鹅小肠上皮细胞的分离培养研究

Study on Isolation and Culture of Intestinal Epithelial Cell in Goose Embryo

取29～30胚龄的鹅胚为试验材料,通过组织块种植法和酶消化法进行原代鹅小肠上皮的体外培养,采用刮除法、相差消化及相差贴壁法对上皮细胞进行纯化,用0.05%Trypsin-EDTA进行消化传代,通过形态学观察及免疫细胞化学法进行鉴定。结果表明:组织块种植法得到的鹅小肠上皮细胞生长状况良好,细胞活性较强,经过纯化,得到了纯度较高的鹅小肠上皮细胞。而联合使用浓度为300U/mL的胶原酶(Ⅺ型)和0.1mg/mL的中性蛋白酶(Ⅰ型)、多次(短时间)消化收获细胞法虽然可获得大量的健全肠绒毛隐窝单位,但传代后其增殖能力明显下降;形态学观察及免疫细胞化学法鉴定为阳性,证明分离出的细胞为小肠上皮细胞,从而为小肠上皮细胞的进一步研究奠定了基础。

The experiment was made to study the isolation and culture of intestinal epithelial cells in goose embryo. The primary intestinal epithelial cells from 29 to 30 days old goose embryo were cultured in vitro by the methods of explants culture or digested tissue filtrate. The epithelial cells were purified by scraping,difference＇ digestion and difference adherence and then digested by the solution of 0.05% trypsin-EDTA before subculture. The epithelial cells were identified by morphological observation and immunocytoehemieal method. The results showed that the epithelial cells of goose embryo grew well and the activity of the cells was also strong by the method of explants culture. Intestinal epithelial cells with high purity could be obtained after purification. Although the villa and crypt units were intact by the method of repeated short time digestion using 300 U/mL collagenase XI and 0.1 mg/mL neutral protease I,the proliferation ability of its future generation was decreased. Both of morphological observation and immunocytochemical method proved that the isolated cells were real intestinal epithelial cells. The results of the experiment could partially contribute to our future study.