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.

p53 gene mutations in neoplastic transformation of C3H 10T1/2 and severe combined immunodeficiency fibroblasts.

The relevance of p53 mutations to the neoplastic malignant transformation of rodent fibroblasts by genotoxic physical and chemical agents is not clear. In the present study, we investigated p53 mutations (in exons 5-8) in non-transformed and neoplastically transformed C3H 10T1/2 and severe combined immunodeficiency (SCID) cells. No p53 mutations were detected in 15 neoplastically transformed (two spontaneous, one 3-methylcholanthrene-induced, seven gamma-ray-induced and five 'hot particle'-induced) and two non-transformed 10T1/2 cells.

Wortmannin sensitizes mammalian cells to radiation by inhibiting the DNA-dependent protein kinase-mediated rejoining of double-strand breaks.

Wortmannin has been shown to be an efficient radiosensitizer. Since wortmannin is able to inhibit DNA-dependent protein kinase (DNA-PK) and double-strand break (DSB) rejoining, it is believed that its mechanism of radiation sensitization is through the inhibition of DNA-PK-mediated repair of DSBs. However, since wortmannin is not a specific inhibitor, the possibility that other kinases are inhibited and thereby may contribute to radiosensitization cannot be ruled out.

Stroke risk factors and stroke prevention.

Stroke is the third leading cause of death and the leading cause of chronic disability in the United States. In the past several decades, case series, case-control studies, and prospective cohort studies have successfully identified nonmodifiable risk markers for stroke, such as age, gender, race, ethnicity, heredity and several well-established modifiable risk factors for ischemic stroke. Hypertension, atrial fibrillation, other cardiac diseases, hyperlipidemia, diabetes, cigarette smoking, physical inactivity, carotid stenosis, and transient ischemic attack (TIA) are all potentially treatable conditions that predispose to stroke.

The protective effect of moderate alcohol consumption on ischemic stroke.

Context: Moderate alcohol consumption has been shown to be protective for coronary heart disease, but the relationship between moderate alcohol consumption and ischemic stroke is more controversial.

Objective: To determine the association between alcohol consumption and risk of ischemic stroke.

Design: Population-based case-control study conducted between July 1993 and June 1997.

Phase transitions in the growth of C3H 10T1/2 cells.

In systems used to express transformation using focus formation as the end point, nontransformed cells generally express a down-regulation of cell growth and division made evident by the formation of a monolayer of cells that completely covers the growth surface. In C3H 10T1/2 cells, down-regulation is thought to be progressively effected principally by cell-to-cell communication via gap junctions. Starting with a sparse population in asynchronous growth--e.

Cerebellar hemorrhage.

Cerebellar hemorrhage may present with a spectrum of clinical manifestations, from a benign course with little to no neurologic deficit to a rapidly fatal course with hydrocephalus and brainstem compression. In patients with clinical deterioration, ventricular drainage and surgical evacuation of clot may be life-saving. Several retrospective studies have attempted to define radiographic indicators of the need for surgery with moderate success.

Cell-cycle sensitivity, recovery from radiation damage and a new paradigm for risk assessment.

Tikvah Alper's interest in science was broad, from scrapie to mammalian cells and cancer. Much of her own work focused on cell lethality, like that of many other radiobiologists, but this was natural because of the simplicity of the endpoint cell survival and its relevance to cancer therapy. Tikvah had broader interests, however, that included the effects of radiation on living systems in general like the induction of cancer and the cellular and molecular processes contributing to it.

Radon-induced cancer: a cell-based model of tumorigenesis due to protracted exposures.

In 1982, results with C3H mouse embryo cells showed that the frequency of neoplastic transformation was enhanced when exposures to fission-spectrum neutrons were protracted in time. This finding was unexpected because the opposite was found with low-LET radiations. Similar neutron enhancements were reported with normal life-span Syrian hamster embryo cells, and with human hybrid cells.

Enhanced cytochrome P450 (Cyp1b1) expression, aryl hydrocarbon hydroxylase activity, cytotoxicity, and transformation of C3H 10T1/2 cells by dimethylbenz(a)anthracene in conditioned medium.

The basal and benz(a)anthracene-induced aryl hydrocarbon hydroxylase activities of C3H 10T1/2 mouse embryo fibroblasts have been shown to vary with population growth. We report here that, in the case of dimethylbenz(a)anthracene, cytotoxicity and transformation (neoplastic/morphological transformation and focus formation) increased as a consequence of population growth and, at high cell densities, DNA adduct formation was elevated. Among the factors that may contribute to these changes, we have found that conditioning of the medium with population growth plays a significant role.

cDNA cloning, sequence analysis, and induction by aryl hydrocarbons of a murine cytochrome P450 gene, Cyp1b1.

C3H mouse embryo fibroblast cells, designated 10T1/2, can be transformed by physical and chemical agents including polycyclic aromatic hydrocarbons. In a previous report (Shen et al., Proc.

Inhibitors of poly (ADP-ribose) synthesis inhibit the two types of repair of potentially lethal damage.

Purpose: The purpose of this study was to examine whether 3-amino-benzamide (3ABA), an inhibitor of poly (ADP-ribose) synthesis, inhibits the two types of potentially lethal damage (PLD) repair, termed slow and fast.

Methods And Materials: The fast-type PLD repair was measured by the decrease in survival of V79 Chinese hamster cells by postirradiation treatment with 3ABA. The slow-type PLD repair was measured by the increase in survival by posttreatment with conditioned medium (CM), which became conditioned by growing a crowed culture of cells and supports the slow-type PLD repair.

Enhanced tumorigenesis by small, protracted doses of densely ionizing radiation.

Starting with observations that were first published in 1982, a series of additional findings led to the discovery of an important property of cells in late G2/mitosis. In addition to being the most sensitive to killing, cells in this age-interval were also shown to be the most sensitive to radiation-induced neoplastic transformation. In this work, C3H mouse cells, designated 10T1/2, were irradiated with fission-spectrum neutrons and assayed in vitro via the endpoint focus formation on a monolayer of normal cells.

Identification of a cytochrome P450 gene by reverse transcription--PCR using degenerate primers containing inosine.

A cytochrome P450-like gene, tentatively named P450CMEF, was amplified by a mixed oligonucleotide-primed amplification of cDNA from C3H mouse embryo fibroblast cells, designated 10T1/2, that had been treated with 7,12-dimethylbenz[a]anthracene (DMBA) or benz[a]anthracene (BA). A set of inosine-containing degenerate primers that were targeted to two conserved regions of known cytochrome P450 cDNAs were used. One primer was coded for the well-described and conserved heme-binding region of P450 enzymes, and the second was designed based upon other considerations of homology among P450 molecules.

Transformation-sensitive cells in G2/M-phase are not promoted by TPA following 137Cs gamma-rays.

Mouse C3H 10T1/2 cells are most sensitive to radiation-induced neoplastic transformation in the G2/M-phase of the cell cycle. When synchronized 10T1/2 cells were exposed to phorbol 12-myristate 13-acetate (TPA) after irradiation, transformation of cells not in the transformation-sensitive window was enhanced, but transformation of cells already in the transformation-sensitive window was not. Earlier work showed that (a) TPA enhances the frequency of transformation of both high and low dose-rate gamma-irradiated cells by about the same factor, but that (b) TPA enhances the transformation of cells exposed to low dose-rate fission spectrum neutrons appreciably less than cells exposed to high dose-rate fission spectrum neutrons.

Cycle sequencing using degenerate primers containing inosines.

A modified protocol for cycle sequencing DNA amplified by polymerase chain reaction (PCR) is described. The method involves two sequential linear PCR amplifications using a small amount of double-stranded DNA as a template and a stringent annealing temperature: 1) alpha-35S-dATP labeling of specific primers initially in degenerate primer mixture and 2) dideoxy-ribonucleotide termination of the extended and alpha-35S-dATP-labeled specific primers. The method does not require end labeling and is useful in sequencing PCR-amplified DNA sequences from highly degenerate primers when the sequences of the regions flanking those to be primed are unknown.

Neoplastic transformation of C3H mouse embryo cells, 10T1/2: cell-cycle dependence for 50 kV X-rays and UV-B light.

The variation of neoplastic transformation induced by 50 kV X-rays, and by solar-simulating UV-B light, was studied through the cell cycle of C3H mouse embryo cells designated 10T1/2. A mitotic shake-off method was used to harvest mitotic cells. The progression through the cell cycle of initially mitotic cells was followed as a function of time by flow cytometry, DNA labelling for passage through S-phase, and growth curves for cell number.

Enhanced sensitivity to neoplastic transformation by 137Cs gamma-rays of cells in the G2-/M-phase age interval.

C3H mouse 10T1/2 cells, exposed to low doses of fission-spectrum neutrons, have an enhanced frequency of neoplastic transformation if protracted exposures are used (Hill et al. 1982, 1984a, 1985). To explain this anomaly, a biophysical model was proposed (Elkind 1991a,b) having the following essential features: (1) a narrow age interval exists in the growth cycle of 10T1/2 cells in which cells have high sensitivities to transformation; (2) in the latter age interval, cells are also sensitive to killing; (3) with increasing dose, cells at ages earlier in the growth cycle are progressively delayed from entering the sensitive age window; and (4) with increasing dose, the transformation sensitivity of cells in the sensitive window is not expressed due to increased killing.

Abrogation of killing and neoplastic transformation of C3H10T1/2 cells due to 7,12-dimethylbenz[a]anthracene.

The combined action of 7,12-dimethylbenz[a]anthracene (DMBA) and alpha-naphthoflavone (alpha NF) on the survival and neoplastic transformation of C3H10T1/2 mouse embryo fibroblasts has been examined and correlated with DNA adduct formation and removal. When a 24 h DMBA treatment of asynchronously growing cells was followed for the next 24 h by a treatment with alpha NF + DMBA, both killing and transformation per viable cell were abrogated to a large extent. In some instances, transformation was completely abrogated--i.

Aryl hydrocarbon hydroxylase activity assayed in whole cell lysates using synchronous fluorescence spectroscopy.

Synchronous fluorescence spectrophotometry has been used to measure induced and constitutive levels of aryl hydrocarbon hydroxylase activity in lysates of C3H 10T1/2 mouse embryo fibroblasts. Without compromising sensitivity, the method was reproducible, eliminated the need to extract metabolites, and made the procedure simpler and less time consuming than other methods. Moreover, since the assay was tailored to directly measure 3-hydroxybenzo(a)pyrene, a metabolite produced by several cytochrome P-450s, it may be more generally applicable than dealkylation assays, which apparently detect only P-450-IA1.