改良的成年大鼠骨髓源性破骨细胞诱导培养体系

为在短时间内获得更多的破骨细胞,在近几年国内成年大鼠骨髓源性破骨样细胞培养的基础上加以改良,以建立一个简便、有效的诱导培养体系.收集3月龄SD大鼠股骨骨髓细胞悬液,置入含有20%胎牛血清、1,25(OH)2VitD3的培养液中.将培养体系分为4组A组为对照组,未加入粒细胞/巨噬细胞集落刺激因子(GM-CSF)和地塞米松(Dexamethasone);B组加入GM-CSF;C组加入Dexamethasone;D组加入GM-CSF和Dexamethasone,观察各组破骨样细胞(osteoclast-like cell,OLC)及骨陷窝形成情况,并计数比较.培养第6 d,便可见所培养细胞衍变为形态不规则、有伪足形成的多核巨细胞,抗酒石酸酸性磷酸酶(TRAP)染色(+),且具有骨吸收作用,符合破骨细胞的鉴定标准.D组所形成OLC及骨陷窝数目较其它3组明显增多(P＜0.01),且OLC出现较早.结果表明成功地建立了一个以1,25(OH)2VitD3、GM-CSF、Dexamethasone为诱导分化条件的培养体系.所获得破骨细胞数量多,特征明显,且培养所需时间短.

Modified Culture System for Induction of Osteoclasts from Bone Marrow Stem Cells in Rat

A more effective culture system for induction of osteoclasts from bone marrow stem cells in rat was explored to obtain more osteoclasts in a short time. Femurs from 3 mouth SD rats were dissected and bone marrow cells collected and cultured in DMEM containing 20 % FCS, 1,25(OH) 2 VitD 3 . The culture system was divided into 4 groups: group A, control group; group B receiving granulocyte macrophage colony stimulating factor (GM CSF); group C receiving dexamethasone; group D receiving GM CSF and dexamethasone. Osteoclast like cells (OLC) and bone resorption pits were observed in all groups and counted. Six days after culture, the bone marrow cells developed into irregular multinucleated cells with pseudopodial projections. It has been confirmed that multinucleated cells were typical osteoclasts which were positive for TRAP staining and formed resorption pits on ivory dentine slices. Comparing with other 3 groups, the number of OLCs and bone resorption pits in the group D were obviously increased ( P <0 01) and cells appeared relatively earlier. The rat osteoclast marrow culture system in the presence of 1,25(OH) 2 VitD 3 , dexamethasone and GM CSF was successfully established.