中华创伤骨科杂志
中華創傷骨科雜誌
중화창상골과잡지
CHINESE JOURNAL OF ORTHOPAEDIC TRAUMA
2010年
6期
551-556
,共6页
姜晓锐%张鑫鑫%肖剑晖%金丹%江汕%王丹%赵培冉%裴国献
薑曉銳%張鑫鑫%肖劍暉%金丹%江汕%王丹%趙培冉%裴國獻
강효예%장흠흠%초검휘%금단%강산%왕단%조배염%배국헌
骨髓细胞%成骨细胞%雪旺细胞%分化%共培养
骨髓細胞%成骨細胞%雪旺細胞%分化%共培養
골수세포%성골세포%설왕세포%분화%공배양
Bone marrow cells%Osteoblasts%Schwann cells%Differentiation%Coculture
目的 研究应用雪旺细胞(SCs)对两种来源的成骨细胞(OB)的增殖与分化的影响,为构建神经化组织工程骨提供体外实验理论依据.方法 通过采用股骨、胫骨髓腔冲洗法获得SD大鼠骨髓基质干细胞(BMSCs),采用胰蛋白酶消化新生鼠颅骨获得OB及采用消化后组织块法获得SCs.将BMSCs在成骨诱导液作用3周,鉴定BMSCs向OB分化.采用96孔共培养板将第2代SCs种植于上室,颅骨来源OB种植下室,共培养为实验组,无SCs干预共培养为对照组.将诱导分化的OB种植于下室,第2代SCs种植于上室,共培养为实验组,无SCs干预共培养为对照组,分别于共培养后1、3、5、7、9d采用甲基噻唑基四唑(MTT)进行增殖比较.采用6孔共培养板,实验组上室种植SCs,下室分别种植颅骨来源及BMSCs来源的OB,不加SCs的两种来源的OB作为对照组,分别共培养后于3、7 d检测两种来源OB的碱性磷酸酶、骨钙素、骨桥蛋白、骨形态发生蛋白-2 mRNA表达.结果 SCs对颅骨来源的OB在共培养3、5、7、9 d 4个时间点均有明显促增殖作用,在碱性磷酸酶、骨桥蛋白、骨形态发生蛋白-2mRNA在各个时间段均起到抑制作用.SCs对大鼠BMSCs来源的OB在共培养1、3 d无明显促增殖作用,在5、7、9 d有明显促增殖作用,在成骨培养基的环境里,SCs可促进BMSCs诱导的OB分化.结论 选择BMSCs来源的OB 与 SCs共培养更适合用于构建神经化组织工程骨.
目的 研究應用雪旺細胞(SCs)對兩種來源的成骨細胞(OB)的增殖與分化的影響,為構建神經化組織工程骨提供體外實驗理論依據.方法 通過採用股骨、脛骨髓腔遲洗法穫得SD大鼠骨髓基質榦細胞(BMSCs),採用胰蛋白酶消化新生鼠顱骨穫得OB及採用消化後組織塊法穫得SCs.將BMSCs在成骨誘導液作用3週,鑒定BMSCs嚮OB分化.採用96孔共培養闆將第2代SCs種植于上室,顱骨來源OB種植下室,共培養為實驗組,無SCs榦預共培養為對照組.將誘導分化的OB種植于下室,第2代SCs種植于上室,共培養為實驗組,無SCs榦預共培養為對照組,分彆于共培養後1、3、5、7、9d採用甲基噻唑基四唑(MTT)進行增殖比較.採用6孔共培養闆,實驗組上室種植SCs,下室分彆種植顱骨來源及BMSCs來源的OB,不加SCs的兩種來源的OB作為對照組,分彆共培養後于3、7 d檢測兩種來源OB的堿性燐痠酶、骨鈣素、骨橋蛋白、骨形態髮生蛋白-2 mRNA錶達.結果 SCs對顱骨來源的OB在共培養3、5、7、9 d 4箇時間點均有明顯促增殖作用,在堿性燐痠酶、骨橋蛋白、骨形態髮生蛋白-2mRNA在各箇時間段均起到抑製作用.SCs對大鼠BMSCs來源的OB在共培養1、3 d無明顯促增殖作用,在5、7、9 d有明顯促增殖作用,在成骨培養基的環境裏,SCs可促進BMSCs誘導的OB分化.結論 選擇BMSCs來源的OB 與 SCs共培養更適閤用于構建神經化組織工程骨.
목적 연구응용설왕세포(SCs)대량충래원적성골세포(OB)적증식여분화적영향,위구건신경화조직공정골제공체외실험이론의거.방법 통과채용고골、경골수강충세법획득SD대서골수기질간세포(BMSCs),채용이단백매소화신생서로골획득OB급채용소화후조직괴법획득SCs.장BMSCs재성골유도액작용3주,감정BMSCs향OB분화.채용96공공배양판장제2대SCs충식우상실,로골래원OB충식하실,공배양위실험조,무SCs간예공배양위대조조.장유도분화적OB충식우하실,제2대SCs충식우상실,공배양위실험조,무SCs간예공배양위대조조,분별우공배양후1、3、5、7、9d채용갑기새서기사서(MTT)진행증식비교.채용6공공배양판,실험조상실충식SCs,하실분별충식로골래원급BMSCs래원적OB,불가SCs적량충래원적OB작위대조조,분별공배양후우3、7 d검측량충래원OB적감성린산매、골개소、골교단백、골형태발생단백-2 mRNA표체.결과 SCs대로골래원적OB재공배양3、5、7、9 d 4개시간점균유명현촉증식작용,재감성린산매、골교단백、골형태발생단백-2mRNA재각개시간단균기도억제작용.SCs대대서BMSCs래원적OB재공배양1、3 d무명현촉증식작용,재5、7、9 d유명현촉증식작용,재성골배양기적배경리,SCs가촉진BMSCs유도적OB분화.결론 선택BMSCs래원적OB 여 SCs공배양경괄합용우구건신경화조직공정골.
Objective To explore the proliferation and differentiation of osteoblasts from 2 sources co-cultured with SD rat Schwann cells(SCs) . Methods Bone marrow stromal cells (BMSCs) were obtained by washing the femoral and tibial bone marrow cavities in SD rats. Osteoblast differentiation of the third passage of BMSCs was induced by incubation in osteogenic medium. Primary rat calvarial osteoblasts were obtained by digestion of the calvarial bone in one day old SD rats. The cells were cultured in DMEM supplemented with 10% fetal bovine serum(FBS) . SCs of passage 2 were obtained by digestion of sciatic nerve. The SCs were identified by S-100. The proliferation of 2 kinds of osteoblasts co-cultured with SCs was tested using 96 co-culture plate by methyl thiazdyl tetrazolium(MTF). Real-time PCR was used to test the osteoblast differentiation through co-culturing with SCs in 3 d and 7 d. The osteoblasts were implanted in the subtus chamber. The SCs were implanted in the superior chamber. Results SCs enhanced significantly the proliferation of calvarial osteoblasts at 7 time points. The expression levels of OPN mRNA, OCN mRNA, ALP mRNA, and BMP-2 mRNA of the osteoblasts were significantly lower in the experiment group than in the control group in 3 d and 7 d. SCs also enhanced significantly the proliferation of the induced osteoblasts in 5 d, 7 d and 9 d. The expression levels of OPN mRNA, OCN mRNA, ALP mRNA, and BMP-2 mRNA of the induced osteoblasts were significantly higher in the experiment group than in the control group in 3 d and 7 d, except the level of ALP mRNA in 7 d.Conclusions The BMSCs-induced osteoblasts cocultured with SCs may be used as seed cells to construct neurotized tissue engineered bone.
