Seglen P O
J Toxicol Environ Health. 1979 Mar-May;5(2-3):551-60. doi: 10.1080/15287397909529766.
Isolation of preneoplastic cell populations would greatly facilitate analysis of the development of liver carcinogenesis. Suspensions of intact single cells can be prepared in an almost quantitative yield by two-step perfusion of the isolated liver. In the first step the liver is perfused with a Ca2+-free buffer (or with EGTA) in order to irreversibly cleave the desmosomes; in the second step perfusion with Ca2+-activated collagenase dissolves the collagenous extracellular matrix. The resulting single-cell suspension will be a mixture of intact normal and preneoplastic hepatocytes, other liver cell types (mostly Kupffer and endothelial cells), damaged cells, and subcellular debris. Intact hepatocytes can be purified--e.g., by differential centrifugation--but separation of preneoplastic from normal cells has not yet been achieved. Density gradient separation or selection in culture on the basis of the unique properties of preneoplastic hepatocytes (e.g., drug resistance) may prove useful. The use of hepatocyte cultures and liver-derived epithelial cell lines as test systems and models for chemical carcinogenesis in vitro is briefly reviewed.
分离癌前细胞群体将极大地促进对肝癌发生发展的分析。通过对离体肝脏进行两步灌注，可几乎定量地制备完整单细胞悬液。第一步，用无钙缓冲液（或EGTA）灌注肝脏，以不可逆地裂解桥粒；第二步，用钙激活的胶原酶灌注以溶解胶原细胞外基质。所得的单细胞悬液将是完整的正常和癌前肝细胞、其他肝细胞类型（主要是库普弗细胞和内皮细胞）、受损细胞及亚细胞碎片的混合物。完整的肝细胞可通过例如差速离心进行纯化，但尚未实现癌前细胞与正常细胞的分离。基于癌前肝细胞的独特特性（如耐药性）进行密度梯度分离或培养选择可能会被证明是有用的。本文简要综述了将肝细胞培养物和肝源性上皮细胞系用作体外化学致癌作用的测试系统和模型的情况。
