碱性成纤维细胞生长因子对兔骨髓基质细胞生物行为的影响

Study of basic fibroblast growth factor in affecting biological behavior of marrow stromal cells

目的:观察兔骨髓基质细胞(MSC)经不同剂量碱性成纤维细胞生长因子(bFGF)刺激后,促血管内皮细胞增生的重要递质血管内皮细胞生长因子(VEGF)在MSC中的表达及分布情况,并与相应成骨情况做一对比,借以预测MSC经bFGF刺激后成骨效能与成血管效能的变化,探讨其对剂效关系。方法:取兔双侧股骨MSC,采用MSC体外培养技术,分别以不同剂量bFGF刺激细胞,并设立空白对照组。培养5d后,进行细胞形态(HE染色)、增殖情况(细胞记数法与MTT法)、碱性磷酸酶(改良钙钴法染色)、钙化结节(图像分析)、VEGF阳性细胞率(免疫组化染色)等项目的检测。结果:不同剂量组间在细胞记数法、MTT法检测、碱性磷酸酶活性、矿化面积百分率、VEGF阳性细胞率5项观测指标上F值分别为65.50,22.47,11.12,8.33和224.37,P<0.01,差别具有显著意义,结合各组均值来看,bFGF剂量为1200U/mL左右时效果最佳。结论:应用bFGF在促进MSC增殖的同时,还可促进促血管内皮细胞增生的重要递质-VEGF的表达,其应用剂量与效果之间存在明显相关关系。bFGF应用剂量为1200U/mL左右时,其促进MSC表达VEGF及促进细胞增殖作用同时达到最佳效果,超过此应用剂量,两者则有下降趋势。

AIM:To observe the expression and distribution of vascular endothelial growth factor (VEGF) in rabbit bone marrow stromal cells (MSC) stimulated with different doses of basic fibroblast growth factor (bFGF), to predict the changes of MSC in osteogenic potential and angiogenesis potential, and to investigate the relationship between applied dose of bFGF and effect.METHODS:MSCs of bilateral femurs of one New Zealand rabbit were procured and cultured in vitro. The samples were stimulated with different dose of bFGF, compared with normal controls. At 5 d post culture, hematoxylin and eosin (HE) staining, cytometry and MTT reduction assay (MTT), modified calcium cobalt staining, image analysis and immunohistochemical staining were performed to detect cell morphology, cell proliferation, alkaline phosphatase activity, calcified nod and ratio of VEGF positive cells respectively.RESULTS:There was statistically significant difference between different dose groups in all the detection items (P< 0.01), and the values of F were 65.50, 22.47, 11.12, 8.33 and 224.37. The group treated with 1 200 U/mL of bFGF had the best effects.