ISOLATION AND PARTIAL CHARACTERIZATIONS OF OVAL AND HYPERPLASTIC BILE DUCTULAR CELL-ENRICHED POPULATIONS FROM THE LIVERS OF CARCINOGEN AND NONCARCINOGEN-TREATED RATS

  • 1 January 1984
    • journal article
    • research article
    • Vol. 44  (8) , 3454-3466
Abstract
An oval cell-enriched population was isolated using 2 isopyknic centrifugation steps in Percoll gradients from the livers of young adult male rats maintained for 6-12 wk on a choline-deficient diet containing 0.05% DL-ethionine. This cell population equilibrated sharply at densities ranging between 1.07-1.08 g/ml, possessed a mean cell diameter in fixed-cell smears of 13.6 .mu.m and showed viabilities of > 95% as judged by trypan blue dye exclusion. Contamination of this population by hepatocytes and Kupffer cells was determined to be < 1% and between 4-14%, respectively. .gamma.-Glutamyl transpeptidase activity was demonstrated both biochemically and histochemically to be the most constant marker for evaluating the oval cell-enriched population isolated at various times over the 6-12 wk of the choline-deficient/DL-ethionine dietary regimen. The percentages of nonhepatocytic cells showing labeling for DNA synthesis and for .alpha.-fetoprotein were both found to be the highest in the oval cell-enriched population isolated at 6 wk and lowest in that obtained at 12 wk of dietary treatment. At 10-11 wk, 19.2% of the nonhepatocytic cells in this population were positive for albumin, while 2.1% were positive for glucose-6-phosphatase activity, indicating some cells to be intermediate in function between the oval cell and the hepatocyte. Hyperplastic bile ductular epithelial cells in tissue preparations isolated from the livers of rats previously subjected to 13 wk of chronic feeding of the noncarcinogenic cholestatic agent, 1-naphthyl isothiocyanate, or at 8-13 wk following bile duct ligation were found to be strongly positive for .gamma.-glutamyl transpeptidase activity, as well as to be positive for alkaline phosphatase activity, but to be essentially negative for glucose-6-phosphatase activity, glycogen content and albumin production. An occasional bile ductular cell in these preparations was found to exhibit a strong cytoplasmic binding of [6,7-3H]estradiol, an indirect measure of .alpha.-fetoprotein production. A low, but demonstrable, amount of DNA synthesis was noted in the bile ductular cells present in these preparations. A viable cell population highly enriched in bile ductular epithelial cells was isolated by isopyknic centrifugation in Percoll following enzymatic dissociation of the hyperplastic tissue preparation from bile duct ligated rats. The peak densities of this cell population ranged between 1.06-1.09 g/ml, and it was composed essentially of small basophilic cells having a mean diameter of .apprx. 15 .mu.m, as well as some intact bile ductules. Greater than 70% of these isolated cells were histochemically positive for .gamma.-glutamyl transpeptidase activity, and this cell population exhibited a specific activity for this enzyme that was at least 3-4 times higher than that shown by the oval cell-enriched population from the carcinogen-treated rats.