Resveratrol ameliorates nutritional steatohepatitis through the mmu‑miR‑599/PXR pathway

Kong,Lingbo; An,Xinyu; Hu,Lingxi; Zhang,Siyu; Liu,Lingdi; Zhao,Suxian; Wang,Rongqi; Nan,Yuemin

doi:10.3892/ijmm.2022.5102

Resveratrol ameliorates nutritional steatohepatitis through the mmu‑miR‑599/PXR pathway

Authors:
- Lingbo Kong
- Xinyu An
- Lingxi Hu
- Siyu Zhang
- Lingdi Liu
- Suxian Zhao
- Rongqi Wang
- Yuemin Nan
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Affiliations: Department of Traditional and Western Medical Hepatology, Third Hospital of Hebei Medical University, Shijiazhuang, Hebei 050051, P.R. China
Published online on: February 8, 2022 https://doi.org/10.3892/ijmm.2022.5102
Article Number: 47
Copyright: © Kong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

The aim of the present study was to elucidate the effect of resveratrol on non‑alcoholic steatohepatitis (NASH), and the molecular basis in mice and Hepa1‑6 cells, in order to verify its therapeutic effect. C57BL/6J mice were fed a methionine‑choline‑deficient (MCD) diet to induce steatohepatitis and were treated with resveratrol. Mouse sera were collected for biochemical analysis and enzyme‑linked immunosorbent assay, and livers were obtained for histological observation, and mmu‑microRNA (miR)‑599 and inflammation‑related gene expression analysis. Hepa1‑6 cells were treated with palmitic acid to establish a NASH cell model, and were then treated with resveratrol, or transfected with mmu‑miR‑599 mimic, mmu‑miR‑599 inhibitor or recombinant pregnane X receptor (PXR) plasmid. Subsequently, the cells were collected for mmu‑miR‑599 and inflammation‑related gene expression analysis. Reverse transcription‑quantitative polymerase chain reaction and western blotting were used to assess mmu‑miR‑599 expression levels, and the mRNA and protein expression levels of PXR and inflammation‑related genes. The binding site of mmu‑miR‑599 in the PXR mRNA was verified by the luciferase activity assay. Mice fed an MCD diet for 4 weeks exhibited steatosis, focal necrosis and inflammatory infiltration in the liver. Resveratrol significantly reduced serum aminotransferase and malondialdehyde levels, and ameliorated hepatic injury. These effects were associated with reduced mmu‑miR‑599 expression, enhanced PXR expression, and downregulated levels of nuclear factor‑κB, tumour necrosis factor‑α, interleukin (IL)‑1β, IL‑6, NOD‑like receptor family pyrin domain‑containing protein 3 and signal transducer and activator of transcription 3. Administration of the mmu‑miR‑599 mimic inhibited PXR expression in Hepa1‑6 cells, whereas the mmu‑miR‑599 inhibitor exerted the opposite effect. A binding site for mmu‑miR‑599 was identified in the PXR mRNA sequence. Furthermore, overexpression of PXR inhibited the expression of inflammatory factors in Hepa1‑6 cells. The present study provided evidence for the protective role of resveratrol in ameliorating steatohepatitis through regulating the mmu‑miR‑599/PXR pathway and the consequent suppression of related inflammatory factors. Resveratrol may serve as a potential candidate for steatohepatitis management.

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1	Friedman SL, Neuschwander-Tetri BA, Rinella M and Sanyal AJ: Mechanisms of NAFLD development and therapeutic strategies. Nat Med. 24:908–922. 2018. View Article : Google Scholar : PubMed/NCBI
2	Eslam M, Valenti L and Romeo S: Genetics and epigenetics of NAFLD and NASH: Clinical impact. J Hepatol. 68:268–279. 2018. View Article : Google Scholar
3	Ryan MC, Itsiopoulos C, Thodis T, Ward G, Trost N, Hofferberth S, O'Dea K, Desmond PV, Johnson NA and Wilson AM: The Mediterranean diet improves hepatic steatosis and insulin sensitivity in individuals with non-alcoholic fatty liver disease. J Hepatol. 59:138–143. 2013. View Article : Google Scholar : PubMed/NCBI
4	Baur JA and Sinclair DA: Therapeutic potential of resveratrol: The in vivo evidence. Nat Rev Drug Discov. 5:493–506. 2006. View Article : Google Scholar : PubMed/NCBI
5	Faghihzadeh F, Adibi P, Rafiei R and Hekmatdoost A: Resveratrol supplementation improves inflammatory biomarkers in patients with nonalcoholic fatty liver disease. Nutr Res. 34:837–843. 2014. View Article : Google Scholar : PubMed/NCBI
6	Tejada S, Capó X, Mascaró CM, Monserrat-Mesquida M, Quetglas-Llabrés MM, Pons A, Tur JA and Sureda A: Hepatoprotective effects of resveratrol in non-alcoholic fatty live disease. Curr Pharm Des. 27:2558–2570. 2021. View Article : Google Scholar
7	Filipowicz W, Bhattacharyya SN and Sonenberg N: Mechanisms of post-transcriptional regulation by microRNAs: Are the answers in sight? Nat Rev Genet. 9:102–114. 2008. View Article : Google Scholar : PubMed/NCBI
8	Wang Y, Du J, Niu X, Fu N, Wang R, Zhang Y, Zhao S, Sun D and Nan Y: MiR-130a-3p attenuates activation and induces apoptosis of hepatic stellate cells in nonalcoholic fibrosing steatohepatitis by directly targeting TGFBR1 and TGFBR2. Cell Death Dis. 8:e27922017. View Article : Google Scholar : PubMed/NCBI
9	Du J, Niu X, Wang Y, Kong L, Wang R, Zhang Y, Zhao S and Nan Y: MiR-146a-5p suppresses activation and proliferation of hepatic stellate cells in nonalcoholic fibrosing steatohepatitis through directly targeting Wnt1 and Wnt5a. Sci Rep. 5:161632015. View Article : Google Scholar : PubMed/NCBI
10	Wang DP, Tang XZ, Liang QK, Zeng XJ, Yang JB and Xu J: MicroRNA-599 promotes apoptosis and represses proliferation and epithelial-mesenchymal transition of papillary thyroid carcinoma cells via down-regulation of Hey2-depentent Notch signaling pathway. J Cell Physiol. 235:2492–2505. 2020. View Article : Google Scholar
11	Zhou JL, Deng S, Fang HS, Du XJ, Peng H and Hu QJ: Circular RNA circANKRD36 regulates Casz1 by targeting miR-599 to prevent osteoarthritis chondrocyte apoptosis and inflammation. J Cell Mol Med. 25:120–131. 2021. View Article : Google Scholar
12	Schwabe RF and Luedde T: Apoptosis and necroptosis in the liver: A matter of life and death. Nat Rev Gastroenterol Hepatol Dec. 15:738–752. 2018. View Article : Google Scholar
13	Okamura M, Shizu R, Abe T, Kodama S, Hosaka T, Sasaki T and Yoshinari K: PXR functionally interacts with NF-κB and AP-1 to downregulate the inflammation-induced expression of chemokine CXCL2 in mice. Cells. 9:E22962020. View Article : Google Scholar
14	Gu X, Ke S, Liu D, Sheng T, Thomas PE, Rabson AB, Gallo MA, Xie W and Tian Y: Role of NF-kappaB in regulation of PXR-mediated gene expression: A mechanism for the suppression of cytochrome P-450 3A4 by proinflammatory agents. J Biol Chem. 281:17882–17889. 2006. View Article : Google Scholar : PubMed/NCBI
15	Du J, Niu X, Wang R, Zhao S, Kong L, Zhang Y and Nan Y: TLR4-dependent signaling pathway modulation: A novel mechanism by which pioglitazone protects against nutritional fibrotic steatohepatitis in mice. Mol Med Rep. 13:2159–2166. 2016. View Article : Google Scholar
16	Li S, Zheng X, Zhang X, Yu H, Han B, Lv Y, Liu Y, Wang X and Zhang Z: Exploring the liver fibrosis induced by deltamethrin exposure in quails and elucidating the protective mechanism of resveratrol. Ecotoxicol Environ Saf. 207:1115012021. View Article : Google Scholar
17	Lv Y, Bing Q, Lv Z, Xue J, Li S, Han B, Yang Q, Wang X and Zhang Z: Imidacloprid-induced liver fibrosis in quails via activation of the TGF-β1/Smad pathway. Sci Total Environ. 705:1359152020. View Article : Google Scholar
18	Han B, Wang X, Wu P, Jiang H, Yang Q, Li S, Li J and Zhang Z: Pulmonary inflammatory and fibrogenic response induced by graphitized multi-walled carbon nanotube involved in cGAS-STING signaling pathway. J Hazard Mater. 417:1259842021. View Article : Google Scholar : PubMed/NCBI
19	Brunt EM, Janney CG, Di Bisceglie AM, Neuschwander-Tetri BA and Bacon BR: Nonalcoholic steatohepatitis: A proposal for grading and staging the histological lesions. Am J Gastroenterol. 94:2467–2474. 1999. View Article : Google Scholar : PubMed/NCBI
20	Kleiner DE, Brunt EM, Van Natta M, Behling C, Contos MJ, Cummings OW, Ferrell LD, Liu YC and Torbenson MS: Unalp-Arida A, et al Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology. 41. pp. 1313–1321. 2005, View Article : Google Scholar
21	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar
22	Zanieri F, Levi A, Montefusco D, Longato L, De Chiara F, Frenguelli L, Omenetti S, Andreola F, Luong TV, Massey V, et al: Exogenous liposomal ceramide-C6 ameliorates lipidomic profile, energy homeostasis, and anti-oxidant systems in NASH. Cells. 9:12372020. View Article : Google Scholar :
23	Pilling D, Karhadkar TR and Gomer RH: A CD209 ligand and a sialidase inhibitor differentially modulate adipose tissue and liver macrophage populations and steatosis in mice on the Methionine and Choline-Deficient (MCD) diet. PLoS One. 15:e02447622020. View Article : Google Scholar
24	Baur JA, Pearson KJ, Price NL, Jamieson HA, Lerin C, Kalra A, Prabhu VV, Allard JS, Lopez-Lluch G, Lewis K, et al: Resveratrol improves health and survival of mice on a high-calorie diet. Nature. 444. pp. 337–342. 2006, View Article : Google Scholar
25	Li Y, Xu S, Mihaylova MM, Zheng B, Hou X, Jiang B, Park O, Luo Z, Lefai E, Shyy JY, et al: AMPK phosphorylates and inhibits SREBP activity to attenuate hepatic steatosis and atherosclerosis in diet-induced insulin-resistant mice. Cell Metab. 13:376–388. 2011. View Article : Google Scholar : PubMed/NCBI
26	Shang J, Chen LL, Xiao FX, Sun H, Ding HC and Xiao H: Resveratrol improves non-alcoholic fatty liver disease by activating AMP-activated protein kinase. Acta Pharmacol Sin. 29:698–706. 2008. View Article : Google Scholar : PubMed/NCBI
27	Nan YM, Han F, Kong LB, Zhao SX, Wang RQ, Wu WJ and Yu J: Adenovirus-mediated peroxisome proliferator activated receptor gamma overexpression prevents nutritional fibrotic steatohepatitis in mice. Scand J Gastroenterol. 46:358–369. 2011. View Article : Google Scholar
28	Hu G, Xu C and Staudinger JL: Pregnane X receptor is SUMOylated to repress the inflammatory response. J Pharmacol Exp Ther. 335:342–350. 2010. View Article : Google Scholar : PubMed/NCBI
29	Mencarelli A, Renga B, Palladino G, Claudio D, Ricci P, Distrutti E, Barbanti M, Baldelli F and Fiorucci S: Inhibition of NF-κB by a PXR-dependent pathway mediates counter-regulatory activities of rifaximin on innate immunity in intestinal epithelial cells. Eur J Pharmacol. 668:317–324. 2011. View Article : Google Scholar : PubMed/NCBI
30	Zhou C, Tabb MM, Nelson EL, Grün F, Verma S, Sadatrafiei A, Lin M, Mallick S, Forman BM, Thummel KE and Blumberg B: Mutual repression between steroid and xenobiotic receptor and NF-kappaB signaling pathways links xenobiotic metabolism and inflammation. J Clin Investig. 116. pp. 2280–2289. 2006, View Article : Google Scholar
31	Huang K, Mukherjee S, DesMarais V, Albanese JM, Rafti E, Draghi Ii A, Maher LA, Khanna KM, Mani S and Matson AP: Targeting the PXR-TLR4 signaling pathway to reduce intestinal inflammation in an experimental model of necrotizing enterocolitis. Pediatr Res. 83:1031–1040. 2018. View Article : Google Scholar : PubMed/NCBI
32	Li J, Jiang H, Wu P, Li S, Han B, Yang Q, Wang X, Han B, Deng N, Qu B and Zhang Z: Toxicological effects of deltamethrin on quail cerebrum: Weakened antioxidant defense and enhanced apoptosis. Environ Pollut. 286:1173192021. View Article : Google Scholar
33	Hess J, Angel P and Schorpp-Kistner M: AP-1 subunits: Quarrel and harmony among siblings. J Cell Sci. 117:5965–5973. 2004. View Article : Google Scholar : PubMed/NCBI
34	Wree A, McGeough MD, Inzaugarat ME, Eguchi A, Schuster S, Johnson CD, Peña CA, Geisler LJ, Papouchado BG, Hoffman HM and Feldstein AE: NLRP3 inflammasome driven liver injury and fibrosis: Roles of IL-17 and TNF in mice. Hepatology. 67:736–749. 2018. View Article : Google Scholar
35	Mridha AR, Wree A, Robertson AAB, Yeh MM, Johnson CD, Van Rooyen DM, Haczeyni F, Teoh NC, Savard C, Ioannou GN, et al: NLRP3 inflammasome blockade reduces liver inflammation and fibrosis in experimental NASH in mice. J Hepatol. 66:1037–1046. 2017. View Article : Google Scholar : PubMed/NCBI
36	Masters SL: Specific inflammasomes in complex diseases. Clin Immunol. 147:223–228. 2013. View Article : Google Scholar : PubMed/NCBI
37	Schroder K and Tschopp J: The inflammasomes. Cell. 140:821–832. 2010. View Article : Google Scholar : PubMed/NCBI
38	Ouyang X, Ghani A and Mehal WZ: Inflammasome biology in fibrogenesis. Biochim Biophys Acta. 1832:979–988. 2013. View Article : Google Scholar
39	Rose-John S, Scheller J, Elson G and Jones SA: Interleukin-6 biology is coordinated by membrane-bound and soluble receptors: Role in inflammation and cancer. J Leukoc Biol. 80:227–236. 2006. View Article : Google Scholar : PubMed/NCBI
40	Grivennikov S and Karin M: Autocrine IL-6 signaling: A key event in tumorigenesis? Cancer Cell. 13:7–9. 2008. View Article : Google Scholar : PubMed/NCBI
41	Imajo K, Fujita K, Yoneda M, Nozaki Y, Ogawa Y, Shinohara Y, Kato S, Mawatari H, Shibata W, Kitani H, et al: Hyperresponsivity to low-dose endotoxin during progression to nonalcoholic steatohepatitis is regulated by leptin-mediated signaling. Cell Metab. 16:44–54. 2012. View Article : Google Scholar : PubMed/NCBI
42	Kessoku T, Imajo K, Honda Y, Kato T, Ogawa Y, Tomeno W, Kato S, Mawatari H, Fujita K, Yoneda M, et al: Resveratrol ameliorates fibrosis and inflammation in a mouse model of nonalcoholic steatohepatitis. Sci Rep. 6:222512016. View Article : Google Scholar :