嗅成鞘细胞条件培养基对PC12细胞促分化作用及对分化后细胞损伤的保护作用  

EFFECT OF OLFACTORY ENSHEATHING CELL CONDITIONED MEDIUM ON DIFFERENTIATION OF PC12 CELLS AND ITS PROTECTIVE EFFECT ON DIFFERENTIATED PC12 CELLS AFTER INJURY IN VITRO

作　　者：杨浩[1] 王春婷[1] 许汉鹏[1] 李静雯[1] 游思维[1] 鞠躬[1] 

机构地区：[1]第四军医大学神经科学研究所,西安710032

出　　处：《解剖学报》2006年第1期43-47,共5页Acta Anatomica Sinica

基　　金：国家自然科学基金资助(30070799)

摘　　要：目的研究嗅成鞘细胞条件培养基(OECCM)对PC12细胞促分化作用及其对-OH自由基损伤分化后细胞的保护作用。方法采用原代分离培养的方法培养和纯化嗅成鞘细胞,收集其培养上清作为OECCM,然后用其培养PC12细胞,培养3 d后,进行细胞形态学观察及-βtubulin免疫细胞化学染色,同时在同一批细胞中加入100μmol/L FeSO4和50μmol/L H2O2作用20 min,再用OECCM继续培养48 h,然后进行MTT对细胞活性进行检测和存活细胞计数。结果用OECCM培养的PC12细胞长有突起,形态酷似神经元,并且-βtubulin免疫细胞化学染色呈阳性,而对照组细胞在同样培养时间里,没有明显的形态变化,-βtubulin染色呈阴性。用FeSO4和H2O2产生的-OH自由基对分化后的PC12细胞进行损伤,发现继续用OECCM培养后,反映细胞活性的A值为0.346 5±0.032,对照组0.201 8±0.034(P<0.01);同时两组细胞存活数目的百分比分别为:实验组为(56.7±5.9)%,对照组为(23.8±7.4)%(P<0.01)。结论嗅成鞘细胞可以分泌有促PC12细胞分化作用的分子和对分化后的细胞在损伤时有保护作用的分子。Objective To investigate whether adult olfactory ensheathing cells（OECs） could release some bioactive factors which affect differentiation of PC12 cells and protect these differentiated cells after hydroxyl free radical-induced injury. Methods Primarily cultured OECs were prepared and purified, and the purified cell conditioned medium was collected and used to culture PC12 ceils. After 3 days in vitro the morphological changes of the PC12 ceils were observed under a phase contrast microscope, β-tubulin immunocytochemical staining was conducted to evaluate differentiation of the PC 12 cells. At mean time, 100μmol/L FeSO4 and 50μmol/L H2O2 were added to cultured PC12 cells with OECCM for 20 min, and these ceils were cultured for 48 hours, then the viability of cells was determined with MTT assays, as well survived cells were counted. Results PC12 cells showed neuron-like morphology with long processes cultured in OECCM. The expression of β-tubulin showed strong positive by means of immunocytochemistry. Whereas the PC12 cells in control group scarcely exhibited morphological changes, and β-tubulin immunocytochemical staining was negative in corresponding period. As for OECCM protective effects on differentiated PC12 cell after hydroxyl free-induced injuries generated by addition of FeSO4 and H2O2, the A value was 0.346 5 ± 0.023, the data represented only 0.201 8 ± 0.034 in control group （ P 〈 0.01 ）. The percentage of survived cell （56.7 ± 5.9） % was higher than that of control group（23.8 ± 7.4） % （ P 〈 0.01 ）. Conclusion These results above suggest that OECs can secrete some factors to either promote the differentiation of PC12 cells or protect differentiated PC12 cells under injury condition in vitro.

关 键 词：嗅成鞘细胞 细胞分化 PC12细胞 损伤 细胞培养 大鼠 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学] R339.12[医药卫生—基础医学]