Frequency Of Spontaneous Transformation Research Articles

Influence of thespxBGene on Competence inStreptococcus pneumoniae

In Streptococcus pneumoniae expression of pyruvate oxidase (SpxB) peaks during the early growth phase, coincident with the time of natural competence. This study investigated whether SpxB influences parameters of competence, such as spontaneous transformation frequency, expression of competence genes, and DNA release. Knockout of the spxB gene in strain D39 abolished spontaneous transformation (compared to a frequency of 6.3 x 10(-6) in the parent strain [P < 0.01]). It also reduced expression levels of comC and recA as well as DNA release from bacterial cells significantly during the early growth phase, coincident with the time of spontaneous competence in the parent strain. In the spxB mutant, supplementation with competence-stimulating peptide 1 (CSP-1) restored transformation (rate, 1.8 x 10(-2)). This speaks against the role of SpxB as a necessary source of energy for competence. Neither supplementation with CSP-1 nor supplementation with the SpxB products H2O2 and acetate altered DNA release. Supplementation of the parent strain with catalase did not reduce DNA release significantly. In conclusion, the pneumococcal spxB gene influences competence; however, the mechanism remains elusive.

Induction of an Adaptive Response against Spontaneous Neoplastic Transformation In Vitro by Low-Dose Gamma Radiation

An adaptive response against spontaneous neoplastic transformation in vitro induced by low-dose gamma radiation has recently been reported using a clone of C3H 10T1/2 cells with a predisposition toward spontaneous transformation (Azzam et al., Radiat. Res. 146, 369-370, 1996). To test the generality of this observation, the HeLa x skin fibroblast human hybrid cell system was used to look for such an adaptive response using a similar experimental protocol. In the experimental protocol used, the frequency of neoplastic transformation of unirradiated cultures (Arm A) was compared to that of cultures which had been irradiated with 1 cGy of gamma radiation and either plated immediately (Arm B) or held for 24 h at 37 degrees C prior to plating (Arm C) for assay of neoplastic transformation. The pooled data from four separate experiments demonstrated a significantly reduced transformation frequency for Arm C compared to Arms A and B. This is indicative of an adaptive response in the case of postirradiation holding, in agreement with the earlier study using C3H 10T1/2 cells. However, with the exception of one experiment, the existence of the adaptive response was not clear on the basis of analysis of individual experiments. This points out the importance of sample size when looking at low-dose effects that are close in magnitude to the inherent variations in spontaneous transformation frequency.

SHE Cell Transformation Assay as a Predictive System for Carcinogenicity of PAH and Non-Genotoxic Carcinogens

Abstract The Syrian hamster embryo (SHE) cell transformation assay is an in vitro method for the detection of genotoxic and non-genotoxic carcinogens. Its reliability as a multistage carcinogen screening method was demonstrated by several authors who showed that cells derived from transformed colonies were capable of tumour formation when inoculated in a suitable host animal. The SHE assay offers the advantage of using normal diploïd cells with metabolizing capacities and low spontaneous transformation frequency. Epithelial cultures can be selected from the mixed cellular suspension obtained from the Syrian hamster embryos in order to develop an in vitro model for the detection of carcinogens inducing carcinomas. Polycyclic aromatic hydrocarbons, such as BaP and DMBA were studied in depth for their cell transforming potency. The synergistic effects of PAH and tumor promoters can be demonstrated by using a sequential treatment consisting in an initiation-promotion scheme1. This protocol also revealed interacting effects between non-genotoxic carcinogens2.

In vitro transformation potential of N-polycyclic aromatic hydrocarbons in rat tracheal epithelial cells

Nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) are environmental contaminants and potential human airway carcinogens. Agents of this class show a wide range of potencies for toxicity, mutagenicity and carcinogenicity that are associated with the structure of the PAH and the position of the nitro group. In order to evaluate the effect of nitro substitution on in vitro biological activity, the cytotoxicity and transformation potential of two parent PAHs, pyrene and chrysene, and a series of nitro derivatives were examined in the rat tracheal epithelial (RTE) cell system. The nitro derivatives, but not pyrene or chrysene, produced dose-dependent decreases in the colony forming efficiency of the RTE cells. The most cytotoxic agents were 1,6-dinitropyrene and 6-nitrochrysene with ED 50s of 1.6 μ m and 5.9 μ m, respectively, followed by 4-nitropyrene and 1-nitropyrene with ED 50s of 26.3 μ m and 44.5 μ m, respectively. These compounds were evaluated for transformation potential at three treatment levels that spanned the cytotoxic range, and the assays were scored for morphologically transformed preneoplastic colonies. The control or spontaneous transformation frequency in this series of experiments was 1.79 ± 0.47 (× 10 −4). 6-Nitrochrysene and 1,6-dinitropyrene were the only compounds that produced transformation frequencies (12.17 × 10 −4 and 9.68 × 10 −4, respectively) that were statistically different from control. The maximum transformation frequencies of the compounds were compared with published data for liver tumorigenicity in the newborn mouse assay. The orders for tumorigenicity and transformation were the same (1,6-dinitropyrene > 4-nitropyrene > 1-nitropyrene ∼- pyrene and 6-nitrochrysene > chrysene), and the relative potencies of the compounds were similar in the two assays. These results suggest that RTE cells are capable of metabolizing nitro-PAHs to reactive products, and that, within this limited class of compounds, in vitro transformation data in the RTE cell system may be correlated with tumorigenicity in animal studies.

Transformation of BALB/c-3T3 cells: I. Investigation of experimental parameters that influence detection of spontaneous transformation.

The frequency of spontaneous morphological transformation is an important variable in measuring chemical-induced transformation in BALB/c-3T3 clone A-31-1-13 cell cultures. Data from 110 experiments, which included benzo[a]pyrene control groups and other chemical treatment groups, were analyzed for factors that influenced spontaneous transformation. Spontaneous transformants demonstrated a continuum of morphological variants (type I, II, and III foci) that fit a normal distribution if converted to log10. The magnitude of transformation depended on the ampule of cryopreserved cells and the serum lot. Although the average frequency was approximately 0.71 x 10(-6) (type III foci/cell that survived and proliferated to confluence), the absolute number of foci/vessel increased in proportion to the surface area of the culture vessel. Thus, the frequency of spontaneous transformation was directly related to the cumulative number of mitoses that occurred in forming the contact-inhibited monolayer. These data are consistent with a hypothesis that spontaneous transformation in BALB/c-3T3 cells is a mutational event or some other single-step phenomenon.

Neoplastic progression of Syrian hamster embryo cells in culture.

AbstractThe transformation of a normal cell into a malignant cell is generally recognized as a multistep process involving genetic alterations in multiple genes in a progressive, stepwise manner (1, 2). The progression of cells toward malignancy can be studied in cell culture models of neoplasia. The advantage of this approach is that it allows one to select and isolate cells at different stages of the process for biochemical, molecular, and cellular studies.One of the best-characterized cell culture models of neoplastic progression utilizes the Syrian hamster embryo cells, which have a low frequency of spontaneous transformation but can be neoplastically transformed by treatment with chemical carcinogens (3). It was shown several years ago that the transformation of the cells in culture is a multistep, progressive process (4), and the cellular, biochemical, and molecular changes of cells at different stages have been described (2, 3, 5-8).

The effect of sodium selenite on cell proliferation and transformation of primary rat tracheal epithelial cells.

The effects of sodium selenite (Na2SeO3) on cell proliferation and the development of preneoplastic transformed variants were studied in primary cultures of rat tracheal epithelial cells. Results revealed a biphasic effect of Na2SeO3 on cell proliferation: at concentrations between 6 x 10(-8) and 6 x 10(-6) M, it stimulated and at concentrations of approximately 2 x 10(-5) and above it inhibited cell proliferation (presumably due to toxicity). Nontoxic concentrations of Na2SeO3 (6 x 10(-8) -6 x 10(-7) M) significantly reduced the spontaneous transformation frequency. Transformation induced by the tobacco-specific nitrosamine 4-(methyl-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK) was effectively inhibited by nontoxic as well as toxic concentrations of Na2SeO3. Treatment of cultures with Na2SeO3 after cessation of NNK exposure, i.e. during the selection period, also significantly reduced the transformation frequency. These experiments show that the inhibition of transformation by Na2SeO3 is not the result of an antiproliferative effect. They further indicate that the inhibitory effect occurs even when the chemical treatment occurs during the 'postinitiation' phase. Thus the inhibition of transformation by Na2SeO3 cannot solely be explained by its effects on drug metabolism.

The effect of ploidy and colcemid on the frequency of spontaneous transformation of cultured cells

The frequency of spontaneous transformation, defined as the acquisition of the ability to form colonies in soft agar (anchorage independence) was compared in rat fibroblasts of recently established lines possessing pseudodiploid to pseudotetraploid numbers of chromosomes. Transformation frequency appeared to be independent on the ploidy level. The effect of colcemid on the transformation frequency of pseudodiploid fibroblasts was also examined. Colcemid treatment resulted in an increased frequency of transformation. In addition, most of the transformed clones, either derived from colcemid-treated cells, or derived from untreated cells, were characterized by significant changes in chromosome number. These results cannot be easily reconciled with the hypothesis that the main cause of spontaneous transformation is the appearance of recessive mutations. They rather support the idea that this conversion is the consequence of alterations in the number of certain chromosomes.

The Characteristics Of Animal Cells Transformed In Vitro

This chapter concerns with the neoplastic transformation of cells in culture. Transformation may occur “spontaneously,” or may be induced by oncogenic viruses and chemical carcinogens. The culture of dissociated animal tissues directly on glass or plastic is established as a simple and valuable technique in many fields of biological research. The methods for the initiation and serial subcultivation of such cultures have remained essentially unchanged since their introduction. During the intervening period, much information has accumulated, although in a rather haphazard fashion, on the growth characteristics of cells derived from various animal species. Some characteristic patterns of behavior have been recognized and cell culture studies have raised many new questions about growth, cell division, metabolism, differentiation, senescence, and transformation. The behavior of cells of different species with regard to their phase of decline and their frequency of spontaneous transformation is represented diagrammatically in the chapter. This representation of behavior of cells from different species is a collation of the results from many published accounts. Some of the studies described in the chapter are related to cells that have spontaneously acquired a high degree of autonomy in vitro . These include BHK2l, NIL-2, and 3T3. These lines retain some of the normal responses to growth controls and lose them when transformed by oncogenic viruses.