Analysis of the radiation-induced loss of testes weight in terms of stem cell survival

To analyze the loss of weight of testes of the mouse 28 days after irradiation to determine the proportion of radiation-insensitive tissue and relate the testes weight loss to the survival curve of stem cells.

The analysis presented above shows that the loss of weight of a certain portion of the testes of the mouse following irradiation can be represented by a curve essentially the same as that for single cell populations exposed to radiation. This result implies that the change in weight of the testes is predominantly determined by the kinetics of death of the stem cells of the spermiogenic line and that the rate of replacement of the stem cell system is sufficiently slow such that the weight loss can reflect changes in the cell population. When the radiation is given, a fraction of the stem cells as measured by the survival curve is killed. Cells more differentiated than the stem cells either die from the radiation or survive and continue differentiation, moving out of the testes as mature spermatozoa. The replacement of these differentiated cells, however, is only proportional to the number of surviving stem cells; hence, the weight of spermiogenic tissue is reduced in proportion to the number of surviving stem cells. The studies of Oakberg have identified the stem cell of the seminiferous epithelium of the mouse as the type A spermatogonium, and he has described the process of differentiation in detail. On the basis of histological observation of mouse testes 72 hours after irradiation, Oakberg made an estimate of survival to type A spermatogonia for a series of gamma ray doses from 7 to 100 R. When fitted by calculation of the regression of log S/so on D in the same manner as was done above for testes weight loss, the computed D37 was 82.5 R and the extrapolation number was 0.85. The histological method is not ideal for estimating cell survival, the dose intervals chosen are not the best for survival curve fitting, and the analysis ignores certain complexities involving differentiation of cells and time of manifestation of lethal injury; nevertheless, the agreement between the data on survival of type A spermatogonia and the loss of weight of the testes is remarkably good. On the joint basis of both results, it can be asserted with reasonable confidence that the loss of weight of the testes of the mouse following irradiation is primarily determined by the killing of type A spermatogonia and that the observed survival curve for the testes weight loss is an estimate of the radiation survival curve of the type A spermatogonia. The establishment of the testes weight loss response as a survival curve for stem cells permits an unambiguous interpretation of changes in radiobiological response. The ratio of D37 for X-ray and fission neutron irradiations is the relative biological effectiveness ratio (R.B.E.) for stem cells of the spermiogenic line. The value (3.75) is somewhat larger than that usually obtained in lethality studies in the mouse (expected value: 2.4 - 2.8), and the R.B.E. for 60Co irradiation of 1.0 is also somewhat larger than that which would be expected from lethality studies (expected value: 0.7 - 0.8). These results suggest that the response of the spermiogenic stem cells is comparable to, but different in some particulars from, the response of bone marrow stem cells to irradiation. The results in Table I indicate that there is no dose-rate effect on loss of testes weight between 14 and 2.8 R/min. Over this range of dose rate, there is a substantial effect on lethality response. In a previous study of testes weight loss, there was no dose-rate effect over a range of dose rate even wider than the one used here. The failure to find a dose-rate effect in the X-ray studies is consistent with the theory that dose-rate effects are dependent upon the presence of a shoulder on the cell survival curve. The extrapolation number of 1.0 indicates no shoulder, and hence, no sublethal injury from which animals may recover during irradiation. The observations with Piromen and AET illustrate the difference between two types of radioprotective drugs. The Piromen is a bacterial polysaccharide which, among several pharmacological actions, appears to 'stimulate' bone marrow, and in Leo studies it confers significant protection on treated mice. The AET is a sulfhydryl-type protective drug. The AET increases the D37 of the testes weight loss curve by an amount comparable to its effect on bone marrow stem cells; the action is equivalent to an effective dose reduction in the primary biological or chemical events leading to injury. The Piromen has no effect on the D37 of the testes weight loss, and it can be inferred that Piromen has little or no intrinsic dose reduction potential, but acts in lethality studies by some form of stimulation of the bone marrow or modification of its differentiation. Piromen had no effect on the weights of testes of irradiated mice. In conclusion, it is shown that loss of weight of the testes of the mouse following irradiation probably reflects closely the destruction of the germinal cell line of the spermatogonial series. Thus, testes weight loss can be used as a mammalian cell survival curve in vivo. Treatment of the data by cell survival analysis allows a comparison of effects of low LET radiation and of radioprotective drugs in terms of the basic biological events common to all radiosensitive tissues. The analysis also provides some comparison of effects of radiation on different in vivo cell lines and leads to better generalization of the nature of radiation response in mammals.