中华创伤骨科杂志
中華創傷骨科雜誌
중화창상골과잡지
CHINESE JOURNAL OF ORTHOPAEDIC TRAUMA
2015年
1期
75-81
,共7页
间质干细胞%创伤与损伤%糖基化%转染%靶向
間質榦細胞%創傷與損傷%糖基化%轉染%靶嚮
간질간세포%창상여손상%당기화%전염%파향
Mesenchymal stem cells%Wounds and injuries%Glycosylation%Transfection%Target
目的 观察糖基化改造的骨髓基质干细胞(BMSCs)在骨缺损模型中的靶向迁移情况.方法 首先选用1只新西兰兔,2个月龄,体质量1.5 kg,进行BMSCs分离、提取、扩增;脂质体法将α-1,3岩藻糖基转移酶Ⅵ(FUT-6)基因转染BMSCs并筛选建立阳性克隆细胞株;ELISA法检测FUT-6基因转染的FUT-6/BMSCs和BMSCs的FUT-6表达量及唾液酸化路易斯寡糖(skX)生成量;流式细胞技术检测FUT-6/BMSCs和BMSCs与E、P选择素的结合力;12只新西兰兔骨缺损造模后分为2组(每组6只):采用FUT-6/BMSCs和BMSCs体外eGFP标记后静脉分别回输入两组相同骨缺损动物模型体内,术后6、12、24 h采用荧光显微镜观察骨缺损处髓腔组织中标记细胞数量,以评估靶向迁移情况.结果 BMSCs可体外培养扩增;FUT-6基因转染后的FUT-6/BMSCs高表达FUT-6,上调sLeX的生成量,差异有统计学意义(P＜0.05);流式细胞仪检测提示:与BMSCs比较,FUT-6/BMSCs与E、P选择素的结合力分别由15.0％、12.7％提高到68.9％、59.7％;术后6、12、24 h荧光显微镜观察FUT-6/BMSCs在骨缺损处髓腔组织中的数量较BMSCs更多,差异均有统计学意义(P＜0.05).结论 糖基化改造的BMSCs具有明显靶向迁移至病损部位的能力.
目的 觀察糖基化改造的骨髓基質榦細胞(BMSCs)在骨缺損模型中的靶嚮遷移情況.方法 首先選用1隻新西蘭兔,2箇月齡,體質量1.5 kg,進行BMSCs分離、提取、擴增;脂質體法將α-1,3巖藻糖基轉移酶Ⅵ(FUT-6)基因轉染BMSCs併篩選建立暘性剋隆細胞株;ELISA法檢測FUT-6基因轉染的FUT-6/BMSCs和BMSCs的FUT-6錶達量及唾液痠化路易斯寡糖(skX)生成量;流式細胞技術檢測FUT-6/BMSCs和BMSCs與E、P選擇素的結閤力;12隻新西蘭兔骨缺損造模後分為2組(每組6隻):採用FUT-6/BMSCs和BMSCs體外eGFP標記後靜脈分彆迴輸入兩組相同骨缺損動物模型體內,術後6、12、24 h採用熒光顯微鏡觀察骨缺損處髓腔組織中標記細胞數量,以評估靶嚮遷移情況.結果 BMSCs可體外培養擴增;FUT-6基因轉染後的FUT-6/BMSCs高錶達FUT-6,上調sLeX的生成量,差異有統計學意義(P＜0.05);流式細胞儀檢測提示:與BMSCs比較,FUT-6/BMSCs與E、P選擇素的結閤力分彆由15.0％、12.7％提高到68.9％、59.7％;術後6、12、24 h熒光顯微鏡觀察FUT-6/BMSCs在骨缺損處髓腔組織中的數量較BMSCs更多,差異均有統計學意義(P＜0.05).結論 糖基化改造的BMSCs具有明顯靶嚮遷移至病損部位的能力.
목적 관찰당기화개조적골수기질간세포(BMSCs)재골결손모형중적파향천이정황.방법 수선선용1지신서란토,2개월령,체질량1.5 kg,진행BMSCs분리、제취、확증;지질체법장α-1,3암조당기전이매Ⅵ(FUT-6)기인전염BMSCs병사선건립양성극륭세포주;ELISA법검측FUT-6기인전염적FUT-6/BMSCs화BMSCs적FUT-6표체량급타액산화로역사과당(skX)생성량;류식세포기술검측FUT-6/BMSCs화BMSCs여E、P선택소적결합력;12지신서란토골결손조모후분위2조(매조6지):채용FUT-6/BMSCs화BMSCs체외eGFP표기후정맥분별회수입량조상동골결손동물모형체내,술후6、12、24 h채용형광현미경관찰골결손처수강조직중표기세포수량,이평고파향천이정황.결과 BMSCs가체외배양확증;FUT-6기인전염후적FUT-6/BMSCs고표체FUT-6,상조sLeX적생성량,차이유통계학의의(P＜0.05);류식세포의검측제시:여BMSCs비교,FUT-6/BMSCs여E、P선택소적결합력분별유15.0％、12.7％제고도68.9％、59.7％;술후6、12、24 h형광현미경관찰FUT-6/BMSCs재골결손처수강조직중적수량교BMSCs경다,차이균유통계학의의(P＜0.05).결론 당기화개조적BMSCs구유명현파향천이지병손부위적능력.
Objective To observe the targeted migration of glycosylated bone marrow mesenchymal stem cells (BMSCs) in rabbit bone defect models.Methods A New Zealand rabbit of 2 months old and 1.5 kg in weight was used to separate,extract and amplify BMSCs.α-1,3 fucosyltransferase Ⅵ (FUT-6) gene was transfected to the BMSCs by liposome method to screen the positive clone cell lines.ELISA was used to detect FUT-6 expression and Sialyl Lewis X (sLeX) of FUT-6/BMSCs and BMSCs.Flow cytometry was used to detect the binding force of FUT-6/BMSCs and BMSCs with E/P selectin.FUT-6/BMSCs and BMSCs were labeled with eGFP in vitro and then intravenously re-infused respectively to 2 equal groups of bone defect models made of 12 New Zealand rabbits.Fluorescence microscopy was used at 6,12 and 24 hours postoperation to observe the number of labeled cells in the medullary cavity tissue at the bone defect sites to evaluate the targeted migration.Results BMSCs were amplified by in vitro culture.Transfected BMSCs expressed a significantly higher level of FUT-6,and increased significantly generation of sLeX (P ＜ 0.05).Compared with BMSCs,FUT-6/BMSCs had an obviously higher binding force with E/P selectin (15.0％ and 12.7％ versus 68.9％ and 59.7％).Fluorescence microscopy at 6,12 and 24 hours showed that there were significantly more FUT-6/BMSCs than BMSCs in the medullary cavity tissue at the bone defect sites (P ＜ 0.05).Conclusion Glycosylated BMSCs possess an obvious ability to migrate to the sites of lesion.
