【摘要】 探讨一种简便实用、成本低、纯度高并且成熟稳定的成人食管上皮原代培养的方法。成人食管上皮细胞来源于食管癌患者手术切除食管的正常黏膜上皮。采用组织块法,分别在DMEM/F12混合培养基和1640培养基中培养成人正常食管上皮细胞。通过直接观察、细胞形态学观察和免疫细胞化学方法观察细胞的生长情况、细胞形态学特征及鉴定所得到的细胞。结果表明,与1640培养基相比,应用DMEM/F12混合培养基具有明显的优越性。正常食管上皮细胞在DMEM/F12混合培养基中生长较好,细胞融合快,成纤维细胞污染少,可以生长成为典型的"铺路石"样,可连续培养21天,组织块最多可以被迁移4次,且细胞生长状态良好。免疫细胞化学染色鉴定证实95%的细胞具有广谱细胞角蛋白,即确定是食管上皮来源的细胞。本研究应用改良的组织块法培养的正常食管上皮细胞纯度高、产量较大,可以满足正常食管上皮细胞恶性转化方面的研究工作,而且方法简便易行、成熟稳定,是一种适合推广应用的正常食管上皮细胞培养方法。更多还原

【Abstract】 To explore the optimal simple and convenient method to culture the highly purified human normal esophageal epithelial cells. The human normal esophageal epithelial cells were isolated from surgically resected normal esophagus of patients with esophageal cancer. The normal esophagus cells were cultured in mixed medium of DMEM/F12 and 1640 medium with 15% fetal calf serum respectively with tissue explant method. The morphological characteristics of the regenerate epithelial cells were observed and identified by direct observation,cell morphology and immunocytochemistry. The results indicated that the regenerate normal esophageal epithelial cells in mixed medium of DMEM/F12 showed faster growth and cellular junction,formed appearance of typical slabstone with less fibroblast than that in 1640 medium. The regenerate epithelial cells could be cultured for at least of 21 days and the tissue explants could be implanted for 4 times with better growth. The result of immunocytochemisty showed that 95% of the epithelial cells cultured in mixed medium of DMEM/F12 expressed CKpan,which confirmed that the cells originated from esophageal epithelium,but only 60% approximately in 1640 medium. In the study,the human normal esophageal epithelial cells cultured in mixed medium of DMEM/F12 with tissue explant could have high purity and abundant yield,which could be applied to the research of esophageal epithelial malignant transformation and be suitable for promotion.更多还原