Effects of diet and folate on levels of micronucleated polychromatic erythrocytes.

We describe a study examining the effects of fried beef consumption on the frequency of micronuclei in RNA-positive polychromatic erythrocytes (PCEs) in humans. 5 splenectomized individuals participated in a 2-week experiment consisting of 3 phases. During the first and last phases the subjects refrained from eating cooked meat for 4-5 days. This was designed to clear their system of mutagens found in cooked-meat products. In the second phase each person ate a similar diet, but also consumed 4 quarter-pound well-done hamburgers per day for 4 days. Erythrocyte-folate levels were also measured for each donor. No association between diet phase and micronucleus frequencies was observed among the 2 subjects with normal levels of folate. However, among the 3 low-folate donors, the frequency of micronucleated PCEs appeared to be associated with diet. Micronucleus frequencies began to increase 1 day following onset of the beef phase, and started to decline 1 day after finishing this phase. These observations are in agreement with erythrocyte maturation kinetics following short-term exposure. A repeat study performed on one of the low-folate donors consisted of two beef phases separated by a vegetarian phase. Beef in one phase was fried (high in mutagenic amino-imidazoazaarenes [AIAs]) while the beef in the other phase was fried after first being microwave-cooked (low in AIAs). Significant increases in the micronucleus frequency were not observed in this experiment, suggesting that AIAs formed during frying were not solely responsible for the induction of micronuclei.

Measurement of neutron-induced genetic damage in mouse immature oocytes.

Recent experimental evidence concerning the nature of radiosensitive targets in mouse immature (resting) oocytes has led to new experimental designs that permit measurement of radiation-induced genetic damage in these important cells. We have previously reported initial results of the detection of genetic damage in mouse immature oocytes using monoenergetic 0.43-MeV neutrons. Here we provide a full report of our data and compare the genetic sensitivity of immature oocytes with those measured by others for maturing oocytes. Until recently, all attempts to detect radiation-induced genetic damage in mouse immature oocytes had failed. This appears to have been because the radiation types and modes of dose delivery used in those studies did not sufficiently spare the hypersensitive lethality target (the plasma membrane) while at the same time deposit enough dose in DNA to produce detectable mutation. Recoil protons from 0.43-MeV neutrons produce short ionization tracks (2.6 micron mean) and can therefore deposit energy in the DNA without simultaneously traversing the plasma membrane. Using these particles, we have obtained dose-response relationships for both chromosome aberrations and dominant lethal mutations in oocytes from females irradiated 8-12 weeks earlier, when oocytes were immature. Results suggest that the intrinsic mutational sensitivity of mouse immature oocytes is not very different from that of maturing oocytes.

Size of lethality target in mouse immature oocytes determined with accelerated heavy ions.

Mouse immature oocytes were irradiated in vivo with highly charged, heavy ions from the Bevalac accelerator at the Lawrence Berkeley Laboratory. The particles used were 670-MeV/nucleon Si14+, 570-MeV/nucleon Ar18+, and 450-MeV/nucleon Fe26+. The cross-sectional area of the lethality target in these extremely radiosensitive cells was determined from fluence-response curves and information on energy deposition by delta rays. Results indicate a target cross-section larger than that of the nucleus, one which closely approximates the cross-sectional area of the entire oocyte. For 450-MeV/nucleon Fe26+ particles, the predicted target cross-sectional area is 120 +/- 16 microns2, comparing well with the microscopically determined cross-sectional area of 111 +/- 12 microns2 for these cells. The present results are in agreement with our previous target studies which implicate the oocyte plasma membrane.

Neutron RBEs and the radiosensitive target for mouse immature oocyte killing.

The highly radiosensitive immature oocytes of mice were irradiated in vivo with graded doses of 252Cf fission radiation, 0.43- or 15-MeV neutrons, or 60Co gamma rays. Comparisons of oocyte survival for neutrons and for gamma rays demonstrate that neutron RBEs for the killing of these important cells do not reach the high values (30-50 or more) at low doses observed for several other biological end points. Rather, neutrons differ little in effectiveness from gamma rays in killing these extremely sensitive murine oocytes. For 0.43-MeV neutrons, RBEs obtained from fitted survival curves reach only 1.7 at 0.1 rad. For 15-MeV neutrons, they are not significantly different from 1 at any dose tested (lowest, 4.5 rad). For 252Cf fission neutrons (E = 2.15 MeV), RBEs are intermediate between those for 0.43- and 15-MeV neutrons. For all neutron energies tested, the RBEs are particularly low in the juvenile period, a time when murine immature oocytes are especially radiosensitive. With exposure just prior to birth, however, when these cells are much less easily killed, higher, more usual RBEs are found. The minimum size of the lethality target in mouse immature oocytes, estimated from the inactivation constant for 0.43-MeV neutrons and microdosimetric values, is larger than the nucleus but not larger than the cell. This and related analytical considerations suggest that the hypersensitive target in these particular oocytes is the plasma membrane, a finding which is in excellent accord with results from other experiments using different, contrasting radiations and dose deliveries (accelerated Si14+ ions, gamma rays, and beta rays from 3HOH compared with those from [3H]thymidine).

The tritium RBE at low-level exposure--variation with dose, dose rate, and exposure duration.

Differing values have been reported for the relative biological effectiveness (RBE) of 3H beta-radiation compared with X- and gamma-rays; but the significance of these differences has not been adequately clarified. Because large quantities of tritium are involved in nuclear energy operations, it is essential to have precise knowledge of its effectiveness compared with a "standard" radiation such as gamma-rays. This is especially true for chronic, low-level exposure since available information on protracted irradiation derives mostly from gamma-ray studies. We have approached this problem experimentally by measuring effects of low-level exposures to HTO and gamma-rays on a very sensitive mammalian system. Microscopic enumeration of oocytes in ovaries of exposed and control mice provided measurements of in vivo cell killing by constant HTO levels in body water and, for comparison, by continuous 60Co gamma-irradiation. For protracted exposure from conception to 14 days after birth, the RBE was found to be greater than 1 and to vary inversely with exposure level. At effective gamma-ray doses of 50 rad, delivered at 3.5 rad/day, the RBE was 1.6. But for 25 rad, at 1.8 rad/day, it increased to 1.9. It continued to rise as dose decreased, reaching a value of 2.5. Corrected for scattered radiation, the RBE was 2.8, close to 2.9 reported previously, but below the maximum value of 3.75 predicted by microdosimetric measurements and the theory of dual radiation action. An RBE close to 3 at low doses was not suprising. But decrease at higher doses, due to downward curvilinearity of the gamma-ray dose-response curve, was unexpected for protracted exposure. It suggested that primary oocytes in the prepubertal mouse (the cells used in this study) have limited capacity for recovery. Survival-curves for constant gamma-ray dose rates of 1 rad/min and 0.002 rad/min were therefore determined. Curvilinearity was seen in both, with greater steepness for the higher dose rate, indicating that oocytes do have some ability to recover, but that even with a dose rate only 3.2 rad/day recovery is incomplete. This in turn suggests that the observed gradient in RBE is due partly to a dose-rate effect and partly to inverse variation with dose itself, as predicted by the theory of dual radiation action for acute exposure (or incomplete recovery). Comparing protracted and shorter, 5-day exposures revealed further contrasts. At 30 rad of gamma-rays the RBE in short exposures was only 1.4, while it was 2 for protracted ones. This could be due at least in part to dose rate; it may to some degree, however, reflect difference in microdistribution of tritium atoms, greater radiobiological effect in protracted exposures resulting from 3H incorporation into especially effective sites. These systematic variations of RBE with dose, dose rate, and exposure duration help to explain some of the existing lack of agreement regarding tritium's RBE and provide a more secure basis for hazard evaluation of chronic, low-level exposure.