成髓鞘細胞研究和應用進展

成髓鞘細胞研究和應用進展陳琳清華大學第二附屬醫(yī)院神經(jīng)外科中心chenlin_china@163.com第一屆全國兒童神經(jīng)修復學術會議暨第四屆全國兒童神經(jīng)康復論壇.深圳.2016.11.25-27.安逸生活pk艱苦卓絕創(chuàng)業(yè)高原如此艱苦的環(huán)境：勇于擔當，發(fā)揮引領作用以苦為樂，人生需要有一股精神特朗普：我拒絕政治正確，我只做正確的事情精英政治2016.10.11.中英脫髓鞘倫敦會議要點一些基本概念與影像學檢查兒童脫髓鞘疾病臨床分類成髓鞘細胞分類神經(jīng)修復的路徑各種細胞近期研究一覽Year1977~201640年demyelinationremyelinationPubmed文獻量趨勢圖髓鞘構成成分CNS髓鞘中60~70%為水分固體成分中脂類占70%,蛋白質占30%構成髓鞘蛋白質主要兩種成分：蛋白脂蛋白(PLP)

和髓鞘堿性蛋白(MBP)髓鞘相關糖蛋白(MAG)髓鞘少突膠質細胞糖蛋白(MOG)Wolfram蛋白脂DM準大小蛋白其他微量蛋白髓鞘功能提供軸突與周圍組織電絕緣，避免干擾相鄰軸突之間軸突與其他結構之間加快動作電位傳遞：“跳躍式傳導”機制引導受損軸突：再生Myelin-relatedcells(Schwanncellsandoligodendrocytes)co-operatewiththeaxonintheformationandmaintenanceofmyelinsheaths.中樞神經(jīng)系統(tǒng)(CNS)有髓神經(jīng)纖維與周圍(PNS)相似結構有髓鞘和郎飛結髓鞘外無基膜（神經(jīng)膜）相鄰神經(jīng)纖維有時融合（箭頭）兒童發(fā)育髓鞘化過程主要集中在：出生后18個月內（1.5年）1歲半接近成人oligodendrocyte少突膠質細胞：大多數(shù)細胞5-10年成熟，存活>50年。每年減少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.髓鞘脫失（脫髓鞘）

一種very常見的臨床神經(jīng)病理變化多指數(shù)T2/磁化轉移MRIprovidesanimportantstepforunderstanding‘typical’myelindevelopmentaswellasprovidingtheabilitytoidentifywhenandwherewhitematterabnormalitiesoccurinneurodevelopmentaldisorders.myelinwatervolumefraction(MVF)髓磷脂體積分數(shù)myelinwaterfraction(MWF)髓磷脂水分脫髓鞘疾病分類（一）兒童脫髓鞘?。褐車院椭袠行灾車悦撍枨什。憾嘁噪p下肢或四肢癱瘓為首發(fā)中樞性脫髓鞘?。阂砸暳φ系K、肢體無力、發(fā)熱、抽搐、頭痛等為首發(fā)脫髓鞘疾病分類（二）經(jīng)典病種：多發(fā)性硬化、急性播散性腦脊髓炎、慢性炎性脫髓鞘性多神經(jīng)根神經(jīng)病其他病種：缺血缺氧性腦病、腦性癱瘓、脊髓損傷、腦外傷、腦白質營養(yǎng)不良等神經(jīng)創(chuàng)傷、變性、遺傳和血管性疾病Remyelinationcanoccurinthedamagedcentralnervoussystem(CNS)Sincethediscoveryinthe1960sthatremyelinationcanoccurinthedamagedCNS(Bungeetal.1961)therehasbeenmuchprogressinunderstandingthecellularandmolecularbiologyofoligodendrogliaandthefactorsthatregulatetheirpropagation,migration,differentiation,maturation,andabilitytomyelinatenerveaxons脊髓損傷脊髓挫傷1個月后，64％的存活纖維發(fā)生脫髓鞘盡管有些軸突保持著解剖上的完整，但已無生理功能損傷部位遠端的軸突出現(xiàn)華勒變性FranklinRJ,etal.1997SpinalcordinjuryisaccompaniedbychronicprogressivedemyelinationTotoiuMO,KeirsteadHS.2005.

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

治療脫髓鞘病理的神經(jīng)修復重要策略和研究方向嗅鞘細胞少突膠質細胞雪旺細胞多能誘導干（iPS）細胞神經(jīng)干細胞胚胎干細胞臍帶血/臍帶間充質細胞骨髓間充質細胞脂肪干細胞皮膚干細胞Olfactoryensheathingcell(OEC)

嗅鞘細胞Remyelination（1）嗅鞘細胞髓鞘化修復類于雪旺細胞

PellitteriR,etal.(2010)OECs能產(chǎn)生多種神經(jīng)營養(yǎng)生長因子體外：能促進軸突生長體內：可形成髓鞘，促進髓鞘再生刺激軸突再生發(fā)芽優(yōu)于雪旺細胞：與星形膠質細胞接觸（1）嗅鞘細胞髓鞘化修復類于雪旺細胞

BabiarzJ,etal.(2011)分離幼年和成年大鼠的嗅球，分析表達GFP的OEC的髓鞘化軸突的能力。OEC：幼鼠OECs能髓鞘化背根神經(jīng)節(jié)（DRG）軸突。嗅鞘細胞與軸突組裝成束需要1周，如果形成可以檢測到的軸突髓鞘，需要1周以上SC：大鼠雪旺細胞不捆束軸突，在1周內能形成P0+和MBP+髓鞘節(jié)段大多數(shù)培養(yǎng)的OEC調寧蛋白（calponin）染色陽性，雪旺細胞為陰性幾乎所有的OEC和雪旺細胞P75NTR和GFAP陽性兩種細胞之間只有細微的免疫標記差異（1）嗅鞘細胞髓鞘化修復類于雪旺細胞

BabiarzJ,etal.(2011)ThediameterofOECgeneratedmyelinwasgreaterthanforSchwanncellmyelinonDRGaxonsOECbutnotSchwanncellsmyelinatedDRGaxonsintheabsenceofvitaminC

（2）嗅鞘細胞修復腦缺血卒中ShiX,etal.(2010)修復大腦中動脈閉塞（MCAO）大鼠腦白質MCAO后56天結果：減少梗死體積，降低死亡率，改善神經(jīng)功能缺損LFB髓鞘染色，NF免疫組織化學，Westernblot：OEC移植大鼠髓鞘和軸突再生（3）嗅鞘細胞修復周圍神經(jīng)損傷RadtkeC,etal.(2010)嗅鞘細胞移植作為輔助治療周圍神經(jīng)損傷嗅鞘細胞移植促進再生過程：介導趨化因子，神經(jīng)營養(yǎng)和神經(jīng)保護作用髓鞘形成：橋接，建立一個允許軸突再生的環(huán)境（4）嗅鞘細胞在體內、體外與其他細胞的相互作用ChuahMI,etal.(2010)與其他類型的細胞：在體外和嗅鞘細胞移植后的膠質疤痕和炎癥環(huán)境下的相互作用嗅鞘細胞和星形膠質細胞克服膠質瘢痕的有害影響不同脊髓損傷的實驗模型：OEC移植相關的膠質瘢痕的形態(tài)學改變在體外：嗅鞘細胞和膠質瘢痕的細胞類型組成之間的相互作用嗅鞘細胞：免疫細胞特性，移植到中樞神經(jīng)系統(tǒng)損傷部位時，調制神經(jīng)炎癥

（5）嗅鞘細胞移植聯(lián)合瘢痕切除修復脊髓挫傷ZhangSX,etal.(2011)疤痕消融+LP/OEC移植促進大鼠脊髓慢性挫傷解剖恢復和P0（髓磷脂糖蛋白,P-zero）陽性髓鞘孟加拉玫瑰紅光毒性方法單獨移植病灶腔：嗅黏膜固有層（LP）或聯(lián)合體外培養(yǎng)的OECs（6）嗅鞘細胞移植修復肌萎縮側索硬化LiY,etal.(2011)OEC移植到脊髓：延長SOD1（G93A）ALS大鼠生存期神經(jīng)保護作用和髓鞘化移植的嗅鞘細胞存活超過4周，在脊髓內遷移4.2毫米（7）嗅鞘細胞移植修復各種脫髓鞘模型SasakiM,etal.(2011)不同的脫髓鞘環(huán)境對OEC髓鞘化修復的影響OECs的遷移和髓鞘形成：炎癥處于活動狀態(tài)炎癥基本平息狀態(tài)（7）嗅鞘細胞移植修復各種脫髓鞘模型(2)

AzimiAlamoutiM,etal.Remyelination

ofthecorpuscallosumby

olfactory

ensheathing

cellinanexperimentalmodelofmultiplesclerosis.(2015)（8）嗅鞘細胞移植修復效果的動物種屬特異性WewetzerK,etal.(2011)

細胞增殖控制：種間差異鼠、狗、豬、猴、人Rodent：

requiremitogensforinvitroexpansionacomplexresponsetoelevatedintracellularcAMP,andundergospontaneousimmortalizationuponprolongedmitogenstimulation

（9）胚胎/新生/成年嗅鞘細胞移植修復效果存在差異CouttsDJ.Embryonic-derivedolfactoryensheathingcellsremyelinatefocalareasofspinalcorddemyelinationmoreefficientlythanneonataloradult-derivedcells.CellTransplant.2013

FormmyelinsheathsOptimaldonorageforOECassociatedremyelinationp75purifiedOECtransplantsfromthreedonoragesolfactorybulbsofembryonic,neonatal,andadultratsandpurifiedbyimmunopanningremyelinatingpotentialwasdirectlycomparedbytransplantationintothesameadultrattoxin-inducedmodelofspinalcorddemyelinationRemyelinationefficiency3weeksaftertransplantationwasassessedmorphologicallyandbyimmunostainingalldonoragesremyelinatethisprocessismostefficientlyachievedbyembryonic-derivedOECs.Oligodendrocyte少突膠質細胞Oligodendrocyteprecursorcells(OPCs),asubpopulationthataccountsfor5to8%ofcellswithinthecentralnervoussystem,arepotentialsourcesofoligodendrocytereplacementafterSCI.OPCsreactrapidlytoinjuries,proliferateatahighrate,andcandifferentiateintomyelinatingoligodendrocytes.However,posttraumaticendogenousremyelinationisrarelycomplete.WangY,etal.(2011)少突膠質祖細胞移植：在成年大鼠脊髓，趨向炎癥區(qū)域存活分化成：可形成髓鞘的少突膠質細胞LiH,etal.(2009)

兩層少突膠質細胞分化的轉錄調控“雙管齊下”的方式：創(chuàng)建一個基因控制的故障安全系統(tǒng)確保在發(fā)育過程、脫髓鞘病變的修復過程中：髓鞘化修復有序進行和有效明確地表達IshiiA,etal.(2009)人類髓鞘蛋白質組學111種已確認的蛋白質/轉錄物：在少突膠質細胞表達，在星形膠質細胞和神經(jīng)元不表達163additionalproteinscomplexityofthismetabolicallyactivemembraneSunF,etal.(2010)軸突變性對少突膠質細胞譜系細胞的影響：背根切斷術喚起：修復反應脊髓挫傷后嘴側軸突變性誘導：修復+細胞凋亡PiatonG,etal.(2010)在發(fā)育髓鞘、脫髓鞘、修復過程中，軸突與少突膠質細胞相互作用髓磷脂的生物合成和髓鞘修復：神經(jīng)元和少突膠質細胞之間的相互溝通必不可少InMS,CNSdemyelinationisoftenfollowedbyspontaneousrepair,mostlyachievedbyadultoligodendrocyteprecursorcells.Extentofthismyelinrepairdiffers,rangingfromverylow,limitedtotheplaqueborder,toextensive,withremyelinationthroughoutthe'shadowplaques.'Inadditiontorestoringneuronalconnectivity,newmyelinisneuroprotective.Itreducesaxonallossandthusdisabilityprogression.

Neuralstemcell(NSC)神經(jīng)干細胞HwangDH,etal.(2009)轉導OLIG2轉錄因子的人類神經(jīng)干細胞（NSCs）脊髓挫傷損傷后大鼠：提高運動功能的恢復增強脊髓白質髓鞘修復再生SherF,etal.(2009)生物發(fā)光成像（bioluminescenceimaging）Olig2-NSCs增加在脫髓鞘小鼠模型的植入效果YangJ,etal.(2010)比較骨髓和腦源性神經(jīng)干細胞在中樞神經(jīng)系統(tǒng)自身免疫性疾病的治療效果類似asimilarabilitytodifferentiateintoneurons,astrocytes,andoligodendrocytesbothinvitroandinvivobothtypesofNSCssuppressedchronicexperimentalautoimmuneencephalomyelitistherapeuticeffectsofNSCsincludeimmunomodulationinthePNSandtheCNS,neuron/oligodendrocyterepopulationbytransplantedcells,andenhancedendogenousremyelinationandaxonalrecoveryCarbajalKS,etal.(2010)多發(fā)性硬化病毒模型神經(jīng)干細胞遷移通過CXCR4介導CXCL12的信號Adiposemesenchymalstemcell脂肪來源間充質干細胞RadtkeC,etal.(2009)脂肪間充質干細胞衍生的神經(jīng)球可分化成周圍神經(jīng)膠質樣細胞ConstantinG,etal.(2009)脂肪來源的間質干細胞改善慢性實驗性自身免疫性腦脊髓炎(EAE)靜脈給藥：顯著降低免疫反應所致EAE的嚴重性、減少脊髓炎和脫髓鞘、軸突損失ASC優(yōu)先歸巢到淋巴器官，遷移中樞神經(jīng)系統(tǒng)雙峰機制的治療潛力：在疾病的早期階段：抑制自身免疫反應誘導內源性祖細胞的神經(jīng)再生AgeoftheDonorReducestheAbilityofHumanAdipose-DerivedStemCellstoAlleviateSymptomsintheExperimentalAutoimmuneEncephalomyelitisMouseModelASCsfromolderdonorsfailedtoamelioratetheneurodegenerationassociatedwithEAE,andmicetreatedwitholderdonorcellshadincreasedCNSinflammation,demyelination,splenocyteproliferationinvitrocomparedwiththemicereceivingcellsfromyoungerdonors.STEMCELLSTRANSLATIONALMEDICINE2013;2:797

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

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