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It is well known that artificial increase in expression of growth factors and their receptors can lead to tumorigenic transformation of cells. Additionally, multiple data on the increased expression of growth factors in many human and animal tumorigenic cells have been published. Nevertheless description of the functional role of endogenous growth factors in maintenance of tumorigenic phenotype remains obscure. Previously, we described a new model for studying neoplastic transformation and dormant metastasis. This model consists of cognate tumorigenic and nontumorigenic cell clones, the latter being obtained as a result of spontaneous reversion of tumorigenic ones. All revertant clones demonstrate the three well known features of normal cells: monolayer growth on a plastic, incapability to grow in soft agar with regular culture media with 10% FCS, and incapability to form tumors in syngeneic animals. Using RT-PCR, we measured expression ratios of main growth factors, commonly believed to be associated with malignization, in tumorigenic and revertant clones of our model. For some of these clones, we also measured an activity of growth factors in conditioned media. The data obtained argue that the levels of growth factor expression, measured by both the methods, are distributed between tumorigenic and revertant clones in a sporadic manner and do not correlate with cell tumorigenicity. Thus, our experimental observations enable us to consider the variability of growth factor expression as insignificant events in the reversion of tumorigenic cells to a nontumorigenic phenotype.

In the present paper we have shown that JB6 and PDV murine skin carcinoma cells, as well as previously described sarcoma B6-4 cells, can revert to a nontumor phenotype. Revertant carcinoma clones could not grow in soft agar conditions and like sarcoma revertants acquired dependence on peptide growth factors, and exhibited a reduced expression of c-jun. Spontaneous revertants were shown to be instable. They could revert back to a transformed phenotype in 1-5 months of in vitro passaging. Being inoculated in syngeneic animals, these transformed cells show a recurrence in 2-5 months, similar to that of a dormant metastasis. Thus, dormant revertant cells are believed to be included in many tumors of different origin. So, spontaneous reversions of tumor cells may play an important role in the dormant metastatic process. The cause of these frequent spontaneous transient reversions and revertant instability appears to be of epigenetic nature. Causes and mechanisms of cell transformations and reversions remain to be clarified.

Karyological analysis of 6 cell lines with distinct tumorigenic properties of mouse strains C3H/He and CBA/Ca has been carried out using differential chromosome staining. All the cell lines are characterized by a decreased number of copies of normal chromosome 7, the increased number of normal copies of chromosome 10 being specific of the cell lines with intermediate tumorigenicity. Cell lines with maximum tumorigenicity differed from all other lines by the increased number of copies of chromosome 5 and by the decreased number of copies of chromosome 6. A wide independent variability was observed in the number of chromosomes and of several types of abnormal chromosomes throughout the neoplastic evolution of cells, to begin from the early immortal passages. But the proportion of normal chromosomes per cell in the studied lines revealed relatively stable values. The potential phenotypical heterogenicity of the lines with maximum tumorigenicity, expressed in their clonal progeny, was associated with the instability in the number of chromosome 15 copies in cells of these lines. It is concluded that multiple genetic events are required in the spontaneous neoplastic evolution of fibroblasts, and only specific traits of the karyotypic instability, associated with the variability of the number of copies of specific chromosomes, may constitute the genetic basis for the above process.

Karyological analysis of 6 lines with distinct tumorigenic properties of mouse strains C3H/He and C57BL/6 has been carried out using a differential staining of chromosomes. The number of normal copies of chromosomes varied in all the investigated cell lines. The more and the less stable chromosomes different from line to line. All the cell lines were characterized by decreased numbers of copies of normal chromosome 7; a decreased number of normal copies of chromosome 2 and 16 was detected in the course of the cell spontaneous neoplastic evolution. The decreased number of normal copies of chromosomes 8, 12 and X, and the increased number of normal copies of chromosome 10 were specific of the cell lines with intermediate tumorigenicity. The maximum tumorigenic cell lines differed from all other lines by increased numbers of copies of chromosomes 4 and 5, and by a decrease in copy number of chromosome 6. The data obtained are discussed in terms of the search of the regularity of karyotypic changes in the course of the cell neoplastic evolution.

Three cell cultures of C3H mouse strain--L, MMT1 and embryonic fibroblasts have different sensitivity to cytolytic action of allogeneic antiserum and to immune lymphocyte action. A sensitivity to cytolytic action of antiserum for these cultures is correlated with the amount of corresponding transplantation antigens. A sensitivity to cytolytic action of immune allogeneic lymphocytes is correlated with the amount of corresponding antigens as well as cell with properties determining the level of lymphocyte sorption on them.

7 continuous cell lines with different degrees of malignancy were obtained from cultures of embryo fibroblasts of 4 strains of mice. Three of them were selected in vivo for enhanced malignancy. A study of regularities of changes in immunologic and some cytologic characteristics of these cells in the processes of transformation and malignization has been started. The data are presented that in these processes the density of H-2 antigens in embryo fibroblasts many vary significantly in both directions. These alterations are stochastic and do not depend on transformation or malignization of cells. It is also shown that the sensitivity of the cells under study to the complement lytic action may significantly vary, and does not always correlate with the density of antigens which are the targets of the complement lytic action.

Data are provided on the development of tumorigenicity in cells of 6 independently produced lines of mouse and rat embryonic fibroblasts during their prolonged passage in vitro. The data on the development of lysis of these cells by macrophages and activated lymphocytes during the same passage indicate that enhanced cell lysis of cells by natural effectors and, hence, cell recognition by them may be regarded as an obligatory manifestation of cell malignization.

The previous report contained the information on obtaining in vitro continuous cell lines of embryonic fibroblasts of mice of 4 genotypes, including C3H. In this paper, data on the degree of lysis of some derivatives of FC3H3 cells by natural killers and macrophages are given. It was shown that normal cells were not lysed by these effectors unlike cells which overcame Hayflick's limit and got minimum malignancy in vitro. However, it was established that the in vivo selection of cells for high tumorigenicity resulted in the formation of tumor cells resistant to lymphocytes--natural killers and macrophages.

Immune allogeneic cells of lymph nodes, spleen and peritoneal exudate lyse in vitro dividing and irradiated non-dividing target cells with the same intensity. The number of target cells lysed during the immune attack are more precisely registered in irradiated non-dividing cells.