Radiation-induced neoplastic transformation of C3H10T1/2 cells is suppressed by ascorbic acid

X-ray induced transformation of C3H10T1/2 cells was suppressed in a concentration-dependent manner by administration of ascorbic acid after irradiation (0.1-20 micrograms/ml for the first week) in the culture medium. The dose-response curve was shifted about 60% downward and was slightly steeper in the presence of ascorbic acid (5 micrograms/ml for the first week) than in its absence. The 1-week treatment procedure revealed that cells initiated by radiation remained susceptible to ascorbic acid until the time of morphological phenotype expression. The neoplastically transformed phenotype expressed after incubation for 8 weeks could no longer be suppressed by ascorbic acid even after culture transfer. Similarly, the neoplastically transformed phenotype suppressed for 8 weeks by ascorbic acid treatment was not subsequently expressed in the absence of ascorbic acid. On the basis of the oxygen-detoxifying nature of ascorbic acid, we postulated that expression of the neoplastically transformed phenotype is promoted by reactive oxygen species and peroxy radicals generated in cells during the whole assay period. The data may be useful as a guide for chemopreventive efforts against radiation carcinogenesis.     

Ultraviolet radiation-induced neoplastic transformation of normal human cells,in vitro

Human foreskin cell cultures in scheduled DNA synthesis (S phase) of the cell cycle were exposed to UV irradiation at a dose of 10 J · m^?2 in the presence of insulin. These treated cell populations, when selectively passaged in a high amino acid supplemented complete growth medium (CM) after 20 Dulbecco's phosphate buffered saline (pH 6.8) (PDL), were able to be grown in soft agar. These treated cell populations were also grown in 1% serum supplemented growth medium and at 41°C in 10% serum supplemented growth medium. Cell populations 4–5 PDL after treatment exhibited altered colony morphology and altered lectin agglutination profiles but would not grow in soft agar. These events appeared to be associated with the early stages in the expression phase of the transformed phenotype. After 20 PDL, we observed that these cells would grow in soft agar at a frequency of 20 colonies/10⁵ cells seeded in soft agar. The cell populations derived from these colonies, when propagated and injected into the nude mice, formed myxofibromas at the injection sites rather than the type of tumor (fibrosarcoma) previously described for chemical carcinogen-induced neoplasms.     

A conundrum in molecular toxicology: Molecular and biological changes during neoplastic transformation of human cells

The process of multistage carcinogenesis lends itself to the concept that the effects of carcinogens are mediated through dose-related, multi-hit, linear changes. Multiplein vitro model systems have been developed that are designed to examine the cellular changes associated with the progression of cells through the different stages in the process; however, these systems may have inherent limitations due to the cell lines used for these studies, the manner of assessing the effects of the carcinogens, and the subsequent growth and differentiation of the exposed cells. Each of these variables results in increasing levels of uncertainty relative to the correlation of the events with the actual process of human tumor development. Therefore, the prediction of the ultimate effect of any carcinogen is difficult. Moreover, relationships between individual biological endpoints resulting from carcinogen treatment appear at best to be approximations. The presence of an activated carcinogen inside the cell can give rise to multiple outcomes, only some of which may be critical events. For example, site-specific modification of the 12th and 13th codons of H-ras is different than that in the adjacent 14th and 15th codons. It is interesting to speculate what effect these differences might have on a biological outcome, e.g., transformation to anchorage-independent growth. The use of different model systems to examine the effects of activated carcinogens also creates additional problems. Comparisons ofin vitro transformed cells with similar cells isolated from human tumors indicate that the culture environment appears to influence the expression of a particular phenotype, in that human tumor cells in culture express many of the same parameters as those found in cells transformed with carcinogensin vitro. If the process of transformation is linear, then less aggressive phenotypes should progress to a more aggressive transformed stage. However, in carcinogen-transformed human cells, the populations exhibit phenotypic diversity in that many of the transformed cells differentiate and fial to continue to divide in culture. Historically, we have assumed only a limited role for epigenetic modulation of molecular changes that occur during progression; however, our data suggest quite strongly that nonmalignant tumor populations can be converted to a more malignant phenotype without additional mutations taking place and, conversely, malignant populations can be downregulated to a nontumorigenic phenotype. Tumor cell plasticity is not only a fundamental characteristic of diverse types of human tumors, but also appears as an integral characteristic of carcinogen-transformed cellsin vitro.Abbreviations AIG anchorage-independent growth - B[a]P benzo[a]pyrene - BPDE-I benzo[a]pyrene diol epoxide I - I-NP 1-nitrosopyrene - PCR polymerase chain reaction - PDL population doubling(s)     

Differentiation and the neoplastic transformation of normal cells

It is suggested that blastomogenic factors can be considered as the means isolating normal cells from each other. Carcinogenic substances caused cell functions to disturb, to inhibit or to intensify the synthesis of specific substances, to change the cell surface disrupted contacts of interactions between cells, pressing cells to become on the way of evolution, and to create heterogenic populations of cells.     

The neoplastic transformation of SCID cells by radiation.

Severe combined immunodeficiency (SCID) cells are hypersensitive to killing by ionizing radiation because of deregulation of DNA-dependent protein kinase (DNA-PK) and a concomitant deficiency in the repair of DNA double-strand breaks. The effect of this condition on the neoplastic transformation of SCID fibroblasts, designated SCID 3T1, has been investigated. The spontaneous transformation rate was approximately 2 x 10(-5) at early passages and increased up to approximately 7 x l0(-3) at later passages. The radiation survival curves of transformed cells had thresholds and therefore appeared to be qualitatively similar to the survival curves of C3H 10T(1/2) mouse fibroblast cells, but the initial slopes were steeper. In contrast, per unit dose, SCID cells were more sensitive to transformation than 10T(1/2) cells. Eight transformed clones were tested for tumorigenicity, and all produced fibrosarcomas in athymic nude mice. Properties associated with the tumor suppressor Trp53 (formerly known as p53) were examined in three of the clones. In these clones, although Trp53 protein was overexpressed, a lower expression of Cdkn1a (formerly known as p21, Cip1) protein was observed compared to parental cells. The expression of Trp53 and Cdkn1a and the G(1)-phase arrest (one set of data on G(1)-phase delay is included as an example) was not induced by ionizing radiation in these transformed clones; each clone carried a point mutation in Trp53. This suggests that the deficiency in the repair of DNA double-strand breaks increased the tumorigenicity and the genomic instability of transformed SCID cells.     

Changes in gene expression accompanying neoplastic transformation of Syrian hamster cells

Proteins solubilized from the chemically transformed, highly tumorigenic Syrian hamster cell line, BP6T, and the untransformed parental embryo cells, have been analyzed by two-dimensional gel electrophoresis. Differences in seven major polypeptides have been identified in cytoplasmic and nuclear cell fractions from these two related cell types. The tumorigenic cells have lost the ability to synthesize detectable amounts of five major polypeptides which are found in untransformed cells; in addition, the tumorigenic cells synthesize two new major polypeptide species not found in the untransformed cells. Butyric acid, an agent which suppresses in vitro cellular properties frequently associated with neoplasia, induces in a reversible fashion synthesis of two of these missing polypeptide species in the tumorigenic cells. The results indicate that a change in the synthesis of less than 1% of the major polypeptide species is associated with a chemical mediated induction of the high tumorigenic state of Syrian hamster cells.     

Serum-induced chromosome damage and neoplastic transformation of mouse cells in vitro

Summary In previous studies, mouse cells grown in medium supplemented with horse serum (HS) developed more chromosomal aberrations and underwent malignant transformation earlier than cells from the same pool grown with fetal bovine serum (FBS) supplement. In the present study cells derived from C3H_f/HeN mouse embryos were grown in medium NCTC-135 supplemented with various combinations of large- and small-molecule fractions of HS and FBS in an effort to determine the effective components. The results indicate that the large-molecule fraction of HS (mare or stallion) produces alterations in chromosome number and structure. HS is also shown to cause chromatid breaks and exchanges at or near the centromere in contrast to fluorescent-light-induced breaks which occur randomly along the chromatid. However, efforts to control completely chromosome stability and malignant transformation through the use of large-and small-molecule fractions of HS and FBS or combinations thereof were unsuccessful. In comnection with this study, diagnosis of malignant transformation in vitro was made by a direct sampling method based on cytologic criteria previously described and documented. With one exception, the diagnoses of 11 different cell lines were consistent with results of in vivo assays.     

Activated leukemic oncogenes responsible for neoplastic transformation of hematopoietic cells

Leukemias are a heterogeneous group of malignant blood diseases that are characterized by expansion of immature blast cells. The point molecular mechanisms of leukemogenesis are still unknown. Leukemia patients frequently have mutations of the genes responsible for normal proliferation and differentiation of hematopoietic stem cells. At present, scientific groups worldwide are engaged in biomedical studies of the structural and functional aspects of leukemic oncogenes and their role in human and animal leukemogenesis. The review describes the current concepts of the molecular properties of oncogenes whose activation may lead to CBF-AML, which results from mutations of the genes for the core binding factors AML1 (CBF6h) and CBFß.     

Radiation-induced changes in the cell membrane of cultured human endothelial cells

We investigated the effect of irradiation on the kinetic characteristics of amino acid and glucose transport, and the effect on the activity of the cell membrane-bound enzyme 5'-nucleotidase and on the receptor-mediated stimulation of cyclic adenosine monophosphate synthesis by prostaglandin E1. Irradiation inhibited the sodium-dependent amino acid transport by a reduced binding of the amino acid to the transport unit. The transport of glucose, which appeared to be a sodium-independent process, was temporarily stimulated by increased maximal velocity of the transport. No effect was found on the binding to the transport unit. Irradiation increased the 5'-nucleotidase activity and decreased the prostaglandin E1-stimulated cyclic adenosine monophosphate synthesis 48 h after exposure to 20 Gy. It is concluded that irradiation decreases sodium-dependent transport by impairment of the transport unit, does not impair a sodium-independent process, and has opposite effects on membrane-bound enzyme activity and a receptor-mediated process.     

Radiation-induced changes in oral carcinoma cells – a multiparametric evaluation

The aim of this study was to see whether serial cytological evaluation of various cellular abnormalities in tumours from patients receiving fractionated radiotherapy can predict radio-response in oral carcinoma. Cytological assessment was carried out in scrape smears collected prior to and during the course of radiotherapy in 68 patients with squamous cell carcinoma of the oral cavity planned for radical radiotherapy with accelerated fraction schedule. Smears were evaluated for a set of 15 radiation-induced cellular abnormalities. The relationship between the cellular alterations and the cumulative radiation dose was analysed by Kruskal-Wallis one-way anova. The results showed that among the various quantifiable changes that occur in irradiated cancer cells, karyolysis, karyorrhexis, pyknosis, cytolysis, multinucleation, micronucleation and nuclear budding show significant increase depending on the dose of radiation. The radio-resistant group of patients exhibited a lesser degree of change compared with the radio-sensitive group. This suggests that radio-resistance may be due to the defective induction of cell damage and that these cytological features may have potential use as predictive markers of radio-sensitivity in oral carcinoma.     

Structural changes occurring during transformation of the labial gland cells to their adult form,in Manduca sexta

The labial gland of the adult sphingid moth, Manduca sexta, is composed of five distinct regions, each made of a single cellular type. Four of these regions are derivatives of the single specialized cellular population that makes up the caterpillar labial duct. Both the larval labial duct and its derivatives are large, polyploid cells with pleiomorphic nuclei. There is a definite cellular continuity between the larval and adult forms of these cells throughout metamorphosis; no mitoses or cell deaths are seen to occur in the gland during transformation. Cytological studies indicate that in the process of cell transformation the ducts first “dedifferentiate,” elongate, then redifferentiate. Intermediates in this process have well defined structures which should make this system useful in studying covert events in the transformation process.     

A novel c-Myc-responsive gene, JPO1, participates in neoplastic transformation

We have identified a novel c-Myc-responsive gene, named JPO1, by representational difference analysis. JPO1 responds to two inducible c-Myc systems and behaves as a direct c-Myc target gene. JPO1 mRNA expression is readily detectable in the thymus, small intestine, and colon, whereas expression is relatively low in spleen, bone marrow, and peripheral leukocytes. We cloned a full-length JPO1 cDNA that encodes a 47-kDa nuclear protein. To determine the role of JPO1 in Myc-mediated cellular phenotypes, stable Rat1a fibroblasts overexpressing JPO1 were tested and compared with transformed Rat1a-Myc cells. Although JPO1 has a diminished transforming activity as compared with c-Myc, JPO1 complements a transformation-defective Myc Box II mutant in the Rat1a transformation assay. This complementation provides evidence for a genetic link between c-Myc and JPO1. Similar to c-Myc, JPO1 overexpression enhances the clonogenicity of CB33 human lymphoblastoid cells in methylcellulose assays. These observations suggest that JPO1 participates in c-Myc-mediated transformation, supporting an emerging concept that c-Myc target genes constitute nodal points in a network of pathways that lead from c-Myc to various Myc-related phenotypes and ultimately to tumorigenesis.     

Glycoprotein electrophoretic patterns have potential to monitor changes associated with neoplastic transformation in oral cancer

Alterations in glycoproteins, important cell surface constituents, have long been associated with various malignancies. The present investigation therefore explored the clinical significance of a glycoproteomics approach in patients with oral precancerous conditions (OPC) and patients with oral cancer. The study included 80 oral cancer patients, 50 patients with OPC, and 84 controls. Native polyacrylamide gel electrophoresis followed by Schiff’s staining was carried out to study the alterations in glycoproteins. The results showed significant elevation (p<0.0001) of 192 kDa, 170 kDa, 116 kDa and 44 kDa glycoproteins in oral cancer patients and patients with OPC compared with controls. The odds ratio indicated a significantly higher risk for oral cancer among users and especially chewers of tobacco. The levels of all the glycoprotein bands (192 kDa, 170 kDa, 116 kDa and 44 kDa) were higher in patients with a habit of tobacco use (WHT) than in patients with no habit of tobacco (NHT) and were also higher in WHT controls than in NHT controls. Moreover, a 230 kDa glycoprotein consistently appeared only in individuals with tobacco habits and an increasing trend was observed from WHT controls to patients with OPC to WHT oral cancer patients. In conclusion, the results indicated the potential utility of glycoprotein alterations in monitoring sequential changes occurring due to tobacco consumption during neoplastic transformation.