 and MMP9 protein expressions in lung tissue were significantly reduced. Additionally, FTY-720 treatment effectively inhibited the expressions of key proteins in TGF-β1/TAK1/P38MAPK pathway and regulated autophagy proteins. Similar results were additionally found in cellular assays with mouse alveolar epithelial cells. Our study provides proof for a new mechanism for FTY-720 to suppress pulmonary fibrosis. FTY-720 is also a target for treating pulmonary fibrosis.

• Tuberculosis and Respiratory Diseases
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• v.79 no.4
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• pp.257-266
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• 2016

Background: Idiopathic pulmonary fibrosis is a common interstitial lung disease; it is a chronic, progressive, and fatal lung disease of unknown etiology. Over the last two decades, knowledge about the underlying mechanisms of pulmonary fibrosis has improved markedly and facilitated the identification of potential targets for novel therapies. However, despite the large number of antifibrotic drugs being described in experimental pre-clinical studies, the translation of these findings into clinical practices has not been accomplished yet. NADH:quinone oxidoreductase 1 (NQO1) is a homodimeric enzyme that catalyzes the oxidation of NADH to $NAD^+$ by various quinones and thereby elevates the intracellular $NAD^+$ levels. In this study, we examined the effect of increase in cellular $NAD^+$ levels on bleomycin-induced lung fibrosis in mice. Methods: C57BL/6 mice were treated with intratracheal instillation of bleomycin. The mice were orally administered with ${\beta}$-lapachone from 3 days before exposure to bleomycin to 1-3 weeks after exposure to bleomycin. Bronchoalveolar lavage fluid (BALF) was collected for analyzing the infiltration of immune cells. In vitro, A549 cells were treated with transforming growth factor ${\beta}1$ (TGF-${\beta}1$) and ${\beta}$-lapachone to analyze the extracellular matrix (ECM) and epithelial-mesenchymal transition (EMT). Results: ${\beta}$-Lapachone strongly attenuated bleomycin-induced lung inflammation and fibrosis, characterized by histological staining, infiltrated immune cells in BALF, inflammatory cytokines, fibrotic score, and TGF-${\beta}1$, ${\alpha}$-smooth muscle actin accumulation. In addition, ${\beta}$-lapachone showed a protective role in TGF-${\beta}1$-induced ECM expression and EMT in A549 cells. Conclusion: Our results suggest that ${\beta}$-lapachone can protect against bleomycin-induced lung inflammation and fibrosis in mice and TGF-${\beta}1$-induced EMT in vitro, by elevating the $NAD^+$/NADH ratio through NQO1 activation.

• Journal of Yeungnam Medical Science
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• v.9 no.2
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• pp.224-238
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• 1992

Hepatic fibrosis was induced in Sprague-Dawley rats to evaluate the ultrastructural changes of fat-storing cells(Ito cells). For experimental induction of liver fibrosis, the rats were administered intraperitoneally with 0.5ml of 50% $Ccl_4$ solution per Kg body weight, twice weekly for 12 weeks. The rats were sacrified every week. The liver tissues were examined under light and eletron microscopes. And the immunohistochemical study of desmin was also performed. The results were summarized as follows : Light microscopic findings : The cellular infiltrations with inflammatory cells and Kupffer cells developed from 1 week after $Ccl_4$ injection, and were the most severe in 4 weeks. The strong immunoreactivity for desmin was also evident in 4 weeks. The centrilobular necrosis and fibrosis developed from 2 weeks after injection, and the necrosis persisted until 8 weeks. The progress of fibrosis was accompanied by decreases in cellular infiltration and reactivity for desmin, and increased gradual nodular formation was also observed. The cirrhosis was developed after 10 weeks. Electron microscopic findings : An increase in number of fat-storing cells was observed from 1 week after injection. Transitional cells characterized by a depletion of lipid droplets and a hypertrophy of the rER appeared after 2 weeks. The number of transitional cells with abundant collagen fibers in the extracellular spaces increased in 4 weeks. With progression of fibrosis the number of fat-storing cells decreased and proliferating fibroblasts with dilated rER were observed. According to these results it was revealed that there was an apparent transition from fat-storing cells to transitional cells and to fibroblasts. These cells had a few similar characteristics and may belong to the same cell population. Thus it was suggested that fat-storing cells might play an important role in hepatic fibrosis.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.2
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• pp.637-642
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• 2014

The pathogenesis of hepatocellular carcinoma (HCC) related to habitual betel quid (BQ) chewing is unclear. Risk of HCCis increased with adverse hepatic fibrosis. This study aimed to assess the impact of chronic viral hepatitis on adverse hepatic fibrosis in HCC related to BQ chewing. This hospital-based case-control study enrolled 200 pairs of age- and gender-matched patients with HCC and unrelated healthy controls. Serologic hepatitis B surface antigen (HBsAg), antibodies to hepatitis C virus (anti-HCV), ${\alpha}$-fetoprotein (AFP), and surrogate markers for significant hepatic fibrosis were measured. Information on substance-use habits was obtained with a questionnaire. By analysis of surrogate markers for hepatic fibrosis, the prevalence of significant hepatic fibrosis in patients chewing BQ was between 45.8% and 91.7%, whereas that for patients without BQ chewing was between 18.4% and 57.9%. The difference was significant (P <0.05 for each surrogate marker). Multivariate analysis indicated that cirrhosis with Child-Pugh C (odds ratio (OR) = 3.28; 95% confidence interval (CI), 1.29-8.37), thrombocytopenia (OR = 3.92, 95% CI, 1.77-8.68), AFP >400 mg/L (OR = 2.21, 95% CI, 1.05-4.66) and male gender (OR = 4.06, 95% CI, 1.29-12.77) were independent factors associated with habitual BQ chewing. In conclusion, adverse hepatic fibrosis and severe liver damage play important roles in the pathogenesis of BQ-related HCC, which could be aggravated by chronic hepatitis B and hepatitis C. BQ-cessation programs and prevention of chronic HBV/HCV infection are needed to prevent HCC related to BQ chewing.

• Korean Journal of Clinical Laboratory Science
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• v.36 no.2
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• pp.163-172
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• 2004

Mast cells (MCs) have been implicated in the pathogenesis of tissue fibrosis. However, the role of MC in the development of liver fibrosis has not been fully elucidated. Stem cell factor (SCF) is known to recruit MCs to the liver following injury as it induces mast cell proliferation, survival and differentiation from resident tissue precursors. This study examines the interaction between activated hepatic stellate cells (HSCs) and MCs in rat fibrotic liver, and SCF production by HSCs during culture in vitro. Rats were studied 4, 7, 14 and 21 days after bile duct ligation (BDL). Fibrogenesis was assessed by a measurement of collagen stained with sirius red F3B. Activated HSCs and MCs were identified by ${\alpha}$-smooth muscle actin (${\alpha}-SMA$) immunohistochemical and alcian blue staining and measured by a computerized image analysis system. SCF production was determined in rat HSC cultures using Western blotting. Mild fibrotic changes were noted in BDL rat livers as early as 4 days after induction of cholestasis. Significant expansion and organization of fibrous tissue has occurred in day 14 BDL rats which progressed to bridging fibrosis by day 21. In BDL rats, both a large number of activated HSCs and MCs were detected in portal tracts and fibrous septa. Both area of activated HSCs infiltration and density of MCs were significantly higher in all BDL group compared with Shams. In BDL rats, both areas of activated HSCs infiltration and density of MCs were no significant difference between day 4 and 7 and were significantly higher in day 14. However, the areas of activated HSCs infiltration were significantly lesser in day 21 and the densities of MCs were significantly higher in day 21 compared with day14 BDL. In BDL rats, both areas of activated HSCs infiltration and density of MCs were highly correlated with areas of fibrosis. Western blotting showed that SCF protein was consistently produced in activated HSCs by culture on plastic and freshly isolated HSCs expressed relatively little 30kD SCF compared to late primary culture activated HSCs (day 14) and passaged HSCs. These results suggest that HSCs activated in vitro produce SCF, and may play an important role in recruiting mast cells to the liver during injury and fibrosis.

• YAKHAK HOEJI
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• v.46 no.2
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• pp.129-136
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• 2002

The pathogenesis of cholestatic liver injury as well as the modulation of hepatic fibrogenesis is causally associated with involvement of reactive oxygen species (ROS) and free radical reactions. In this study, we investigated whether dried extracts of oriental medicine (LH) have antioxidant and antifibrotic effect under the biliary liver fibrosis (cirrhosis) c ondition. The female Sprague-Dawley rats were divided in six groups (Normal, N-LH, op-2, op-4, opLH-2, opLH-4) and were observed in 2 weeks or 4 weeks. For this purpose the rats were operated by bile duct ligation/scission (BDL/S), which induced to liver fibrosis and cirrhosis. After surgery, the prepared LH was administered p.o. 2 mι/day/rat in 2 weeks or 4 weeks for opLH groups. During the observation period, jaundices appeared in eyes, ears and tail of all BDL/S operated rats. And at the time of sacrifice, cholestasis was observed in proximal bile duct, especially the color of bile juice and urine in opLH-4 group showed more clear than op-2, op-4 and opLH-2 group. The value of clinical parameters and product of lipid peroxidation (MDA) in sera and the hydroxyproline (hyp) content in liver tissue were significantly increased in all liver fibrosis (cirrhosis) developed rats (p＜0.001~0.05). Among the clinical parameters of sera, value of BUN, ALP in opLH-4 group showed significantly lower than in op-4 group (p＜0.05, p＜0.001). The content of hyp in opLH-2, opLH-4 group (478.0 $\pm$ 134.3 $\mu\textrm{g}$/g 897.5 $\pm$ 118.2 $\mu\textrm{g}$/g) showed lower than in op-2, op-4 group (528.9 $\pm$ 220.7 $\mu\textrm{g}$/g, 1023.8 $\pm$ 277.1 $\mu\textrm{g}$/g) and then the value of MDA in opLH-4 was also significantly reduced to 59.4% of that in op-4 group (p＜0.001). The histological change (bile duct proliferation, fibrosis, collagen bundle) was similarly observed in op-2 group and in opLH-2 group, but the weak fibrosis and bile duct proliferation were observed in opLH-4 group compared with in op-4 group. Our data indicate that the 4 weeks treatment with LH extract suppressed lipid peroxidation and inhibited fibrotic (cirrhosis) process, and experimental cholestatic liver disease is associated with increased lipid peroxidation in BDL/S operated rats. Hence we concluded that the measurement of MDA and hyp can be useful monitor for the screening of antioxidant and antifibrotic effect in experimental liver fibrosis (cirrhosis), and LH has been shown to have hepatoprotective effect, antifibrotic effect and antioxidant effect.

• YAKHAK HOEJI
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• v.45 no.5
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• pp.513-521
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• 2001

The chronic cholestasis induce to biliary liver fibrosis (cirrhosis) and the increased products of ROS(reactive oxygen species) cause to the liver damage. In this study ; the antioxidant and antifibrotic effect of dried extracts of oriental medicine (DW) was investigated under the liver fibrotic (cirrhotic) condition. The female Sprague-Dawley rats were divided in 5 groups (Normal, Op-2, Op-4, OpDW-2, OpDW-4). Except for normal group, the rats were induced to biliary liver fibrosis (cirrhosis) by the operation of bile duct ligation/scission (BDU/S) and were observed in 2 weeks or 4 weeks. And the prepared DW was treated p.o.2 ml/day/rats in 2 weeks or 4 weeks for OpDW groups. At the time of sacrifice, the liver, kidney, spleen were weighed and the ratio of organ weight/body weight was calculated. The MDA, the hyp and biochemical parameters (GOT GTP, ALP, t-bili) were measured in sera and liver tissue of rats. The biochemical change was observed on liver tissue. In the result, the hepatomegaly and spleenomegaly appeared in all BDL/S operated rats, and significantly lower liver weight was observed in OpDW-4 group compared with in Op-4 group (p<0.05). The level of clinical parameters in sera of all liver fibrosis (cirrhosis) developed rats was higher than in normal group. Especial1y, the value of GOT in OpDW-2 group and ALP in OpDW-4 group showed significantly lower than in Op-2 group and Op-4 group (p<0.01, p<0.005). The content of hyp in all operation groups was significantly higher than in normal group (p<0.05∼<0.005), and showed significantly lower value in the OpDW-4 group than in Op-4 group (p<0.05). The product of lipid peroxidationUDA) increased significantly under the fibrotic(cirrhotic) condition (p<0.05∼ <0.005), and the MDA value in OpDW-4 group decreased significantly in Op-4 group (p<0.005). The histological change (bile duct proliferation, fibrosis, collagen bundle) was similarly observed in Op-2 group and in OpDW-2 group, but the weak fibrosis and bile duct proliferation were observed in OpDW-4 group compared with in Op-4 group. In conclusion, lipid peroxidation and severe liver damage were activated by bile duct obstruction, and the measurement of MDA and hap can be useful monitor for the screening of antioxidant and antifibrotic effect in experimental liver fibrosis (cirrhosis). The 4 weeks treatment with DW extracts suppressed lipid peroxidation and inhibited fibrotic (cirrhotic) process in BDL/S operated rats.

• Journal of Ginseng Research
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• v.47 no.4
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• pp.534-542
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• 2023

Background: Ginsenoside Rg1, a bioactive component of Ginseng, has demonstrated anti-inflammatory, anti-cancer, and hepatoprotective effects. It is known that the epithelial-mesenchymal transition (EMT) plays a key role in the activation of hepatic stellate cells (HSCs). Recently, Rg1 has been shown to reverse liver fibrosis by suppressing EMT, although the mechanism of Rg1-mediated anti-fibrosis effects is still largely unclear. Interestingly, Smad7, a negative regulator of the transforming growth factor β (TGF-β) pathway, is often methylated during liver fibrosis. Whether Smad7 methylation plays a vital role in the effects of Rg1 on liver fibrosis remains unclear. Methods: Anti-fibrosis effects were examined after Rg1 processing in vivo and in vitro. Smad7 expression, Smad7 methylation, and microRNA-152 (miR-152) levels were also analyzed. Results: Rg1 significantly reduced the liver fibrosis caused by carbon tetrachloride, and reduced collagen deposition was also observed. Rg1 also contributed to the suppression of collagenation and HSC reproduction in vitro. Rg1 caused EMT inactivation, reducing Desmin and increasing E-cadherin levels. Notably, the effect of Rg1 on HSC activation was mediated by the TGF-β pathway. Rg1 induced Smad7 expression and demethylation. The over-expression of DNA methyltransferase 1 (DNMT1) blocked the Rg1-mediated inhibition of Smad7 methylation, and miR-152 targeted DNMT1. Further experiments suggested that Rg1 repressed Smad7 methylation via miR-152-mediated DNMT1 inhibition. MiR-152 inhibition reversed the Rg1-induced promotion of Smad7 expression and demethylation. In addition, miR-152 silencing led to the inhibition of the Rg1-induced EMT inactivation. Conclusion: Rg1 inhibits HSC activation by epigenetically modulating Smad7 expression and at least by partly inhibiting EMT.

• Korean Journal of Radiology
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• v.21 no.12
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• pp.1317-1325
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• 2020

Objective: The aim of this study was to prospectively evaluate whether liver stiffness (LS) assessments, obtained by two-dimensional (2D)-shear wave elastography (SWE) with a propagation map, can evaluate liver fibrosis stage using histopathology as the reference standard. Materials and Methods: We prospectively enrolled 123 patients who had undergone percutaneous liver biopsy from two tertiary referral hospitals. All patients underwent 2D-SWE examination prior to biopsy, and LS values (kilopascal [kPa]) were obtained. On histopathologic examination, fibrosis stage (F0-F4) and necroinflammatory activity grade (A0-A4) were assessed. Multivariate linear regression analysis was performed to determine the significant factors affecting the LS value. The diagnostic performance of the LS value for staging fibrosis was assessed using receiver operating characteristic (ROC) analysis, and the optimal cut-off value was determined by the Youden index. Results: Reliable measurements of LS values were obtained in 114 patients (92.7%, 114/123). LS values obtained from 2D-SWE with the propagation map positively correlated with the progression of liver fibrosis reported from histopathology (p < 0.001). According to the multivariate linear regression analysis, fibrosis stage was the only factor significantly associated with LS (p < 0.001). The area under the ROC curve of LS from 2D-SWE with the propagation map was 0.773, 0.865, 0.946, and 0.950 for detecting F ≥ 1, F ≥ 2, F ≥ 3, and F = 4, respectively. The optimal cut-off LS values were 5.4, 7.8, 9.4, and 12.2 kPa for F ≥ 1, F ≥ 2, F ≥ 3, and F = 4, respectively. The corresponding sensitivity and specificity of the LS value for detecting cirrhosis were 90.9% and 88.4%, respectively. Conclusion: The LS value obtained from 2D-SWE with a propagation map provides excellent diagnostic performance in evaluating liver fibrosis stage, determined by histopathology.