成髓鞘细胞研究和应用进展

成髓鞘细胞研究和应用进展陈琳清华大学第二附属医院神经外科中心chenlin_china@163.com第一届全国儿童神经修复学术会议暨第四届全国儿童神经康复论坛.深圳.2016.11.25-27.安逸生活pk艰苦卓绝创业高原如此艰苦的环境：勇于担当，发挥引领作用以苦为乐，人生需要有一股精神特朗普：我拒绝政治正确，我只做正确的事情精英政治2016.10.11.中英脱髓鞘伦敦会议要点一些基本概念与影像学检查儿童脱髓鞘疾病临床分类成髓鞘细胞分类神经修复的路径各种细胞近期研究一览Year1977~201640年demyelinationremyelinationPubmed文献量趋势图髓鞘构成成分CNS髓鞘中60~70%为水分固体成分中脂类占70%,蛋白质占30%构成髓鞘蛋白质主要两种成分：蛋白脂蛋白(PLP)

和髓鞘碱性蛋白(MBP)髓鞘相关糖蛋白(MAG)髓鞘少突胶质细胞糖蛋白(MOG)Wolfram蛋白脂DM准大小蛋白其他微量蛋白髓鞘功能提供轴突与周围组织电绝缘，避免干扰相邻轴突之间轴突与其他结构之间加快动作电位传递：“跳跃式传导”机制引导受损轴突：再生Myelin-relatedcells(Schwanncellsandoligodendrocytes)co-operatewiththeaxonintheformationandmaintenanceofmyelinsheaths.中枢神经系统(CNS)有髓神经纤维与周围(PNS)相似结构有髓鞘和郎飞结髓鞘外无基膜（神经膜）相邻神经纤维有时融合（箭头）儿童发育髓鞘化过程主要集中在：出生后18个月内（1.5年）1岁半接近成人oligodendrocyte少突胶质细胞：大多数细胞5-10年成熟，存活>50年。每年减少1-300OLSThefinalstageofoligodendrocytedevelopmentismyelination.Unliketheperipheralnervoussystemwherethereisastrictsize-dependentbiasformyelination(onlyaxons>1umdiameterenjoytheprivilege),intheCNSaxonsassmallas300nmaresometimesmyelinated.Nevertheless,CNSmyelinationisalsoinfluencedbyaxonalsize.Surprisingly,culturedoligodendrocyteswillbeginmyelinatingevensyntheticaxon-liketubeslackingtheusualneuron-gliasignalingcues.Inthistypeof‘blind’myelination,onlytubeswithadiameterof≥400nmweremyelinated,suggestingthismodeofmyelinationmaybeparticularlyimportantforlargeraxons.Moreover,oncedifferentiating,itappearsthatoligodendrocytesonlyhaveaverynarrowwindowofopportunitytoselectwhichadjacentaxonstomyelinate(~5hinthedevelopingzebrafishand~12hinmyelinatingculturesofrodentderivedcells),irrespectiveofthetotalnumberofsheathsbeingmade.Thisimpliesthatthesignalingpathwaysmediatingthisprocessarelikelytoberelativelyrobust.Althoughinvitrostudieshaveimplicatedseveralkeypathways,todatenoonesinglemoleculehasbeenshowntobeindispensableformyelinationofCNSaxonsinvivo,highlightingthegreatdegreeofredundancyinthecontrolofthisvitalprocess.Despitethis,recentinvivostudieshaverevealedagreatdealabouthowthesevarioussignalingpathwaysconvergetocontroltheextentortimingofmyelination,ifnotoverallmyelinationperse.髓鞘脱失（脱髓鞘）

一种very常见的临床神经病理变化多指数T2/磁化转移MRIprovidesanimportantstepforunderstanding‘typical’myelindevelopmentaswellasprovidingtheabilitytoidentifywhenandwherewhitematterabnormalitiesoccurinneurodevelopmentaldisorders.myelinwatervolumefraction(MVF)髓磷脂体积分数myelinwaterfraction(MWF)髓磷脂水分脱髓鞘疾病分类（一）儿童脱髓鞘病：周围性和中枢性周围性脱髓鞘病：多以双下肢或四肢瘫痪为首发中枢性脱髓鞘病：以视力障碍、肢体无力、发热、抽搐、头痛等为首发脱髓鞘疾病分类（二）经典病种：多发性硬化、急性播散性脑脊髓炎、慢性炎性脱髓鞘性多神经根神经病其他病种：缺血缺氧性脑病、脑性瘫痪、脊髓损伤、脑外伤、脑白质营养不良等神经创伤、变性、遗传和血管性疾病Remyelinationcanoccurinthedamagedcentralnervoussystem(CNS)Sincethediscoveryinthe1960sthatremyelinationcanoccurinthedamagedCNS(Bungeetal.1961)therehasbeenmuchprogressinunderstandingthecellularandmolecularbiologyofoligodendrogliaandthefactorsthatregulatetheirpropagation,migration,differentiation,maturation,andabilitytomyelinatenerveaxons脊髓损伤脊髓挫伤1个月后，64％的存活纤维发生脱髓鞘尽管有些轴突保持着解剖上的完整，但已无生理功能损伤部位远端的轴突出现华勒变性FranklinRJ,etal.1997SpinalcordinjuryisaccompaniedbychronicprogressivedemyelinationTotoiuMO,KeirsteadHS.2005.

Extentofdemyelinationandremyelinationupto450daysfollowingcontusivespinalcordinjuryinadultrats1daypostinjury:theoverallnumberofdemyelinatedaxonspeaked7-14dayspostinjury:declinedthenprogressivelyincreasedupto450dayspostinjuryOligodendrocyteandSchwanncellremyelinatedaxonsappearedby14dayspostinjuryremyelinatedaxonswerepresentfrom14to450dayspostinjury,remyelinationwasincompletespinalcordinjuryisaccompaniedbychronicprogressivedemyelinationoligodendrocyteprogenitorcellproliferationgenerationofnewoligodendrocytesformationofthinnermyelinDominguesetal.2016.Dominguesetal.2016.Firstresearchontransplantationofmyelin-formingcellsintothedemyelinatedspinalcordThecellssurvive,migrate,andfindaxonsthatneedmyelininsulation,andremyelinatethemtherebyrestoringabilitytoconductimpulsesStephenWaxmanandJefferyKocsis细胞移植髓鞘化修复再生

治疗脱髓鞘病理的神经修复重要策略和研究方向嗅鞘细胞少突胶质细胞雪旺细胞多能诱导干（iPS）细胞神经干细胞胚胎干细胞脐带血/脐带间充质细胞骨髓间充质细胞脂肪干细胞皮肤干细胞Olfactoryensheathingcell(OEC)

嗅鞘细胞Remyelination（1）嗅鞘细胞髓鞘化修复类于雪旺细胞

PellitteriR,etal.(2010)OECs能产生多种神经营养生长因子体外：能促进轴突生长体内：可形成髓鞘，促进髓鞘再生刺激轴突再生发芽优于雪旺细胞：与星形胶质细胞接触（1）嗅鞘细胞髓鞘化修复类于雪旺细胞

BabiarzJ,etal.(2011)分离幼年和成年大鼠的嗅球，分析表达GFP的OEC的髓鞘化轴突的能力。OEC：幼鼠OECs能髓鞘化背根神经节（DRG）轴突。嗅鞘细胞与轴突组装成束需要1周，如果形成可以检测到的轴突髓鞘，需要1周以上SC：大鼠雪旺细胞不捆束轴突，在1周内能形成P0+和MBP+髓鞘节段大多数培养的OEC调宁蛋白（calponin）染色阳性，雪旺细胞为阴性几乎所有的OEC和雪旺细胞P75NTR和GFAP阳性两种细胞之间只有细微的免疫标记差异（1）嗅鞘细胞髓鞘化修复类于雪旺细胞

BabiarzJ,etal.(2011)ThediameterofOECgeneratedmyelinwasgreaterthanforSchwanncellmyelinonDRGaxonsOECbutnotSchwanncellsmyelinatedDRGaxonsintheabsenceofvitaminC

（2）嗅鞘细胞修复脑缺血卒中ShiX,etal.(2010)修复大脑中动脉闭塞（MCAO）大鼠脑白质MCAO后56天结果：减少梗死体积，降低死亡率，改善神经功能缺损LFB髓鞘染色，NF免疫组织化学，Westernblot：OEC移植大鼠髓鞘和轴突再生（3）嗅鞘细胞修复周围神经损伤RadtkeC,etal.(2010)嗅鞘细胞移植作为辅助治疗周围神经损伤嗅鞘细胞移植促进再生过程：介导趋化因子，神经营养和神经保护作用髓鞘形成：桥接，建立一个允许轴突再生的环境（4）嗅鞘细胞在体内、体外与其他细胞的相互作用ChuahMI,etal.(2010)与其他类型的细胞：在体外和嗅鞘细胞移植后的胶质疤痕和炎症环境下的相互作用嗅鞘细胞和星形胶质细胞克服胶质瘢痕的有害影响不同脊髓损伤的实验模型：OEC移植相关的胶质瘢痕的形态学改变在体外：嗅鞘细胞和胶质瘢痕的细胞类型组成之间的相互作用嗅鞘细胞：免疫细胞特性，移植到中枢神经系统损伤部位时，调制神经炎症

（5）嗅鞘细胞移植联合瘢痕切除修复脊髓挫伤ZhangSX,etal.(2011)疤痕消融+LP/OEC移植促进大鼠脊髓慢性挫伤解剖恢复和P0（髓磷脂糖蛋白,P-zero）阳性髓鞘孟加拉玫瑰红光毒性方法单独移植病灶腔：嗅黏膜固有层（LP）或联合体外培养的OECs（6）嗅鞘细胞移植修复肌萎缩侧索硬化LiY,etal.(2011)OEC移植到脊髓：延长SOD1（G93A）ALS大鼠生存期神经保护作用和髓鞘化移植的嗅鞘细胞存活超过4周，在脊髓内迁移4.2毫米（7）嗅鞘细胞移植修复各种脱髓鞘模型SasakiM,etal.(2011)不同的脱髓鞘环境对OEC髓鞘化修复的影响OECs的迁移和髓鞘形成：炎症处于活动状态炎症基本平息状态（7）嗅鞘细胞移植修复各种脱髓鞘模型(2)

AzimiAlamoutiM,etal.Remyelination

ofthecorpuscallosumby

olfactory

ensheathing

cellinanexperimentalmodelofmultiplesclerosis.(2015)（8）嗅鞘细胞移植修复效果的动物种属特异性WewetzerK,etal.(2011)

细胞增殖控制：种间差异鼠、狗、猪、猴、人Rodent：

requiremitogensforinvitroexpansionacomplexresponsetoelevatedintracellularcAMP,andundergospontaneousimmortalizationuponprolongedmitogenstimulation

（9）胚胎/新生/成年嗅鞘细胞移植修复效果存在差异CouttsDJ.Embryonic-derivedolfactoryensheathingcellsremyelinatefocalareasofspinalcorddemyelinationmoreefficientlythanneonataloradult-derivedcells.CellTransplant.2013

FormmyelinsheathsOptimaldonorageforOECassociatedremyelinationp75purifiedOECtransplantsfromthreedonoragesolfactorybulbsofembryonic,neonatal,andadultratsandpurifiedbyimmunopanningremyelinatingpotentialwasdirectlycomparedbytransplantationintothesameadultrattoxin-inducedmodelofspinalcorddemyelinationRemyelinationefficiency3weeksaftertransplantationwasassessedmorphologicallyandbyimmunostainingalldonoragesremyelinatethisprocessismostefficientlyachievedbyembryonic-derivedOECs.Oligodendrocyte少突胶质细胞Oligodendrocyteprecursorcells(OPCs),asubpopulationthataccountsfor5to8%ofcellswithinthecentralnervoussystem,arepotentialsourcesofoligodendrocytereplacementafterSCI.OPCsreactrapidlytoinjuries,proliferateatahighrate,andcandifferentiateintomyelinatingoligodendrocytes.However,posttraumaticendogenousremyelinationisrarelycomplete.WangY,etal.(2011)少突胶质祖细胞移植：在成年大鼠脊髓，趋向炎症区域存活分化成：可形成髓鞘的少突胶质细胞LiH,etal.(2009)

两层少突胶质细胞分化的转录调控“双管齐下”的方式：创建一个基因控制的故障安全系统确保在发育过程、脱髓鞘病变的修复过程中：髓鞘化修复有序进行和有效明确地表达IshiiA,etal.(2009)人类髓鞘蛋白质组学111种已确认的蛋白质/转录物：在少突胶质细胞表达，在星形胶质细胞和神经元不表达163additionalproteinscomplexityofthismetabolicallyactivemembraneSunF,etal.(2010)轴突变性对少突胶质细胞谱系细胞的影响：背根切断术唤起：修复反应脊髓挫伤后嘴侧轴突变性诱导：修复+细胞凋亡PiatonG,etal.(2010)在发育髓鞘、脱髓鞘、修复过程中，轴突与少突胶质细胞相互作用髓磷脂的生物合成和髓鞘修复：神经元和少突胶质细胞之间的相互沟通必不可少InMS,CNSdemyelinationisoftenfollowedbyspontaneousrepair,mostlyachievedbyadultoligodendrocyteprecursorcells.Extentofthismyelinrepairdiffers,rangingfromverylow,limitedtotheplaqueborder,toextensive,withremyelinationthroughoutthe'shadowplaques.'Inadditiontorestoringneuronalconnectivity,newmyelinisneuroprotective.Itreducesaxonallossandthusdisabilityprogression.

Neuralstemcell(NSC)神经干细胞HwangDH,etal.(2009)转导OLIG2转录因子的人类神经干细胞（NSCs）脊髓挫伤损伤后大鼠：提高运动功能的恢复增强脊髓白质髓鞘修复再生SherF,etal.(2009)生物发光成像（bioluminescenceimaging）Olig2-NSCs增加在脱髓鞘小鼠模型的植入效果YangJ,etal.(2010)比较骨髓和脑源性神经干细胞在中枢神经系统自身免疫性疾病的治疗效果类似asimilarabilitytodifferentiateintoneurons,astrocytes,andoligodendrocytesbothinvitroandinvivobothtypesofNSCssuppressedchronicexperimentalautoimmuneencephalomyelitistherapeuticeffectsofNSCsincludeimmunomodulationinthePNSandtheCNS,neuron/oligodendrocyterepopulationbytransplantedcells,andenhancedendogenousremyelinationandaxonalrecoveryCarbajalKS,etal.(2010)多发性硬化病毒模型神经干细胞迁移通过CXCR4介导CXCL12的信号Adiposemesenchymalstemcell脂肪来源间充质干细胞RadtkeC,etal.(2009)脂肪间充质干细胞衍生的神经球可分化成周围神经胶质样细胞ConstantinG,etal.(2009)脂肪来源的间质干细胞改善慢性实验性自身免疫性脑脊髓炎(EAE)静脉给药：显著降低免疫反应所致EAE的严重性、减少脊髓炎和脱髓鞘、轴突损失ASC优先归巢到淋巴器官，迁移中枢神经系统双峰机制的治疗潜力：在疾病的早期阶段：抑制自身免疫反应诱导内源性祖细胞的神经再生AgeoftheDonorReducestheAbilityofHumanAdipose-DerivedStemCellstoAlleviateSymptomsintheExperimentalAutoimmuneEncephalomyelitisMouseModelASCsfromolderdonorsfailedtoamelioratetheneurodegenerationassociatedwithEAE,andmicetreatedwitholderdonorcellshadincreasedCNSinflammation,demyelination,splenocyteproliferationinvitrocomparedwiththemicereceivingcellsfromyoungerdonors.STEMCELLSTRANSLATIONALMEDICINE2013;2:797

GhasemiN.Transplantationofhumanadipose-derivedstemcellsenhancesremyelinationinlysolecithin-inducedfocaldemyelinationofratspinalcord.MolBiotechnol.2014.

transplantedhumanADSCs(hADSCs)intoalysophosphatidylcholine(lysolecithin)modelofmultiplesclerosis(MS)anddeterminedtheefficiencyofthesecellsinremyelinationprocess.Fortyadultratswererandomlydividedintocontrol