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H Johns

Publications and source records attributed to H Johns.

15 recordsLinked to original sources
PubMed1995

Radiosensitization of mouse skin by oxygen and depletion of glutathione.

PURPOSE: To determine the oxygen enhancement ratio (OER) and shape of the oxygen sensitization curve of mouse foot skin, the extent to which glutathione (GSH) depletion radiosensitized skin, and the dependence of such sensitization on the ambient oxygen tension. METHODS AND MATERIALS: The feet of WHT mice were irradiated with single doses of 240 kVp x-rays while mice were exposed to carbogen or gases with oxygen/nitrogen mixtures containing 8-100% O2. The anoxic response was obtained by occluding the blood supply to the leg of anesthetized mice with a tourniquet, surrounding the foot with nitrogen, and allowing the mice to breathe 10% O2. Further experiments were performed to assess the efficacy of this method to obtain an anoxic response. Radiosensitivity of skin was assessed using the acute skin-reaction assay. Glutathione levels were modified using two schedules of DL-buthionine sulphoximine (BSO) and diethylmaleate (DEM), which were considered to produce extensive and intermediate levels of GSH depletion in the skin of the foot during irradiation. RESULTS: Carbogen caused the greatest radiosensitization of skin, with a reproducible enhancement of 2.2 relative to the anoxic response. The OER of 2.2 is lower than other reports for mouse skin. This may indicate that the extremes of oxygenation were not produced, although there was no direct evidence for this. When skin radiosensitivity was plotted against the logarithm of the oxygen tension in the ambient gas, a sigmoid curve with a K value of 17-21% O2 in the ambient gas was obtained. Depletion of GSH caused minimal radiosensitization when skin was irradiated under anoxic or well-oxygenated conditions. Radiosensitization by GSH depletion was maximal at intermediate oxygen tensions of 10-21% O2 in the ambient gas. Increasing the extent of GSH depletion led to increasing radiosensitization, with sensitization enhancement ratios of 1.2 and 1.1, respectively, for extensive and intermediate levels of GSH depletion. In mice exposed to 100% O2, a significant component of skin radiosensitivity was due to diffusion of oxygen directly through the skin. Pentobarbitone anesthesia radiosensitized skin in mice exposed to 100% O2 by a factor of 1.2, but did not further sensitize skin in mice exposed to carbogen. CONCLUSIONS: Glutathione levels and the local oxygen tension at the time of irradiation were important determinants of mouse foot skin radiosensitivity. The extent to which GSH levels altered the radiosensitivity of skin was critically dependent on the local oxygen tension. These results have significant implications for potential clinical application of GSH depletion.

Animals↗
PubMed1995

Time evolution of the number of functional murine eccrine sweat glands after irradiation: a quantitative analysis of experimental data using a model of proliferative and functional organization.

The function of eccrine sweat glands in the beta-pad of the mouse foot after irradiation was followed over time. Graded doses of X-rays were given to the foot, either as single doses or in two equal fractions separated by a 24-h interval. A quantitative, non-invasive, functional assay was used allowing repeated evaluation of the animals. Sweat gland function was assessed once a week for the first 6 weeks, and at 8, 10, 14, 18, 28, 38 and 45 weeks after irradiation. The beta-pad of the unirradiated foot was used as a control. The function dropped to a nadir within 8-10 weeks after irradiation, whereafter it gradually recovered, reaching a stable level 20-25 weeks after irradiation. These data were analyzed using a mathematical model of proliferative and functional organization of the sweat pore. The model provides a description of the time evolution of pore function, and its basic features were as follows. The functional subunit is a single sweat pore, which will be assayed as functional provided that it contains a number of functional cells above a critical threshold. The functional cells are capable of self renewal (a so-called 'flexible' tissue), and the proportion of proliferating cells is subject to homeostatic control. Irradiation is assumed to transfer a certain fraction of the cells into a state with a limited probability of successful division. This fraction is assumed to have a linear-quadratic dependence on dose. The values of all free model parameters were optimized by a maximum-likelihood fit to the experimental data. With optimized parameter values, the initial decrease, nadir, and long-term level of tissue function estimated from the model were in close agreement with the experimental observations for all the 28 dose groups. Some of the estimated model parameters are: growth fraction 4.2 +/- 0.2%; cell cycle time, 0.95 +/- 0.04 days; number of functional cells in a single pore in the unirradiated animal, 9.9 +/- 0.5; and alpha/beta ratio, 4.3 Gy (95% confidence interval 3.1-5.0 Gy). It is concluded that the present model, despite its relative mathematical simplicity, provides a close description of the postirradiation kinetics of functional cells in the mouse sweat gland.

Animals↗
PubMed1994

The effect of BW12C on the radiosensitivity and necrosis of murine tissues and tumours.

BW12C is a drug that has the potential to induce normal tissue and tumour hypoxia by binding to haemoglobin, increasing its affinity for oxygen and thereby reducing oxygen availability to tissues. Initial results suggested that BW12C administration caused significant radioprotection of normal tissues and induced tumour necrosis, but variable results have been reported subsequently. This work was carried to extend the range of observations concerning the ability of BW12C to radioprotect normal tissues and tumours and to induce necrosis of tumours of the mouse. BW12C was administered as 70 mg/kg i.v. 15 min before irradiation of jejunum in CBA mice and of foot skin in WHT mice with single doses of 240 kVp X-rays while mice breathed gases of varying oxygen tensions. The radiosensitivities of these tissues were assessed by the crypt survival assay and the acute skin reaction, respectively. The radiosensitivity of CaNT tumours to single fraction irradiation was assessed by the regrowth delay assay following administration of single or multiple doses of BW12C at varying times to air-breathing CBA mice. The radiation response was compared to the radiosensitivity of clamped tumours. The effect of BW12C alone on tumours was assessed by regrowth delay and histological examination for necrosis. BW12C did not change the radiosensitivity of jejunal crypts irradiated while mice breathed air or 10% O2, or of foot skin when mice breathed 12% O2. BW12C protected foot skin by a factory of 1.1 when mice breathed air. Single or multiple doses of BW12C did not influence the radiosensitivity of CaNT tumours, although marked radioprotection could be induced by clamping the tumours during irradiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

A test of equal effect per fraction in the kidney of the mouse.

Measurements of renal damage in the mouse were made to determine if there was an equal effect per fraction during a course of repeated 240-kVp X-ray doses. An X-ray dose of 2 Gy was given 2, 8, 14, or 20 times with interfraction intervals of 12 h. Some animals were also irradiated with twenty 2-Gy doses using a 5-h interfraction interval. The underlying effect per fraction (-logeSF of the notional target cell population) was determined from the additional top-up dose of d(4)-Be neutrons needed to produce measurable renal impairment assessed by decreased clearance from the plasma of [51Cr]EDTA and by a reduction in the hematocrit at 25, 29, 33, and 39 weeks after treatment. There was no significant influence of the time of assay on the values of underlying effect measured. A mean value of underlying effect was therefore calculated for the two different assays of each mouse, from the measurements at the four times. This gave approximately 40 estimates (one for each animal assessed) with each assay of the effectiveness of 2-Gy fractions in each of the four fractionation schedules, a total of 321 determinations in the study with 12-h intervals. Regression analysis showed that there was no significant trend in underlying effect per fraction with number of fractions, i.e., the damage per fraction was constant regardless of the number of fractions used. With underlying effect normalized to 1 unit of damage for a single 2-Gy dose, the slope of this plot was -0.0013 per fraction2 +/- 0.0097 (95% CL). The assumption of equal effect per fraction was therefore not invalidated in the kidney of the mouse. With a 5- instead of a 12-h interfraction interval, the 20-fraction schedule was 7% more effective as measured by the two assays analyzed together; this was significant at P = 0.0001. This shows that 5 h is not sufficient time between fractions for full repair to occur in the kidney, and underlines the need for intervals of at least 6 h between the doses in clinical radiotherapy using more than one fraction per day. The data are consistent with an alpha/beta ratio approximately 1.6 Gy, with a repair half-time approximately 1.3 h. However, these experiments were not designed to determine these parameters and their values should be regarded only as rough estimates.

Animals↗
PubMed1991

A therapeutic benefit from combining normobaric carbogen or oxygen with nicotinamide in fractionated X-ray treatments.

The ability of normobaric oxygen and carbogen (95% O2 + 5% CO2) combined with nicotinamide to enhance the radiosensitivity of two rodent adenocarcinomas and of mouse skin and kidneys, using a 10 fraction radiation schedule, was compared with the effect of radiation in air with and without the drug. Tumour response was assayed using local control and regrowth delay, and compared with acute skin reactions, decreased renal 51Cr-EDTA clearance and reduction in haematocrit. Nicotinamide increased the radiation sensitivity of CaNT tumours under all three different oxygen concentrations tested (21, 95 and 100% oxygen). The effect was statistically significant for oxygen and carbogen but not for air; the combination of nicotinamide with carbogen gave the greatest increase in tumour radiosensitivity. Relative to treatments in air without the drug, the enhancement ratios (ER) at the TCD50 level were 1.17, 1.65 and 1.83 for CaNT tumours irradiated in air, oxygen or carbogen and injected with nicotinamide 1 h before each fraction. The ER in CaRH tumours irradiated in carbogen plus the drug was 1.83, which was greater, but statistically not significantly different, to that seen with carbogen alone (ER = 1.68). In skin, relative to air without the drug, the increase in radiosensitivity by nicotinamide was greater in oxygen and carbogen than in air (1.29, 1.36 and 1.08, respectively). The ERs for both assays of renal damage were similar and lower than those in skin: less than or equal to 1.07, less than or equal to 1.13 and less than or equal to 1.16 for irradiations done in air, oxygen and carbogen plus nicotinamide, relative to air alone. A comparison of these results in the tumours and normal tissues showed that a significant therapeutic benefit was obtained with normobaric oxygen and carbogen combined with nicotinamide. This benefit is greater than observed with other radiosensitizers tested so far. Toxic side effects of the treatment are unlikely in a clinical situation, since prolonged administration of nicotinamide is well tolerated in man. The combination of normobaric carbogen with nicotinamide could be an effective method of enhancing tumour radiosensitivity in clinical radiotherapy where hypoxia limits the outcome of treatment.

Adenocarcinoma↗
PubMed1991

Early detection of damage following bilateral renal irradiation in the mouse.

The rate and early pattern of development of radiation-induced renal damage has been determined in the mouse by measuring reductions in both haematocrit and excretion of 51Cr-EDTA, and increases in both urination frequency and urine volume. Kidneys of CBA mice were irradiated bilaterally with 2 fractions of X-rays, one week apart. Renal function was determined immediately prior to irradiation and at 3-4 weekly intervals to 22 weeks post-irradiation. Onset of damage was detected as early as 3-6 weeks using the urination frequency assay. This was confirmed by estimating the volume of urine excreted. A significant fall in haematocrit was not detected until 6-9 weeks post-treatment and a fall in isotope clearance was not detected significantly until 12 weeks. This early detection of damage was consistent with reports using both mouse and other species. The time at which damage was detected first was independent of radiation dose for the frequency and haematocrit assays. For 51Cr-EDTA clearance, there was the suggestion of earlier functional loss for the higher doses. Following the onset of damage, a steady, dose-dependent decline in renal function was measured by all assays. The latency period is defined as the time required to reach a given level of functional damage. This time decreased with increasing radiation dose, to a minimum value set by the time of onset of damage, which varied from 3 to 12 weeks, depending on the assay used. The differences in response measured prior to 12 weeks post-irradiation represent the first occasion on which a dissociation between these 3 assays has been detected.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Recovery kinetics of X-ray damage in mouse skin: the influence of dose per fraction.

The rate of recovery from radiation damage, as a function of dose per fraction, was investigated in mouse skin. Two different experimental designs were used, both incorporating the neutron top-up technique which enables the X-ray dose per fraction to be kept constant whilst changing the interfraction interval. Either equally spaced X-ray fractions (concertina design) or single or multiple pairs of X-ray doses (single and multiple split-dose designs) were given at varying intervals, followed by graded doses of neutrons. A wide range of X-ray doses per fraction were investigated (from 1 to 10.5 Gy) and the data were analysed using the Thames Incomplete Repair (IR) model modified for use with neutron top-up doses. Analyses of the data, obtained from five different experiments, indicate that the rate of recovery from radiation damage is significantly faster at doses per fraction between 1 and 4.4 Gy than at 10.5 Gy. These data appear not to support the assumption, made by most recovery models, that the rate of recovery is independent of dose.

Animals↗
PubMed1991

A simple method for fitting curves to dose-effect data for functional damage.

Dose-effect curves are used extensively to assess how tissues respond to radiation. One method of obtaining these is to fit a curve to the values of some measured effect plotted against dose using non-linear least-squares regression. This paper reports the use of a generalized (four-parameter) sigmoid equation fitted to all the individual data points, rather than to the mean values for each dose group, which eliminates the need to incorporate weighting of the data. The equation allows an analytical solution for values of isoeffect doses, which can be used, for example, to determine dose enhancement ratios, or equivalent remembered doses in top-up experiments. The regression approach can also determine both standard errors and 95% confidence limits on the mean predicted effect values from the fit to the data at all doses, and these define a uniform envelope of errors about the best-fit line, from which an error in an isoeffect dose can be assessed. This approach has been used to fit dose-effect data from a variety of normal tissues and tumours with highly satisfactory results.

Animals↗
PubMed1991

Repopulation in murine skin after X-ray treatments with multiple fractions per day.

The kinetics of repopulation of clonogens in skin after fractionated X-ray exposures was studied in a series of experiments using a top-up design. The feet of mice were exposed to small X-ray doses (1.5 or 2 Gy), given two or three times a day on consecutive days with a minimum interfraction interval of 8 h. A single top-up dose of d(4)-Be neutrons was then given at various intervals after the last X-ray fraction, typically on Days 1,4,8, 15, and 19. The acute skin reaction produced was scored an analyzed by both a standard 23-day averaging and a 7-day averaging procedure. Either method gave similar results and led to the same conclusions. The amount of top-up dose needed to produce a fixed skin reaction was used as a measure of the net effect of the X-ray treatments. This net effect is a result of the initial reduction in skin clonogens due to X rays, and their repopulation before the top-up dose was given. Repopulation was not detected during any of these courses of fractionated treatment, up to an overall time of at least 12 and possibly 16 days. On completion of X-ray schedules lasting 6-16 days, repopulation started 4 days later. In contrast, this delay lengthened to approximately 8 days for shorter overall treatment times of 3-4 days. Once repopulation started, it proceeded rapidly over 11 days so that by 15 days after the cessation of X rays, the skin was restored almost to its normal state with respect to radiosensitivity. The residual damage from Day 15 to Day 19 postirradiation was 3-13% of a full-effect level. The rate of repopulation can be expressed as a clonogen doubling time (Tclon), assuming that an average skin reaction of 1.5 is equivalent to a clonogen surviving fraction of 1.7 x 10(-5). Tclon varied inversely with the amount of initial damage inflicted by the X rays, with the shortest values (1-1.3 days) seen following X-ray doses that gave an initial damage level of 60-80% of full effect. These data are consistent with a hypothesis that damage is "sensed" only 10-12 days after the first X-ray fraction, which provides the stimulus for repopulation of the target cells in the basal layer, the keratinoblasts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Recovery kinetics in mouse skin and CaNT tumours.

Recovery kinetics and recovery capacity were studied in a fast proliferating normal tissue, skin, and in an anaplastic mouse mammary carcinoma, CaNT. Three fractions per day of X-rays, repeated over 5 days, were given at varying interfraction intervals from 0 to 8 h. The rate of recovery in tumours (t1/2 = 0.31 +/- 0.15 h for local control) was significantly faster than in skin (t1/2 = 0.96 +/- 0.10 h). By contrast, the fractionation sensitivity of CaNT tumours was less than that of skin (alpha/beta = 43.3 +/- 8.5 Gy vs. alpha/beta = 7.9 +/- 0.2 Gy). Tissues with recovery half-times similar to or longer than that determined for skin would be at risk if interfraction intervals less than 6 h are used in regimes which involve the use of two or more fractions per day. This would be especially true for tissues that show a greater sensitivity to dose fractionation, and hence more sparing of radiation damage with hyperfractionation.

Adenocarcinoma↗
PubMed1989

Role of glutathione peroxidase in the radiation response of mouse kidney.

Glutathione peroxidase (GSH-Px) has been implicated in mediating the radioprotective effects of glutathione (GSH). This hypothesis was tested in vivo by determining the effect of GSH-Px depletion on the radiation response of murine kidneys. Renal GSH-Px levels were depleted to 17% of control values by feeding animals a selenium deficient diet for 6 weeks; this had no significant effect on renal levels of GSH or GSH-S-transferase (GST). However, we also tested the effect of direct depletion of GSH to 3-4% of control values, using a combination of DL-buthionine sulphoximine (BSO) and diethyl maleate (DEM). Kidneys with normal or depleted levels of GSH-Px and/or GSH were irradiated with 240kVp X rays (2 fractions, 7 days apart to minimize intestinal injury). Mice breathed 7% oxygen during irradiation. Renal damage was assessed at 20, 25, and 32 weeks after the first fraction of X rays, in terms of reduced hematocrit and renal clearance of 51Cr-EDTA. Depletion of GSH-Px levels to 17% of control did not alter renal radiosensitivity, but depletion of GSH to 3-4% of control values radiosensitized the kidney by a factor of 1.4. Depletion of both GSH and GSH-Px together did not radiosensitize the kidney any more than was achieved by GSH depletion alone.

Animals↗
PubMed1989

Does the repair capacity of skin change with repeated exposure to X-rays?

A sensitive experimental design and data analysis were used to test rigorously whether the repair capacity in the skin of the mouse foot changes during a course of repeated 240 kVp X-ray doses. Any such changes might reflect saturation or induction of repair enzymes resulting from progressive radiation damage, but most importantly this assumption of equal effect per dose fraction is central to all analyses of multiple-fraction radiation treatments, and remained to be demonstrated conclusively in skin. An X-ray dose of 2.5 Gy was given two, eight, 14 or 20 times with an interfraction interval of 8 h. Individual skin reactions for each mouse were analysed separately, giving 139 estimates of the effectiveness of 2.5 Gy (approximately 35 in each of the four fractionation schedules). Regression analysis of effect per fraction versus number of fractions showed that there was no significant trend, i.e. the damage per fraction was constant regardless of the number of fractions used. The mean damage per fraction was 3.75 +/- 0.15 per cent (95 per cent CL) of the full underlying damage equivalent to transient moist desquamation, and the slope of this plot was 0.0075 per cent +/- 0.022 per cent (95 per cent CL) per fraction. It was concluded that the assumption of equal effect per fraction was not invalidated in mouse skin. Shorter interfraction intervals would not allow full repair between fractions, and this could be misinterpreted as a progressive loss of repair capacity in this type of experiment. This was tested in skin by giving 2.5 Gy doses two, eight and 14 times with a 1-h interfraction interval. Effect per fraction increased with number of fractions, by an extra 37 per cent from two to eight fractions and by a further 14 per cent from eight to 14 fractions, giving the illusion of loss of repair as predicted. This confirms the need to check that where loss of repair capacity is suspected, this is not due artifactually to incomplete repair between fractions in slowly repairing systems.

Animals↗
PubMed1988

Renal damage in the mouse: the response to very small doses per fraction.

Experiments were undertaken to study the effect on the mouse kidney of repeated X-ray doses in the range 0.2 to 1.6 Gy per fraction and neutron doses in the range 0.05 to 0.25 Gy per fraction. A top-up design of experiment was used, so that additional graded doses of d(4)-Be neutrons (EN = 2.3 MeV) were given to bring the subthreshold damage produced by these treatments into the measurable range. This approach avoided the necessity to use a large number of fractions to study low doses per fraction. Renal damage was assessed using three methods: 51Cr-EDTA clearance, urine output, and hematocrit at 16-50 weeks postirradiation. The dose-response curves obtained were resolved best at 29 weeks. However, the results were also examined by fitting second-order polynomials to the data for response versus time postirradiation and using interpolated values from these functions at 29 weeks to construct dose-response curves. This method reduced slightly the variation in the dose-response data, but the interrelationship between the dose-response curves remained the same. The data were used to test the linear-quadratic (LQ) description of the underlying X-ray dose-fractionation relationship. The model fits well down to X-ray doses per fraction of approximately 1 Gy, but lower X-ray doses were more effective per gray than predicted by LQ, as seen previously in skin [M. C. Joiner et al., Int. J. Radiat. Biol. 49, 565-580 (1986)]. This increased X-ray effectiveness and deviation from LQ are reflected directly in a decrease in the RBE of d(4)-Be neutrons relative to X-rays at low doses, since the underlying response to these neutrons is linear in this low-dose region. The RBE decreases from 9.9 to 4.7 as the X-ray dose per fraction is reduced below 0.8 Gy to 0.2 Gy, reflecting an increase in X-ray effectiveness by a factor of 2.1. A model is discussed which attempts to explain this behavior at low doses per fraction.

Animals↗
PubMed1987

Renal damage in the mouse: the effect of d(4)-Be neutrons.

A further study on the response of the mouse kidney to d(4)-Be neutrons (EN = 2.3 MeV) is described. The results confirm and augment the work published previously by Stewart et al. [Br. J. Radiol. 57, 1009-1021 (1984)]; the present paper includes the data from a "top-up" design of experiment which extends the measurements of neutron RBE (relative to 240 kVp X rays) down to X-ray doses of 0.75 Gy per fraction. The mean RBE for these neutrons increases from 5.8 to 7.3 as X-ray dose per fraction decreases from 3.0 to 1.5 Gy in the kidney. This agrees with the predictions from the linear quadratic (LQ) model, based on the renal response to X-ray doses above 4 Gy per fraction. The mean RBE estimate from a single dose group at 0.75 Gy per fraction of X rays is, however, 3.9. This is below the LQ prediction and may indicate increasing X-ray sensitivity at low doses. Data from this study and from those published previously have been used to determine more accurately the shape of the underlying response to d(4)-Be neutrons; an alpha/beta ratio of 20.5 +/- 3.7 Gy was found. The best value of alpha/beta for X rays determined from these experiments was 3.04 +/- 0.35 Gy, in agreement with previous values.

Animals↗