重组人促红细胞生成素对宫内炎症致新生大鼠脑损伤髓鞘少突细胞糖蛋白表达变化与髓鞘修复关系的研究

doi:10.11852/zgetbjzz2018-26-04-09

摘要/Abstract

摘要： 目的通过观察新生大鼠脑组织内髓鞘少突细胞糖蛋白(MOG) mRNA及MOG蛋白动态变化情况,探讨重组人促红细胞生成素(rhEPO)与宫内炎症致脑组织髓鞘损伤、修复的关系,为临床干预新生儿脑白质损伤提供依据。方法 1)选取孕15 d的Wistar大鼠分别经腹腔注射脂多糖及无菌生理盐水后分为脂多糖感染组和对照组,随机选取感染组经苏木素-伊红染色证实胎盘和新生大鼠脑组织存在炎性损伤的同胎新生大鼠64只,分为rhEPO治疗组和感染对照组,随机选取对照组中新生大鼠32只为空白对照组,分别于生后每日腹腔注射rhEPO(5 000 U/kg)或等量生理盐水。2)三组新生大鼠均分别于生后当日、3 d、7 d及14 d行左心室甲醛灌注取脑组织,利用RT-PCR检测脑组织中MOG mRNA,Western blot检测MOG蛋白水平变化。结果 1)空白对照组MOG mRNA水平在3 d[(2.05±0.75)倍]、7 d[(2.37±0.57)倍]及14 d[(3.36±0.81)倍]新生大鼠脑组织中均较当日组[(1.01±0.16)倍]明显升高,差异有统计学意义(P＜0.05);2) 感染对照组MOG mRNA水平在当日、3 d、7 d及14 d新生大鼠脑组织中均较同日龄空白对照组下降,且3 d和7 d组差异具有统计学意义(P＜0.05);3)rhEPO治疗组3 d[(2.18±0.89)倍]、7 d[(3.53±1.55)倍]及14 d[(3.30±1.12)倍]新生大鼠脑组织MOG mRNA水平较同日龄感染对照组均显著上升,差异有统计学意义(P＜0.05);4)rhEPO治疗组MOG蛋白水平在3 d(12.63±0.65 vs. 18.23±0.37)、7 d(13.59±1.10 vs. 22.72±2.77)及14 d(15.63±1.32 vs. 25.85±2.70)时均低于感染对照组,差异有统计学意义(P＜0.05)。结论宫内炎症致新生大鼠脑组织损害中存在通过抑制髓鞘MOG基因及蛋白表达的机制。早期应用rhEPO能够促进MOG基因及蛋白表达发挥神经修复保护作用,14 d的疗程目前为最佳疗程。

关键词: 宫内炎症, 新生, 脑损伤, 促红细胞生成素, 髓鞘少突胶质细胞糖蛋白

Abstract: Objective To study the neuroprotective effect of rhEPO on brain tissue of newborn rats with intrauterine infection by detecting the changes of MOG expression,in order to provide reference for clinical intervention. Methods 1)Wistar rats that had been pregnant for 15 days were selected and divided into LPS infection group and control group through intraperitoneal injection of LPS (0.3 mg/kg) and equivalent stroke-physiological saline solution.Totally 64 newborn rats in the same fetus of the LPS infection group with brain inflammatory injury proved by HE staining were randomly selected and were divided into rhEPO treatment group (n=32) and infection control group(n=32).Meantime,32 newborn rats in the control group were randomly enrolled as the blank control group.Three groups of rats were conducted intraperitoneal injection of rhEPO (5000IU/kg) or equivalent saline solution simultaneously promptly every day after the birth.2) Brain tissues of three groups were took out by perfusing formaldehyde on 0h,the 3rd,7th and 14th of their birth respectively.And the levels of MOG mRNA and MOG protein were tested by RT-PCR and western blot respectively. Results 1)The levels of MOG mRNA of blank control group were significantly higher on the 3rd (2.05±0.75),7th (2.37±0.57) and 14th (3.36±0.81) day than those on 0 hour (1.01±0.16)(P＜0.05).2)The levels of MOG mRNA on 0 hour,the 3rd,7th and 14th day of infection control group were lower than those of blank control group,and the differences were significant on the 3rd and 7th day.3)The levels of MOG mRNA on the 3rd(2.18±0.89),7th(3.53±1.55) and 14th(3.30±1.12) were significantly higher in rhEPO treatment group than those in infection control group(P＜0.05).4)The level of MOG protein in rhEPO treatment group was significantly higher than that in the infection control group with the same age on the 3rd(18.23±0.37 vs12.63±0.65),7th(22.72±2.77 vs 13.59±1.10),and 14th day(25.85±2.70 vs15.63±1.32). Conclusion The mechanism of inhibiting the expression of MOG gene and protein in the myelin sheath was found in the brain tissue damage induced by intrauterine inflammation in neonatal rats.Early application of rhEPO can promote the expression of MOG gene and protein,to play a protective role in nerve repairment.The best treatment may last for 14 days.

Key words: intrauterine inflammation, newborn, brain injury, erythropoietin, myelin oligodendrocyte glycoprotein

中图分类号:

R179

孙梦雅, 罗刚, 王倩, 姜红. 重组人促红细胞生成素对宫内炎症致新生大鼠脑损伤髓鞘少突细胞糖蛋白表达变化与髓鞘修复关系的研究[J]. 中国儿童保健杂志, 2018, 26(4): 381-385.

SUN Meng-ya, LUO Gang, WANG Qian, JIANG Hong. Effect of rhEPO treatment on MOG expression in brain tissue of newborn rats with intrauterine inflammation of remyelination and its correlation with myelin repair[J]. journal1, 2018, 26(4): 381-385.

参考文献

[1] Merelli A,Czornyj L.Erythropoietin as a new therapeutic opportunity in brain inflammation and neurodegenerative diseases[J].Int J Neurosci,2015,125(11):793-797.
[2] Strunk T,Inder T,Wang X,et al.Infection-induced inflammation and cerebral injury in preterm infants[J].Lancet Infect Dis,2014,14(8):751-762.
[3] Platt MJ,Cans C,Johnson A,et al.Trends in cerebral palsy among infants of very low birthweight (<1500 g) or born prematurely (<32 weeks) in 16 European centres:a database study[J].Lancet (London,England),2007,369(9555):43-50.
[4] Larroque B,Ancel PY,Marret S,et al.Neurodevelopmental disabilities and special care of 5-year-old children born before 33 weeks of gestation (the EPIPAGE study):a longitudinal cohort study[J].Lancet,2008,371(9615):813-820.
[5] Malaeb S.Fetal inflammatory response and brain injury in the preterm newborn[J].J Child Neurol,2009,24(9):1119-1126.
[6] Chau V,Poskitt KJ,McFadden DE,et al.Effect of chorioamnionitis on brain development and injury in premature newborns[J].Ann Neurol,2009,66(2):155-164.
[7] Rees S,Harding R.The biological basis of injury and neuroprotection in the fetal and neonatal brain[J].Int J Dev Neurosci,2011,29(6):551-563.
[8] Shatrov JG,Birch SC,Lam LT,et al.Chorioamnionitis and cerebral palsy:a meta-analysis[J].Obstet Gynecol,2010,116(2):387-392.
[9] Hemmer B,Archelos JJ.New concepts in the immunopathogenesis of multiple sclerosis[J].Nat Rev Neurosci,2002,3(4):291-301.
[10] Lee DH.The role of myelin oligodendrocyte glycoprotein in autoimmune demyelination:a target for multiple sclerosis therapy?[J].Expert Opin Ther Targets,2012,16(5):451-462.
[11] Girard S,Kadhim H,Roy M,et al.Role of perinatal inflammation in cerebralpalsy[J].Pediatr Neurol,2009,40(3):168-174.
[12] Inder TE,Anderson NJ,Spencer C,et al.White matter injury in the premature infant:a comparison between serial cranial sonographic and MR findings at term[J].AJNR,2003,24(5):805-809.
[13] Dale RC,Tantsis EM,Merheb V,et al.Antibodies to MOG have a demyelination phenotype and affect oligodendrocyte cytoskeleton[J].Neurol Neuroimmunol Neuroinflamm,2014,1(1):e12.
[14] Rostasy K.Role of autoantibodies in acquired inflammatory demyelinating diseases of the central nervous system in children[J].Neuropediatrics,2013,44(6):297-301.
[15] Reindl M,Di Pauli F,Rostásy K.The spectrum of MOG autoantibody-associated demyelinating diseases[J].Nat Rev Neurol,2013,9(8):455-461.
[16] Tegetmeyer H.Anti-myelin oligodendrocyte glycoprotein antibodies in paediatric patients with optic neuritis[J].Klin Monatsbl Augenheilkd,2017,234(10):1243-1249.
[17] Baumann M,Hennes EM,Schanda K,et al.Children with multiphasic disseminated encephalomyelitis and antibodies to the myelin oligodendrocyte glycoprotein (MOG):Extending the spectrum of MOG antibody positive diseases[J].Multiple Sclerosis (Houndmills,Basingstoke,England),2016,22(14):1821-1829.
[18] Hacohen Y,Absoud M,Deiva K,et al.Myelin oligodendrocyte glycoprotein antibodies are associated with a non-MS course in children[J].Neurol Neuroimmunol Neuroinflamm,2015,2(2):e81-e87.
[19] Hino-Fukuyo N,Haginoya K,Nakashima I,et al.Clinical features and long-term outcome of a group of Japanese children with inflammatory central nervous system disorders and seropositivity to myelin-oligodendrocyte glycoprotein antibodies[J].Brain & Development,2015,37(9):849-852.
[20] Jantzie LL,Miller RH.Erythropoietin signaling promotes oligodendrocyte development following prenatal systemic hypoxic-ischemic brain injury[J].Pediatr Res,2013,74(6):658-667.
[21] Rangarajan V.Erythropoietin:emerging role of erythropoietin in neonatal neuroprotection[J].Pediatr Neurol,2014,51(4):481-488.
[22] Gonzalez FF,Abel R,Almli CR,et al.Erythropoietin sustains cognitive function and brain volume after neonatal stroke[J].Dev Neurosci,2009,31(5):403-411.
[23] Cervellini I,Annenkov A,Brenton T,et al.Erythropoietin (EPO) increases myelin gene expression in CG4 oligodendrocyte cells through the classical EPO receptor[J].Mol Med,2013,19:223-229.
[24] Wang R,Zhao H,Li J,et al.Erythropoietin attenuates axonal injury after middle cerebral artery occlusion in mice[J].Neurol Res,2017,39(6):545-551.
[25] 华威,吴鹤,周敏,等.重组人促红细胞生成素对脑梗死后少突胶质细胞的保护作用[J].中华病理学杂志,2015,(5):323-328.
[26] 沈盈.促红细胞生成素治疗宫内感染致新生大鼠脑白质损伤的研究[D].杭州:浙江大学,2009.
[27] 李婷.EPO对宫内感染脑白质损伤新生大鼠脑组织MBP、CD68表达的影响及神经行为学观察[D].青岛:青岛大学,2014.