Long-term cultivation of two diploid epithelial cell lines derived from normal rat liver cells

To obtain a useful rat liver cell line for in vitro carcinogenesis, two rat diploid epithelial cell lines were established from a 7-day-old male rat by the repeated colonial clone method. More than 80% of cells from each cell line have maintained normal diploid karyotype for over 30 months in vitro. The diploid cells were identi. fied as normal diploid karyotype by conventional Giemsa and trypsin. Giemsa techniques. They showed little difference in morphology and growth rate between early and late passages. Without cloning, they tended to be heterogenous in cell morphology, became heteroploid in chromosome and showed increased growth potential with time. Highly heteroploid cells which were derived from one of the lines produced ascites and solid tumors when inoculated into syngeneic rats intraperitoneally. Histologically, the tumors were diagnosed as poorly differentiated hepatocarcinomas. One of these diploid epithelial cell lines in early passage contained some activity of tyrosine transaminase and liver type aldolase and .glycokinase. Therefore, it is suggested that these epithelial cell lines represent liver parenchymal cells.</p

Cytological alteration of cultured rat liver cells by 3'-methyl-4-dimethylaminoazobenzene with special reference to chromosome changes, changes of growth patterns at a colony level and alpha-fetoprotein production.

A near diploid clone derived from a rat liver cell line was continuously treated with various concentrations of 3'-methyl-4-dimethylaminoazobenzene (3'-Me-DAB) in culture. By treatment with 2.8 micrograms/ml, cells with 41 chromosomes formed a mode and which then shifted to 39. The chromosome numbers of cells treated with 5.4 micrograms/ml were widely distributed at early stages, but later the mode shifted to hypotetraploid region. Untreated control cells were confirmed as near diploid. Increased plating efficiency by 3'-Me-DAB as well as the appearance of large sized colonies was obtained. The production of alpha-fetoprotein (AFP) by the cells was slightly enhanced by treatment with 3'-Me-DAB. The cells treated with and without 3'-Me-DAB did not produce any tumor in rats 6 months after their intraperitoneal injection.</p

Effect of 3'-methyl-N,N-dimethyl-4-aminoazobenzene in the presence or absence of liver microsomes on the liver cells having low tumor-producing capacity in culture.

A cell strain having low tumor-producing capacity was exposed in culture to 3'-methyl-N,N-dimethyl-4-aminoazobenzene (3'-Me-DAB) in the presence or absence of liver microsomes, and whether or not the cells will progress to those having high tumor-producing capacity was examined. When transplanted into rats, the cells treated with 3'-Me-DAB four (Exp-I) or thirteen times (Exp-II) formed larger tumors than untreated control cells, the latter treatment being more efficient in this regard. Furthermore, the tumors formed by the cells treated with 3'-Me-DAB in the presence of liver microsomes were considerably larger than those formed by the cells treated with 3'-Me-DAB alone. The subcutaneous tumors produced by the cells treated with 3'-Me-DAB with S-15 Mix showed poorly differentiated histology compared with those produced by control and 3'-Me-DAB-treated cells. The frequency of lung metastasis tended to increase by 3'-Me-DAB with S-15 Mix. The cells treated with 3'-Me-DAB in the presence or absence of liver microsomes differed from untreated control cells in vitro in some properties, including the size of aggregates in rotation culture, plating efficiency in liquid medium and morphology. These observations suggest that cell malignancy was promoted by 3'-Me-DAB alone as well as by 3'-Me-DAB in the presence of liver microsomes.</p

Decreased albumin secretion in serum-free primary cultures of adult rat hepatocytes during proliferation induced by epidermal growth factor and insulin.

Upon addition of epidermal growth factor (EGF, 0.1 microgram/ml) and insulin (0.1 microM), adult rat hepatocytes proliferated and increased 120-134% in number in serum-free primary culture. However, in the absence of the growth factors, hepatocytes decreased in number with time. The average albumin secretion per cell was much lower in the proliferating cultures than in the non-proliferating cultures. The results suggest that albumin production in hepatocytes decreases during cell proliferation.</p

Reactivities of azoxybenzenes (I) : Wallach rearrangement of 2,2'-dimetlyl azoxybenzene

　In this paper we authors studied on Wallach rearrangement reaction of 2,2'-dimethyl azoxybenzene (DMAB) using sulfuric acid, chlorosulfonic acid and some other weak acids as catalysts, and obtained the following results ; (1) We could obtain 4-hydroxy-2,2'-dimethyl azobenzene as a rearrangement product from DMAB in 80% sulfuric acid, under the reaction temperature 80℃ and time 30 minutes. The yield of 64% obtained in this experiment was better than that of 31% at which Gore et al had obtained the product. (2) In case where chlorosulfonic acid was used in the rearrangement, It is a contradictory results against Wallach rearrangement by ordinary acid catalysts. The yield of o-isomer in the reaction products was higher than that of p-isomer in rearrangement. (3) We also tried Wallach rearrangement by weak acids for the first taime, and obtained p-isomer at yield of max. 38.8% with trichloroacetic acid in acetic anhydride

Non-perturbative Evaluation of the Effective Potential of λϕ4\lambda\phi^4 Theory at Finite Temperature under the Super-Daisy Approximation

We calculate the effective potential of the scalar theory at finite temperature under the super-daisy approximation, after expressing its derivative with respect to mass square in terms of the full propagator. This expression becomes the self-consistent equation for the derivative of the effective potential. We find the phase transition is first order with this approximation. We compare our result with others.Comment: 12 page, 8 figure

Chromosome 8-14 translocation in a non-African Burkitt's lymphoma with leukemic conversion.

A specific chromosome translocation, t(8q-; 14q+), was observed in a 43-year-old female with non-African Burkitt's lymphoma in which leukemic conversion had occurred. The chromosome studies used cells from ascites. The ascites was apparently the result of a primary tumor involving the ovaries and contained 68% of lymphoma cells. The frequent occurrence of abnormalities related to chromosomes 1, 8 and 14 in African and non-African Burkitt's lymphomas was emphasized.</p

Effect of anti-alpha-fetoprotein serum on growth and plating efficiency of alpha-fetoprotein-producing hepatoma cells in vitro

The effect of a specific rabbit antiserum to rat alpha-fetoprotein (AFP) was examined on the growth and the plating efficiency of AFP-producing rat hepatoma cells (AH70Btc Clone 10-5) in cultures. The addition of anti-AFP serum to the culture medium inhibited cell growth moderately and inhibited plating efficiency markedly, although no inhibitory effect of complexes of AFP and antibody to AFP was observed on cell growth. Anti-AFP globulin in the immune serum was demonstrated on the cell surface by fluorescent antibody technique. Several clones producing low levels of AFP were obtained by long-term treatment of the original Clone 10-5 cells with anti-AFP serum. These treated clones showed characteristics that differed from the untreated original clone 10-5 cells: The relative plating efficiency of the treated clones on agar plates containing 5% anti-AFP serum was higher than the original Clone 10-15 cells and the amount of AFP secreted by the treated clones was lower.</p

An attempt to eliminate fibroblast-like cells from primary cultures of fetal human livers.

The elimination of fibroblast-like cells from primary cultures of fetal human livers was studied. A fibroblast-like cell line (HuF), which was obtained by subculturing fetal human liver cells 4 or more times, was briefly treated with hydrocortisone (HC) or putrescine (PUT). The growth of HuF cells was inhibited by HC at a concentration of 10(-2) M and by PUT at a concentration higher than 10(-3) M. Long-term treatment of HuF cells with 10(-3) M HC inhibited the growth of the cells. Primary cultures of fetal human livers were made in medium containing HC or PUT, and morphological and functional examinations were made. The cultures were predominantly composed of epithelial-like cells, with few fibroblast-like cells, when the HC concentration was 10(-5)M to 10(-3) M. A high amount of albumin was secreted at these concentrations of HC. On the other hand, at 10(-3) M PUT, many epithelial-like cells were seen, but albumin was undetectable. The present results indicate that albumin-producing epithelial-like cells can be selectively maintained in medium containing HC, in primary cultures of fetal human livers.</p

Effects of laminin and collagen type I on the morphology and secretion of proteins in human hepatoblastoma and hepatoma cell lines.

The effects of laminin (LAM) and collagen type I (C-I) on human hepatoblastoma (HuH-6) and hepatoma (HuH-7) cell lines were investigated. C-I was superior to LAM in supporting the attachment of the cells, especially of HuH-6, to plastic surfaces. No effect of LAM and C-I on cellular morphology was recognizable by phase contrast microscopy. By scanning electron microscopy (SEM), much more microvilli were found on the cell surface of HuH-6 on LAM substrate than on C-I substrate. In HuH-7 cells, however, these microvilli were rarely found on either LAM substrate or C-I substrate. The gel profile of the proteins secreted by HuH-6 and HuH-7 cells was not affected by the culture substrate except for the major band, though the amount of alpha-fetoprotein (AFP) secreted was larger when the cells were cultured on LAM substrate than on C-I substrate. These results indicate that the ability of LAM or C-I to enhance attachment is different from that to enhance AFP production or microvilli expression in HuH-6 cells and probably in HuH-7 cells.</p