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. 2017 May 11:8:256.
doi: 10.3389/fphar.2017.00256. eCollection 2017.

SRT1720 Alleviates ANIT-Induced Cholestasis in a Mouse Model

Linxi Yu  1 , Xiaoxin Liu  1 , Zihang Yuan  1 , Xiaojiaoyang Li  1 , Hang Yang  1 , Ziqiao Yuan  1 , Lixin Sun  1 , Luyong Zhang  1   2   3 , Zhengzhou Jiang  1   4
Affiliations

SRT1720 Alleviates ANIT-Induced Cholestasis in a Mouse Model

Linxi Yu et al. Front Pharmacol. .

Abstract

Intrahepatic cholestasis is a kind of clinical syndrome along with hepatotoxicity which caused by intrahepatic and systemic accumulations of bile acid. There are several crucial generating factors of the pathogenesis of cholestasis, such as inflammation, dysregulation of bile acid transporters and oxidative stress. SIRT1 is regarded as a class III histone deacetylase (HDAC). According to a set of researches, SIRT1 is one of the most important factors which can regulate the hepatic bile acid metabolism. SRT1720 is a kind of activator of SIRT1 which is 1000 times more potent than resveratrol, and this paper is aimed to study its protective influence on hepatotoxicity and cholestasis induced by alpha-naphthylisothiocyanate (ANIT) in mice. The findings revealed that SRT1720 treatment increased FXR and Nrf2 gene expressions to shield against hepatotoxicity and cholestasis induced by ANIT. The mRNA levels of hepatic bile acid transporters were also altered by SRT1720. Furthermore, SRT1720 enhanced the antioxidative system by increasing Nrf2, SOD, GCLc, GCLm, Nqo1, and HO-1 gene expressions. In conclusion, a protective influence could be provided by SRT1720 to cure ANIT-induced hepatotoxicity and cholestasis, which was partly through FXR and Nrf2 activations. These results indicated that SIRT1 could be regarded as a therapeutic target to cure the cholestasis.

Keywords: ANTI; FXR; Nrf2; SRT1720; cholestasis.

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Figures

FIGURE 1
FIGURE 1
The chemical structure of SRT1720.
FIGURE 2
FIGURE 2
Protective effects of SRT1720 on cholestasis and hepatotoxicity induced by alpha-naphthylisothiocyanate (ANIT). Biochemical indicators in mice treated with vehicle or, 10 or, 20 mg/kg of SRT1720, were determined at the time points of 24 and 48 h after ANIT or vehicle administration. (A) Serum ALT, (B) AST, (C) ALP, and (D) γ-GGT activity, as well as (E) serum total bilirubin, (F) serum total bile acid (TBA), and (G) hepatic TBA. (H) Bile acid output decreased in mice due to ANIT and was significantly ameliorated in SRT1720-treated mice. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 3
FIGURE 3
SRT1720 attenuated liver injury induced by ANIT in mice. Images of H&E stained liver sections (200× magnification) at 48 h after ANIT administration were shown. (A) Vehicle: no histological change was observed; ANIT: necrotic and degenerative changes were observed. SRT1720 10mg/kg+ANIT: small necrotic and degenerative changes were observed; SRT1720 20 mg/kg+ANIT: small degenerative changes were observed. Areas of severe liver necrosis were marked by triangle arrows, and degenerative changes were marked by normal arrows. (B) Graph showed the quantitative analysis of necrotic lesions and degenerative changes. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 4
FIGURE 4
SRT1720 altered the gene expressions of hepatic transporters involved in bile acid transport in mice total livers. Quantitative real-time PCR analysis was performed to measure the gene expression levels (A) HNF1α and FXR, (B) Ntcp and Oatp1b2, (C) Bsep and Mrp2, (D) Mrp3 and Mrp4, (E) Cyp7A1 and Shp. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 5
FIGURE 5
SRT1720 altered the gene expressions of hepatic enzymes involved in bile acid metabolism in mice total livers. (A) Bile acid metabolizing enzymes, including the phase I enzymes Cyp3a11 and Cyp2b10 were shown. (B) Bile acid metabolizing enzymes, including the phase II enzymes Ugt1a1 and Sult2a1, were shown. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 6
FIGURE 6
SRT1720 restored the protein expressions of FXR, Bsep, and Mrp2 in mice total livers. (A) Western blot analysis was used to measure FXR, Bsep, and Mrp2 expressions. (B) Specific band intensity was quantified, normalized to GAPDH. (C) Immunofluorescence staining of frozen liver sections showing Bsep and Mrp2 expressions. (D) Fluorescent intensities of Bsep and Mrp2 were measured by Image-Pro Plus software.
FIGURE 7
FIGURE 7
Effects of SRT1720 on FXR, Bsep, and Mrp2 expressions in vitro. (A) Immunofluorescence was used to investigate the effects of SRT1720 on Bsep; after treatment with ANIT for 24 h, Bsep decreased in fluorescence intensity, and SRT1720 (5 μM, 10 μM) induced up-regulation of Bsep, which was observed compared with groups treated with ANIT. (B) Fluorescent intensity of Bsep was measured by Image-Pro Plus software. (C) Mouse primary hepatocytes were treated with ANIT (40 μM), and SRT1720 (5 μM, 10 μM); ANIT inhibited FXR, Bsep, and Mrp2 expression at the mRNA level, and SRT1720 increased all of them. (D) HNF1α silencing efficiency was measured by Western blot. (E) HNF1α silencing abrogated the regulation of FXR by SRT1720 (10 μM) in mice primary hepatocytes. ∗P < 0.05 versus DMSO alone; #P < 0.05 versus SRT1720 alone.
FIGURE 8
FIGURE 8
Effects of SRT1720 on antioxidant system and inflammatory factors in vivo. (A) Effects of SRT1720 on hepatic T-AOC, SOD, GSH, and MDA activities at 48 h after ANIT administration in mice. (B) Effects of SRT1720 on MPO activity, TNF-α and IL-6 levels at 48 h after ANIT administration in mice. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 9
FIGURE 9
SRT1720 altered gene expressions of Nrf2/ARE signalling in ANTI-induced liver hepatotoxicity and cholestasis in mice. (A) Nrf2, (B) SOD, (C) GCLm, (D) GCLc, (E) Nqo1, and (F) HOO-1. Data are the mean ± SD (n = 6). ∗P < 0.05 versus vehicle; #P < 0.05 versus vehicle +ANIT.
FIGURE 10
FIGURE 10
The possible mechanism of SRT1720 attenuated ANIT-induced cholestasis and liver injury.
See this image and copyright information in PMC

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