Effect of miR-582-5p targeting regulation of FOXO1 on neuronal damage in neonatal rats with hypoxic ischemic encephalopathy

doi:10.11958/20230591

Abstract

Abstract:

Objective To investigate the effect of microRNA-582-5p (miR-582-5p) on neuronal damage in neonatal rats with hypoxic ischemic encephalopathy (HIE) through targeted regulation of forkhead box transcription factor O1 (FOXO1). Methods Newborn rats (n=90) were randomly grouped into the control (NC) group, the model (HIE) group, the miRNA non-specific control (LV-miRNA-NC) group, the miR-582-5p overexpression (LV-miR-582-5p) group, the miR-582-5p overexpression+mRNA control (LV-miR-582-5p+LV-NC) group and the miR-582-5p overexpression+FOXO1 overexpression (LV-miR-582-5p+LV-FOXO1) group. HIE model was established in all groups except the NC group, and neurological deficits were scored on rats. TTC staining was applied to measure the volume of cerebral infarction. Real-time PCR was applied to detect miR-582-5p and FOXO1 expression. Dual fluorescence reporter gene experiment was applied to detect the targeting relationship between miR-582-5p and FOXO1. HE staining method was applied to observe pathological changes in hippocampal tissue. TUNEL and NeuN fluorescence dual labeling co localization were applied to detect neuronal apoptosis in hippocampal tissue. Immunohistochemistry was applied to detect expressions of FOXO1 and Caspase-3 proteins. Results There was a targeting relationship between MiR-582-5p and FOXO1. Compared with the NC group, the neurological deficit score, cerebral infarction volume, FOXO1 expression, neuronal apoptosis rate, FOXO1 and Caspase-3 protein expression were increased in the HIE group, and the miR-582-5p expression obviously decreased, the hippocampal tissue showed obvious pathological damage (P<0.05). Compared with the LV-miRNA-NC group, the neurological deficit score, cerebral infarction volume, FOXO1 expression, neuronal apoptosis rate, FOXO1 and Caspase-3 protein expression obviously decreased in the LV-miR-582-5p group, and the miR-582-5p expression obviously increased, the pathological damage of hippocampus tissue was obviously improved (P<0.05). LV-FOXO1 was able to reverse the protective effect of LV-miR-582-5p on neuronal damage in HIE rats. Conclusion MiR-582-5p can directly target FOXO1, negatively regulate FOXO1 expression, reduce neuronal apoptosis in HIE neonatal rats, and have a protective effect on neural injury.

Key words: hypoxia-ischemia, brain, trauma, nervous system, forkhead box protein O1, microRNA-582-5p

CLC Number:

R342，R722.1

Figures/Tables 8

References 18

[1]	LI B, DASGUPTA C, HUANG L, et al. MiRNA-210 induces microglial activation and regulates microglia-mediated neuroinflammation in neonatal hypoxic-ischemic encephalopathy[J]. Cell Mol Immunol, 2020, 17(9):976-991. doi:10.1038/s41423-019-0257-6.
[2]	曾高, 刘雨桐, 孙澎, 等. 儿童脑动静脉畸形治疗策略[J]. 中国现代神经疾病杂志, 2023, 23(5):388-392.
	ZENG G, LIU Y T, SUN P, et al. Treatment of pediatric cerebral arteriovenous malformation[J]. Chin J Contemp Neurol Neurosurg, 2023, 23(5):388-392.
[3]	CHEN R, WANG M, FU S, et al. MicroRNA-204 may participate in the pathogenesis of hypoxic-ischemic encephalopathy through targeting KLLN[J]. Exp Ther Med, 2019, 18(5):3299-3306. doi:10.3892/etm.2019.7936.
[4]	NIU R Z, WANG L Q, YANG W, et al. MicroRNA-582-5p targeting Creb1 modulates apoptosis in cardiomyocytes hypoxia/reperfusion-induced injury[J]. Immun Inflamm Dis, 2022, 10(11):e708-e721. doi:10.1002/iid3.708.
[5]	WANG D, WANG Y, ZOU X, et al. FOXO1 inhibition prevents renal ischemia-reperfusion injury via cAMP-response element binding protein/PPAR-γ coactivator-1α-mediated mitochondrial biogenesis[J]. Br J Pharmacol, 2020, 177(2):432-448. doi:10.1111/bph.14878.
[6]	HE F, ZHANG N, LV Y, et al. Low-dose lipopolysaccharide inhibits neuronal apoptosis induced by cerebral ischemia/reperfusion injury via the PI3K/Akt/FoxO1 signaling pathway in rats[J]. Mol Med Rep, 2019, 19(3):1443-1452. doi:10.3892/mmr.2019.9827.
[7]	刘鑫, 霍世芳, 马中岭, 等. 微小RNA-126靶向调控血管内皮生长因子对新生大鼠缺氧-缺血性脑病神经元损伤的影响[J]. 解剖学报, 2021, 52(6):875-881.
	LIU X, HUO S F, MA Z L, et al. Effect of targeting vascular endothelial growth factor by microRNA-126 on neuronal damage in neonatal rats with hypoxic-ischemic encephalopathy[J]. Acta Anatomica Sinica, 2021, 52(6):875-881. doi:10.16098/j.issn.0529-1356.2021.06.006.
[8]	XIONG Q, LI X, XIA L, et al. Dihydroartemisinin attenuates hypoxic-ischemic brain damage in neonatal rats by inhibiting oxidative stress[J]. Mol Brain, 2022, 15(1):36. doi:10.1186/s13041-022-00921-y.
[9]	DIAO M, QU Y, LIU H, et al. Effect of carbamylated erythropoietin on neuronal apoptosis in fetal rats during intrauterine hypoxic-ischemic encephalopathy[J]. Biol Res, 2019, 52(1):28. doi:10.1186/s40659-019-0234-7.
[10]	AL-WARD H, LIU N, OMOROU M, et al. The relationship between miRNA-210 and SCN1B in fetal rats with hypoxic-ischemic brain injury[J]. Biosci Rep, 2023, 43(1):BSR20222016. doi:10.1042/BSR20222016.
[11]	WU Z, BAI Y, QI Y, et al. lncRNA NEAT1 downregulation ameliorates the myocardial infarction of mice by regulating the miR-582-5p/F2RL2 axis[J]. Cardiovasc Ther, 2022, 2022:4481360. doi:10.1155/2022/4481360.
[12]	ZHANG Y F, LI X X, CAO X L, et al. MicroRNA-582-5p contributes to the maintenance of neural stem cells through inhibiting secretory protein FAM19A1[J]. Front Cell Neurosci, 2022, 16:866020. doi:10.3389/fncel.2022.866020.
[13]	MEI Z G, HUANG Y G, FENG Z T, et al. Electroacupuncture ameliorates cerebral ischemia/reperfusion injury by suppressing autophagy via the SIRT1-FOXO1 signaling pathway[J]. Aging (Albany NY), 2020, 12(13):13187-13205. doi:10.18632/aging.103420.
[14]	LIU Y, JIANG J, WANG X, et al. miR-582-5p is upregulated in patients with active tuberculosis and inhibits apoptosis of monocytes by targeting FOXO1[J]. PLoS One, 2013, 8(10):e78381. doi:10.1371/journal.pone.0078381.
[15]	MA Q, ZHANG L, PEARCE W J. MicroRNAs in brain development and cerebrovascular pathophysiology[J]. Am J Physiol Cell Physiol, 2019, 317(1):C3-C19. doi:10.1152/ajpcell.00022.2019.
[16]	TOADER A M, HOTEIUC O, BIDIAN C, et al. Neuronal apoptosis can be prevented by the combined therapy with melatonin and hypothermia in a neonatal rat model of hypoxic-ischemic encephalopathy[J]. Med Pharm Rep, 2021, 94(2):197-207. doi:10.15386/mpr-1903.
[17]	RODRIGUEZ J, LI T, XU Y, et al. Role of apoptosis-inducing factor in perinatal hypoxic-ischemic brain injury[J]. Neural Regen Res, 2021, 16(2):205-213. doi:10.4103/1673-5374.290875.
[18]	WANG X X, ZHANG B, XIA R, et al. Inflammation, apoptosis and autophagy as critical players in vascular dementia[J]. Eur Rev Med Pharmacol Sci, 2020, 24(18):9601-9614. doi:10.26355/eurrev_202009_23048.

组别	神经功能缺损评分/分（n=15）	脑梗死体积/ %（n=5）
NC组	0.00±0.00	0.00±0.00
HIE组	2.54±0.42^a	31.14±4.41^a
LV-miRNA-NC组	2.32±0.34	32.02±3.95
LV-miR-582-5p组	1.51±0.32^b	15.32±2.33^b
LV-miR-582-5p+LV-NC组	1.32±0.36	16.10±2.46
LV-miR-582-5p+LV-FOXO1组	2.15±0.40^c	23.01±4.40^c
F	114.531^＊＊	64.997^＊＊

组别	神经功能缺损评分/分（n=15）	脑梗死体积/ %（n=5）
NC组	0.00±0.00	0.00±0.00
HIE组	2.54±0.42^a	31.14±4.41^a
LV-miRNA-NC组	2.32±0.34	32.02±3.95
LV-miR-582-5p组	1.51±0.32^b	15.32±2.33^b
LV-miR-582-5p+LV-NC组	1.32±0.36	16.10±2.46
LV-miR-582-5p+LV-FOXO1组	2.15±0.40^c	23.01±4.40^c
F	114.531^＊＊	64.997^＊＊

组别	FOXO1	miR-582-5p
NC组	1.00±0.02	1.00±0.02
HIE组	1.54±0.12^a	0.38±0.04^a
LV-miRNA-NC组	1.52±0.13	0.35±0.05
LV-miR-582-5p组	0.70±0.08^b	0.89±0.08^b
LV-miR-582-5p+LV-NC组	0.68±0.07	0.76±0.06
LV-miR-582-5p+LV-FOXO1组	1.12±0.09^c	0.73±0.04
F	84.133^＊＊	131.981^＊＊

组别	FOXO1	miR-582-5p
NC组	1.00±0.02	1.00±0.02
HIE组	1.54±0.12^a	0.38±0.04^a
LV-miRNA-NC组	1.52±0.13	0.35±0.05
LV-miR-582-5p组	0.70±0.08^b	0.89±0.08^b
LV-miR-582-5p+LV-NC组	0.68±0.07	0.76±0.06
LV-miR-582-5p+LV-FOXO1组	1.12±0.09^c	0.73±0.04
F	84.133^＊＊	131.981^＊＊

组别	FOXO1	Caspase-3
NC组	8.52±1.01	6.25±1.14
HIE组	26.21±3.42^a	21.32±3.02^a
LV-miRNA-NC组	25.34±3.05	21.03±3.42
LV-miR-582-5p组	11.02±1.78^b	8.77±1.25^b
LV-miR-582-5p+LV-NC组	11.55±1.56	9.02±1.44
LV-miR-582-5p+LV-FOXO1组	19.01±2.44^c	17.32±1.74^c
F	52.857^＊＊	47.163^＊＊