两种豚鼠听毛细胞分离法比较  

COMPARISON OF TWO METHODS FOR ISOLATING HAIR CELLS FROM GUINEA PIGS

作　　者：曹恒明[1] 钱雪冶[1] 李华伟[1] 迟放鲁[1] 张勤修[1] 

机构地区：[1]上海医科大学眼耳鼻喉科医院

出　　处：《徐州医学院学报》1999年第2期92-95,共4页Acta Academiae Medicinae Xuzhou

摘　　要：目的对耳蜗灌注法和酶解加机械分离法的结果进行比较，并观察单离毛细胞的形态和存活情况。方法用耳蜗灌注法及酶解加机械分离法分离豚鼠听毛细胞，并在倒置相差显微镜下观察单离毛细胞。结果用耳蜗灌注法，７只豚鼠，７耳，平均每耳可分离出长于３０μｍ的外毛细胞１７３个（１１６～３３９），短于３０μｍ的外毛细胞１３０个（５０～４４７），内毛细胞０～１０个。室温下短期培养６ｈ后，长于３０μｎ的外毛细胞有１２３个（５８～３３６），短于３０μｍ的外毛细胞有１０３个（２０～２９６）。用酶解加机械分离法，７只豚鼠，７耳，平均每耳可分离出长于３０μｍ的外毛细胞７２个（２１～１５２），而短于３０μｍ的外毛细胞及内毛细胞则未能分离出。室温下短期培养６ｈ后，长于３０μｍ的外毛细胞有３０个（８～５６）。毛细胞损伤表现为细胞扭曲，细胞顶部表皮板脱落。细胞变性表现为细胞膜不平，细胞水肿，随时间变化逐渐加重。死亡表现为细胞核不被中性红染色。结论用耳蜗灌注法分离毛细胞简便易行，能提供足够量的存活时间较长的外毛细胞，而用酶解加机械分离法分离的毛细胞数相对较少，存活时间相对较短。Objective To compare the methods for isolating hair cells and study the viability and morphology of the isolated cells.Methods Hair cells were isolated by cochlear perfusion method and proteolytic enxyme digestion and micromanipulation method. The appearance and viability of the cells after incubation at room temperature for 6h were observed under inverted phase contrast microscope.Results By using the cochlear perfusion method in 7 guinea pigs (1 ear each animal), the yield consisted of an average of 173 (116～339) longer outer hair cells (>30μm), 130(50～447) shorter outer hair cells (<30μm) and 0～10 inner hair cells. After incubation, 123 (58～336) longer outer hair cells and 103 (20～296) shorter hair cells were still alive. By using the proteolytic enzyme digestion and micromanipulation method in 7 guinea pigs, only an average of 72 (21～125) longer hair cells were obtained, without shorter outer hair cells or inner hair cells, and 30 (8～56) of the isolated hair cells were alive after incubation. The mechanically traumatized hair cells showed cell body distortion and fall of the cuticular plate. Cell degeneration appeared as increasing swelling and deformity of cell membranes. The dead cells were unable to take the neutral red stain.Conclusion The cochlear perfusion method can produce better yields of isolated outer hair cells with satisfactory viability than the proteolytic enzyme digestion and micromanipulation method.

关 键 词：毛细胞分离 耳蜗灌注法 豚鼠 分离 耳蜗 

分 类 号：R764.35[医药卫生—耳鼻咽喉科] R329.492.3[医药卫生—临床医学]