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Biomedical subjects

A C Begg

Publications and source records attributed to A C Begg.

At least 91 records · Page 5Linked to original sources
PubMed1985

A method to measure the duration of DNA synthesis and the potential doubling time from a single sample.

A method is described whereby the DNA synthesis time, Ts, can be calculated using data of a single sample of cells taken several hours after labelling with bromodeoxyuridine (BrdUrd). The method involves a simple calculation using flow cytometry data of BrdUrd incorporation (green fluorescence, FITC-labelled anti-BrdUrd-DNA antibody) and total DNA content (red fluorescence, propidium iodide). The movement of BrdUrd-labelled cells through the S phase can be quantified by measuring their mean red fluorescence relative to that of G1 and G2 cells. Assuming the movement of the labelled cells toward G2 is linear with time, Ts can be calculated by measuring their relative movement at any one time. The method was tested on cells in vitro and on bone marrow and tumor cells in vivo. Reasonable agreement was seen with published estimates of Ts for these tissues.

Animals↗
PubMed1985

Differences in chemosensitivity between subcutaneous and pulmonary tumours.

The response of a murine tumour to cyclophosphamide has been studied in two sites (subcutis and lungs) over a wide range of tumour sizes. The site of tumour growth has been shown to have a marked influence on chemosensitivity for tumours of equivalent size. Pulmonary metastases are much more sensitive than subcutaneous implants. In the lung there is a simple decrease in curability as the tumour grows, presumably reflecting the increase in clonogenic cells per tumour. In the subcutis the pattern is more complex. An initial sensitivity is followed by a decline during the avascular phase of growth. As the vascular network develops the tumours regain their chemosensitivity. There is no correlation between volume doubling time and chemosensitivity. This study indicates that it is impossible to predict the response of pulmonary deposits from a study of subcutaneous implants.

Animals↗
PubMed1985

Influence of cis-diamminedichloroplatinum (II) on mouse duodenal crypt stem cell survival after multifraction X ray treatment.

The mechanism of interaction of cis-platinum and X rays was investigated in mouse duodenal crypts using the microcolony assay. Mice were exposed to 1, 2, 5, 10, or 15 fractions of X rays, either alone or preceded by a single i.p. injection of cis-platinum, 8 mg/kg, one-half hour before the first fraction. In all fractionation regimens, cisplatinum caused a shift of the X ray survival curve for crypt cells towards lower doses. The vertical distances between the survival curves after X rays and those in combination with cis-platinum were about the same, giving a mean value of 0.89 +/- 0.12 log10 cells. After cis-platinum treatment alone, a crypt cell survival curve was established in the high dose range. The estimated cell kill by 8 mg/kg of cis-platinum, obtained by extrapolation of this curve, was 1 log10 cell number. These data imply independent cell killing mechanisms for cis-platinum and X rays. However, even after correction for cell kill by the drug, cis-platinum tended to inhibit slightly sublethal damage repair after X rays. This was supported by linear quadratic analyses, in which the alpha/beta value after combined treatment was found to be slightly higher than after X rays alone (20 +/- 4 Gy versus 13.0 +/- 1.7 Gy).

Animals↗
PubMed1985

Cell cycle kinetics of aerated, hypoxic and re-aerated cells in vitro using flow cytometric determination of cellular DNA and incorporated bromodeoxyuridine.

The effects of extreme hypoxia on cell cycle progression were studied by simultaneous determination of DNA and bromodeoxyuridine (BrdU) contents of individual cells. V79-379A cells were pulse-labelled with BrdU (1 microM, 20 min, 37 degrees C) and then incubated for up to 12 hr in BrdU-free medium under either aerated or extremely hypoxic conditions. After the incubation interval (0-12 hr), the cells were trypsinized and fixed in 50% EtOH. Propidium iodide and a fluorescein-labelled monoclonal antibody to BrdU were then used to quantify DNA content and incorporated BrdU, respectively. Measurements in individual cells were made by simultaneous detection of green and red fluorescence upon excitation at 488 nm using flow cytometry. Bivariate analysis revealed progression of BrdU-labelled cells in aerated cultures out of S phase, into G2 and cell division, with halving of mean fluorescence, and back into S phase by approximately 9 hr after the BrdU pulse. Hypoxia immediately arrested cells in all phases of the cell cycle. Both the DNA distribution and the bivariate profile of cells that were fixed from 2 to 12 hr after induction of hypoxia were identical to the 0 hr controls. The percent of cells with green fluorescence in a mid-S phase window remained 100% and the mean fluorescence of these cells remained at control (0 hr) levels. This indicates that, under hypoxic conditions, cells were moving neither into nor out of S phase. Cultures that had been hypoxic for 12 hr exhibited an increasing rate of BrdU uptake with time after re-aeration. Re-aerated cells were able to complete or initiate DNA synthesis, but their rates of progression through the cell cycle were markedly reduced. A large fraction of cells appeared unable to divide up to 12 hr following release from hypoxia.

Aerobiosis↗
PubMed1985

Fluorescent markers of hypoxic cells: a comparison of two compounds on three cell lines.

Two compounds, nitroakridin 3582 (NA) and a 3-nitro-naphthalimide (DM113), have been tested as potential fluorescent markers for hypoxic cells. Cellular fluorescence in three cell lines (V79-379A, WHF1B, EMT6) was measured by flow cytometry and high-performance liquid chromatography (HPLC) after incubating the cells with the drugs for various times in air or hypoxia. In all three cell lines, both drugs showed greater fluorescence in hypoxic than in oxic cells. There were, however, differences between the cell lines in respect of the magnitudes of hypoxic and oxic cell fluorescence and in the ratio of hypoxic to oxic fluorescence. Differences in hypoxic cell fluorescence were due to differences in the rate and extent of nitroreduction. Drug uptake and DNA content per cell were relatively unimportant factors in determining the magnitude of fluorescence. There was not a good correlation between cytotoxicity of the drugs and hypoxic fluorescence. Nitroakridin was more toxic to hypoxic than to aerated cells but the reverse was true for DM113. The dependence of fluorescence and radiosensitivity on oxygen concentration were compared for the three cell lines and only small differences between the "K"-curves for the two end-points were found. Two problems with the present compounds which should be addressed in designing future fluorogenic compounds as hypoxic markers were oxic cell fluorescence and leakage of fluorescent products from hypoxic cells.

Aminacrine↗
PubMed1985

Effects of hypoxia, pH, and growth stage on cell killing in Chinese hamster V79 cells in vitro by activated cyclophosphamide.

Several factors which influence the sensitivity of Chinese hamster V79 cells to cyclophosphamide (CY) have been studied in vitro in both suspension and monolayer cultures. Activated CY was obtained from the blood of mice 15 to 30 min after i.p. injection of CY (400 mg/kg). At pH 7.4, hypoxia rendered the cells more sensitive to activated CY. At lower values of pH (6.6 and 7.0), there was no difference between the sensitivities of oxic and hypoxic cells, although cells in both conditions were more sensitive to CY than at pH 7.4. Drug sensitivity was markedly affected by the stage of cell growth. Monolayer cultures were most sensitive to CY within a few hours of plating. Cultures then rapidly became less sensitive, with maximum resistance occurring between 24 and 48 h after plating, while the cells were still exhibiting rapid exponential growth. This development of resistance parallelled the formation of small colonies (2 to 4 cells), implying that intercellular contact may confer resistance to killing by activated CY.

Animals↗
PubMed1984

The sensitivity of normal stroma to fractionated radiotherapy measured by a tumour growth rate assay.

The growth rate of implanted tumours has been used as an assay for radiation injury in normal stroma. The subcutaneous tissue was irradiated in an unstimulated state and was then stimulated to produce new blood vessels by the inoculation of a syngeneic tumour 3 days after the last irradiation. Steep dose-response curves were obtained with a dose resolution of approximately 1 Gy. The time between irradiation and stimulation of the stroma by the tumour implant was shown to have no effect on tumour growth rate for times ranging from 1 h to 14 days after single doses. This functional assay of stromal damage has been used after irradiation with 1, 2, 5, 10 or 20 fractions of X-rays. Isoeffect data were well fitted by a linear-quadratic equation, for which the ratio of linear to quadratic coefficients (alpha/beta) was 6.2 +/- 0.6 Gy. This is on the high end of the range of published alpha/beta values for late reacting tissues and other stromal/vascular assays, but lower than those for all early reacting tissues. Overall treatment times ranging from 1 to 11 days were tested with the 2 and 5 fraction schedules. No effect attributable to slow repair or repopulation could be demonstrated over this period.

Animals↗
PubMed1984

A bioassay for cyclophosphamide in blood, lung and tumour.

A bioassay has been developed to detect and quantify the concentration of cytotoxic metabolites of cyclophosphamide (CY) in blood, tumour, and lungs of mice. Extracts were made of blood or solid tissues taken from mice given CY and these were used to treat log phase Chinese Hamster V79 cells in culture for up to 24 h. The amount of cell killing was tested by colony formation 7 days later. The effects of incubation time, CY dose, and the time of tissue sampling after CY injection were investigated. The bioassay could detect cytotoxic metabolites in blood after doses as low as 10 mg kg-1 CY given i.p. The half life of these metabolites in blood after giving 400 mg kg-1 i.p. decreased over a 2 h period from 14 to 9 min. The method was then modified to define the pharmacokinetics of CY metabolites in two different types of tumour and in lung. The half life of the cytotoxic metabolites in the lung was longer than in blood, falling from 35 to 11 min over a 2 h period. In tumours, the half lives were longer again, i.e. approximately 61 min. The maximum metabolite levels achieved were similar in the two tumour types, although these differed markedly in their therapeutic response to CY. The bioassay for CY is a relatively simple and rapid procedure, and the extension of its application from body fluids to solid tissues makes it a useful tool in experimental pharmacokinetic studies.

Animals↗
PubMed1983

Modification of stromal radiosensitivity by misonidazole and WR-2721.

The radiosensitivity of normal vascular connective tissue has been assessed by measuring the growth rate of untreated tumours implanted into pre-irradiated sites. Dose response curves have been constructed from two end-points, the latent period (time for the tumours to reach a diameter of 2 mm) and the growth rate in the macroscopic range (above 4 mm). Well defined dose response curves were obtained for both end-points, with a lower dose threshold for the macroscopic growth rate. A third end-point which combined the first two was also tested, i.e., time to reach 8 mm (23 to 76 days over the dose range tested). This was found to give the steepest dose response curves overall and was therefore judged to be the best assay. Modification of the radiosensitivity of stromal tissue by misonidazole and by WR-2721 was investigated using this assay. The dose response curve for X rays plus misonidazole was similar to that for X rays alone indicating little or no radiosensitisation. WR-2721, however, significantly reduced the X-ray response indicating radioprotection by a factor of 1.8 (range 1.4-2.7). These data show that tumour implantation is an effective way of eliciting a quantifiable response in previously irradiated stromal tissue. The assay allows resolution of dose differences as small as 1 to 2 Gy.

Adenocarcinoma↗
PubMed1980

Analysis of growth delay data: potential pitfalls.

Several theoretical aspects of growth delay curves which can affect the relationship between delay and cell killing have been considered and compared with examples of real data. (1) Damage to the tumour "bed", presumably vasculature, can result in slower growth compared with untreated tumours; the so called "tumour bed effect". As a consequence the delay in growth depends on the end-point size. In addition, agents which modify tumour and stromal cell damage to a different degree may lead to different conclusions depending upon the assay. (2) A slow rate of clearance of dead cells and debris after treatment can lead to slower growth of tumours in the initial stages of regrowth above treatment size. This can appear to be a tumour bed effect even when stromal damage is absent. (3) Average growth curves for a group of tumours should not be used instead of the delay for each individual animal because this can also lead to an apparently slower regrowth in the absence of a tumour bed effect. (4) Tumour size at treatment should be standardized. If the normal growth pattern before and after treatment is Gompertzian, treatments which produce equal killing will produce more delay in larger tumours.

Animals↗
PubMed1980

The effects of misonidazole during continuous low dose rate irradiation.

Hypoxic cell sensitizers have been shown to enhance the radiation effects on tumors for single and multifractionated doses of radiation given in acute exposures. Low dose rate brachytherapy provides an opportunity for short-term high dose drug administration and continuous drug exposure throughout the entire brachytherapy course. To determine the effects of misonidazole during continuous low dose rate irradiation we have exposed the EMT6/SF tumor to continuous irradiation at 4.2 and 1.64 rads/minute with and without misondiazole (at doses of 1 mg/g or 0.2 mg/g), in bilaterally nephrectomized mice, and compared the results to those obtained wih acute exposures. Radiosensitization by misonidazole at a clinically achievable plasma concentration was observed during continuous low dose rate irradiation. The DMF (dose-modifying factor) varied with drug dose and dose rate. At a dose of 1 mg/g the DMF was 2.4 at 221 rads/minute and 1.6 at 4.2 rads/minute; at 0.2 mg/g it was 1.5 at 221 rads/minute and 4.2 rads/minute and 1.4 at 1.64 rads/minute. Cytotoxicity was also observed with drug exposure greater than 6 hours. Our results suggest that it would be advantageous to sue misonidazole during low dose rate brachytherapy.

Animals↗