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【目的】探讨科学评价深低温保存角膜内皮细胞活性的方法 ,弥补直接形态学染色法的缺陷。【方法】体外培养新鲜及深低温保存牛角膜内皮细胞 ,观察其贴壁时间 ,融合时间及贴壁率。用MTT法检测细胞吸光度 (A )及用3 H tdR法检测放射性核素计数 ,评价细胞活性。【结果】原代培养时 ,深低温保存角膜内皮细胞的贴壁时间及融合时间分别为 (5 5± 1 8)d及 (15 8±3 2 )d ,均较新鲜角膜内皮细胞长 [分别为 (2 5± 1 3)d及 (8 6± 2 4)d];贴壁率、吸光度及放射计数分别为 71 8%± 6 6 %、0 46± 0 11及 (9 6± 1 1)Bq,也均较新鲜角膜内皮细胞低 [分别为 90 2 %± 4 5 %、1 74± 0 75及 (70 1± 5 4)Bq]。但其余各代培养细胞二组间无显著性差异 (P >0 0 5 )。【结论】通过体外培养 ,研究深低温冻存的角膜内皮细胞生物学特性 ,能更科学、准确地判定细胞活性。深低温保存对角膜内皮细胞活性有抑制作用 ,但这种抑制是可以恢复的
【Objective】 To investigate the method of scientifically evaluating the activity of preserved corneal endothelial cells at cryogenic temperature to make up for the defects of direct morphological staining. 【Method】 The cultured bovine corneal endothelial cells were cultured in fresh and deep hypothermia. The attachment time, fusion time and attachment rate were observed. Cell absorbance (A) was measured by MTT assay and radionuclide counts were measured by 3 H tdR method to evaluate cell viability. 【Result】 The results showed that the preservation time and fusion time of preserved corneal endothelial cells in deep culture were (5 5 ± 1 8) d and (15 8 ± 3 2) d, respectively, compared with fresh corneal endothelial cells [ (2 5 ± 1 3) d and (8 6 ± 2 4) d], respectively. The rates of adherence, absorbance and radiation were 71 8% ± 6 6%, 0 46 ± 0 11 and 96 6 1 1 ) Bq were also lower than fresh corneal endothelial cells [90 2% ± 45%, 1 74 ± 0 75 and (70 1 ± 5 4) Bq, respectively]. However, there was no significant difference between the other groups of cultured cells (P> 0.05). 【Conclusion】 The biological characteristics of corneal endothelial cells cryopreserved in vitro can be used to determine the cell activity more scientifically and accurately. Cryopreservation has an inhibitory effect on corneal endothelial cell activity, but this inhibition is regressible