Thymosin β4 suppresses CCl(4) -induced murine hepatic fibrosis by down-regulating transforming growth factor β receptor-II.

The present study aimed to clarify the effects of thymosin β4 (Tβ4) on CCl4 -induced hepatic fibrosis in mice and to further explore the underlying mechanisms. Expression of Tβ4 in fibrotic liver tissues was assessed by a quantitative real time-reverse transcriptase polymerase chain reaction and immunohistochemistry. The effects of intraperitoneal adeno-associated virus-Tβ4 (AAV-Tβ4) on CCl4 -induced hepatic fibrosis were observed by the evaluation of collagen deposition, hepatic stellate cell (HSC) activation and pro-fibrotic cytokine expression. In vitro tests with HSCs and hepatocytes were performed to confirm the effects of Tβ4. The expression of Tβ4 was down-regulated in fibrotic mouse livers but was rapidly up-regulated by CCl4 -induced acute injury. AAV-Tβ4 pre-treatment significantly attenuated liver injury, collagen deposition, HSC activation and pro-fibrotic cytokine over-expression, such as transforming growth factor β1 (TGF-β1), platelet-derived growth factor B (PDGF-B), connective tissue growth factor (CTGF) and plasminogen activator inhibitor-1 (PAI-1) in CCl4 -intoxicated mouse livers. In vitro experiments showed that Tβ4 suppressed HSC proliferation, blunted TGF-β1-induced HSC activation and reduced TGF-β1-induced TGF-β1, PDGF-B, CTGF and PAI-1 expression in both HSCs and hepatocytes. Ectopic Tβ4 ameliorated the over-expression of TGF-β receptor-II (TGF-βRII) in the fibrotic mouse livers. Exogenous Tβ4 down-regulated TGF-βRII expression, whereas neutralizing endogenous extracellular Tβ4 with a specific antibody up-regulated TGF-βRII expression in cultured HSCs and hepatocytes. Tβ4 possesses anti-fibrotic activity in the liver, which is attributable, at least partly, to down-regulating TGF-βRII and thereby blunting TGF-β1-mediated fibrogenetic signaling in both HSCs and hepatocytes.

Li H ，Li Q ，Zhang X ，Zheng X ，Zhang Q ，Hao Z ... -  《-》

Hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein (HIP/PAP) confers protection against hepatic fibrosis through downregulation of transforming growth factor β receptor II.

Li Q ，Li H ，Lv Y ，Zhang Q ，Zhang X ，Li S ，Zheng X ，Wang Y ，Hao Z ... -  《-》

Epigenetically-Regulated MicroRNA-9-5p Suppresses the Activation of Hepatic Stellate Cells via TGFBR1 and TGFBR2.

Recently, microRNAs (miRNAs) have been demonstrated to act as regulators of activation of hepatic stellate cells (HSCs). It is well known that the main profibrogenic inducer transforming growth factor-β1 (TGF-β1) contributes to HSC activation, which is a key event in liver fibrosis. Increasing studies show that miR-9-5p is down-regulated in liver fibrosis and restoration of miR-9-5p limits HSC activation. However, the role of miR-9-5p in TGF-β1-induced HSC activation is still not clear. miR-9-5p expression was quantified using real-time PCR in chronic hepatitis B (CHB) patients and TGF-β1-treated LX-2 cells. In CHB patients, histological activity index (HAI) and fibrosis stages were assessed using the Ishak scoring system. Effects of miR-9-5p on liver fibrosis in vivo and in vitro were analyzed. Luciferase activity assays were performed to examine the binding of miR-9-5p to the 3'-untranslated region of type I TGF-β receptor (TGFBR1) as well as TGFBR2. Compared with healthy controls, miR-9-5p was reduced in CHB patients. There was a lower miR-9-5p expression in CHB patients with higher fibrosis scores or HAI scores. miR-9-5p was down-regulated by TGF-β1 in a dose-dependent manner. TGF-β1-induced HSC activation including cell proliferation, α-SMA and collagen expression was blocked down by miR-9-5p. Notably, miR-9-5p ameliorates carbon tetrachloride-induced liver fibrosis. As determined by luciferase activity assays, TGFBR1 and TGFBR2 were targets of miR-9-5p. Further studies demonstrated that miR-9-5p inhibited TGF-β1/Smads pathway via TGFBR1 and TGFBR2. Interestingly, promoter methylation was responsible for miR-9-5p down-regulation in liver fibrosis. The relationship between miR-9-5p expression and methylation was confirmed in CHB patients and TGF-β1-treated cells. Our results demonstrate that miR-9-5p could inhibit TGF-β1-induced HSC activation through TGFBR1 and TGFBR2. Loss of miR-9-5p is associated with its methylation status in liver fibrosis.

Yu F ，Chen B ，Fan X ，Li G ，Dong P ，Zheng J ... -  《-》

Involvement of the nuclear high mobility group B1 peptides released from injured hepatocytes in murine hepatic fibrogenesis.

This study investigated the pro-fibrogenic role of high mobility group box 1 (HMGB1) peptides in liver fibrogenesis. An animal model of carbon tetrachloride (CCl4)-induced liver fibrosis was used to examine the serum HMGB1 levels and its intrahepatic distribution. The increased serum HMGB1 levels were positively correlated with elevation of transforming growth factor-β1 (TGF-β1) and collagen deposition during fibrogenesis. The cytoplasmic distribution of HMGB1 was noted in the parenchymal hepatocytes of fibrotic livers. In vitro studies confirmed that exposure to hydrogen peroxide and CCl4 induced an intracellular mobilization and extracellular release of nuclear HMGB1 peptides in clone-9 and primary hepatocytes, respectively. An uptake of exogenous HMGB1 by hepatic stellate cells (HSCs) T6 cells indicated a possible paracrine action of hepatocytes on HSCs. Moreover, HMGB1 dose-dependently stimulated HSC proliferation, up-regulated de novo synthesis of collagen type I and α-smooth muscle actin (α-SMA), and triggered Smad2 phosphorylation and its nuclear translocation through a TGF-β1-independent mechanism. Blockade with neutralizing antibodies and gene silencing demonstrated the involvement of the receptor for advanced glycation end-products (RAGE), but not toll-like receptor 4, in cellular uptake of HMGB1 and the HMGB1-mediated Smad2 and ERK1/2 phosphorylation as well as α-SMA up-regulation in HSC-T6 cells. Furthermore, anti-RAGE treatment significantly ameliorated CCl4-induced liver fibrosis. In conclusion, the nuclear HMGB1 peptides released from parenchymal hepatocytes during liver injuries may directly activate HSCs through stimulating HSC proliferation and transformation, eventually leading to the fibrotic changes of livers. Blockade of HMGB1/RAGE signaling cascade may constitute a therapeutic strategy for treatment of liver fibrosis.

Kao YH ，Lin YC ，Tsai MS ，Sun CK ，Yuan SS ，Chang CY ，Jawan B ，Lee PH ... -  《-》

Thymosin β4 attenuates liver fibrosis via suppressing Notch signaling.

Thymosin β4 (Tβ4) has been shown to have beneficial effects in a number of pathological processes. Although there was research reporting the endogenous expression of Tβ4 influences hepatic stellate cells (HSCs) activation, the effect of exogenous administration of Tβ4 in hepatic fibrosis remains unclear. In the current study, we used CCl4-induced liver fibrosis model mice to investigate the effect of Tβ4 administration on fibrosis in vivo and the underlying mechanism. Our study indicates that Tβ4 attenuates hepatic fibrosis and down-regulates the expression of fibrogenic genes in hepatic liver. In addition, Tβ4 inhibits the expression of pro fibrogenic and proliferation genes in activated HSCs. Further study revealed that Tβ4 attenuates liver fibrosis through inhibition of the Notch signaling, as Tβ4 significantly reduces the expression of Notch2 and Notch 3 that were increased in hepatic liver. Our data indicate that Tβ4 might be an effective anti-fibrotic drug for the treatment of liver fibrosis.

Hong Y ，Yao Q ，Zheng L 《-》