广西医科大学学报2011，Vol.28Issue(4)：506-511,6.
BrdU标记胎兔骨髓间充质干细胞及对细胞生物学行为影响的体外研究
IN VIRTO BROMODEOXYURIDINE LABELING OF BONE MARROW MESENCHYMAL STEM CELLS FROM BREPHO-RABBIT
摘要
Abstract
Objective: We explored the optimal dosage, timing and cytotoxicity for bromodeoxyurident (Br-dU) labeling of brepho-rabbit bone marrow mesenchymal stem cells (BMSCs) in vitro, in order to confirm its feasibility for stem cells labeling and tracer means. Methods:Brepho-rabbit bone marrow was isolated and cultured the BMSCs using the density gradient centrifugation method in vitro. The three passages of BMSCs were incubated with BrdU at different concentrations (5,10,15 and 20 mg/L ) for incubating time (12,24, 48 and 72 h), to identify the optimal BrdU concentration and incubating time for cell labeling. Immunohis-tochemistry and trypanblau strain were performed respectively to calculate the labeling index (positive rate) and the cells' activity for different time after BrdU labeling. Results:The observation of bmSCs' culture: the main appearance of primary BMSCs is wide, applanate and short fusiform shape, and long fusiform shape is the maim appearance of passage 3 BMSCs. The passage 3 of BMSCs can be differentiated into osteoblast and adipocyte under correspoonding inductive medium. The observation of immunohistoch-emistry:the BMSCs' nucleus showed green fluor under fluorescence microscope after labeled by the BrdU. The labeling ratio increasingly elevated of Groups A, B,C and D after the BMSCs were incubated 12 h,their mean labeling ratio was (33. 2±4. 3) %,(34. 5±3. 9) % , (47. 9±4. 8)% and (52. 8±5. 1) % , respectively, and the labeling ratio of group E is zero, which were significantly difference among the groups A,B and D ( P ＜0. 01=. The labeling ratio of groups A,B,C and D was (48. 1±4. 0)%,(83. 4±5. 3)%, (91. 7 ±4. 9)% and (92. 6±4. 2)% .respectively after the BMSCs were incubated 24 h, and the labeling ratio of group E is zero. The results of all groups after incubating 48 h,72 h were similar to those of the 24 h. The rate of label increased gradually with time and concentration, more than 90% BMSCs were labelled after 48 h and when the concentration of BrdU beyond 10 mg/L, at the same time, the living cell more than 90%. Conclusion:The optimal time for labeling BMSCs is 48h,the most appropriate concentration is 10 mg/L. The labeling rate of BrdU is very high and its cytotoxicity is very little, so it is safety.关键词
骨髓间充质干细胞/组织工程/尿嘧啶核苷酸类/细胞标记/细胞毒性Key words
bone marrow mesenchymal stem cells (BMSCs) / tissue engineering/ uracil mucleotides/ stem cell label/ cytotoxicity分类
生物科学引用本文复制引用
李德绘,黎洪棉,梁自乾,柳大烈..BrdU标记胎兔骨髓间充质干细胞及对细胞生物学行为影响的体外研究[J].广西医科大学学报,2011,28(4):506-511,6.基金项目
广西自然科学基金资助项目(No.挂科自0991112) （No.挂科自0991112）
