体外培养人牙囊细胞表达的OPG和RANKL对破骨细胞表型分化的影响  

Effects of OPG and RANKL expressed of human dental follicle cells on osteoclastogenesis

作　　者：孙海燕[1] 金作霖 张永宽[3] 林珠 

机构地区：[1]解放军第307医院口腔科,北京100071 [2]第四军医大学口腔医院正畸科,陕西西安710032 [3]解放军第150医院口腔科,河南洛阳471031

出　　处：《中国美容医学》2009年第8期1121-1125,共5页Chinese Journal of Aesthetic Medicine

摘　　要：目的:建立人牙囊细胞(Human dental follicle cells,HDFCs)与人外周血单核细胞(Peripheral blood monocytes,PBMCs)共培养体系,研究体外培养人牙囊细胞表达的骨保护素(Osteoprotegerin,OPG)和核心因子kappB受体激活剂(Receptor activator of nuclear factor-kappa B ligand,RANKL)对破骨细胞表型分化的影响。方法:取健康成人外周血分离出单核细胞,同时分离12～14岁青少年第三磨牙牙囊进行原代培养,建立两种细胞共同培养系统。实验分七组:A、单核细胞;B、单核细胞+牙囊细胞(接触);C、单核细胞+牙囊细胞(接触)+集落刺激因子-1(Colony stimulating factor-1,CSF-1)+甲状旁腺相关蛋白(Parat hyroidhormone-related protein,PTHrP);D、单核细胞+牙囊细胞(不接触);E、单核细胞+RANKL+CSF-1;F、单核细胞+RANKL+CSF-1+牙囊细胞(不接触);G、单核细胞+RANKL+CSF-1+牙囊细胞(不接触)+anti-OPG。把每组细胞接种到预置玻片和牙本质片的24孔板中培养,观察细胞变化。培养第10天,取出玻片,进行抗酒石酸酸性磷酸酶(TRAP)染色,计数每组TRAP染色阳性细胞;第14天取出牙本质片,扫描电镜下观察骨陷窝形成情况。结果:B、F组可见少量破骨样细胞,A、D组无破骨样细胞,E组破骨样细胞分化最显著,C、G组破骨样细胞较E组略少,但无显著性差别(P>0.05),与其他各组差别显著(P<0.05)。结论:实验结果证实体外培养人牙囊细胞表达的RANKL和OPG分别对破骨细胞表型分化具有正向和负向调节作用。Objective To establish the co-culture system of human dental follicle cells （HDFCs） and human peripheral blood mononuclear cells （PBMCs）, and study the effects of OPG and RANKL expressed by human dental follicle cells on osteoclastogenesis. Methods Mononuclear cells were separated from healthy human peripheral blood and the human dental follicles cells dissected from the lower third impacted molars of adolescence.The study includes 7 groups： A, PBMCs; B,PBMCs ＋ HDFCs （contacted）; C,PBMCs ＋ HDFCs （contacted） ＋CSF-1 ＋PTHrP; D,PBMCs ＋ HDFCs （uncontacted）; E,PBMCs ＋ CSF-1 ＋ RANKL; F, PBMCs ＋ CSF-1 ＋ RANKL ＋ HDFCs （uncontacted） ; G,PBMCs ＋ CSF-1 ＋ RANKL ＋ HDFCs （uncontacted） ＋ anti-OPG..The cells of each group were seeded on the coverslips and dentin slices, placed in the wells in advance, and cultured in 1640 medium. At 10 days the PBMCs, growing on coverslips, were treated for TRAP staining.At 14 days the dentin slices were taken out and the resorption pits were observed by scaning electron microscope. Results There were few osteoclast like cells （OLCs） in group B and F, and no OLCs were found in group A and C. The OLCs in group E were most ,followed by group C and group G. Conclusions We establish the co-culture system of two types of cells and prove the effects of OPG and RANKL expressed of human dental follicle cells on osteoclastogenesis.

关 键 词：牙囊 牙齿萌出 集落刺激因子-1 甲状旁腺相关蛋白 骨保护素 核心因子kappB受体激活剂 破骨细胞形成 

分 类 号：Q813.1[生物学—生物工程]