Chromosome aberration yields in human lymphocytes induced by fractionated doses of x-radiation.

Unstimulated (G0) human peripheral blood lymphocytes were exposed at 37 degrees C to doses of 200 or 500 rad of X-rays delivered in two equal fractions. The dose fractions were separated by intervals of up to 7 h in the 200 rad study and up to 48 h for 500 rad. In both studies the mean levels of dicentrics and total unstable aberrations began to decline when fractions were delivered with intervals of greater than 2 h. With 200 rad the yield had decreased to an additive baseline (i.e. equal to only twice the yield of a single 100-rad fraction) by an interval of 4 h. Following 500 rad the yield declined until 8 h and then remained 20% above the additive baseline even when 48 h separated the fractions. Possible explanations for this discrepancy are discussed. In a second experiment PHA stimulated lymphocyte cultures were exposed to 2 doses of 125 rad of X-rays up to 7 h apart in an attempt to demonstrate the late peak in aberration yields originally reported by Lane [5]. Control cultures received unsplit doses of 250 rad at the time of the corresponding second 125-rad fraction. No evidence of a late peak in dicentric yield was observed. The yield remained approximately the same irrespective of the time interval between fractions but these split dose yields were significantly different from the accompanying unsplit controls.     

Radiation-induced chromosome aberrations in lymphocytes from man and crab-eating monkey: The dose-response relationships at low doses

To obtain information on the relation beteween yiels of chrmosome aberrations and dose at low-dose levels, experiments were conducted with 5, 10, 20, 30 and 50 rad of ¹³⁷Cs γ-rays, on lymphocytes from man and crab-eating monkey (Macaca fascicularis). The dose-response relationship for dicentrics was obtained from the combined data of these low-dose experiments with those of our previous onse at high doses (100–400 rad) When the difference between observed yields and those expected from the linear-quadratic model were computed, the dose-response curve had a good fit for man, but not for the monkey. The linear regression lines between 0 and 30 rad were calculated, because the expected values of α/β for man and monkey would be about 100 and 60 rad.The human date gave a satisfactory fit to a linear model, i.e., a linear increase in aberration frequency with dose, whereas this was not so for those of the monkey. Furtyhermose, there was some suggestive evidence for the existence of plateau in dicentrics yields between 10 and 30 rad for the monkey and between 20 and 30 rad for human lymphocytes, but more data would be needed to verify this suggestion, particularly for human lymphocytes.     

Dicentric yields induced by gamma-radiation and chromosome arm number in primates.

To evaluate the effect of the chromosome arm number on the yield of dicentric chromosomes, frequencies of gamma-ray-induced chromosome aberrations were examined with peripheral lymphocytes from three different primate species, Saimiri sciureus (arm number, 77), Macaca fascicularis (arm number, 83) and Nycticebus coucang (arm number, 99). Irradiated blood samples were cultured by the same standard technique as that commonly used for human lymphocytes. The yields of dicentrics and dicentrics plus rings at doses of 100, 200 and 300 rad of gamma-irradiation were not significantly different among the three species, in spite of the difference in the chromosome arm number. Furthermore, dose-response relationships for these species were consistent with that for man. Statistical analysis indicated that the expected dicentric yields calculated from the arm number model were significantly different from the observed yields at 200 and 300 rad doses (P less than 0.01). From these results it can be pointed out that there is no correlation between the yield of dicentrics and the effective chromosome arm number, and that the chromosomal radiosensitivity of these primates is essentially the same as that of man, at least in the lymphocyte system.     

A comparative study of the frequencies of radiation-induced chromosome aberrations in somatic and germ cells of the rhesus monkey (Macaca mulatta)

Frequencies of radiation-induced chromosome aberrations in spermatogonia, peripheral blood lymphocytes and bone-marrow cells of the rhesus monkey (Macaca mulatta) and in human blood lymphocytes, were determined at different exposures of X-rays. The dose-response curve for the induction of reciprocal translocations in spermatogonia suggested a “hump” at about 200 rad. The absolute frequencies of chromosome aberrations in somatic and germ cells of the rhesus monkey were low in comparison with most other mammalian species and the ratio between aberrations in the two tissues was 25 to 1 at the 100 rad level. Although the numbers of “effective chromosome arms” in man and rhesus monkey are similar (81 vs. 83), the rhesus monkey showed a lower rate of induction of dicentrics in blood lymphocytes than man at all doses, reaching statistical significance at the 300 rad level.     

Life shortening in BALB/c mice following brief, protracted, or fractionated exposures to neutrons

The effects of acute, protracted, or fractionated exposures to fission neutrons on survival times of female BALB/c mice were examined and compared. Mice were given single, brief exposures or exposures given in equal fractions at either 1- or 30-day intervals to doses of 0, 2.5, 5, 10, 20, 50, and 200 rad at the Health Physics Research Reactor (HPRR) or protracted exposures at rates ranging from 0.1 to 10 rad/day using a moderated 252Cf source to doses of 0, 2.5, 5, 10, 20, and 40 rad. The 252Cf source was moderated to have a similar spectron to that of the HPRR facility. After single or fractionated exposures the extent of life shortening increased rapidly over the 0-50 rad range and then began the plateau. No simple model adequately described the dose response over this entire dose range. Over the 0-50 rad dose range for exposures at the HPRR and over the 0-40 rad dose range for protracted exposures the dose response could be adequately described by either a linear model or a square root of the dose regression model except when the dose was fractionated using a 30-day interval. In this instance a linear model provided an adequate fit while a square root of the dose model could be rejected. No increase in effectiveness after fractionation or protraction was observed for neutron-induced life shortening at doses below 50 rad, while at 50 and 200 rad an increase in effectiveness was observed in this and in previous studies. These data were interpreted to suggest that in the dose range below 20-40 rad the dose-effect curve for life shortening may be linear and begins to flatten at higher doses rather than continuously bending at low doses.     

Aberrations induced by fission neutrons in human peripheral lymphocytes

Analysis of dose-response relationship was carried out for chromosome aberrations produced in human peripheral lymphocytes by fission neutrons at doses of 25, 50, 100 or 200 rad.Statistical treatment showed experimental data to be fitted by a regression curve described by the mathematical model Y = a+bD. A linear relation to dose characterized both one-break and two-break aberration yields. Numerical values of coefficients are reported for yields of dicentrics, chromosome fragments, minutes, aberrant cells, total number of aberrations, and total breakage.Based on chromosome fragments and aberrant cells, relative biological efficiency (RBE) value derived for fission neutrons relative to 180 kV X-rays for chromosome fragments was 2.53, and for aberrant cells it was 2.80.     

The effect of repeated small doses of radiation on recovery from sub-lethal damage by Chinese hamster cells irradiated in the plateau phase of growth.

Unfed plateau-phase cells have been irradiated with either single doses or up to ten fractions of X-rays 6 hours apart. The single-dose survival curve had an extrapolation number of 11-4, and the oxygen-enhancement ratio (o.e.r.) was 3-1. Cells were exposed to multiple fractions of 200 rad or 150 rad in air and 600 rad or 450 rad in hypoxia. The resulting survival curves did not fit a multi-target, single-hit model of cell survival, being much steeper than that would predict. The curves were exponential up to five fractions of X-rays, but tended to bend downwards with increasing number of fractions. Cells that had survived five fractions of 200 rad (or 600 rad in hypoxia) 6 hours apart, were less able to absorb damage as sub-lethal than those which had not previously been exposed to radiation. The ratio of the initial slopes of the fractionated survival curves for irradiation in air and hypoxia was 2-1, implying that the o.e.r. "on the shoulder" may be less than that in the exponential region of survival.     

Dose—response relationship for X-ray-induced reciprocal translocations in stem-cell spermatogonia of the rhesus monkey (Macaca mulatta)

The induction of reciprocal translocation in stem-cell spermatogonia of the rhesus monkey (Macaca mulatta) was studied after testicular X-irradiation of mature males (50, 100 and 200 rad) or whole-body irradiation of young males (200 and 300 rad). After the recovery of the germinal epithelium, cytogenetic analysis was carried out on spermatocytes descended from irradiation spermatogonia. Preparations of C-banded diakinesis-metaphase I were screened for translocation configurations. The frequencies of abberations obtained were 0% at 0 rad, 0.36% at 50% rad, 0.86% at 100 rad, 0.99% at 200 rad and 0.68% at 300 rad, suggesting a humped dose—response relationship. There was no evidence for the contribution of a quadratic component to the yield in the lower dose range. A comparison of these results with those obtained for other mammals by a number on investigators shows that the frequencies of translocations in the rhesus monkey are much lower than those published for most other mammalian species.     

Effects of dose rate and dose fractionation of irradiation on pulmonary alveolar macrophage colony-forming cells

We have studied the effects of dose rate and dose fractionation on murine pulmonary alveolar macrophage colony-forming cells (AL-CFC). The dose-response curve of AL-CFC to ionizing irradiation has a Dq of about 100 rad, reflecting the cells' ability to repair sublethal damage. For comparison, we investigated the effect of dose schedule on the committed bone marrow stem cells for both granulocytes and monocytes (GM-CFC) since their dose-response curve has a very small shoulder. We compared the results of dose rates of 3 and 10 rad/min to those obtained with a dose rate of 85 rad/min. We determined survival after giving 100, 300, and 500 rad either in vivo or in vitro. A significant dose rate effect was observed. To study the effect of dose fractionation, a total of 600 rad was given either as a single fraction, three fractions of 200 rad on 3 consecutive days, or six fractions of 100 rad in 3 days. The most dramatic effect was seen in the group that received six 100-rad fractions. No reduction in the number of AL-CFC was seen in this group. In sharp contrast, only a minimal dose schedule effect was observed with GM-CFC.     

Dose-response curves for radiation-induced gene mutations in mouse oocytes and their interpretation

Previous work, in which female mice had been given fractionated doses of 20 X 10 rad X-rays, had confirmed and extended Russell's observations that the dose-response relationship for specific-locus mutations in mature-mouse oocytes is curved at low doses. The present work was intended to study the relationship at relatively high doses. Adult female mice were given doses of 200, 400 or 600 rad x-rays at 52 or 72 rad/min, and mated immediately. Offspring conceived in the first 7 days (i.e. using oocytes which were mature at time of treatment) were scored for specific-locus mutations. The data indicate that the departure from linearity of the dose-response curve is marginally significant at the 5% level. A quadratic dose-response curve (y = c + aD + bD2) and a square-law relationship (y = c + bD2) both give a good fit to the data. Both curves fit data of other authors obtained at low doses or dose-rates. These results could be interpreted either in terms of dose-dependent repair phenomena, or by considering specific-locus mutations as two-track events. In view of knowledge of other phenomena concerning mutation and cell killing in mouse oocytes, such as the variation in sensitivity of different cell stages, the interpretation in terms of repair phenomena is preferred.     

Radiosensitivity of vascular tissue. II. Differential radiosensitivity of aortic cells in vitro

The cellular outgrowths from three layers of rabbit and monkey aorta were used as primary cultures. Irradiation of the tissue fragments at the time of explanation resulted in a reduction in outgrowth of 50% with a dose of 200 rad, and in a reduction of over 90% with doses of 300 rad and above. When comparable cultures were irradiated after 2 months in vitro as a mature actively metabolizing but slowly proliferating cell population, radioresistance was increased. Subcultures of medial smooth muscle cells irradiated during their logarithmic growth phase showed a linear dose response in the cell number parameter up to 150 rad. A dose of 250 rad resulted in complete flattening of the growth curve, with a reduction in labeling index, after a 3-hr terminal [3H]TdR pulse. On the other hand, the labeling index indicated some recovery 3 days after irradiation in cultures receiving less than 250 rad. Under the same experimental conditions, cells derived from the intima of the same aorta showed no recovery when increase in cell numbers over time, or the number of labeled cells per area, were used as parameters. Cells derived from adventitia showed a relative increase in the number of labeled cells per area 4 and 7 days after irradiation following an initial decrease on Day 1.     

Muscle activation and the isokinetic torque-velocity relationship of the human triceps surae

The influence of muscle activation and the time allowed for torque generation on the angle-specific torque-velocity relationship of the triceps surae was studied during plantar flexion using supramaximal electrical stimulation and a release technique on six male subjects [mean (SD) age 25 (4) years]. Torque-velocity data were obtained under different levels of constant muscle activation by varying the stimulus frequency and the time allowed for isometric torque generation prior to release and isokinetic shortening. To eliminate the effects of the frequency response on absolute torque the isokinetic data were normalized to the maximum isometric torque values at 0.44 rad. There were no significant differences in the normalized torques generated at any angular velocity using stimulus frequencies of 20, 50 or 80 Hz. When the muscle was stimulated at 50 Hz the torques obtained after a 400 ms and 1 s pre-release isometric contraction did not differ significantly. However, with no pre-release contraction significantly less torque was generated at all angular velocities beyond 1.05 rad · s^–1 when compared with either the 200, 400 ms or 1 s condition. With a 200 ms pre-release contraction significantly less torque was generated at angular velocities beyond 1.05 rad · s^–1 when compared with the 400 ms or 1 s conditions. It would seem that the major factor governing the shape of the torque-velocity curve at a constant level of muscle activation is the time allowed for torque generation.     

Product formation and kinetic simulations in the pH range 1-14 account for a free-radical mechanism of peroxynitrite decomposition

The yields of nitrate and nitrite from decomposition of peroxynitrite in phosphate buffer at 37 degrees C were determined in the pH range 1-14. The NO(2)(-)/NO(3)(-) yields showed a stepwise variation with pH, with inflection points at approximately pH 3.1, 5.8, 6.8, 8.0, and 11.9. Nitrite formation increased strongly above pH 7 at the expense of nitrate, but above pH 12 nitrate again became the major product (80% at pH 14). At this pH, the Arrhenius parameters were E(a)=24.1+/-0.2kcal mol(-1) and A=(4.9+/-1.3)x10(12)s(-1). The yields of NO(2)(-), NO(3)(-), and O(2) measured at pH 5.8, 7.4, and 8.5 as a function of the initial peroxynitrite concentration (50-1000 microM) were linear only at pH 5.8. In the presence of carbon dioxide, oxygen production at pH 7.5 and pH 10 was found to be linear on the CO(2) concentration. The experimental observations were satisfactorily reproduced by kinetic simulations including principal component analyses. These data strongly suggest that the chemistry of peroxynitrite is exclusively mediated by z.rad;NO(2) and HO(z.rad;) radicals in the absence, and by z.rad;NO(2) and CO(3)(z.rad;-) radicals in the presence of CO(2).     

X-ray-induced translocations in marmoset (Callithrix jacchus) stem-cell spermatogonia

The induction of reciprocal translocations in spermatogonial stem cells of marmosets (Callithrix jacchus) was studied after irradiation with different doses of X-rays (50, 100 and 200 rad) via spermatocyte analysis many cell generations later. The obtained results show a dose-effect relationship with clear saturation effects at 200 rad. The recorded frequencies of translocations were much lower than those reported for closely related marmosets (Saguinus fuscicollis and Saguinus oepidus). Possible reasons for this difference are discussed.     

Is the mean inactivation dose a good measure of cell radiosensitivity?

The relationship between the mean inactivation dose, D, and cell radiosensitivity, particularly in the low-dose region, was investigated for linear-quadratic (LQ) and two-component (TC) cell survival curves. Although D is approximately equal to D40 for most survival curves, the results showed that there is no consistent relationship, in theory, between D and the extent of cell killing in the low-dose region. Curves with equal D values could potentially represent cell populations with markedly different levels of survival after 200 rad, and conversely, curves with equal surviving fractions at 200 rad could have D values that differ by a factor of two or more. The variability in replicate estimates of D was also investigated and compared with that of SF200, the surviving fraction at 200 rad. The results of a simulation experiment suggest that estimates of D are somewhat more variable than those of SF200 and are more dependent on the choice of cell survival model and the range of available data used for the estimation. It is concluded that SF200 is a more reliable and numerically stable parameter of cell radiosensitivity than D.     

Radiation effect on the proliferating capillaries in rat transparent chambers.

The revascularization of the devascularized area in rat transparent chambers by the proliferating blood vessels was studied quantitatively on the changes of vascularity. On the vascularizing border, a hypervascular zone about 0-5 mm wide was formed. The border advanced constantly at 0-020 cm a day. Vascular density was fixed in the repaired area. The vascular length and repaired area increased on parabolic curves. The chambers were irradiated with single doses of 50, 100, 200, 400, 1000, 2000, and 4000 rad of 30Co gamma-rays, and the effects on the advance of the border were examined. With 200 rad and more, significant inhibition, greater with bigger doses, was seen in the advance of the borders. Regression of the borders also occurred. Irradiation with over 1000 rad destroyed the hypervascular zones.     

The effect of multiple small doses of x rays on skin reactions in the mouse and a basic interpretation

Multiple-fraction experiments have been carried out to determine the response to repeated small doses of 240 kV X rays down to 45 rad per fraction, using the mouse skin reaction system. A method of irradiating without anesthetic was developed so that up to 64 fractions could be given within 8 days; over this time, proliferation was negligible. It was found that the total dose required to produce a given reaction continued to rise with the number of fractions above 30 fractions, in contradiction to the recent conclusions of Dutreix and colleagues. The plot of reciprocal total dose against size of each fraction was shown to be linear. This finding led to an analysis in terms of a function F(e), which is proportional to the slope of the chord of the appropriate cell survival curve from the origin to the dose per fraction used. The cell survival curve derived here was well fitted by an equation of the form [Formula: see text]The initial slope was 1/690 rad and the slope at 2340 rad was 1/126 rad. Thus, 1 rad at a dose approaching 0 rad has 18% of the effect of 1 rad at a single dose of 2340 rad for mouse skin reactions. A cell survival theory based on Neary's theory of chromosome aberrations is presented and the current results are consistent with the postulate that cell death results from the formation of chromosome aberrations.     

Induction of neoplastic transformation by low-dose-rate exposure to tritiated water

Exponentially growing mouse BALB/3T3 cells seeded at low density were incubated with various concentrations of tritiated water (3HOH). A dose-dependent increase in the yield of transformants occurred in cells incubated for 100 h with 25 to 500 micrograms/ml of 3HOH. The dose-response curve rose linearly at low doses, with no evidence of a threshold or quadratic component. The transformation frequencies were similar when the same total dose of radiation was given over periods of 5 to 168 h, although survival was slightly lower with 5 h exposure compared with longer intervals. Acute X-ray exposure appeared to be more efficient in inducing transformation than protracted exposure to 3HOH at doses of 200-400 rad, but slightly less effective in the 25-100 rad range. When normalized for survival, 3HOH was more effective than X rays even at the higher doses. This result is very similar to previous findings with [3H]thymidine. A comparison of [3H]thymidine and 3HOH results suggest that the transmutational effect of tritium decay for radioactivity incorporated into DNA may be a significant factor in cytotoxicity but not in transformation; the enhanced effectiveness of both types of tritium exposure for transformation and mutagenesis may result from a higher relative biological effectiveness for the tritium beta particle for these end points.     

Voltage-current relation and K+ transport in tobacco hornworm (Manduca sexta) midgut

Summary Voltage-current curves for the isolated midgut of the tobacco hornworm were determined by transient and steady voltage clamping over the range of 200 to –200 mV. Over this range the transient method yields a linear relation while the steady method usually yields a curve consisting of two lines of differing slope which intersect at zero voltage. The difference between the results of the methods is due to a slow decline in total conductance which accompanies steady voltage clamping.Holding the midgut at short circuit increases the total conductance of the tissue in a manner consistent with increasing shunt conductance; this effect was seen in both diet-reared and leaf-reared animals.When potassium transport is inhibited by substitution of choline or sodium for potassium in bathing solution the total conductance decreases and the voltage-current curve intersects the normal curve in the hyperpolarizing region. Applying a simple equivalent circuit analysis to the results from partial or total potassium replacement suggests that the electromotive force of the potassium transport system is of the order of 140–190 mV. The conductance decrease during inhibition of potassium transport by transient anoxia is of similar magnitude, suggesting that a major effect of metabolic inhibition is to decrease the active conductance of the potassium transport pathway.     

Combined application of misonidazole and piotron pions on mouse embryos

Mouse embryos on day 8 of gestation were irradiated with negative pions (12.5-100 rad) or 200 kV X-rays (12.5-150 rad). Misonidazole (MISO), a hypoxic cell radiosensitizer, was applied 30 min before exposure. On day 13 the fetuses were examined for lethality, growth retardation and malformation. No significant embryolethal effects were observed after irradiation alone in the dose range of 12.5-100 rad (X-rays or pions). However, MISO alone and in combination with radiation led to high rates of lethality. The frequency of growth retardation was significantly increased at 100 rad and in combined treatments at low radiation doses. MISO and irradiation with 50 rad and more induced complex damages consisting of multiple and severe malformations and growth retardation. The relative biological effectiveness (RBE) for teratogenic effects was 1.6. In conclusion, the combined application of MISO and radiation of different LET revealed a strong enhancing action compared to single treatments. The extent of enhancement depends on both radiation quality and dose.