Nakamura T, Nagao M, Ichihara A
Exp Cell Res. 1987 Mar;169(1):1-14. doi: 10.1016/0014-4827(87)90218-7.
Immunocytochemical staining of tryptophan 2, 3-dioxygenase (TO, EC 1.13.11.11), which is a typical marker of terminal differentiation of rat hepatocytes, showed that TO was expressed as early as the first day after birth by a few hepatocytes, and that the number of hepatocytes expressing TO gradually increased during early neonatal development. In contrast, immature hepatocytes grew actively in culture even without growth factors and serum, showing a labelling index with [3H]thymidine of over 80% just after birth, but their ability to show autonomous growth decreased rapidly during postnatal development. Double staining of hepatocytes from neonatal rats indicated that hepatocytes showing DNA synthesis were distinct from those expressing TO, suggesting that immature cells proliferate, but do not express TO, and then become fully differentiated expressing TO, but not proliferating any more. On the contrary, albumin was fully expressed in most hepatocytes of 0-day-old rats, in which the hepatocytes were still growing. When immature hepatocytes without TO from 0-day-old rats were cultured on a feeder layer of adult rat hepatocytes for 3.5 days, their expression of TO increased rapidly to almost the adult level (over 70% of the cells became TO-positive). Conversely, this feeder layer strongly inhibited autonomous growth of the neonatal rat hepatocytes. Other feeder layers, such as non-parenchymal liver cells of adult rats, Reuber hepatoma, and Swiss 3T3 fibroblasts, had no effect on the reciprocal changes of terminal differentiation and autonomous growth of immature rat hepatocytes. A feeder layer of dead adult rat hepatocytes, obtained by treating the cells with cytosine arabinoside for 24 h or drying them at 37 degrees C, had the same ability as a feeder layer of living cells to induce cytodifferentiation of immature cells. When the dead feeder layer was treated with 1 N HCl or digested with 0.1% trypsin, its ability to induce differentiation was almost completely lost. These results suggest that a cell surface component of adult rat hepatocytes, probably an acid-labile protein, controls terminal differentiation and growth of immature hepatocytes.
色氨酸2,3-双加氧酶（TO，EC 1.13.11.11）是大鼠肝细胞终末分化的典型标志物，免疫细胞化学染色显示，早在出生后第一天就有少数肝细胞表达TO，且在新生儿早期发育过程中表达TO的肝细胞数量逐渐增加。相反，未成熟肝细胞即使在没有生长因子和血清的情况下也能在培养中活跃生长，出生后即刻其[3H]胸腺嘧啶核苷标记指数超过80%，但其自主生长能力在出生后发育过程中迅速下降。对新生大鼠肝细胞进行双重染色表明，显示DNA合成的肝细胞与表达TO的肝细胞不同，这表明未成熟细胞增殖但不表达TO，然后分化为完全成熟的表达TO但不再增殖的细胞。相反，白蛋白在出生0天大鼠的大多数肝细胞中充分表达，而此时肝细胞仍在生长。将出生0天大鼠不表达TO的未成熟肝细胞在成年大鼠肝细胞饲养层上培养3.5天，其TO表达迅速增加至几乎成年水平（超过70%的细胞变为TO阳性）。相反，该饲养层强烈抑制新生大鼠肝细胞的自主生长。其他饲养层，如成年大鼠的非实质肝细胞、鲁伯肝癌细胞和瑞士3T3成纤维细胞，对未成熟大鼠肝细胞的终末分化和自主生长的相互变化没有影响。用阿糖胞苷处理细胞24小时或在37℃干燥获得的成年大鼠死亡肝细胞饲养层，与活细胞饲养层诱导未成熟细胞向细胞分化的能力相同。当死亡饲养层用1N盐酸处理或用0.1%胰蛋白酶消化时，其诱导分化的能力几乎完全丧失。这些结果表明，成年大鼠肝细胞的一种细胞表面成分，可能是一种酸不稳定蛋白，控制着未成熟肝细胞的终末分化和生长。
