摘要
目的探讨携带骨形态发生蛋白-2（BMP-2）／Tet-on基因的骨髓基质干细胞（BMSCs）微囊与磷酸钙骨水泥（CPC）多孔支架复合后分泌BMP-2及胶原蛋白的能力。方法以携带BMP－2／Tet-on基因的腺病毒转染大鼠BMSCs，应用微胶囊技术包裹BMSCs。采用食盐造孔法使CPC形成多孔支架。实验分为3组（每组4个样本）：实验组在CPC多孔支架上复合BMSCs-海藻酸钠-聚赖氨酸-海藻酸钠（APA）微囊复合体，阳性对照组直接培养含BMP-2／Tet-on基因的BMSCs，单纯A-PA-CPC组在CPC支架上接种空壳微胶囊。培养第3、6、9、12天取3组培养基用酶联免疫吸附测定法（ELISA）测定BMP-2的浓度。实验组固定后做硬组织切片染色观察胶原蛋白的分泌情况。结果培养第3、6、9、12天，实验组BMP-2浓度平均分别为（4713．98±178．50）、（3288．85±194．38）、（1292．25±300．11）、（337．19±84．49）μg／mL，阳性对照组BMP-2浓度平均分别为（4663．87±242．99）、（3250．67±293．72）、（1276．74±157．10）、（293．65±92．48）μg／mL，单纯APA-CPC组BMP-2浓度平均分别为（105．14±10．93）、（91．42±18．00）、（89．63±12．99）、（108．72±23．90）μg／mL，同一时间点3组间比较差异有统计学意义（P〈0．05），其中实验组、阳性对照组分别与单纯APA．CPC组比较差异均有统计学意义（P〈0．05），而实验组与阳性对照组比较差异无统计学意义（P〉0．05）。实验组硬组织切片染色后可观察到淡黄色类骨质、绿色胶原纤维。结论微囊化BMSCs在CPC中可以存活并预期表达BMP-2和胶原蛋白，可以进行下一步体内实验。
Objective To observe whether the bone marrow stromal cells （BMSCs） which carry BMP-2/Tet-on genes and are microcapsulated by alginate polylysine alginate （APA） and seeded onto calcium phosphate cement （CPC） scaffolds can express bone morphogenetic proteins-2 （BMP-2） and secrete collagen proteins. Methods The BMSCs from rats were prepared, transfected with adenoviruses carrying BMP-2/Tet-on genes, and then microcapsulated by APA. Eight porous CPC scaffolds were prepared and mounted onto a 12-hole plate. The experiment was conducted in 3 groups, with 4 parallel specimens in each group. Microcapsulated BMSCs with the target gene were seeded onto the CPC scaffolds in the experimental group; the transfected BMSCs were seeded into the plate holes as positive controls; APA microcapsules without cells were seeded onto the CPC scaffolds as blank controls. The culture medium in each group was taken for ELISA examination on 3, 6, 9 and 12 days to evaluate the BMP-2 concentrations respectively. The samples in the experimental group were fixed, sliced and dyed to observe secretion of collagen. Results On 3, 6, 9 and 12 days after culture, the BMP-2 concentrations were 4713.98 ± 178.50 txg/mL, 3288.85 ± 194.38 [μg/mL, 1292.25 ± 300. 11 μg/mL and 337. 19 ± 84.49 μg/mL respectively in the experimental group; 4663.87± 242.99 μg/mL, 3250. 67 ±293.72 μg/mL, 1276.74 ± 157. 10 μg/mL and 293.65 ±92. 48 μg/mL respectively in the positive controls; 105. 14 ± 10.93 μg/mL, 91.42 ± 18.00 μg/mL, 89. 63 ± 12.99 μg/mLand 108.72 ± 23.90 μg/mL respectively in the blank controls. There were significant differences between the 3 groups at each same time point （ P 〈 0. 05）. The blank controls were significantly different from both the experimental group and the positive controls （ P 〈 0. 05）, but there were no significant differences between the experimental group and the positive controls （ P 〉 0. 05） . Collagen fibrils were observed by optic mi- croscopy in the sections of the experimental
出处
《中华创伤骨科杂志》
CAS
CSCD
北大核心
2012年第11期978-981,共4页
Chinese Journal of Orthopaedic Trauma
基金
广东省自然科学基金（06022696）
关键词
骨形态发生蛋白质类
间质干细胞
磷酸钙类
胶原
Bone morphogenetic proteins
Mesenchymal stem cells
Calcium phosphates
Collagen
