MicroRNA‑218 aggravates H<sub>2</sub>O<sub>2</sub>‑induced damage in PC12 cells via spred2‑mediated autophagy

Chen,Duoping; Li,Chunmei; Lv,Rui

doi:10.3892/etm.2021.10787

MicroRNA‑218 aggravates H₂O₂‑induced damage in PC12 cells via spred2‑mediated autophagy

Authors:
- Duoping Chen
- Chunmei Li
- Rui Lv
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Affiliations: Second Department of Orthopaedics, Zhang Ye People's Hospital Affiliated to Hexi University, Zhangye, Gansu 734000, P.R. China, Image Center, Zhang Ye People's Hospital Affiliated to Hexi University, Zhangye, Gansu 734000, P.R. China
Published online on: September 23, 2021 https://doi.org/10.3892/etm.2021.10787
Article Number: 1352
Copyright: © Chen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

The current study aimed to investigate the effects and underlying mechanism of miR‑218 in H₂O₂‑induced neuronal injury. The impacts of miR‑218 knockdown on cell viability, apoptosis and autophagy‑associated proteins were detected by Cell Counting Kit‑8 assay, flow cytometry and western blotting in H₂O₂‑injured PC12 cells, respectively. Reverse transcription‑quantitative PCR (RT‑qPCR) and western blotting was executed to explore the expression level of miR‑218 and sprouty‑related EVH1 domainprotein2 (spred2) in H₂O₂‑stimulated cells. Besides, the regulatory association between miR‑218 and spred2 was explored through bioinformatics and luciferase reporter assay. Following knockdown of miR‑218 and spred2, the functions of miR‑218 and spred2 in H₂O₂‑injured cells were further studied. High expression level of miR‑218 was observed in H₂O₂‑disposed PC12 cells, while spred2 expression level was downregulated. Knockdown of miR‑218 expression alleviated H₂O₂-induced PC12 cell injury by increasing cell proliferation, and decreasing apoptosis and autophagy. Furthermore, spred2 was identified as a direct target of miR-218 and was negatively regulated by miR-218. Moreover, suppression of spred2 abrogated the protective effects of miR‑218 inhibition on H₂O₂‑injured PC12 cells. Depletion of miR‑218 protected PC12 cells against H₂O₂‑induced cell injury via the upregulation of spred2, which provided a promising therapeutic strategy for spinal cord injury.

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1	Schwab JM, Maas AIR, Hsieh JTC and Curt A: Raising awareness for spinal cord injury research. Lancet Neurol. 17:581–582. 2018.PubMed/NCBI View Article : Google Scholar
2	Jendelova P: Therapeutic strategies for spinal cord injury. Int J Mol Sci. 19(3200)2018.PubMed/NCBI View Article : Google Scholar
3	Venkatesh K, Ghosh SK, Mullick M, Manivasagam G and Sen D: Spinal cord injury: Pathophysiology, treatment strategies, associated challenges, and future implications. Cell Tissue Res. 377:125–151. 2019.PubMed/NCBI View Article : Google Scholar
4	Wang JL, Luo X and Liu L: Targeting CARD6 attenuates spinal cord injury (SCI) in mice through inhibiting apoptosis, inflammation and oxidative stress associated ROS production. Aging (Albany NY). 11:12213–12235. 2019.PubMed/NCBI View Article : Google Scholar
5	Guan B, Chen R, Zhong M, Liu N and Chen Q: Protective effect of oxymatrine against acute spinal cord injury in rats via modulating oxidative stress, inflammation and apoptosis. Metab Brain Dis. 35:149–157. 2020.PubMed/NCBI View Article : Google Scholar
6	Lu TX and Rothenberg ME: MicroRNA. J Allergy Clin Immunol. 141:1202–1207. 2018.PubMed/NCBI View Article : Google Scholar
7	Vishnoi A and Rani S: miRNA biogenesis and regulation of diseases: An overview. Methods Mol Biol. 1509:1–10. 2017.PubMed/NCBI View Article : Google Scholar
8	Jiao S, Liu Y, Yao Y and Teng J: miR-124 promotes proliferation and neural differentiation of neural stem cells through targeting DACT1 and activating Wnt/β-catenin pathways. Mol Cell Biochem. 449:305–314. 2018.PubMed/NCBI View Article : Google Scholar
9	Yuan Z, Zhong L, Liu D, Yao J, Liu J, Zhong P, Yao S, Zhao Y, Li L, Chen M, et al: miR-15b regulates cell differentiation and survival by targeting CCNE1 in APL cell lines. Cell Signal. 60:57–64. 2019.PubMed/NCBI View Article : Google Scholar
10	Dai J, Xu LJ, Han GD, Sun HL, Zhu GT, Jiang HT, Yu GY and Tang XM: miR-137 attenuates spinal cord injury by modulating NEUROD4 through reducing inflammation and oxidative stress. Eur Rev Med Pharmacol Sci. 22:1884–1890. 2018.PubMed/NCBI View Article : Google Scholar
11	Jiao G, Pan B, Zhou Z, Zhou L, Li Z and Zhang Z: MicroRNA-21 regulates cell proliferation and apoptosis in H₂O₂-stimulated rat spinal cord neurons. Mol Med Rep. 12:7011–7016. 2015.PubMed/NCBI View Article : Google Scholar
12	Liu D and Bao F: Hydrogen peroxide administered into the rat spinal cord at the level elevated by contusion spinal cord injury oxidizes proteins, DNA and membrane phospholipids, and induces cell death: Attenuation by a metalloporphyrin. Neuroscience. 285:81–96. 2015.PubMed/NCBI View Article : Google Scholar
13	Zhu H, Wang X and Chen S: Downregulation of miR-218-5p protects against oxygen-glucose deprivation/reperfusion-induced injuries of PC12 cells via upregulating N-myc downstream regulated gene 4 (NDRG4). Med Sci Monit. 26(e920101)2020.PubMed/NCBI View Article : Google Scholar
14	Zhu L, Tu H, Liang Y and Tang D: miR-218 produces anti-tumor effects on cervical cancer cells in vitro. World J Surg Oncol. 16(204)2018.PubMed/NCBI View Article : Google Scholar
15	Cao Q, Dong P and Wang Y, Zhang J, Shi X and Wang Y: miR-218 suppresses cardiac myxoma proliferation by targeting myocyte enhancer factor 2D. Oncol Rep. 33:2606–2612. 2015.PubMed/NCBI View Article : Google Scholar
16	Li L and Zhao G: Downregulation of microRNA-218 relieves neuropathic pain by regulating suppressor of cytokine signaling 3. Int J Mol Med. 37:851–858. 2016.PubMed/NCBI View Article : Google Scholar
17	Yin D, Zheng X, Zhuang J, Wang L, Liu B and Chang Y: Downregulation of long noncoding RNA Sox2ot protects PC-12 cells from hydrogen peroxide-induced injury in spinal cord injury via regulating the miR-211-myeloid cell leukemia-1 isoform2 axis. J Cell Biochem. 119:9675–9684. 2018.PubMed/NCBI View Article : Google Scholar
18	Guo Z, Li L, Gao Y, Zhang X and Cheng M: Overexpression of lncRNA ANRIL aggravated hydrogen peroxide-disposed injury in PC-12 cells via inhibiting miR-499a/PDCD4 axis-mediated PI3K/Akt/mTOR/p70S6K pathway. Artif Cells Nanomed Biotechnol. 47:2624–2633. 2019.PubMed/NCBI View Article : Google Scholar
19	Zhang Y, Sun B, Zhao L, Liu Z, Xu Z, Tian Y and Hao C: Up-regulation of miRNA-148a inhibits proliferation, invasion, and migration while promoting apoptosis of cervical cancer cells by down-regulating RRS1. Biosci Rep. 39(BSR20181815)2019.PubMed/NCBI View Article : Google Scholar
20	Eckert MJ and Martin MJ: Trauma: Spinal cord injury. Surg Clin North Am. 97:1031–1045. 2017.PubMed/NCBI View Article : Google Scholar
21	Li Z, Wu F, Xu D, Zhi Z and Xu G: Inhibition of TREM1 reduces inflammation and oxidative stress after spinal cord injury (SCI) associated with HO-1 expressions. Biomed Pharmacother. 109:2014–2021. 2019.PubMed/NCBI View Article : Google Scholar
22	Abou-El-Hassan H, Bsat S, Sukhon F, Assaf EJ, Mondello S, Kobeissy F, Wang KKW, Weiner HL and Omeis I: Protein degradome of spinal cord injury: Biomarkers and potential therapeutic targets. Mol Neurobiol. 57:2702–2726. 2020.PubMed/NCBI View Article : Google Scholar
23	Pinchi E, Frati A, Cantatore S, D'Errico S, Russa R, Maiese A, Palmieri M, Pesce A, Viola RV, Frati P and Fineschi V: Acute spinal cord injury: A systematic review investigating miRNA FAMILIES INVolved. Int J Mol Sci. 20(1841)2019.PubMed/NCBI View Article : Google Scholar
24	Lu W, Wan X, Tao L and Wan J: Long non-coding RNA HULC promotes cervical cancer cell proliferation, migration and invasion via miR-218/TPD52 axis. Onco Targets Ther. 13:1109–1118. 2020.PubMed/NCBI View Article : Google Scholar
25	Li XC, Hai JJ, Tan YJ, Yue QF and Liu LJ: miR-218 suppresses metastasis and invasion of endometrial cancer via negatively regulating ADD2. Eur Rev Med Pharmacol Sci. 23:1408–1417. 2019.PubMed/NCBI View Article : Google Scholar
26	Wen YD, Wang H, Kho SH, Rinkiko S, Sheng X, Shen HM and Zhu YZ: Hydrogen sulfide protects HUVECs against hydrogen peroxide induced mitochondrial dysfunction and oxidative stress. PLoS One. 8(e53147)2013.PubMed/NCBI View Article : Google Scholar
27	Ravanan P, Srikumar IF and Talwar P: Autophagy: The spotlight for cellular stress responses. Life Sci. 188:53–67. 2017.PubMed/NCBI View Article : Google Scholar
28	Parzych KR and Klionsky DJ: An overview of autophagy: Morphology, mechanism, and regulation. Antioxid Redox Signal. 20:460–473. 2014.PubMed/NCBI View Article : Google Scholar
29	Ray SK: Modulation of autophagy for neuroprotection and functional recovery in traumatic spinal cord injury. Neural Regen Res. 15:1601–1612. 2020.PubMed/NCBI View Article : Google Scholar
30	Yoshimura A: Regulation of cytokine signaling by the SOCS and Spred family proteins. Keio J Med. 58:73–83. 2009.PubMed/NCBI View Article : Google Scholar
31	Takahashi S, Yoshimura T, Ohkura T, Fujisawa M, Fushimi S, Ito T, Itakura J, Hiraoka S, Okada H, Yamamoto K and Matsukawa A: A novel role of spred2 in the colonic epithelial cell homeostasis and inflammation. Sci Rep. 6(37531)2016.PubMed/NCBI View Article : Google Scholar
32	Jiang K, Liu M, Lin G, Mao B, Cheng W, Liu H, Gal J, Zhu H, Yuan Z, Deng W, et al: Tumor suppressor spred2 interaction with LC3 promotes autophagosome maturation and induces autophagy-dependent cell death. Oncotarget. 7:25652–25667. 2016.PubMed/NCBI View Article : Google Scholar
33	Ullrich M, Aßmus B, Augustin AM, Häbich H, Abeßer M, Martin Machado J, Werner F, Erkens R, Arias-Loza AP, Umbenhauer S, et al: Spred2 deficiency elicits cardiac arrhythmias and premature death via impaired autophagy. J Mol Cell Cardiol. 129:13–26. 2019.PubMed/NCBI View Article : Google Scholar
34	Mardakheh FK, Yekezare M, Machesky LM and Heath JK: Spred2 interaction with the late endosomal protein NBR1 down-regulates fibroblast growth factor receptor signaling. J Cell Biol. 187:265–277. 2009.PubMed/NCBI View Article : Google Scholar
35	Chen KC, Liao YC, Wang JY, Lin YC, Chen CH and Juo SH: Oxidized low-density lipoprotein is a common risk factor for cardiovascular diseases and gastroenterological cancers via epigenomical regulation of microRNA-210. Oncotarget. 6:24105–24118. 2015.PubMed/NCBI View Article : Google Scholar
36	Xu Y, Ito T, Fushimi S, Takahashi S, Itakura J, Kimura R, Sato M, Mino M, Yoshimura A and Matsukawa A: Spred-2 deficiency exacerbates lipopolysaccharide-induced acute lung inflammation in mice. PLoS One. 9(e108914)2014.PubMed/NCBI View Article : Google Scholar
37	Hu L, Ye H and Liao J: LncRNA TUG1 reverses LPS-induced cell apoptosis and inflammation of macrophage via targeting miR-221-3p/SPRED2 axis. Biosci Biotechnol Biochem. 84:2458–2465. 2020.PubMed/NCBI View Article : Google Scholar
38	Peng W, Li J, Chen R, Gu Q, Yang P, Qian W, Ji D, Wang Q, Zhang Z, Tang J and Sun Y: Upregulated METTL3 promotes metastasis of colorectal cancer via miR-1246/SPRED2/MAPK signaling pathway. J Exp Clin Cancer Res. 38(393)2019.PubMed/NCBI View Article : Google Scholar
39	Sung YJ, Cheng CL, Chen CS, Huang HB, Huang FL, Wu PC, Shiao MS and Tsay HJ: Distinct mechanisms account for beta-amyloid toxicity in PC12 and differentiated PC12 neuronal cells. J Biomed Sci. 10:379–388. 2003.PubMed/NCBI View Article : Google Scholar