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The use of the number of nucleoli as a criterion allowed us to identify significant differences in the organization of nucleoli in newly formed hepatocytes under the conditions of dipine-induced carcinogenesis 8-10 weeks after its initiation, as compared with normal liver cells of adult mice. The number of nucleoli in tetraploid new hepatocytes was equal to that in the diploid nuclei of normal liver (on average 3.5 and 3.6, respectively) and the number in the new octaploid nuclei was equal to that in tetraploid (6.2 and 6.4, respectively). A higher extent of association of nucleolar organizers under the conditions of endoreproduction or the association of homologous chromosomes in G1 period of the cell cycle is discussed as a possible mechanism responsible for the decrease in the number of nucleoli.

It has been shown that a population of the oval cells is formed in mouse liver during the dipin-induced carcinogenesis (Radaeva, Factor, 1990b). This paper deals with the origin of the oval cells and their proliferation potential depending on localization in the liver lobule. Series of semithin liver sections were studied under the light microscope and detected labeled cells analyzed under electron microscope on serial ultrathin sections. We found that proliferation of cells of terminal bile ductules (Hering [correction of Gering] canals) takes place at the early stages of liver carcinogenesis. These cells and first labeled oval cells had similar size and morphology and jointly formed the ducts. Oval cell population was heterogeneous in terms of proliferative potential. Proportion of proliferating cells (38-45%) in the oval cells of Hering [correction of Gering] canals and small ducts surrounding portal tracts remained similar throughout the period of formation of the oval cell population. In the oval cells infiltrating the parenchyma, the proportion of proliferating cells appeared to depend on the intensity of the oval cell response: it attained the maximum (62%) on intermediate stage and decreased to the minimum (22%) at the peak of the reaction. These data suggest that Hering [correction of Gering] canals probably give origin to the ducts formed by oval cells.

Hepatocytes and the fraction of non parenchymal cells enriched with oval cells were extracted from the preneoplastic mouse liver at the stage of hyperplastic node formation and implanted into the spleen. In 14-16 months after the transplantation, multiple islets of hepatocytes which replaced up to 25% of the spleen cut area, were found in 57% (4 of 7) and 22% (8 of 36) of recipients respectively. The hepatocytes formed 2-3-cell bulks or solid masses organized into multicellular trabecules, and expressed biliary capillary antigen, albumin and transferrin. In the inoculation of nonparenchymal cell fraction, the growth of hepatic tissue in the spleen depended on the magnitude of hepatocyte admixture to be undetectable in absence of hepatocytes in the donor suspension. The growth of hepatic tissue in spleen was observed following the injection of a small number (3 x 10(-4)-6 x 10(-5)) of live hepatocytes. This fact evidences an extremely high clonogenic potency of clonogenic potency of preneoplastic hepatocytes.

This paper considers the problem of epithelial stem cells of the liver and possibilities of its experimental solution. Authors' own data obtained with the model of induced hepatocarcinogenesis in mice are discussed; the experiments were performed using electron microscopy, autoradiography and immunochemistry. In accordance with these data, Gering cells are stem cells of the liver, and oval cells correspond to committed precursors capable to differentiation in either hepatocellular or cholangiolar direction under the conditions of periportal microenvironment. We have also compared hepatocyte differentiation in preneoplastic mouse liver and regenerating pancreas of adult rats (Rao et al. Amer. J. Pathol. 1989. V. 134. P. 1069-1086). We also discuss stem cell compartment organization in organs having glandular structure and the possibility of existence in the adult of non-committed multipotent cells capable of producing various types of differentiation in tissues having common origin during embryogenesis.

Monoclonal antibodies (MAb) were produced against antigens (Ag) of oval cells isolated from the preneoplastic murine liver. To suppress the immune response to major antigens common with hepatocytes, the principle of anti-idiotype immunization was employed. Characteristics of three MAb reacting selectively with the foci of oval cell proliferation are described. MAb A6 and G7 detected two different antigens (Ag A6 and Ag G7, respectively) common for oval cells and cholangiocytes. Ag A6 was also found in normal parenchyma (in membranes of single hepatocytes adjacent to portal veins), in the preneoplastic liver (in hepatocytes formed de novo) and in some hepatoma cells. Ag G7 was not detected in hepatocytes. MAb E5 stained the matrix in the areas adjacent to oval cells and large bile ducts. All the three Ag were widely distributed in normal tissues of mice. The significance of the detected Ag as markers of murine liver epithelial cell lines and stages of their differentiation is discussed as well as the possible relationship between Ag A6 and Ag of human blood groups.

It has been shown that dipin in combination with partial hepatectomy (model of dipin hepato-carcinogenesis) induces massive oval cell proliferation. The main stages of oval cells development examined by means of light microscopy of half-thin slices are the following: 1) appearance around portal tracts--1-3 weeks; 2) migration into parenchyma along terminal branches of portal vessels--3-8 weeks; 3) maximum development--8-10 weeks; 4) induction focuses of growth of new formed hepatocytes around portal tracts--8-10 weeks; 5) forcing of oval cells towards the centre of liver lobules and their elimination. There was close correlation between oval cells development, fibrosis and inflammation. Oval cells remained around portal tracts until the development of numerous hepatomas in 10 months after treatment.

An electron microscopic study of murine oval cells, induced by a single injection of genotoxic agent dipin and by a partial hepatectomy, has shown that their ultrastructure and direction of differentiation depend on localization in the liver lobule. Oval cells around portal tracts go through three stages of development: low differentiated cells 4.40 +/- 0.51 mu in diameter with ovoid nuclei 3.43 +/- 0.44 mu, intermediate cells, and young hepatocytes. They form common ducts surrounded by a basal lamina, and produce bile canaliculi-like structures and intermediate junctions between them. Another part of the oval cell population is organized similar to the bile duct epithelium. It consists of cells 9.37 +/- 1.1 mu in diameter with nuclei 7.28 +/- 1.16 mu in diameter and form a system of branching and anastomosing ducts widespread along the parenchyma from the portal to the central veins. Our data indicate that the oval cells can differentiate into hepatocytes, and support a hypothesis according to which the cells of terminal bile ductules are liver epithelial stem cells which can differentiate into a hepatocyte or a bile duct cell lineage in periportal microenvironment.

The object of this investigation was the frequency of dominant lethal mutations (DLM) induced by ethylene imine (EI) in spermatozoa, spermatids, spermatocytes I and spermatogonia of Drosophila melanogaster. The DLM frequency was shown to increase up to a certain level in case of storage of EI-treated spermatozoa in spermathecas of intact females. No such increase of the DLM frequency was observed for treated spematids. No differences in the induced DLM frequency between treated spermatozoa and spermatids were observed. The treatment of spermatocytes I resulted in the sterility of males; however, after the elapse of some time their fertility was restored at the expense of dividing spermatozoa. The sterility of males in case of the EI treatment of spermatocytes I is presumably caused by a great number of chromosome aberrations. The data obtained are discussed from the standpoint of the hypothesis of potential chromosome lesions. The realization of these potential chromosome damages in the groups of cells studied is assumed to depend on the activity of the protein synthesis or of the metabolism in the cells. The more active is the chromosome, the more rapidly are realized the potential chromosome damages.

Supplementary effect of high temperature (37 degrees C) an hour after the treatment of mature sperms of Drosophila with ethylene imine resulted in an increased level of the inactivation (the frequency of dominant lethal mutation). The frequency of complete mutations (recessive sex-linked lethal mutations) increased by the supplementary effect of high temperature at low doses of E1, and it did not change under a comparatively high dose of the mutagen. The frequency of mosaic mutations decreased under the effect of high temperature at both doses of E1. No effect of high temperature was observed in 4 and 24 h after the E1 treatment. The results obtained are discussed in connection with the proportion of inactivation and mutagenesis under the effect of chemical mutagens on Drosophila germ cells.

A simple method of determination (with a high reproducibility) of concentrations of the alkylating compound in the culture medium is described. Thiophosphamide failed to decompose in the course of 24 hours either in the culture mixture or in the human lymphocyte culture. As to phosphamide, dipine and fotrin, these drugs disintegrated significantly in the culture during 24 hours, but not in the course of the first 6 hours.

By means of affinity labelling with N,N-dimethyl-2-phenylaziridinium ion (DPA) two forms of acetylcholinesterase were synthesized that contained one or two molecules of the label covalently attached to the enzyme. The reaction of native and covalently modified acetylcholinesterases with n-alkane sulfonyl chlorides CnH2n + 1SO2Cl at n = 1 -4 was used to characterize the reactivity and properties of the enzymes. It was found that labelling of acetylcholinesterase with one molecule of DPA did not affect the enzyme's reactivity. Acetylcholinesterase containing two labels (the second one presumably located at the anionic centre of the enzyme) displayed enhanced and more specific reactivity towards alkane sulfonyl chlorides. It was found that the phenomenon of acceleration caused by affinity modification is analogous to the influence of n-tetraalkylammonium ions on the same reaction. Therefore, the mechanism of regulation of the properties of the esteric centre, caused by affinity labelling of the enzyme at the anionic centre, is the same as in the case of n-tetralkylammonium ions.

Rubomycin or dipin in combination with diacarb had higher toxicity and antitumor activity as compared to administration of the cytostatic agents alone to mice with transplanted lymphosarcoma, strain L10-1. The chemotherapeutic indices of the combinations did not differ or were somewhat lower as compared to the same indices of the cytostatic agents used alone.

Ethylene imine induces mitotic crossing-over between the locus "white" and the centromere by treatment of embryonic and post-embryonic development stages of Drosophila melanogaster. Embryos at the age of 0-4 hours are less sensitive to the recombinogenic effect of ethylene imine than the late embryos at the age of 16-20 hours and early larvae at the age of 24-28 hours after the egg deposition. In the latter case the frequency of somatic mosaics reaches 30%. Mitotic crossing-over is considered as one of the sensitive test-methods of evaluation of genetic activity of some environmental factors.