中华小儿外科杂志
中華小兒外科雜誌
중화소인외과잡지
CHINESE JOURNAL OF PEDIATRIC SURGERY
2014年
3期
218-223
,共6页
马丽丽%袁正伟%蒋俊红%陈海琛%杨兆辉%姜艳%徐延波
馬麗麗%袁正偉%蔣俊紅%陳海琛%楊兆輝%薑豔%徐延波
마려려%원정위%장준홍%진해침%양조휘%강염%서연파
间质干细胞%脊柱裂,先天性%细胞分化%神经元
間質榦細胞%脊柱裂,先天性%細胞分化%神經元
간질간세포%척주렬,선천성%세포분화%신경원
Mesenchymal stem cells%Spina bifida,congenital%Cell differentiation%Neurons
目的 胚胎期移植骨髓间充质干细胞(bone marrow stem cell,BMSCs)至先天性脊柱裂模型胎鼠脊髓中,观察其在显性脊柱裂胎鼠脊髓中存活及分化情况,并探讨其对脊髓神经元修复的治疗潜能.方法 利用胎仔外科和显微注射技术将绿色荧光蛋白(eGFP)转染的BMSCs分别在胚胎第16、17和18天移植到83只脊柱裂和20只正常大鼠胚胎中(group E16、E17and E18),胚胎20 d的胎鼠脊髓被取出并固定.经冰冻切片后,计数eGFP阳性细胞数目,并利用免疫荧光方法检测脊髓中eGFP阳性的BMSCs中神经干细胞(Nestin)、神经元(Neurofilament)和神经胶质细胞(GFAP)的分布与表达.结果 本研究成功的将MSC在宫内移植入胎鼠体内,孕鼠的存活率为94％(81/86),胎鼠的存活率为78.1％(136/174);实验中孕16d的胎鼠存活率为75.8％(91/120)低于孕17 d的80.6％(25/31)和孕18d的87.0％(20/23).计数脊髓中eGFP阳性细胞数,范围在166～735个/根,其中P0代MSCs存活率最低,为10.8％;P3～6代为22.6％、P10～14代为20.1％;P10～14代MSC在母鼠孕16d时移植得到最佳分化率;统计eGFP阳性细胞表达Nestin、Neurofilament和GFAP的阳性率分别为4％～5％、1％～2％和5％～7％.结论 BMSCs移植到显性脊柱裂胎鼠脊髓中能够存活,并在脊髓中表达Nestin、Neurofilament和GFAP,提示移植骨髓间充质干细胞具有向神经干细胞、神经元和神经胶质细胞分化的可能.
目的 胚胎期移植骨髓間充質榦細胞(bone marrow stem cell,BMSCs)至先天性脊柱裂模型胎鼠脊髓中,觀察其在顯性脊柱裂胎鼠脊髓中存活及分化情況,併探討其對脊髓神經元脩複的治療潛能.方法 利用胎仔外科和顯微註射技術將綠色熒光蛋白(eGFP)轉染的BMSCs分彆在胚胎第16、17和18天移植到83隻脊柱裂和20隻正常大鼠胚胎中(group E16、E17and E18),胚胎20 d的胎鼠脊髓被取齣併固定.經冰凍切片後,計數eGFP暘性細胞數目,併利用免疫熒光方法檢測脊髓中eGFP暘性的BMSCs中神經榦細胞(Nestin)、神經元(Neurofilament)和神經膠質細胞(GFAP)的分佈與錶達.結果 本研究成功的將MSC在宮內移植入胎鼠體內,孕鼠的存活率為94％(81/86),胎鼠的存活率為78.1％(136/174);實驗中孕16d的胎鼠存活率為75.8％(91/120)低于孕17 d的80.6％(25/31)和孕18d的87.0％(20/23).計數脊髓中eGFP暘性細胞數,範圍在166～735箇/根,其中P0代MSCs存活率最低,為10.8％;P3～6代為22.6％、P10～14代為20.1％;P10～14代MSC在母鼠孕16d時移植得到最佳分化率;統計eGFP暘性細胞錶達Nestin、Neurofilament和GFAP的暘性率分彆為4％～5％、1％～2％和5％～7％.結論 BMSCs移植到顯性脊柱裂胎鼠脊髓中能夠存活,併在脊髓中錶達Nestin、Neurofilament和GFAP,提示移植骨髓間充質榦細胞具有嚮神經榦細胞、神經元和神經膠質細胞分化的可能.
목적 배태기이식골수간충질간세포(bone marrow stem cell,BMSCs)지선천성척주렬모형태서척수중,관찰기재현성척주렬태서척수중존활급분화정황,병탐토기대척수신경원수복적치료잠능.방법 이용태자외과화현미주사기술장록색형광단백(eGFP)전염적BMSCs분별재배태제16、17화18천이식도83지척주렬화20지정상대서배태중(group E16、E17and E18),배태20 d적태서척수피취출병고정.경빙동절편후,계수eGFP양성세포수목,병이용면역형광방법검측척수중eGFP양성적BMSCs중신경간세포(Nestin)、신경원(Neurofilament)화신경효질세포(GFAP)적분포여표체.결과 본연구성공적장MSC재궁내이식입태서체내,잉서적존활솔위94％(81/86),태서적존활솔위78.1％(136/174);실험중잉16d적태서존활솔위75.8％(91/120)저우잉17 d적80.6％(25/31)화잉18d적87.0％(20/23).계수척수중eGFP양성세포수,범위재166～735개/근,기중P0대MSCs존활솔최저,위10.8％;P3～6대위22.6％、P10～14대위20.1％;P10～14대MSC재모서잉16d시이식득도최가분화솔;통계eGFP양성세포표체Nestin、Neurofilament화GFAP적양성솔분별위4％～5％、1％～2％화5％～7％.결론 BMSCs이식도현성척주렬태서척수중능구존활,병재척수중표체Nestin、Neurofilament화GFAP,제시이식골수간충질간세포구유향신경간세포、신경원화신경효질세포분화적가능.
Objective To observe the survival and differentiation status of transplanted bone marrow mesenchymal stem cells (BM-MSCs) in spinal cord of dominant spina bifida fetal rats and discuss their therapeutic potentials for repairing spinal cord neurons.Methods Fetal surgery and micro-injection technology were employed to transplant green fluorescent protein (eGFP)-transfected BMSCs into 83 spina bifida and 20 normal rat embryos respectively at embryonic days 16,17 and 18 (groups E16,E17 & E18).And,at Day 20,spinal cord was removed and fixed.The number of eGFP-positive cells was counted.And immunofluorescence method was used to detect the distribution and expression of Nestin,Neurofilament and GFAP in eGFP-positive MSCs.Results After transplantation,the maternal survival rate was 94％ and the fetal survival rate 78.1％.The overall survival rates were tabulated in different operative groups.Better survival was achieved in the operative groups performed during later development stages (E17/E18) as compared with E16 group.The count of eGFP-positive cells in each spinal cord was 166-735.Secondly,we investigated if prolonged culture of MSCs affected the survival of transplanted MSCs.P3-P6 and P10-P14 cells had similar survival (22.6％ vs.20.1％) and it was higher than that of P0 cells (10.8％).In line with the increased neuronal differentiation potentials of MSCs in prolonged culture,MSCs generated more neurons in spinal column.Development stages of recipients also affected neural differentiation of MSCs.MSCs in E16 spinal column displayed significantly higher neuronal marker expression as compared with E18 and E17 groups.The eGFP-positive cells expressed nestin,neurofilament and glial fibrillary acidic protein at a positive rate of 4-5 ％,1-2 ％ and 5-7 ％ respectively.Conclusions After transplantation,the eGFP-positive BMSCs survive and differentiate in spinal cord of dominant spina bifida fetal rats.And they may express Nestin,Neurofilament and GFAP.
