体外培养颌下腺细胞分泌功能的实验研究

Excretion Function of Submandibular Gland Cells Cultured in Vitro

目的 :获取成活率高且保持分化及分泌功能的颌下腺细胞。方法 :以改良合成培养基DMEM进行颌下腺细胞的原代及传代培养 ,观察颌下腺培养细胞生长状况。透射电镜观察颌下腺细胞的超微结构。免疫组化染色鉴定细胞来源及检测颌下腺培养细胞淀粉酶分泌功能。结果 :免疫组化染色显示培养的颌下腺细胞特异性抗体 :细胞角蛋白及角蛋白染色 (+++) ,胞浆内可见棕黄色点状分布的颗粒。透射电镜下腺泡上皮细胞表面可见到微绒毛、胞浆皱褶和细胞顶部的酶原颗粒 ,胞核大卵圆形 ,胞质内见线粒体和粗面内质网。颌下腺细胞培养上清淀粉酶含量测定显示不同代次颌下腺细胞淀粉酶含量随传代次数的增加而逐渐减弱 ,原代培养和传代培养第 2代的培养上清淀粉酶含量较高并且可维持一定水平。传代培养第 4代淀粉酶含量下降。结论 :DMEM体外培养颌下腺细胞原代培养和传代培养第 1代、第 2代细胞分泌功能较强 。

Objective: Our aim was to obtain a large number of submandibular gland cells with high survival rate and function of proliferation, differentiation, and excretion. Methods: Submandibular gland cells were cultured in fetal bovine serum (10%) on DMEM. Shape and structure of cultured cells were observed with phase contrastmicroscopeand transmission electron microscope. The cultured submandibular epithelial cells were subjected to immunohistochemistry research . The secretion function of cultured cells was evaluated by assay of amylase activity. Results: Immunohistochemistry staining that the CK8.13 and keratin antibodies were epithelium specific positive. Microvilli, plasm crease, and zymogen granules could be seen on the surface of acinus epithelial cells. Amylase content assay indicated cellular function of submandibular gland.The amylase quantity of cells in primary and secondary culture was higher than that of the forth generation at 96 hours. Conclution: Submandibular gland cells of primary and secondary culture had function of secretion.