骨细胞分离培养及其与成骨细胞鉴别比较的实验研究  

Experimental study on the isolated culture of osteocytes and identification of osteoblasts in rats

作　　者：汤小康[1] 程婉[1] 许兵[1] 应航[1] 童培建[1,2] 肖鲁伟[1] 

机构地区：[1]浙江中医药大学第一临床医学院,浙江杭州310053 [2]浙江省中医院,浙江杭州310006

出　　处：《中国骨伤》2013年第3期227-231,共5页China Journal of Orthopaedics and Traumatology

基　　金：浙江省自然基金(编号:Y2110928);浙江省中医药科学研究基金计划(编号:2010ZA020)~~

摘　　要：目的:建立比较稳定的骨细胞实验室分离培养方法,并将其部分生物学特性与实验室培养的成骨细胞进行比较研究,明确两者区别。方法:采用序列酶消化法,分别从3只3dSD乳鼠骨骼中分离培养骨细胞和成骨细胞,培养24h后进行细胞形态学观察。第1代细胞用碱性磷酸酶试剂盒采用重氮盐法(改良Kaplow氏法)染色,采用免疫细胞化学法对细胞的骨钙素(BGP)染色,测定碱性磷酸酶并计算其活性。结果:骨细胞多呈星状或树枝状,且有很多的突触;成骨细胞呈长梭形,有少量的突触。骨细胞碱性磷酸酶染色,胞浆内cAKP颗粒不明显;成骨细胞碱性磷酸酶染色,胞浆内可见众多cAKP颗粒。骨细胞BGP染色阳性明显,成骨细胞BGP染色阳性不如骨细胞明显。ALP在骨细胞中分泌较成骨细胞低,且有统计学差异。结论:实验室条件下能培养出骨细胞,该类细胞和成骨细胞有明显区别。Objective： To establish a more stable method to isolate osteocytes in vitro, and then to find the differences with osteoblast biological characteristics. Methods ： Osteocytes and osteoblasts were isolated from the bone tissue of 3-day-old rats using sequential collagenase digestion. The cells were identified through cell morphology after 24 hours later. Alkaline phos- phatase （ALP） kit was used to stain the first generation cells by Kaplow-way,the bone gla protein （BGP） of the cells were stained by immunocytochemitry. Measured ALP and computed its activity. Results：Osteocytes and osteoblasts showed obvi- ously differences in cell morphology. Osteocytes were star-shaped or dendrite-shaped within more dendrites,while osteoblasts were spindle-shaped with short dendrites. Osteocytes were negative for ALP,but osteoblasts were positive; Osteocytes were more positive for BGP, and osteoblasts were less positive. The secretion of ALP in osteocytes was lower than that of osteoblasts. Conclusion：Osteocytes can be isolated and cultured in vitro. These characteristics of osteocytes are apparently difference with those of osteoblasts.

关 键 词：骨细胞 成骨细胞 细胞分离 

分 类 号：R446.5[医药卫生—诊断学]