Gli3 is a novel downstream target of miR‑200a with an anti‑ﬁbrotic role for progression of liver fibrosis in vivo and in vitro

Li,Li; Ran,Jianghua; Li,Lan; Chen,Gang; Zhang,Shengning; Wang,Yingjia

doi:10.3892/mmr.2020.10997

Gli3 is a novel downstream target of miR‑200a with an anti‑ﬁbrotic role for progression of liver fibrosis in vivo and in vitro

Authors:
- Li Li
- Jianghua Ran
- Lan Li
- Gang Chen
- Shengning Zhang
- Yingjia Wang
View Affiliations

Affiliations: Department of Hepatobiliary Surgery, First People's Hospital of Kunming City, Kunming, Yunnan 650034, P.R. China
Published online on: February 21, 2020 https://doi.org/10.3892/mmr.2020.10997
Pages: 1861-1871
Copyright: © Li et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

GLI family zinc finger 3 (Gli3), as the upstream transcriptional activator of hedgehog signaling, has previously been demonstrated to participate in the process of liver fibrosis. The present study aimed to investigate the potential functions of microRNA (miR)‑200a and Gli3 in the progression of liver fibrosis. The expression levels of miR‑200a and Gli3 in cells and tissues were determined by PCR and western blotting; the interaction of Gli3 and miR‑200a was evaluated by bioinformatics analysis and dual‑luciferase reporter assay. miR‑200a was significantly reduced in serum samples from clinical patients, liver tissues of a carbon tetrachloride (CCl4)‑induced rat model and activated LX2 cells. The expression of α‑smooth muscle actin (α‑SMA) and albumin at the mRNA and protein levels was increased and decreased in LX2 cells, respectively. However, the expression levels of α‑SMA and albumin were reversed and Gli3 expression was markedly decreased in LX2 cells when transfected with miR‑200a mimics. In addition, the dual‑luciferase reporter assay confirmed the target interaction between miR‑200a and Gli3. Finally, following the administration of miR‑200a mimics to CCl4‑induced rats, it was revealed that the alterations of α‑SMA, albumin and Gli3 presented a similar trend to that in LX2 cells with miR‑200a mimics transfection. Taken together, these results indicated that downregulation of miR‑200a might enhance the formation of liver fibrosis, probably by targeting Gli3, and elevated miR‑200a may attenuate the progression of liver fibrosis by suppressing Gli3. These findings suggested that miR‑200a may function as a novel anti‑ﬁbrotic agent in liver fibrosis via inhibition of the expression of Gli3.

View Figures

View References

1	Lee UE and Friedman SL: Mechanisms of hepatic fibrogenesis. Best Pract Res Clin Gastroenterol. 25:195–206. 2011. View Article : Google Scholar : PubMed/NCBI
2	Elpek GÖ: Cellular and molecular mechanisms in the pathogenesis of liver fibrosis: An update. World J Gastroenterol. 20:7260–7276. 2014. View Article : Google Scholar : PubMed/NCBI
3	Friedman SL: Mechanisms of disease: Mechanisms of hepatic fibrosis and therapeutic implications. Nat Clin Pract Gastroenterol Hepatol. 1:98–105. 2004. View Article : Google Scholar : PubMed/NCBI
4	Coletta M, Nicolini D, Benedetti Cacciaguerra A, Mazzocato S, Rossi R and Vivarelli M: Bridging patients with hepatocellular cancer waiting for liver transplant: All the patients are the same? Transl Gastroenterol Hepatol. 2:782017. View Article : Google Scholar : PubMed/NCBI
5	Lim R, Ricardo SD and Sievert W: Cell-based therapies for tissue fibrosis. Front Pharmacol. 8:6332017. View Article : Google Scholar : PubMed/NCBI
6	Treiber T, Treiber N, Plessmann U, Harlander S, Daiß JL, Eichner N, Lehmann G, Schall K, Urlaub H and Meister G: A compendium of RNA-binding proteins that regulate MicroRNA biogenesis. Mol Cell. 66:270–284 e13. 2017. View Article : Google Scholar : PubMed/NCBI
7	Liu X, Fortin K and Mourelatos Z: MicroRNAs: Biogenesis and molecular functions. Brain Pathol. 18:113–121. 2008. View Article : Google Scholar : PubMed/NCBI
8	Murakami Y and Kawada N: MicroRNAs in hepatic pathophysiology. Hepatol Res. 47:60–69. 2017. View Article : Google Scholar : PubMed/NCBI
9	Roderburg C, Urban GW, Bettermann K, Bettermann K, Vucur M, Zimmermann H, Schmidt S, Janssen J, Koppe C, Knolle P, et al: Micro-RNA profiling reveals a role for miR-29 in human and murine liver fibrosis. Hepatology. 53:209–218. 2011. View Article : Google Scholar : PubMed/NCBI
10	Sekiya Y, Ogawa T, Yoshizato K, Ikeda K and Kawada N: Suppression of hepatic stellate cell activation by microRNA-29b. Biochem Biophys Res Commun. 412:74–79. 2011. View Article : Google Scholar : PubMed/NCBI
11	Ogawa T, Iizuka M, Sekiya Y, Yoshizato K, Ikeda K and Kawada N: Suppression of type I collagen production by microRNA-29b in cultured human stellate cells. Biochem Biophys Res Commun. 391:316–321. 2010. View Article : Google Scholar : PubMed/NCBI
12	Li J, Ghazwani M, Zhang Y, Lu J, Li J, Fan J, Gandhi CR and Li S: miR-122 regulates collagen production via targeting hepatic stellate cells and suppressing P4HA1 expression. J Hepatol. 58:522–528. 2013. View Article : Google Scholar : PubMed/NCBI
13	Zhao J, Tang N, Wu K, Dai W, Ye C, Shi J, Zhang J, Ning B, Zeng X and Lin Y: MiR-21 simultaneously regulates ERK1 signaling in HSC activation and hepatocyte EMT in hepatic fibrosis. PLoS On. 9:e1080052014. View Article : Google Scholar
14	He Y, Huang C, Zhang SP, Sun X, Long XR and Li J: The potential of microRNAs in liver fibrosis. Cell Signal. 24:2268–2272. 2012. View Article : Google Scholar : PubMed/NCBI
15	Yang S, Banerjee S, de Freitas A, Sanders YY, Ding Q, Matalon S, Thannickal VJ, Abraham E and Liu G: Participation of miR-200 in pulmonary fibrosis. Am J Pathol. 180:484–493. 2012. View Article : Google Scholar : PubMed/NCBI
16	Xu M, Wang G, Zhou H, Cai J, Li P, Zhou M, Lu Y, Jiang X, Huang H, Zhang Y and Gong A: TGF-β1-miR-200a-PTEN induces epithelial-mesenchymal transition and fibrosis of pancreatic stellate cells. Mol Cell Biochem. 431:161–168. 2017. View Article : Google Scholar : PubMed/NCBI
17	Yang JJ, Tao H, Hu W, Liu LP, Shi KH, Deng ZY and Li J: MicroRNA-200a controls Nrf2 activation by target Keap1 in hepatic stellate cell proliferation and fibrosis. Cell Signal. 26:2381–2389. 2014. View Article : Google Scholar : PubMed/NCBI
18	Sun X, He Y, Ma TT, Huang C, Zhang L and Li J: Participation of miR-200a in TGF-β1-mediated hepatic stellate cell activation. Mol Cell Biochem. 388:11–23. 2014. View Article : Google Scholar : PubMed/NCBI
19	Riobo NA and Manning DR: Pathways of signal transduction employed by vertebrate Hedgehogs. Biochem J. 403:369–379. 2007. View Article : Google Scholar : PubMed/NCBI
20	Bloomsmith MA, Perlman JE, Hutchinson E and Sharpless M: Behavioral management programs to promote laboratory animal welfare. Weichbrod RH, Thompson GAH and Norton JN: Management of animal care and use programs in research, education, and testing. 2nd. Boca Raton (FL): CRC Press/Taylor & Francis. Chapter 5. 2018
21	Prestigiacomo V, Weston A, Messner S, Lampart F and Suter-Dick L: Pro-fibrotic compounds induce stellate cell activation, ECM-remodelling and Nrf2 activation in a human 3D-multicellular model of liver fibrosis. PLoS One. 12:e01799952017. View Article : Google Scholar : PubMed/NCBI
22	Park EK, Jung HS, Yang HI, Yoo MC, Kim C and Kim KS: Optimized THP-1 differentiation is required for the detection of responses to weak stimuli. Inflamm Res. 56:45–50. 2007. View Article : Google Scholar : PubMed/NCBI
23	Zhou L, Dong X, Wang L, Shan L, Li T, Xu W, Ding Y, Lai M, Lin X, Dai M, et al: Casticin attenuates liver fibrosis and hepatic stellate cell activation by blocking TGF-β/Smad signaling pathway. Oncotarget. 8:56267–56280. 2017.PubMed/NCBI
24	Knight V, Lourensz D, Tchongue J, Correia J, Tipping P and Sievert W: Cytoplasmic domain of tissue factor promotes liver fibrosis in rat. World J Gastroenterol. 23:5692–5699. 2017. View Article : Google Scholar : PubMed/NCBI
25	Ishak K, Baptista A, Bianchi L, Callea F, De Groote J, Gudat F, Denk H, Desmet V, Korb G, MacSween RN, et al: Histological grading and staging of chronic hepatitis. J Hepatol. 22:696–699. 1995. View Article : Google Scholar : PubMed/NCBI
26	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 : PubMed/NCBI
27	Rasheed SA, Teo CR, Beillard EJ, Voorhoeve PM and Casey PJ: MicroRNA-182 and microRNA-200a control G-protein subunit α-13 (GNA13) expression and cell invasion synergistically in prostate cancer cells. J Biol Chem. 288:7986–7995. 2013. View Article : Google Scholar : PubMed/NCBI
28	Kim JY, An HJ, Kim WH, Gwon MG, Gu H, Park YY and Park KK: Anti-fibrotic effects of synthetic oligodeoxynucleotide for TGF-β1 and smad in an animal model of liver cirrhosis. Mol Ther Nucleic Acids. 8:250–263. 2017. View Article : Google Scholar : PubMed/NCBI
29	King A, Houlihan DD, Kavanagh D, Haldar D, Luu N, Owen A, Suresh S, Than NN, Reynolds G, Penny J, et al: Sphingosine-1-phosphate prevents egress of hematopoietic stem cells from liver to reduce fibrosis. Gastroenterology. 153:233–248.e16. 2017. View Article : Google Scholar : PubMed/NCBI
30	Xu F, Han Y, Zhu D, Tian H, Zhu H, Ren J, Gu D and Duan Y: Construction of a recombinant pIRES2-EGFP-ARTS plasmid and its effect on LX-2 cells. Mol Med Rep. 16:4737–4743. 2017. View Article : Google Scholar : PubMed/NCBI
31	Jiang XP, Ai WB, Wan LY, Zhang YQ and Wu JF: The roles of microRNA families in hepatic fibrosis. Cell Biosci. 7:342017. View Article : Google Scholar : PubMed/NCBI
32	Zhou DD, Wang X, Wang Y, Xiang XJ, Liang ZC, Zhou Y, Xu A, Bi CH and Zhang L: MicroRNA-145 inhibits hepatic stellate cell activation and proliferation by targeting ZEB2 through Wnt/β-catenin pathway. Mol Immunol. 75:151–160. 2016. View Article : Google Scholar : PubMed/NCBI
33	Dong R, Zheng Y, Chen G, Zhao R, Zhou Z and Zheng S: miR-222 overexpression may contribute to liver fibrosis in biliary atresia by targeting PPP2R2A. J Pediatr Gastroenterol Nutr. 60:84–90. 2015. View Article : Google Scholar : PubMed/NCBI
34	Tsuchida T and Friedman SL: Mechanisms of hepatic stellate cell activation. Nat Rev Gastroenterol Hepatol. 14:397–411. 2017. View Article : Google Scholar : PubMed/NCBI
35	Zhang CY, Yuan WG, He P, Lei JH and Wang CX: Liver fibrosis and hepatic stellate cells: Etiology, pathological hallmarks and therapeutic targets. World J Gastroenterol. 22:10512–10522. 2016. View Article : Google Scholar : PubMed/NCBI
36	Friedman RC, Farh KK, Burge CB and Bartel DP: Most mammalian mRNAs are conserved targets of microRNAs. Genome Res. 19:92–105. 2009. View Article : Google Scholar : PubMed/NCBI
37	Xia H, Ng SS, Jiang S, Cheung WK, Sze J, Bian XW, Kung HF and Lin MC: miR-200a-mediated downregulation of ZEB2 and CTNNB1 differentially inhibits nasopharyngeal carcinoma cell growth, migration and invasion. Biochem Biophys Res Commun. 391:535–541. 2010. View Article : Google Scholar : PubMed/NCBI
38	Ding XQ, Wu WY, Jiao RQ, Gu TT, Xu Q, Pan Y and Kong LD: Curcumin and allopurinol ameliorate fructose-induced hepatic inflammation in rats via miR-200a-mediated TXNIP/NLRP3 inflammasome. Pharmacol Res. 137:64–75. 2018. View Article : Google Scholar : PubMed/NCBI
39	Saha S, Choudhury J and Ain R: MicroRNA-141-3p and miR-200a-3p regulate insulin-like growth factor 2 during mouse placental development. Mol Cell Endocrinol. 414:186–193. 2015. View Article : Google Scholar : PubMed/NCBI
40	Hyun J, Wang S, Kim J, Rao KM, Park SY, Chung I, Ha CS, Kim SW, Yun YH and Jung Y: MicroRNA-378 limits activation of hepatic stellate cells and liver fibrosis by suppressing Gli3 expression. Nat Commun. 7:109932016. View Article : Google Scholar : PubMed/NCBI
41	Trnski D, Sabol M, Gojević A, Martinić M, Ozretić P, Musani V, Ramić S and Levanat S: GSK3β and Gli3 play a role in activation of Hedgehog-Gli pathway in human colon cancer-Targeting GSK3β downregulates the signaling pathway and reduces cell proliferation. Biochim Biophys Acta. 1852:2574–2584. 2015. View Article : Google Scholar : PubMed/NCBI
42	Shen X, Peng Y and Li H: The injury-related activation of hedgehog signaling pathway modulates the repair-associated inflammation in liver fibrosis. Front Immunol. 8:14502017. View Article : Google Scholar : PubMed/NCBI
43	Machado MV and Diehl AM: Hedgehog signalling in liver pathophysiology. J Hepatol. 68:550–562. 2018. View Article : Google Scholar : PubMed/NCBI