Abstract
Neonatal hypoxic–ischemic encephalopathy (HIE) is a major cause of lifelong disabilities worldwide, without effective therapies and clear regulatory mechanisms. MicroRNAs (miRNAs) act as a significant regulator in neuroregeneration and neuronal apoptosis, thus holding great potential as therapeutic targets in HIE. In this study, we established the hypoxia–ischemia (HI) model in vivo and oxygen–glucose deprivation (OGD) model in vitro. Zea-longa score and magnetic resonance imaging were applied to verify HI-induced neuronal dysfunction and brain infarction. Subsequently, a miRNA microarray analysis was employed to profile miRNA transcriptomes. Down-regulated miR-124 was found 24 h after HIE, which corresponded to the change in PC12, SHSY5Y, and neurons after OGD. To determine the function of miR-124, mimics and lentivirus-mediated overexpression were used to regulate miR-124 in vivo and in vitro, respectively. Our results showed that miR-124 overexpression obviously promoted cell survival and suppressed neuronal apoptosis. Further, the memory and neurological function of rats was also obviously improved at 1 and 2 months after HI, indicated by the neurological severity score, Y-maze test, open field test, and rotating rod test. Our findings showed that overexpression of miR-124 can be a promising new strategy for HIE therapy in future clinical practice.
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28 December 2021
A Correction to this paper has been published: https://doi.org/10.1007/s10571-021-01178-0
Abbreviations
- DAPI:
-
4′, 6-Diamidino-2-phenylindole
- DMSO:
-
Dimethyl sulphoxide
- HTS:
-
High-throughput sequencing
- HIE:
-
Hypoxic–ischemic encephalopathy
- HI:
-
Hypoxia–ischemia
- OGD:
-
Oxygen glucose deprivation
- PBS:
-
Phosphate-buffered saline
- Q-PCR:
-
Quantitative polymerase chain reaction
- SD:
-
Sprague–Dawley
- TUNEL:
-
Terminal deoxynucleotidyl transferase-mediated nick end labeling
- MRI:
-
Magnetic resonance imaging
- miRNAs:
-
MicroRNAs
- MTT:
-
Methyl thiazolyltetrazolium
- NSS:
-
Neurological severity score
References
Åkerblom M, Sachdeva R, Barde I, Verp S, Gentner B, Trono D, Jakobsson J (2012) MicroRNA-124 is a subventricular zone neuronal fate determinant. J Neurosci. 32:8879–8889. https://doi.org/10.1523/JNEUROSCI.0558-12.2012
Al Mamun A, Yu H, Romana S, Liu F (2018) Inflammatory responses are sex specific in chronic hypoxic-ischemic encephalopathy. Cell Transplant 27:1328–1339. https://doi.org/10.1177/0963689718766362
Ashwal S, Tone B, Tian HR, Chong S, Obenaus A (2006) Serial magnetic resonance imaging in a rat pup filament stroke model. Exp Neurol 202(2):294–301. https://doi.org/10.1016/j.expneurol.2006.06.004
Azzopardi DV, Strohm B, Edwards AD, Dyet L, Halliday HL, Juszczak E, Kapellou O, Levene M, Marlow N, Porter E, Thoresen M, Whitelaw A, Brocklehurst P, TOBY Study Group (2009) Moderate hypothermia to treat perinatal asphyxial encephalopathy. N Engl J Med 361:1349–1358. https://doi.org/10.1056/NEJMoa0900854
Bhalala OG, Srikanth M, Kessler JA (2013) The emerging roles of microRNAs in CNS injuries. Nat Rev Neurol 9:328–339. https://doi.org/10.1038/nrneurol.2013.67
Chen W, Hartman R, Ayer R, Marcantonio S, Kamper J, Tang J, Zhang JH (2009) Matrix metallo proteinases inhibition provides neuroprotection against hypoxia-ischemia in the developing brain. J Neurochem. 111:726–736. https://doi.org/10.1111/j.1471-4159.2009.06362.x
Chen Z, Liu S, Tian L, Wu M, Ai F, Tang W, Zhao L, Ding J, Zhang L, Tang A (2015) miR-124 and miR-506 inhibit colorectal cancer progression by targeting DNMT3B and DNMT1. Oncotarget 6:38139–38150. https://doi.org/10.18632/oncotarget.5709
Di Y, Lei Y, Yu F, Changfeng F, Song W, Xuming M (2014) MicroRNAs expression and function in cerebral ischemia reperfusion injury. J Mol Neurosci 53:242–250. https://doi.org/10.1007/s12031-014-0293-8
Dixon BJ, Chen D, Zhang Y, Flores J, Malaguit J, Nowrangi D, Zhang JH, Tang J (2016) Intranasal administration of interferon beta attenuates neuronal apoptosis via the JAK1/STAT3/BCL-2 pathway in a rat model of neonatal hypoxic-ischemic encephalopathy. ASN Neuro 8:1759091416670492. https://doi.org/10.1177/1759091416670492
Dobbing J, Sands J (1979) Comparative aspects of the brain growth spurt. Early Hum Dev 3:79–83. https://doi.org/10.1016/0378-3782(79)90022-7
Doeppner TR, Doehring M, Bretschneider E, Zechariah A, Kaltwasser B, Müller B, Koch JC, Bähr M, Hermann DM, Michel U (2013) MicroRNA-124 protects against focal cerebral ischemia via mechanisms involving Usp14-dependent REST degradation. Acta Neuropathol 126:251–265. https://doi.org/10.1007/s00401-013-1142-5
Feng T, Shao F, Wu Q, Zhang X, Xu D, Qian K, Xie Y, Wang S, Xu N, Wang Y, Qi C (2016) miR-124 downregulation leads to breast cancer progression via LncRNA-MALAT1 regulation and CDK4/E2F1 signal activation. Oncotarget 7:16205–16216. https://doi.org/10.18632/oncotarget.7578
Fernández-López D, Natarajan N, Ashwal S, Vexler ZS (2014) Mechanisms of perinatal arterial ischemic stroke. J Cereb Blood Flow Metab 34:921–932. https://doi.org/10.1038/jcbfm.2014.41
Gould TD, Dao DT, Kovacsics CE (2009) The open field test. In: Gould T (ed) Mood and anxiety related phenotypes in mice. Neuromethods, vol 42. Humana Press, Totowa, NJ, p 1–20. https://doi.org/10.1007/978-1-60761-303-9_1
Hamm RJ, Pike BR, O'Dell DM, Lyeth BG, Jenkins LW (1994) Therotarod test: an evaluation of its effectiveness in assessing motor deficits following traumatic brain injury. J Neurotrauma 11:187–196. https://doi.org/10.1089/neu.1994.11.187
Higgins RD, Raju T, Edwards AD (2011) Eunice Kennedy Shriver National Institute of Child Health and Human Development Hypothermia Workshop Speakers and Moderators. Hypothermia and other treatment options for neonatal encephalopathy: an executive summary of the Eunice Kennedy Shriver NICHD workshop. J Pediatr 159:851–858. https://doi.org/10.1016/j.jpeds.2011.08.004
Huang Y, Chuang AY, Ratovitski EA (2011) Phospho-DeltaNp63alpha/miR-885-3p axis in tumor cell life and cell death upon cisplatin exposure. Cell Cycle 10:3938–3947. https://doi.org/10.4161/cc.10.22.18107
Huang W, Liu X, Cao J, Meng F, Li M, Chen B, Zhang J (2015) miR-134 regulates ischemia/reperfusion injury-induced neuronal cell death by regulating CREB signaling. J Mol Neurosci 55:821–829. https://doi.org/10.1007/s12031-014-0434-0
Jacobs SE, Morley CJ, Inder TE, Stewart MJ, Smith KR, McNamara PJ, Wright IM, Kirpalani HM, Darlow BA, Doyle LW (2011) Infant Cooling Evaluation Collaboration. Whole-body hypothermia for term and near-term newborns with hypoxic-ischemic encephalopathy: a randomized controlled trial. Arch Pediatr Adolesc Med 165:692–700. https://doi.org/10.1001/archpediatrics.2011.43
Ji S, Li S, Zhao X, Kang N, Cao K, Zhu Y, Peng P, Fan J, Xu Q, Yang S, Liu Y (2019) Protective role of phenylethanoid glycosides, Torenoside B and Savatiside A, in Alzheimer's disease. Exp Ther Med 17:3755–3767. https://doi.org/10.3892/etm.2019.7355
Jiang D, Du J, Zhang X, Zhou W, Zong L, Dong C, Chen K, Chen Y, Chen X, Jiang H (2016) miR-124 promotes the neuronal differentiation of mouse inner ear neural stem cells. Int J Mol Med 38:1367–1376. https://doi.org/10.3892/ijmm.2016.2751
Kim HY (2016) Statistical notes for clinical researchers: Sample size calculation 3. Comparison of several means using one-way ANOVA. Restor Dent Endod 41:231–234. https://doi.org/10.5395/rde.2016.41.3.231
Koukos G, Polytarchou C, Kaplan JL, Morley-Fletcher A, Gras-Miralles B, Kokkotou E, Baril-Dore M, Pothoulakis C, Winter HS, Iliopoulos D (2013) MicroRNA-124 regulates STAT3 expression and is down-regulated in colon tissues of pediatric patients with ulcerative colitis. Gastroenterology 145:842–852.e842. https://doi.org/10.1053/j.gastro.2013.07.001
Lai MC, Yang SN (2011) Perinatal hypoxic-ischemic encephalopathy. J Biomed Biotechnol 2011:609813. https://doi.org/10.1155/2011/609813
Li Y, Gonzalez P, Zhang L (2012) Fetal stress and programming of hypoxic/ ischemic-sensitive phenotype in the neonatal brain: mechanisms and possible interventions. Prog Neurobiol 98:145–165. https://doi.org/10.1016/j.pneurobio.2012.05.010
Li X, Yang Z, Han Z, Wen Y, Ma Z, Wang Y (2018) Niclosamide acts as a new inhibitor of vasculogenic mimicry in oral cancer through upregulation of miR-124 and downregulation of STAT3. Oncol Rep 39:827–833. https://doi.org/10.3892/or.2017.6146
Li M, Cui L, Feng X, Wang C, Zhang Y, Wang L, Ding Y, Zhao T (2019a) Losmapimod protected epileptic rats from hippocampal neuron damage through inhibition of the MAPK pathway. Front Pharmacol 10:625. https://doi.org/10.3389/fphar.2019.00625
Li Z, Zhang B, Yao WL, Zhang CH, Wang L, Zhang Y (2019b) APC-Cdh1 regulates neuronal apoptosis through modulating glycolysis and pentose-phosphate pathway after oxygen-glucose deprivation and reperfusion. Cell Mol Neurobiol 39:123. https://doi.org/10.1007/s10571-018-0638-x
Liu R, Zhao W, Zhao Q, Liu SJ, Liu J, He M, Xu Y, Wang W, Liu W, Xia QJ, Li CY, Wang TH (2014) Endoplasmic reticulum protein 29 protects cortical neurons from apoptosis and promoting corticospinal tract regeneration to improve neural behavior via caspase and Erk signal in rats with spinal cord transection. Mol Neurobiol 50:1035–1048. https://doi.org/10.1007/s12035-014-8681-1
Longa EZ, Weinstein PR, Carlson S, Cummins R (1989) Reversible middle cerebral artery occlusion without craniectomy in rats. Stroke 20:84–91. https://doi.org/10.1161/01.STR.20.1.84
Ma Q, Dasgupta C, Li Y, Bajwa NM, Xiong F, Harding B, Hartman R, Zhang L (2016) Inhibition of microRNA-210 provides neuroprotection in hypoxic-ischemic brain injury in neonatal rats. Neurobiol Dis 89:202–212. https://doi.org/10.1016/j.nbd.2016.02.011
Maguire S, Strittmatter R, Chandra S, Barone FC (2004) Stroke-prone rats exhibit prolonged behavioral deficits without increased brain injury: an indication of disrupted post-stroke brain recovery of function. Neurosci Lett 354(3):229–233. https://doi.org/10.1016/j.neulet.2003.09.079
Martinez B, Peplow PV (2016) Blood microRNAs as potential diagnostic and prognostic markers in cerebral ischemic injury. Neural Regener Res 11:1375–1378. https://doi.org/10.4103/1673-5374.191196
Miao W, Bao TH, Han JH, Yin M, Zhang J, Yan Y, Zhu YH (2016) Neuroprotection induced by post-conditioning following ischemia/reperfusion in mice is associated with altered microRNA expression. Mol Med Rep 14:2582–2588. https://doi.org/10.3892/mmr.2016.5576
Panneerdoss S, Viswanadhapalli S, Abdelfattah N, Onyeagucha BC, Timilsina S, Mohammad TA, Chen Y, Drake M, Vuori K, Kumar TR, Rao MK (2017) Cross-talk between miR-471–5p and autophagy component proteins regulates LC3-associated phagocytosis (LAP) of apoptotic germ cells. Nat Commun 8:598. https://doi.org/10.1038/s41467-017-00590-9
Parikh P, Juul SE (2018) Neuroprotective strategies in neonatal brain injury. J Pediatr 192:22–32. https://doi.org/10.1016/j.jpeds.2017.08.031
Pei Z, Cheung RT (2003) Melatonin protects SHSY5Y neuronal cells but not cultured astrocytes from ischemia due to oxygen and glucose deprivation. J Pineal Res 34:194–201. https://doi.org/10.1034/j.1600-079X.2003.00026.x
Rice JE 3rd, Vannucci RC, Brierley JB (1981) The influence of immaturity on hypoxic-ischemic brain damage in the rat. Ann Neurol 9:131–141. https://doi.org/10.1002/ana.410090206
Shankaran S, Pappas A, McDonald SA et al (2012) Eunice Kennedy Shriver NICHD Neonatal Research Network. Childhood outcomes after hypothermia for neonatal encephalopathy. N Engl J Med 366:2085–2092. https://doi.org/10.1056/NEJMoa1112066
Shohami E, Novikov M, Bass R (1995) Long-term effect of HU-211, a novel non-competitive NMDA antagonist, on motor and memory functions after closed head injury in the rat. Brain Res 674:55–62. https://doi.org/10.1016/0006-8993(94)01433-i
Wang W, Kwon EJ, Tsai LH (2012) MicroRNAs in learning, memory, and neurological diseases. Learn Mem 19:359–368. https://doi.org/10.1101/lm.026492.112
Xing Y, Hou J, Guo T, Zheng S, Zhou C, Huang H, Chen Y, Sun K, Zhong T, Wang J, Li H, Wang T (2014) microRNA-378 promotes mesenchymal stem cell survival and vascularization under hypoxic-ischemic conditions in vitro. Stem Cell Res Ther 5:130. https://doi.org/10.1186/scrt520
Yang D, Kuan CY (2015) Anti-tissue plasminogen activator (tPA) as an effective therapy of neonatal hypoxia-ischemia with and without inflammation. CNS Neurosci Ther 21:367–373. https://doi.org/10.1111/cns.12365
Yang J, Zhang X, Chen X, Wang L, Yang G (2017) Exosome mediated delivery of miR-124 promotes neurogenesis after ischemia. Mol Ther Nucleic Acids 7:278–287. https://doi.org/10.1016/j.omtn.2017.04.010
Zhang W, Ma J, Danzeng Q, Tang Y, Lu M, Kang Y (2017a) Safety of moderate hypothermia for perinatal hypoxic-ischemic encephalopathy: a meta-analysis. Pediatrneurol 74:51–61. https://doi.org/10.1016/j.pediatrneurol.2017.04.023
Zhang X, Liu F, Wang Q, Geng Y (2017b) Overexpressed microRNA-506 and microRNA-124 alleviate H2O2-induced human cardiomyocyte dysfunction by targeting kruppel-like factor 4/5. Mol Med Rep 16:5363–5369. https://doi.org/10.3892/mmr.2017.7243
Zhang G, Zhang T, Li N et al (2018) Tetramethylpyrazinenitrone activates the BDNF/Akt/CREB pathway to promote post-ischaemic neuroregeneration and recovery of neurological functions in rats. Br J Pharmacol 175:517–531. https://doi.org/10.1111/bph.14102
Zhang ZB, Tan YX, Zhao Q et al (2019) miRNA-7a-2-3p Inhibits neuronal apoptosis in oxygen-glucose deprivation (OGD) model. Front Neurosci. 13:16. https://doi.org/10.3389/fnins.2019.00016
Zhao F, Qu Y, Liu J, Liu H, Zhang L, Feng Y, Wang H, Gan J, Lu R, Mu D (2015) Microarray profiling and co-expression network analysis of LncRNAs and mRNAs in neonatal rats following hypoxic-ischemic brain damage. Sci Rep 5:13850. https://doi.org/10.1038/srep13850
Acknowledgements
This study was supported by Research Fund for the Doctoral Program of Liu-Liu Xiong (201903) and the Program Innovative Research Team in Science and Technology in Yunnan province (2017HC007). It was also supported by grant from the National Natural Science Foundation of China (Grant Nos. 81471268, 81960214 and 81660193). We would like to thank Professor Zhao-Qiong Zhu from Department of Anesthesiology, the Affiliated Hospital of Zunyi Medical University and Professor Fei Liu from Department of Anesthesiology, West China Hospital, Sichuan University for their technical support.
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All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by LX, HZ, QZ, JH, LX, MW, YZ, JD, MH, and MY. The first draft of the manuscript was written and revised by LX, HZ, and TW ,and all authors commented on previous versions of the manuscript. The whole manuscript was revised and improved by MAH and MY. All authors read and approved the final manuscript.
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Xiong, L., Zhou, H., Zhao, Q. et al. Overexpression of miR-124 Protects Against Neurological Dysfunction Induced by Neonatal Hypoxic–Ischemic Brain Injury. Cell Mol Neurobiol 40, 737–750 (2020). https://doi.org/10.1007/s10571-019-00769-2
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DOI: https://doi.org/10.1007/s10571-019-00769-2