MicroRNA-29a suppresses the invasion and migration of osteosarcoma cells by regulating the SOCS1/NF-κB signalling pathway through negatively targeting DNMT3B

Gong,Hao‑Li; Tao,Ye; Mao,Xin‑Zhan; Song,De‑Ye; You,Di; Ni,Jiang‑Dong

doi:10.3892/ijmm.2019.4287

MicroRNA-29a suppresses the invasion and migration of osteosarcoma cells by regulating the SOCS1/NF-κB signalling pathway through negatively targeting DNMT3B

Authors:
- Hao‑Li Gong
- Ye Tao
- Xin‑Zhan Mao
- De‑Ye Song
- Di You
- Jiang‑Dong Ni
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Affiliations: Department of Orthopedics, The Second Xiangya Hospital, Central South University, Changsha, Hunan 410008, P.R. China, Department of Radiology, The Third Xiangya Hospital, Central South University, Changsha, Hunan 410013, P.R. China
Published online on: July 24, 2019 https://doi.org/10.3892/ijmm.2019.4287
Pages: 1219-1232
Copyright: © Gong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

The present study aimed to investigate the roles of the microRNA‑29a/DNA methyltransferase 3B/suppressor of cytokine signalling 1 (miR‑29a/DNMT3B/SOCS1) axis in the invasion and the migration of osteosarcoma (OS). The expression levels of miR‑29a, DNMT3B and SOCS1 were determined in tissue samples and OS cell lines by reverse transcription‑quantitative polymerase chain reaction (PCR). Apoptosis was measured using flow cytometry analysis. Transwell and wound healing assays were conducted to measure the invasion and migration abilities of OS cells, respectively. A dual‑luciferase reporter assay was also conducted to determine the interaction between DNMT3B and miR‑29a, while methylation‑specific PCR was used to detect the methylation of SOCS1. Western blotting was performed to determine the protein levels of DNMT3B and SOCS1, as well as the levels of proteins associated with epithelial‑mesenchymal transition (EMT), apoptosis and the nuclear factor (NF)‑κB signalling pathway. The results demonstrated that miR‑29a and SOCS1 were downregulated, and DNMT3B was upregulated in both OS tissues and cell lines. The expression of miR‑29a and SOCS1 was found to be associated with advanced clinical stage and distant metastasis. In addition, the dual‑luciferase reporter assay revealed that DNMT3B was a direct target of miR‑29a. Overexpression using miR‑29a mimics decreased DNMT3B expression and the methylation level of SOCS1, which resulted in the upregulation of SOCS1 in U2OS and MG‑63 cells, while miR‑29a inhibition led to the opposite results. Transfection with miR‑29a mimics also promoted the apoptosis, and inhibited the invasion, migration and EMT process of OS cells, while it markedly reduced the nuclear translocation of p65 and IκB‑α degradation. Treatment with 5‑aza‑2'‑deoxycytidine worked together with miR‑29a mimics to synergistically enhance the aforementioned effects. By contrast, the effects induced by miR‑29a were partly reversed upon co‑transfection with SOCS1 siRNA. In conclusion, miR‑29a promoted the apoptosis, and inhibited the invasion, migration and EMT process of OS cells via inhibition of the SOCS1/NF‑κB signalling pathway by directly targeting DNMT3B.

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1	Mirabello L, Troisi RJ and Savage SA: Osteosarcoma incidence and survival rates from 1973 to 2004: Data from the surveillance, epidemiology, and end results program. Cancer. 115:1531–1543. 2009. View Article : Google Scholar : PubMed/NCBI
2	Luetke A, Meyers PA, Lewis I and Juergens H: Osteosarcoma treatment-where do we stand? A state of the art review. Cancer Treat Rev. 40:523–532. 2014. View Article : Google Scholar
3	Broadhead ML, Clark JC, Myers DE, Dass CR and Choong PF: The molecular pathogenesis of osteosarcoma: A review. Sarcoma. 2011:9592482011. View Article : Google Scholar : PubMed/NCBI
4	Kansara M, Teng MW, Smyth MJ and Thomas DM: Translational biology of osteosarcoma. Nat Rev Cancer. 14:722–735. 2014. View Article : Google Scholar : PubMed/NCBI
5	Di Fiore R, Drago-Ferrante R, Pentimalli F, Di Marzo D, Forte IM, D'Anneo A, Carlisi D, De Blasio A, Giuliano M, Tesoriere G, et al: MicroRNA-29b-1 impairs in vitro cell proliferation, self-renewal and chemoresistance of human osteosarcoma 3AB-OS cancer stem cells. Int J Oncol. 45:2013–2023. 2014. View Article : Google Scholar : PubMed/NCBI
6	Durfee RA, Mohammed M and Luu HH: Review of osteosarcoma and current management. Rheumatol Ther. 3:221–243. 2016. View Article : Google Scholar : PubMed/NCBI
7	Lin S and Gregory RI: MicroRNA biogenesis pathways in cancer. Nat Rev Cancer. 15:321–333. 2015. View Article : Google Scholar : PubMed/NCBI
8	Xie B, Ding Q, Han H and Wu D: miRCancer: A microRNA-cancer association database constructed by text mining on literature. Bioinformatics. 29:638–644. 2013. View Article : Google Scholar : PubMed/NCBI
9	Dinh TK, Fendler W, Chałubińska-Fendler J, Acharya SS, O'Leary C, Deraska PV, D'Andrea AD, Chowdhury D and Kozono D: Circulating miR-29a and miR-150 correlate with delivered dose during thoracic radiation therapy for non-small cell lung cancer. Radiat Oncol. 11:612016. View Article : Google Scholar : PubMed/NCBI
10	Pasqualini L, Bu H, Puhr M, Narisu N, Rainer J, Schlick B, Schäfer G, Angelova M, Trajanoski Z, Börno ST, et al: miR-22 and miR-29a are members of the androgen receptor cistrome modulating LAMC1 and Mcl-1 in prostate cancer. Mol Endocrinol. 29:1037–1054. 2015. View Article : Google Scholar : PubMed/NCBI
11	Zhang H, Bai M, Deng T, Liu R, Wang X, Qu Y, Duan J, Zhang L, Ning T, Ge S, et al: Cell-derived microvesicles mediate the delivery of miR-29a/c to suppress angiogenesis in gastric carcinoma. Cancer Lett. 375:331–339. 2016. View Article : Google Scholar : PubMed/NCBI
12	Sampson VB, Yoo S, Kumar A, Vetter NS and Kolb EA: MicroRNAs and potential targets in osteosarcoma: Review. Front Pediatr. 3:692015. View Article : Google Scholar : PubMed/NCBI
13	He DX, Gu XT, Li YR, Jiang L, Jin J and Ma X: Methylation- regulated miR-149 modulates chemoresistance by targeting GlcNAc N-deacetylase/N-sulfotransferase-1 in human breast cancer. FEBS J. 281:4718–4730. 2014. View Article : Google Scholar : PubMed/NCBI
14	Li X, Pan Q, Wan X, Mao Y, Lu W, Xie X and Cheng X: Methylation-associated Has-miR-9 deregulation in paclitaxel- resistant epithelial ovarian carcinoma. BMC Cancer. 15:5092015. View Article : Google Scholar : PubMed/NCBI
15	Liu YL, Lensing SY, Yan Y, Webster C and Emanuel PD: Overexpression Of DNMT3a and DNMT3b is related to down-regulation of Mir-29a in juvenile myelomonocytic leukemia (JMML). Blood. 122:48902013.
16	Babon JJ and Nicola NA: The biology and mechanism of action of suppressor of cytokine signaling 3. Growth Factors. 30:207–219. 2012. View Article : Google Scholar : PubMed/NCBI
17	Beaurivage C, Champagne A, Tobelaim WS, Pomerleau V, Menendez A and Saucier C: SOCS1 in cancer: An oncogene and a tumor suppressor. Cytokine. 82:87–94. 2016. View Article : Google Scholar : PubMed/NCBI
18	Zhang XH, Yang L, Liu XJ, Zhan Y, Pan YX, Wang XZ and Luo JM: Association between methylation of tumor suppressor gene SOCS1 and acute myeloid leukemia. Oncol Rep. 40:1008–1016. 2018.PubMed/NCBI
19	Huang FJ, Steeg P, Price J, Sawaya R and Huang S: Suppressor of cytokine signaling-1 (SOCS-1) negatively regulates angiogenesis, invasion, and brain metastasis of melanoma cells. Cancer Res. 68:2008. View Article : Google Scholar
20	Shao N, Ma G, Zhang J and Zhu W: miR-221-5p enhances cell proliferation and metastasis through post-transcriptional regulation of SOCS1 in human prostate cancer. BMC Urol. 18:142018. View Article : Google Scholar : PubMed/NCBI
21	Hashimoto M, Ayada T, Kinjyo I, Hiwatashi K, Yoshida H, Okada Y, Kobayashi T and Yoshimura A: Silencing of SOCS1 in macrophages suppresses tumor development by enhancing antitumor inflammation. Cancer Sci. 100:730–736. 2009. View Article : Google Scholar : PubMed/NCBI
22	Gaspericampani A, Pancotti F and Roncuzzi L: Abstract 4396: Caveolin-1 as an oncopromoter in solidtumors: A role mediated by STAT3 in vitro. Cancer Res. 73(8 Suppl): S4396. 2014.
23	Zhang H, Zhao Z, Pang X, Yang J, Yu H, Zhang Y, Zhou H and Zhao J: Genistein protects against Ox-LDL-induced inflammation through microRNA-155/SOCS1-mediated repression of NF-ĸB signaling pathway in HUVECs. Inflammation. 40:1450–1459. 2017. View Article : Google Scholar : PubMed/NCBI
24	Lee YJ, Han JY, Byun J, Park HJ, Park EM, Chong YH, Cho MS and Kang JL: Inhibiting Mer receptor tyrosine kinase suppresses STAT1, SOCS1/3, and NF-κB activation and enhances inflammatory responses in lipopolysaccharide-induced acute lung injury. J Leukoc Biol. 91:921–932. 2012. View Article : Google Scholar : PubMed/NCBI
25	Ren D, Yang Q, Dai Y, Guo W, Du H, Song L and Peng X: Oncogenic miR-210-3p promotes prostate cancer cell EMT and bone metastasis via NF-κB signaling pathway. Mol Cancer. 16:1172017. View Article : Google Scholar
26	Liu X, Li J, Peng X, Lv B, Wang P, Zhao X and Yu B: Geraniin inhibits LPS-induced THP-1 macrophages switching to M1 phenotype via SOCS1/NF-κB pathway. Inflammation. 39:1421–1433. 2016. View Article : Google Scholar : PubMed/NCBI
27	Maier HJ, Schmidt-Strassburger U, Huber MA, Wiedemann EM, Beug H and Wirth T: NF-kappaB promotes epithelial-mesenchymal transition, migration and invasion of pancreatic carcinoma cells. Cancer Lett. 295:214–228. 2010. View Article : Google Scholar : PubMed/NCBI
28	Wu Y and Zhou BP: TNF-alpha/NF-kappaB/Snail pathway in cancer cell migration and invasion. Br J Cancer. 102:639–644. 2010. View Article : Google Scholar : PubMed/NCBI
29	Meister P, Konrad E, Lob G, Janka G, Keyl W and Stürz H: Osteosarcoma: Histological evaluation and grading. Arch Orthop Trauma Surg. 94:91–98. 1979. View Article : Google Scholar : PubMed/NCBI
30	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
31	Brunet Vega A, Pericay C, Moya I, Ferrer A, Dotor E, Pisa A, Casalots A, Serra-Aracil X, Oliva JC, Ruiz A and Saigí E: microRNA expression profile in stage III colorectal cancer: Circulating miR-18a and miR-29a as promising biomarkers. Oncol Rep. 30:320–326. 2013. View Article : Google Scholar : PubMed/NCBI
32	Zhang W, Qian JX, Yi HL, Yang ZD, Wang CF, Chen JY, Wei XZ, Fu Q and Ma H: The microRNA-29 plays a central role in osteosarcoma pathogenesis and progression. Mol Biol (Mosk). 46:622–627. 2012. View Article : Google Scholar
33	Lv Y, Song G and Li P: Correlation of SOCS-1 gene with onset and prognosis of breast cancer. Oncol Lett. 16:383–387. 2018.PubMed/NCBI
34	Wang X, Liu S, Cao L, Zhang T, Yue D, Wang L, Ping Y, He Q, Zhang C, Wang M, et al: miR-29a-3p suppresses cell proliferation and migration by downregulating IGF1R in hepatocellular carcinoma. Oncotarget. 8:86592–86603. 2017.PubMed/NCBI
35	Qin S, Ai F, Ji WF, Rao W, Zhang HC and Yao WJ: miR-19a promotes cell growth and tumorigenesis through targeting SOCS1 in gastric cancer. Asian Pac J Cancer Prev. 14:835–840. 2013. View Article : Google Scholar : PubMed/NCBI
36	Agarwal S, Amin KS, Jagadeesh S, Baishay G, Rao PG, Barua NC, Bhattacharya S and Banerjee PP: Mahanine restores RASSF1A expression by down-regulating DNMT1 and DNMT3B in prostate cancer cells. Mol Cancer. 12:992013. View Article : Google Scholar : PubMed/NCBI
37	Chen Q, Dan Y, Zhang Y, Yu L, Li XD, Zhou ZJ, Zhou SL, Gao DM, Hu J, Jin C, et al: MicroRNA-29a induces loss of 5-hydroxymethylcytosine and promotes metastasis of hepatocellular carcinoma through a TET-SOCS1-MMP9 signaling axis. Cell Death Dis. 8:e29062017. View Article : Google Scholar : PubMed/NCBI
38	Maugeri M, Barbagallo D, Barbagallo C, Banelli B, Di Mauro S, Purrello F, Magro G, Ragusa M, Di Pietro C, Romani M and Purrello M: Altered expression of miRNAs and methylation of their promoters are correlated in neuroblastoma. Oncotarget. 7:83330–83341. 2016. View Article : Google Scholar : PubMed/NCBI
39	Fu X, Song X, Li Y, Tan D and Liu G: Hepatitis B virus X protein upregulates DNA methyltransferase 3A/3B and enhances SOCS-1CpG island methylation. Mol Med Rep. 13:301–308. 2016. View Article : Google Scholar
40	Gebeshuber CA, Zatloukal K and Martinez J: miR-29a suppresses tristetraprolin, which is a regulator of epithelial polarity and metastasis. EMBO Rep. 10:400–405. 2009. View Article : Google Scholar : PubMed/NCBI
41	Kogure T, Kondo Y, Kakazu E, Ninomiya M, Kimura O and Shimosegawa T: Involvement of miRNA-29a in epigenetic regulation of transforming growth factor-β-induced epithelial-mesenchymal transition in hepatocellular carcinoma. Hepatol Res. 44:907–919. 2014. View Article : Google Scholar
42	Tang B, Li X, Ren Y, Wang J, Xu D, Hang Y, Zhou T, Li F and Wang L: MicroRNA-29a regulates lipopolysaccharide (LPS)-induced inflammatory responses in murine macrophages through the Akt1/NF-κB pathway. Exp Cell Res. 360:74–80. 2017. View Article : Google Scholar : PubMed/NCBI