犬纤维化肝贮脂细胞的分离、培养和鉴定

ISOLATION AND IDENTIFICATION OF FAT-STORING CELLS FROM FIBROUS DOG LIVER WITH A MODIFIED CENTRIFUGATION

改良一种分离肝贮脂细胞（Ｉｔｏ）的方法。结扎犬胆总管，制备肝纤维化模型；用 ６０ ｇ／ Ｌ聚蔗糖校正比重为１．０７７的淋巴细胞分离液，使其比重为１．０５３，采用单层密度梯度离心法分离犬纤维化肝贮脂细胞。结果：所获得的该种细胞在３２８ ｎｍ紫外光激发下出现自发荧光，呈ｄｅｓｍｉｎ免疫组化阳性染色，电镜观察显示其胞质中有较多脂滴，细胞核形状极不规则。结论：分离所得的细胞为贮脂细胞；本工作简化了贮脂细胞的分离方法，使其既方便又经济实用，分离效率也较高。

PURPOSE Rat fat-storing (Ito) cell is widely used in study of hepatic fibrosis, while the isolation of the cell is quite complicated, the author would simplify the procedures and get Ito cell from fibrous dog liver. METHODS Three male dogs of one year old, with ligation of common bile duct for three months were sacrificed. The dog Ito cell was purified with a single-step density gradient centrifugation with a modified medium: commercial lyphocyte separation medium adjusted with ficoll 400 with final density 1 .053. RESULTS More than eighty per cents of isolated cells were proved to be Ito cells which showed cytoplasmic lipid droplets with autofluorescence under 328 urn wave length and positive immunoreactivity of desmin. Under TEM the cell showed lipid droplets in cytoplasm. The yield of Ito cell was 5.7 × 108-3.2 ×10 9 per liver. CONCLUSIONS The successful isolation of Ito cells from fibrous dog liver with modified medium for a single-step density centrifugation has the advantages of simple procedure, low expense, high efficiency and usable value.