组织块培养法及胶原酶消化法分离培养豚鼠耳蜗微血管内皮细胞

目的：建立稳定的豚鼠耳蜗微血管内皮细胞的培养方法。方法：采用显微解剖技术分离出豚鼠耳蜗血管纹,分别用组织块法及酶消化法进行培养及纯化。结果：耳蜗血管纹组织块培养后2天,部分组织块边缘有散在的细胞生长,这些细胞增逐渐增殖形成大片细胞集落;耳蜗血管纹组织经胶原酶消化后1－3天,可观察到培养皿底由一些细胞组成的细胞岛;细胞纯化后大多呈多边形,致密融合时具有内皮细胞培养时典型的“铺路石样”外观,经免疫组化检测其内皮细胞标志性抗原Ⅷ因子,95％以上的培养细胞的胞浆中呈棕黄色阳性反应。结论：组织块法及酶消化法能够获得体外培养的豚鼠耳蜗微血管内皮细胞。

Isolation and Cultivation of Cochlear Microvascular Endothelial Cells from Guinea Pigs by Tissue Nubbles Cultivation and Collagenase Digestion.

Objective To set up reliable cell culture methods of cochlear microvascular endothelial cells in guinea pigs.Methods The cochlear striae vascularies were acquired from guinea pigs with micrergy, the cells were cultured and pured by tissue nubbles cultivation and collagenase digestion.Results Two days after tissue nubbles cultivation of striae vascularis, cells grew at the edge of some tissue nubbles,and proliferated to massive colonies composed of more cells;Some islands made up of a few cells on the bottom of Petri dish were found one to three days after striae vascularies tissue was digested by collagenase;These cells were mainly polygon after purification, monolayer confluent culture cells showed typical "flagstone" appearance as cul tured endothelial cells. According to immunohistochemical detection of factor VIII, which is the mark antigen of endothelial cell, we had observed that the positive reaction cells with cytoplasmic brown pigment were over 95%.Conclusion Cochlear microvascular endothelial cells from guinea pigs can be acquired by tissue nubbles cultivation and collagenase digestion .