切除卵巢后大鼠骨小梁重建组织学特征的电镜观察

切除卵巢后大鼠骨小梁重建组织学特征的电镜观察 切除卵巢后大鼠骨小梁重建组织学特征的 电镜观察 J 中 . 国 urn 组 a 织 lof 工 C 程 lin 研 ica 究 lR 与 e 临 ha 床 bil 康 ita 复 tive 第 Tiss ,, u 卷 eE 第ngi'nee朋 ring 2 R O e O s 7 e — a O r 8ch一1 A 2 u 出 gu 版 st1.2一,2DD7.V.OL71一.N—O.32zLIult?,,2 Electro-microscopicobservationoftrabecularbone remodelinginovariectomizedrats? .】uChuan—guang,MaQing—jun,DangGeng'ding,WangXiao'ying Abstract BACKGROUND:Thechangesintrabecularbonemicroarchitectureinosteoporosishavearousedmuchattention.The decreaseinthenumberoftrabecularnodesandincreaseinthenumberoffreeendsarefoundinosteoporosis,butthe mechanismisstilIunclear. OB]ECTIVE:Toobservethetrabecularremodelingprocessinovariectomizedratsastheosteoporosismodelselectron microscopically,andtoexplorethereasonsforthedecreaseinthenumberoftrabecularnodesandincreaseinthe numberoffreeends. DESIGN:RandomizedandcontrolledanimaItriaI SETTING:DepartmentofOrthopedics,ThirdHospitalofPekingUniversity MATERIALS:TheexpenmentwasconductedintheAnimalLaboratory.ThirdHospitalofPekingUniversityfr0m September1999toFebmary2000.Thirty-sixfemaleWistarratsof3monthsoldand240-280gwereselectedand randomlydividedintoovariectomizedgroupandcontrolgroupwith18ratsineachgroup.Theratswereobservedat4, 8,and12weeksp0st0peratiVeIywith6ratsateachtimepoint. METHODS:Theratsofovariectomizedgroupweresubjectedtoovariectomy1weekafterfeeding.butthecontrolgroup wasnot.Thechangesofproximaltibiatrabecularmicroarchitecturewasobservedunderscanningelectronmicroscope at4,8and12weeks,respectively,andtheosteoclast,osteoblast,andstructureofcellorganswereobservedunder transmissionelectronmicroscope. MAINOUTCOMEMEASURES:? Theremodelingprocessafterovadectomybyelectronmicroscope;?morphological changesoftrabecularbone. 'Wendeng OrthopedicsHospital, Wendeng264400, ShandongProvince. China;2Departmentof Orthopedics.Third HospitalofPeking University.Beijing 264400.China JuChuan-guang?. Master.Associate Chietphys~c!an. WendengOrthopedics Hospita1.Wendeng 264400,Shandong Province.China wfsw@163cem Received:2007-03-05 Accepted:2007-06-04 (06-50-12-9467/H) JuCG.MaQJ.Dang GD.WangXY Electro-microscopic observationof trabecularbone remodelingin ovadectomizedra协 ZhongguoZuzhi GongchengYanjiuyu LinchuangKangfu 2007;11(32): 6509-6512(China) RESULTS:Scanningelectronmicroscopeobservationshowedthattrabecularboneremodelingwasdistributedin.zglckfcom/ everyregionoftrabecularmicroarchitecture,especiallyStandNd— Stregion.Afterovariectomy.thetransversetrabecularzglckf/ejoumal/ waseaS.e? 0perforatedandbroken;thetrabecularnetworkwasaim0stintactat4weekslbulgraduallydamageaat:2pfiles/0097(-ps)32/. pdf]weeks8and12:moreover ,thecollagenfibersonthesurfaceoftrabecularbonewerescrappy.disorderandthinner.?……' Bythetransmissionelectronmicroscopicstudy.thetibiaIosteoclastwerefoundactiveat12weeks.Whenabsorbing cancellousbone,osteoclastcloselyadheredtoitssurface,anddigitationsstretchedintothecancellousbone.Theshape andsizeofdigitationsweresignificantlydliferent,andaroundthem,Iucentareawasobserved.Osteoclastwas pelynucleationwithabundantkytoplasm,andtherewereplentyofGolgicomplex,smoothendoplasmicreticulumand mitochondnum.Lysosomeinclusioncompoundswithdifferentsizesandelectrondensitywerefoundincells.Osteoblast wasrarelyfound,andcelledgewasrough,withbonelacuna. CONCLUSION:BoneremodelingissignificantlyactiveinStandNd-Stregionoftrabecularboneinovanectomizedrats Thismaybethereasonforthedecreaseinthenumberoftrabecularnodesandincreaseinthenu mberoffreeends. INTRoDUCTIoN Atpresent,thechangesintrabecularmicroarchitecture dunngboneremodelinghavebecomeafocus.Inthis study,theosteoporosismode1wasestablishedby ovadectomytofindoutthereasonsforthedecreaseinthe numberoftrabecularnodesandIncreaseinthenumberof freeendsbyobservingtheboneremodelingprocessand morphologicalchangesintrabecularboneunderelectron microscopes.soastooffersomeheIpfulinformationfor thepreventionandtreatmentofosteoporosis. MATERIALSANDMETH0DS Materials TheexperimentwasconductedintheAnimalLaboratory 沈阳1200邮政信箱110004kf23385083@sina.comwww.zglckf.com ThirdHospitalofPekingUniversityfr0mSeptember1999 toFebruary2000.Thirty-six3-month-oldfemaleWistarrats ofcleangradeand240—280g(animallaboratory.MedicaI DepartmentofPekingUniversity.No.SYXK1999.025) wereselectedandrandomlydividedintoovariectomized groupandcontrolgroupwith18ratsineachgroup.The ratswereobservedat4,8,and12weekspostoperatively Ih6ratsateachtimepoint.A?animalswerefeedwith standarddiet,anddrankfreely. Methods Grouping:Theratsofovadectomizedgroupwere subjectedtoovadectomy1weekafterfeeding,butthe controlgroupwasnot.Thentheratswerefeedseparately withnoantibiotics.Four,eightandtwelveweeksafter ovariectomy,theanimalswereexecutedandsampled. Samplepreparationandobservationbyscanning electronmicroscope:TheIeftproximaltibiawasincised 6509 ISSN1673.8225CN21-1539/R 鞠传厂,等切除日巢后大鼠骨小梁重建组织学特征的电镜观察 ww~zg/ckf.comkf23385083@sina,com alongcoronalaxis.andwashedwithnormalsalineuntilthe trabecularbonecouldbeobservedclearly.Cancellousbonewas takenandcutintospecimensof1mm×2mm×3mm.The specimenswerefixed,dehydrated.displaced,driedandvacuum sprayed,thenobservedunder$450scanningelectronmicroscope andshot. Samplepreparationandobservationbytransmissionelectron microscope:Therightproximaltibiawasincisedalongthecoronal axis.washedwithnormalsaline.andcutintospecimensof1mmx 1mmx1mm.Thespecimensweresubjectedtofixation. dehydration,embedding,semithinsectionpreparation,ultrathin sectionpreparationandstaining,thenobservedunder SEM-JEOL5600electronmicroscopeandshot. DivisionoftrabecularmicroarchitecturewasshowninFigure1 1:Node(Nd):2:Node-Strut(Nd-st):3:Strut(st);4:Strut-Terminus (St-Tm);5:Terminus(Tm) Figure1Microarchitectureareaoftrabecularbone RESULTS Scanningelectronmicroscopeobservation lncontrolgroup.thecancellousboneatproximaltibiawascomposed ofcolumnorslabshapedtrabecularbonehorizontaltothey-axisof tibiaandverticaItotheX-axisoftibia.Thetrabecularbonewas arrangedregularlyandintenselywiththickverticaloneandthin horizontalone.formingathree-dimensionalnetworkstructure.Ofthe areasoftrabecularbone,Ndareawasthick,smoothandglossy; Nd-Stareawaslittlebreak.andsmooth;Stareawasperfectlyround, andsmoothwithuniformthickness;Tmareapresentedregular borderlinewithrarespaceormicro-fracture.Thetrabecularbonewas coveredwithun-mineralizedregularanddensecollagenfibrils, arrangingparallellyatthesameplate,andcrossingbetweentwo plates.Boneabsorptionandboneformationsurfacewereobserved locallyinsomeareas.withshallowabsorptionlacuna.Osteoblast coveredonthesurfaceofreconstructedbone,andtheirappearance andstructureinlacuneweredifferentateachstageTheywerefullat productivesecretionstage,packagingbyprocollagenandmineralized crystalgranules,andtherewereplentyofprocollagen,tropocollagen andcrystalgranulesandsmallamountofcollagenicfibrils(Figure2). Repairedcollagenfibrilswereparalleltothesameplate,andfew osteoclastswerefound. Figure2Osteoblastofproductive (Scanningelectronmicroscope,x4 6510 functionincontrolgroup(T) 500) lnovariectomizedgroup,thetrabecularbonewascolumnorslab shaped.but4,8.and12weeksafterovariectomy,trabecularbone turnedsparsegradually,'andtrabecularnetworkstructurewas damagedandfinallydisappeared.Theverticaltrabecularbonewent thinner.andsomeofthehorizontaltrabecularboneturnedinto ice—coneormicro-rodorshortconeshapedevendisappeared (Figure3) a:Ice—conoidtrabecularbone(t) b:Micro—rodtrabecularbone(t) c:Shortconoidtrabecularbone(t) Figure3Ultramicrostructureofbonetrabecular12weeksafter ovariectomy(Scanningelectronmicroscope,x300) BoneremodelingandabsorptionIacunawerefoundatthesurfaceof trabecularboneat4weeks.TheIacunawithclearstructurewas coveredwithMR—mineralizeddensecollagenfibrilslayer.foIlowingby thebrokenendsofabsorbedlayeroflamellarbonematrix.Theedge ofbrokenendswasuneven.andthesidelinewasindifferentshapes. ThelayerareaoflacunawasdiminishedgraduallywiththedeDlh0f lacuna.Abonelacunawasfoundatthebottomofabsorptionlacuna. withnoosteocyte.andscrambledandscarcefibrilswerefoundonits surface.Therewasmicro-fractureinlamellaroneatdifferentlayers. stretchingintothebottomofabsorptionlacuna.Thebreak—overof Nd-Stareawasunevenorobviouslysunken;atStarea.thesurface oftrabecularbonewasrough,andsunken:thespacebetweenTm wasincreased,butstillwithcompletenetworkstructureuntilthe8 week.Atthe12weekthenetworkstructurehadbeendamaged severelyandintopieces;trabeculararchitecturearoundnodes disappeared.theverticaltrabecularbonewasobviouslythinner.and horizontalonebecamethinner,perforatedevenbroken(Figure4a). Sometrabecularbonespresentedmicro-rod.ice-conoid.shortconoid orshortrodshapewithnomineralizedcollagenfibrils.The un-mineralizedcollagenfibrilsonthesurfaceoftrabecularbone turnedsparsegraduallyuntiIdisappearedatweeksof4.8.and12 (Figure4b).Exposedmineralizedmatrixwasfoundinlargeareaat P.O.Box1200.Shenyang110004kf23385083@sina.comwww.zglckf.com ISSN1673.8225CN21-1539/R 鞠传广.等.切除卵巢后大鼠骨小梁重建组织学特征的电镜观察 P,w~zg/ckf.cornkf23385083~sina.com endswereincreased.Thereasonsforthechangesincollagenfibrils onthesurfaceoftrabecularbonemightberelatedtotheincreasein osteoclastactivationbutdecreaseinosteoblastfunction.When osteoblastrepairingabsorptionlacuna,thecollagenfibrilswere arrangedinasameorientationtothelayer.Therepairedtissue coveredonlacunaexceededthesurface.1nthisstudy.thebottomof eachabsorptionlacunawasconnectedwithonebonelacuna.Butifit iscommonnessandwhatisthesignificanceofthiscommonnessstill needfurtherinvestigation. REFERENCES 1SeemanE.Thestructurebasisofbonefragilityinmen.Bone1999;25: 143_147 2JensenKS.MosekildeLI.AmodeIofvertebratrabecularbone architectureanditsmechanicaIproperties.Bone1990:11:417-423 30dgoadA.GundersonHJG.QuanUtationofconnectivityincancellous bone.withspeciaIemphasison3.Dreconstructions.Bone1993;14:173 4DaiKR.Tang-i-r.xueWD.eta1.Thechangesofparametersof cancellousboneconnectivityofosteoporosisandcomputer-aided mP_.asurem~t.ZhonggcoGuzhishusongZazhi2O00;6(1):19-22 5CompstonJE.Connectivityofcancellousboneassessmentand mechanicaIimplications.Bone1994:15(6):463-466 6HahnM.VogelM,Pompesius-KempaM,eta1.Trabecularbonepaffem factor:.anewparameterforsimplequant~cationofbone microarohitecture.Bone1992;13:327 切除卵巢后大鼠骨小梁重建组织学特征的 电镜观察? 鞠传广',马庆军,党耕町,王晓英' '文登整骨医院,山东省文登市264400;北京大学第三医院骨科,北京 市264400 鞠传广?,男,1967年生,山东省文登市人,汉族,2000年北京大学毕 业,硕士,副主任医师,主要从事脊柱脊髓疾病及骨质疏松等的研究. 摘要 背景:骨小梁微构筑在骨质疏松中的变化已引起广泛关注,骨质疏松后 骨小梁的节点数减少,游离末端数增加,但机制尚不明确. 目的:观察卵巢切除大鼠骨质疏松模型骨小梁重建的电镜变化,分析骨 小梁节点数减少而游离末端数增加的原因. :随机对照动物实验. 单位:北京大学第三医院骨科. 材料:实验于1999—09/2000—02在北京大学第三医院动物实验室完成. 选用36只3月龄雌性Wistar大鼠,体质量240-280g.采用随机摸球 法分为卵巢切除组和对照组,每组18只,分别于术后4,8,12周3个时 间点进行观察,每时间点6只. 方法:卵巢切除组饲养1周后双侧卵巢切除.对照组不切除卵巢.于术后 4,8,12周采用扫描电镜观察胫骨近干骺端骨小梁各区形态结构的变 化;采用透射电镜对术后12周胫骨近干骺端骨小梁面破骨细胞,成骨 细胞及细胞器的结构变化进行观察. 主要观察指标:?通过电镜对骨小梁微构筑的分区观察分析去卵巢后骨 的重建过程.(观察骨小梁的形态结构变化. 结果:(量)扫描电镜显示:骨小梁骨重建活动分布于骨小梁微构筑各个部 位,但以st,Nd-st区最显着:卵巢切除后骨小梁穿孔,断裂多见于水平 骨小梁,骨小梁网状结构4周时完整,第8周和第12周后逐渐被破坏, 12周最严重:卵巢切除后骨小梁表面的胶原纤维逐渐变得杂乱,稀薄.? 透射电镜显示:卵巢切除后12周大鼠胫骨破骨细胞的形态结构观察发 现,功能活跃的破骨细胞较多见.破骨细胞对骨质进行吸收时,紧紧贴附 于骨质表面,将指状突起伸向骨质深处,指状突起形状,大小相差悬殊. 指状突起周围可见透亮区.破骨细胞多核,胞浆丰富,胞浆内含有发达的 高尔基复合体,滑面内质网和大量的线粒体.细胞体内见大小不等,电子 密度不均的溶酶体包含物.成骨细胞少见,胞体边缘不光滑,周围出砚骨 陷窝轮廓. 结论:卵巢切除后骨小梁st,Nd-st区骨重建最活跃,这可能是骨质疏松 时骨小梁节点数减少而游离末端数增加的原因. 关键词:卵巢切除术;骨质疏松;电子显微镜检查;动物,试验 中图分类号:R587.1文献标识码:B文章编号:1673-8225(2007)32-06509-04 鞠传广,马庆军,党耕町,王晓英.切除卵巢后大鼠骨小梁重建组织学特征的电镜观 察【J】.中国组织工程研究与临床康复,2007,11(32):6509-6512 【WWW.zglckf.comlzglckflejoumallupfileslO7-32132k-6509(ps).嗍 (EditedbyMaQ.ZhouCS/SuLL/WangL) 壁匿王堡丝查 骨髓间质干细胞是存在于骨 髓,脐带血,肌肉,骨骼,软骨,脂 肪,血管等的多能干细胞.在体外 培养时,细胞贴壁生长,呈成纤维样 细胞表型,在一定条件下可分化内 胚层,中胚层和外胚层细胞.因骨 髓间质干细胞具有易分离扩增,基 因转染,支持体外造血,体内造血 6512 缉织I程研究与绉床康复》杂志社所 重建以及免疫调节作用等特点,在 临床上有很大的潜在应用价值.骨 髓问质干细胞能分化成骨,脂肪, 软骨和其他种类细胞. 移植物收缩到70%并失去了原有 的外形和体积.研究者曾试图应用 脂肪抽吸术后游离的脂肪细胞,但 是这些细胞的寿命也很短. 宣筮壁蕉垒皇 现今,外科医生经常应用自体(信息来源:) 组织移植,这会导致供区的损伤.(编辑:李眷会) 移植的细胞经常不能存活,这使得 P.o.Box1200.Shenyang110004kf23385083@sina.comv~vlglckf.com