Silencing of long non‑coding antisense RNA brain‑derived neurotrophic factor attenuates hypoxia/ischemia‑induced neonatal brain injury

Qiao,Li‑Xing; Zhao,Rui‑Bin; Wu,Ming‑Fu; Zhu,Li‑Hua; Xia,Zheng‑Kun

doi:10.3892/ijmm.2020.4625

Silencing of long non‑coding antisense RNA brain‑derived neurotrophic factor attenuates hypoxia/ischemia‑induced neonatal brain injury

Authors:
- Li‑Xing Qiao
- Rui‑Bin Zhao
- Ming‑Fu Wu
- Li‑Hua Zhu
- Zheng‑Kun Xia
View Affiliations

Affiliations: Department of Pediatrics, Zhongda Hospital, Southeast University, Nanjing, Jiangsu 210029, P.R. China, Department of Pediatrics, Jinling Hospital, Nanjing University School of Medicine, Nanjing, Jiangsu 210002, P.R. China, Department of Pediatrics, Affiliated Hospital of Yang Zhou University, Yangzhou, Jiangsu 225000, P.R. China, Institute of Clinical Science, Jiangsu Health Vocational College, Nanjing, Jiangsu 210029, P.R. China
Published online on: June 2, 2020 https://doi.org/10.3892/ijmm.2020.4625
Pages: 653-662
Copyright: © Qiao 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

Hypoxic/ischemic (HI) brain damage (HIBD) is a major cause of acute neonatal brain injury, leading to high mortality and serious neurological deficits. The antisense RNA of brain‑derived neurotrophic factor (BDNF‑AS) is transcribed from the opposite strand of the BDNF gene. The aim of the present study was to investigate the role of BDNF‑AS in HI‑induced neuronal cell injury in vivo and in vitro. Reverse transcription‑quantitative PCR (RT‑qPCR) assays indicated that BDNF‑AS expression was significantly upregulated in HI‑injured neonatal brains and hippocampal neurons. However, BDNF expression was downregulated in HI‑injured neonatal brains and hippocampal neurons. Cell Counting Kit‑8 assays, Hoechst staining, calcein‑AM/PI staining, immunostaining, water maze tests and rotarod tests demonstrated that BDNF‑AS silencing protected against hypoxia‑induced primary hippocampal neuron injury in vitro and HI‑induced brain injury in vivo. Mechanistically, RT‑qPCR assays and western blotting indicated that BDNF‑AS silencing led to increased expression of BDNF and activated the BDNF‑mediated signaling pathway, as demonstrated by increased expression levels of BDNF, phosphorylated‑Akt and phosphorylated‑tropomyosin receptor kinase B. Collectively, the present study provides important insights into the pathogenesis of HIBD, and it was indicated that BDNF‑AS silencing may be a promising approach for the treatment of neonatal HIBD.

View Figures

View References

1	Zhu C, Kang W, Xu F, Cheng X, Zhang Z, Jia L, Ji L, Guo X, Xiong H, Simbruner G, et al: Erythropoietin improved neurologic outcomes in newborns with hypoxic-ischemic encephalopathy. Pediatrics. 124:e218–e226. 2009. View Article : Google Scholar : PubMed/NCBI
2	Rocha-Ferreira E and Hristova M: Antimicrobial peptides and complement in neonatal hypoxia-ischemia induced brain damage. Front Immunol. 6:562015. View Article : Google Scholar : PubMed/NCBI
3	Johnston MV, Trescher WH, Ishida A and Nakajima W: Neurobiology of hypoxic-ischemic injury in the developing brain. Pediat Res. 49:735–741. 2001. View Article : Google Scholar : PubMed/NCBI
4	Blackmon LR and Stark AR: American Academy of Pediatrics Committee on Fetus and Newborn: Hypothermia: A neuroprotective therapy for neonatal hypoxic-ischemic encephalopathy. Pediatrics. 117:942–948. 2006. View Article : Google Scholar : PubMed/NCBI
5	Tagin MA, Woolcott CG, Vincer MJ, Whyte RK and Stinson DA: Hypothermia for neonatal hypoxic ischemic encephalopathy: An updated systematic review and meta-analysis. Arch Pediat Adol Med. 166:558–566. 2012. View Article : Google Scholar
6	Quinn JJ and Chang HY: Unique features of long non-coding RNA biogenesis and function. Nat Rev Genet. 17:47–62. 2016. View Article : Google Scholar
7	Mercer TR, Dinger ME and Mattick JS: Long non-coding RNAs: Insights into functions. Nat Rev Genet. 10:155–159. 2009. View Article : Google Scholar : PubMed/NCBI
8	Fatica A and Bozzoni I: Long non-coding RNAs: New players in cell differentiation and development. Nat Rev Genet. 15:7–21. 2014. View Article : Google Scholar
9	Zhang J, Yuan L, Zhang X, Hamblin MH, Zhu T, Meng F, Li Y, Chen YE and Yin KJ: Altered long non-coding RNA transcriptomic profiles in brain microvascular endothelium after cerebral ischemia. Exp Neurol. 277:162–170. 2016. View Article : Google Scholar : PubMed/NCBI
10	Zhao F, Qu Y, Liu J, Liu H, Zhang L, Feng Y, Wang H, Gan J, Lu R and Mu D: Microarray profiling and co-expression network analysis of LncRNAs and mRNAs in neonatal rats following hypoxic-ischemic brain damage. Sci Rep. 5:138502015. View Article : Google Scholar : PubMed/NCBI
11	Mehta SL, Kim T and Vemuganti R: Long noncoding RNA FosDT promotes ischemic brain injury by interacting with REST-associated chromatin-modifying proteins. J Neurosci. 35:16443–16449. 2015. View Article : Google Scholar : PubMed/NCBI
12	Zhao RB, Zhu LH, Shu JP, Qiao LX and Xia ZK: GAS5 silencing protects against hypoxia/ischemia-induced neonatal brain injury. Biochem Biophys Res Commun. 497:285–291. 2018. View Article : Google Scholar : PubMed/NCBI
13	Zhang X, Tang X, Liu K, Hamblin MH and Yin KJ: Long noncoding RNA Malat1 regulates cerebrovascular pathologies in ischemic stroke. J Neurosci. 37:1797–1806. 2017. View Article : Google Scholar : PubMed/NCBI
14	Rossi C, Angelucci A, Costantin L, Braschi C, Mazzantini M, Babbini F, Fabbri ME, Tessarollo L, Maffei L, Berardi N and Caleo M: Brain-derived neurotrophic factor (BDNF) is required for the enhancement of hippocampal neurogenesis following environmental enrichment. Eur J Neurosci. 24:1850–1856. 2006. View Article : Google Scholar : PubMed/NCBI
15	Angelucci F, Mathe AA and Aloe L: Neurotrophic factors and CNS disorders: Findings in rodent models of depression and schizophrenia. Prog Brain Res. 146:151–165. 2004. View Article : Google Scholar : PubMed/NCBI
16	Li Y, Xu F, Xiao H and Han F: Long noncoding RNA BDNF-AS inversely regulated BDNF and modulated high-glucose induced apoptosis in human retinal pigment epithelial cells. J Cell Biochem. 119:817–823. 2018. View Article : Google Scholar
17	Zheng X, Lin C, Li Y, Ye J, Zhou J and Guo P: Long noncoding RNA BDNF-AS regulates ketamine-induced neurotoxicity in neural stem cell derived neurons. Biomed Pharmacother. 82:722–728. 2016. View Article : Google Scholar : PubMed/NCBI
18	Zhang Y, Yan L, Cao Y, Kong G and Lin C: Long noncoding RNA BDNF-AS protects local anesthetic induced neurotoxicity in dorsal root ganglion neurons. Biomed Pharmacother. 80:207–212. 2016. View Article : Google Scholar : PubMed/NCBI
19	Guo CC, Jiao CH and Gao ZM: Silencing of lncRNA BDNF-AS attenuates Aβ25-35-induced neurotoxicity in PC12 cells by suppressing cell apoptosis and oxidative stress. Neurol Res. 40:795–804. 2018. View Article : Google Scholar : PubMed/NCBI
20	Cetin A, Komai S, Eliava M, Seeburg PH and Osten P: Stereotaxic gene delivery in the rodent brain. Nat Protoc. 1:3166–3173. 2006. View Article : Google Scholar
21	Klaunberg BA, O'malley J, Clark T and Davis JA: Euthanasia of mouse fetuses and neonates. Contemp Top Lab Anim Sci. 43:29–34. 2004.PubMed/NCBI
22	Artwohl J, Brown P, Corning B and Stein S: Report of the ACLAM task force on rodent euthanasia. J Am Assoc Lab Anim Sci. 45:98–105. 2006.PubMed/NCBI
23	Kramer M, Dang J, Baertling F, Denecke B, Clarner T, Kirsch C, Beyer C and Kipp M: TTC staining of damaged brain areas after MCA occlusion in the rat does not constrict quantitative gene and protein analyses. J Neurosci Methods. 187:84–89. 2010. View Article : Google Scholar : PubMed/NCBI
24	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
25	Merritt JR and Rhodes JS: Mouse genetic differences in voluntary wheel running, adult hippocampal neurogenesis and learning on the multi-strain-adapted plus water maze. Behav Brain Res. 280:62–71. 2015. View Article : Google Scholar
26	Yin KJ, Hamblin M and Chen YE: Non-coding RNAs in cerebral endothelial pathophysiology: Emerging roles in stroke. Neurochem Int. 77:9–16. 2014. View Article : Google Scholar : PubMed/NCBI
27	Xin JW and Jiang YG: Long noncoding RNA MALAT1 inhibits apoptosis induced by oxygen-glucose deprivation and reoxygenation in human brain microvascular endothelial cells. Exp Ther Med. 13:1225–1234. 2017. View Article : Google Scholar : PubMed/NCBI
28	Zhang H, Li D, Zhang Y, Li J, Ma S, Zhang J, Xiong Y, Wang W, Li N and Xia L: Knockdown of lncRNA BDNF-AS suppresses neuronal cell apoptosis via downregulating miR-130b-5p target gene PRDM5 in acute spinal cord injury. RNA Biol. 15:1071–1080. 2018.PubMed/NCBI
29	Hagg T: Molecular regulation of adult CNS neurogenesis: An integrated view. Trends Neurosci. 28:589–595. 2005. View Article : Google Scholar : PubMed/NCBI
30	Waterhouse EG, An JJ, Orefice LL, Baydyuk M, Liao GY, Zheng K, Lu B and Xu B: BDNF promotes differentiation and maturation of adult-born neurons through GABAergic transmission. J Neurosci. 32:14318–14330. 2012. View Article : Google Scholar : PubMed/NCBI
31	Lu B, Nagappan G and Lu Y: BDNF and synaptic plasticity, cognitive function, and dysfunction. Handb Exp Pharmacol. 220:223–250. 2014. View Article : Google Scholar : PubMed/NCBI
32	Yoshii A and Constantine-Paton M: Postsynaptic BDNF-TrkB signaling in synapse maturation, plasticity, and disease. Dev Neurobiol. 70:304–322. 2010.PubMed/NCBI
33	Vannucci SJ and Hagberg H: Hypoxia-ischemia in the immature brain. J Exp Biol. 207:3149–3154. 2004. View Article : Google Scholar : PubMed/NCBI
34	Kumral A, Gonenc S, Acikgoz O, Sonmez A, Genc K, Yilmaz O, Gokmen N, Duman N and Ozkan H: Erythropoietin increases glutathione peroxidase enzyme activity and decreases lipid peroxidation levels in hypoxic-ischemic brain injury in neonatal rats. Biol Neonate. 87:15–18. 2005. View Article : Google Scholar
35	Vexler ZS and Ferriero DM: Molecular and biochemical mechanisms of perinatal brain injury. Semin Neonatol. 6:99–108. 2001. View Article : Google Scholar : PubMed/NCBI