BMP2活性多肽体外定向诱导大鼠BMSCs 向成骨方向分化实验探究.docVIP

BMP2活性多肽体外定向诱导大鼠BMSCs 向成骨方向分化实验探究 　【摘要】 骨形成蛋白2（BMP2）具有较强的诱导大鼠骨髓间充质干细胞（BMSCs）向成骨方向定向分化的能力。天然的BMP2数量有限，结构复杂，限制了其广泛 应用 。本实验设计合成了BMP2活性多肽，体外试验探讨其对BMSCs向成骨方向定向诱导成骨分化的能力，评价BMP2活性多肽的诱导成骨效应。［ 方法 ］实验分2组，诱导组和非诱导组。取4周的Wistar大鼠分离培养BMSCs，传至第3代时，实验组，改用成骨诱导培养基（DMEM完全培养基+BMP2活性多肽200 μg／ml）。非诱导组，仍采用DMEM完全培养基。继续培养2~4周，采用细胞爬片培养、碱性磷酸酶活性和钙含量测定、实时定量PCR检测Ⅰ型胶原（ColⅠ）及骨桥蛋白（OPN）mRNA的表达，评价BMP2活性多肽的体外诱导成骨能力。［结果］诱导组BMSCs生长良好，表现出与典型的成骨细胞相似的形态特征和生物学特性，ALP活性上升，钙含量增加，ColⅠ和OPN的mRNA呈高表达。非诱导组，成骨特性不明显。［结论］合成的BMP2活性多肽能有效地促进BMSCs向成骨方向分化，是组织工程化骨中理想的诱导成骨的细胞因子。具有与天然BMP2类似的骨诱导活性，具有广阔的应用前景。 【关键词】 BMP2活性多肽 骨髓间充质干细胞 导成骨 Abstract：［Objective］To investigate the capability of synthesis BMP2derived peptide on osteogenic induction in bone marrow stem cells(BMSEs),and to evaluate the osteoinductivity of BMP2derived peptide in vitro.［Method］After segregating and cultivating four weeks Wistar rats marrow stromal cells in induction group were induced by osteogenasis medium containing 200 gg/ml BMP2derived peptide,and in noninductional group BMSCs were still culture with DMEM medium.Following continue culture for 2~4 weeks,cells film preparation,alkaline phosphatase activity and calcium deposition were measured.Type I collagen（CoM）and osteopontin（OPN）mRNA expression were measured using realtime fluorescent quantitative polymerase chain reaction（FQPCR）technique. The capability of synthesis BMP2derived peptide on osteogenic induction of BMSCs was investigated.［Result］After inductived with BMP2derived peptide.BMSCs cultured survived well greatly changed in cell morphology,and showed a biological and morphologie characteristics similar to those of osteoblasts.The lever of alkaline phosphatase activity and calcium deposition increased.ColⅠand OPN mRNA were expressed at higher level.In noninductional group no conspicuous osteogenic induction was shown.［Conclusion］BMP2derived peptide can induce BMSCs to differentiate into osteoblasts.It has the similar capacity of osteogenic induction as nat