高原体外肝细胞分离方法

A METHOD OF ISOLATION HEPATOCYTES AT HIGH ALTITUDE

目的 :探索高原环境下动物肝细胞的体外分离方法 ;方法 :采用胶原酶体外两步灌流法先进行肝叶灌流 ,再将所得肝组织悬液分别置 3 7℃温育不同时间 ,对比观测肝细胞产量、存活率、纯度及细胞接种后生长情况的差异 ;结果 :未经温育组及温育 1 0min、3 0min、60min组肝组织悬液的肝细胞产量分别为 ( 0 .85、2 .67、3 .84、4.2 9)× 1 0 6/ g湿重肝组织 ;肝细胞存活率分别为 97.6%、96.9%、88.3 %及 71 .5% ;肝细胞纯度分别为 96.2 %、95.4%、96.7%及 95.8% ;前两组细胞贴壁生长和增殖情况均较好 ,后两组则较差 ;结论 :在高原环境下先采用肝叶体外两步灌流后再行胶原酶短时间 ( 1 0min左右 )温育处理是一种既可获得较满意的细胞活性 ,同时又可显著提高细胞产量的分离肝细胞方法。

To grope for a method of isolation hepatocytes from animal liver at high altitude.Methods:After perfusion liver with collagenase,the liver tissue and collagenase solutions were incubated at 37℃ for different period,and the yield,viability,and purity of hepatocytes and growth status of cells cultured were observed.Results:The yield of hepatocytes form the liver tissue and collagenase solution incubated at 37℃ for 0,10,30 and 60 minutes were (0.85,2.67,3.84 and 4.29)×10 6 cells/g liver tissue,the viability was 97.6%,96.9%,88.3% and 71.5%,the purity was 96.2%,95.4%,96.7% and 95.8%,respectively.The growth status of the former two groups of cells cultured was better than that of the latter.Conclusion:Incubating the liver tissue and collagenase solution obatined by the two-step collagenase perfusion liver at 37℃ for short period(about 10 minutes)is a method of isolation hepatocytes from animal liver at high altitude,by which,a large yield as well as good viability can be obtained.