A lifelong, wide-range radiation biodosimeter: erythrocytes with transferrin receptors.

Rats exposed to 0.1-600 cGy x-rays developed significantly prolonged increases (1.9-11%) of normoblasts (red cell precursors) in the bone marrow as well as a subpopulation of transferrin receptor marked erythrocytes (E-Tr, 1.9-20%) in the blood. Several characteristics are shared in common by these two progenies of the hematopoietic stem cell (HSC): a heretofore unrecognized dose-response curve extending from 0.1-600 cGy composed of logarithmic and quadratic dose response segments, the joining of the segments at approximately 150 cGy, and an identical 29-wk period for one-half repair of the quantified cell damage. This suggests that both of these cells could be surrogates for assessing radiation injury. Preliminary findings suggest that this phenomenon is identifiable in humans.

Low dose radiation: what is it? A personal perspective.

In any retrospective, there is an uneven mixture of personal and impersonal recollections. But in the end, what is personal for one is necessarily impersonal for another. In this case, what low dose x-radiation is, is in the eyes of the beholder.

Iron kinetics effects of 88 millirads. Partial-versus-total body X-irradiation.

Rats were irradiated with one tibia shielded (95% marrow exposure), total body exposed (TBI, 100%), and only one tibia exposed (5%), or they were sham irradiated (SI, 0%). Plasma Fe-59 clearance time (T1/2) and Fe-59 content ratio in the right and left tibia (RT/LT) were assayed to determine the erythroid activity of the overall marrow of the animals and the relative marrow activity in the exposed and shielded tibias, respectively. When a major fraction of the overall marrow was shielded or irradiated, the overall erythroid activity levels were identical to those of the SI and TBI animals, respectively. Interestingly, enhanced normoblastosis was observed in the marrow of the exposed tibia of individual animals exhibiting normal erythroid activity in 95% of the marrow. Conversely, localized marrow with normal erythroid activity was found in a shielded tibia of individual rats, demonstrating an enhanced erythroid activity in a major fraction of the total body. It was concluded that 88 mrad can alter marrow functions in a small isolated skeletal region as effectively as in the whole body, and tandem assays of the Fe-59 T1/2 and Fe-59 RT/LT can facilitate ultra-low-dose X-ray studies involved with partial body exposures.

Direct evidence for a stem cell common to hematopoiesis and its in vitro microenvironment: studies on syngeneic (inbred) Wistar Furth rats.

When injected into a group of lethally irradiated syngeneic (inbred) Wistar Furth (WF) rats, suspensions of stromal cells grown in monolayer culture from the marrow of WF rats produced hemopoietic colonies in the spleen and rescued 50% of the rats, while 90% of the non-injected (control) rats died within 30 days and had no hemopoietic colonies in the spleens. Fifty percent of the injected (test) rats which died between days 6 and 22 showed hemopoietic regeneration in the bone marrow, while little or no evidence of hemopoietic regeneration was seen in the control animals. Our results suggest that the marrow stroma grown in vitro contain cells with hemopoietic potential and are transplantable.

The distribution and kinetics of nuclei in rat osteoclasts.

Osteoclast development and growth were studied by determining the number of labelled nuclei in osteoclasts of different sizes (based on the number of nuclei per osteoclast, N/O) and the number of osteoclasts with labelled nuclei at various intervals after tritiated thymidine [( 3H]TdR) injection in young rats. The osteoclast smears were made from the cellular periosteum of the proximal tibia. The frequency distribution of the N/O osteoclasts types in the smears had profiles similar to that of in situ osteoclasts in whole mounts of proximal tibia, which indicates that the osteoclast population of the smears was representative of that on the bone surface. A vast majority of the osteoclasts had a 1-6 N/O, and a number of the cells had as many as 26 or more nuclei. Furthermore, profiles of N/O frequency distributions were similar over the course of the study. Nuclei with [3H]TdR label were initially observed in osteoclasts between 4 and 12 hr after isotope injection. However, fusion of labelled nuclei to osteoclasts continued for at least 150 hr. In general, the labelled osteoclasts exhibited a significantly larger number of nuclei than the unlabelled osteoclasts. The probability of an osteoclast incorporating one or more labelled nuclei increased with time after injection and with an increase in N/O. Labelling intensity decreased with time post injection and with an increase in N/O. The data suggest that turnover of nuclei is more rapid in osteoclasts with high N/O values.

A comparative study of osteoblasts: in situ versus isolated specimens.

Osteoblasts and osteoclasts are known to adhere tenaciously to bone surfaces even when the fibrous periosteal covering is removed. Brush-smear isolates and in situ, whole bone mounts of the osteogenic periosteum on the inner and outer calvarial tables of young adult rats were examined at the light microscopic level after exposing the cells by removing the fibrous periosteum. Based on staining, enzymatic activity, and overall morphology, most of the cells could be classified as osteogenic in nature. However, detailed analysis revealed that some cells exhibited certain unique morphologic appearances that suggested the existence of subpopulations or variant forms of the conventional or prototypic osteoblast. Three variant forms were the large nucleate, small nucleate, and multinucleate osteoblast-like cells, the latter with nuclear numbers ranging from 2-10. The finding of all such forms in smears, short-term cell cultures, and in situ specimens greatly lessened the probability that the unusual forms were artifactual. Phase contrast microscopic studies and sectioning of cellular isolates for study at both the light and electron microscopic levels provided additional support for the existence of the multinucleate osteoblast-like cell. The small size of these osteoblast-like subpopulations and the orientation of the cells to the commonly used plane of section of most histologic bone specimens, particularly with the multinucleate type, could explain why they had escaped earlier detection.

The effects of low dose (less than 1 rad) X-rays on the erythropoietic marrow.

Adult female Sprague-Dawley rats were exposed (200 kvp X-rays) to whole body doses of 22-1320 mrad and examined for changes in the level of red blood cell precursors (RBCp) in the marrow at 5-30 weeks post-irradiation, under nonbled and phlebotomy-induced anemic stress conditions. Increases in the RBCp %, RBCp/mg marrow, and RBCp/skeleton under nonbled conditions, and a suppressed erythroid response to an induced anemia, were found after acute doses in the range of at least 70 mrad. Dosages of 22 or 44 mrad that induced no measurable changes when applied only once were found to be effective when they were employed 4 or 2 times/week, respectively. The results suggested the presence of a linear-quadratic dose-response relationship in which the quadratic function exists between 88 and 981 mrad, and the linear dependency, below 88 mrad.

A comparative study of osteoclasts: in situ versus smear specimens.

Osteoclasts residing on rims and walls of bony concavities on remodeling proximal tibia from growing rats were examined by light microscopy following stripping of the periosteal connective tissue layer. Comparison of these cells in situ and after transfer to glass slides revealed the presence of numerous mononucleate osteoclasts, as well as typical multinucleate forms, all exhibiting a ruffled border and acid phosphatase and succinic dehydrogenase activity. Macrophage-like cells were situated adjacent to osteoclasts in situ. Osteoblasts were relatively inconspicuous. The possibility that the basic functional osteoclast unit is a mononucleate cell is discussed.

Recurrent decreases of erythrocyte electrophoretic mobility in rats exposed to 1.0 R whole-body x-radiation.

Circulating erythrocytes from rats were examined up to 30 weeks post whole-body exposures of 1.0 R for alterations in the expression of net negative surface charge as measured by whole-cell microelectrophoresis in saline sorbitol. Erythrocyte electrophoretic mobility was determined in an apparatus composed of a horizontal transilluminated cylindrical chamber, equipped with a reversible, blacked platinum electrode, immersed in a water bath maintained at 25.0 +/- 1.0 degree C (Rank Brothers). In two separate experiments, recurrent decreases in the expression of net negative surface charge occurred at 10, 17, and 30 weeks post-irradiation. At these times distributional analyses of recorded erythrocyte electrophoretic mobility (EEPM) values revealed a skewing of the normally distributed EEPM population values to lower EEPM. Total sialic acid content released from hydrolyzed erythrocyte membrane preparations revealed no significant differences between erythrocytes from sham and irradiated animals. In vivo post-irradiation labeling of erythrocytes with diisopropyl-[32P] phosphorofluoridate at 4 and 33 weeks (separate experiments) indicated only a minor abbreviated erythrocyte life span at 33 weeks. Therefore, effects from low dose (1.0 R) whole-body irradiation would appear to include a recurrent defect in the expression of the net negative surface charge.

Infusion at constant rate in vivo.

Infusion can be maintained at a constant rate over an extended period of time in vivo by the use of an implanted diffusion chamber. Plasma 59Fe was maintained at a constant level for 10 days when infused from a s.c. implant. Injected isotope was cleared exponentially with a half-clearance time of about 8 h.

Endosteal marrow: a rich source of hematopoietic stem cells.

Hematopoietic cells isolated from the endosteal bone surface, that is,the endosteal marrow cells, were found to consist mainly (60 to 80 percent) of lymphoid and late-stage normoblast-like cells. Unlike the cells they resemble, the endosteal marrow cells showed an affinity for Sudan black, demonstrable nucleoli (Feulgen reaction), and an absence of hemoglobin. Assays showed that over one-half of the endosteal marrow cell population may be the colony-forming units, the CFU-S of Till and McCulloch. Thus, high concentrations of stem cells could be obtained from the endosteal bone surface by means of the present isolation technique.

Anemic stress as a trigger of myelogenous leukemia in the unirradiated RF mouse.

Ninety-six percent of mice that were bled of 50 percent of their blood volume when they were 9 weeks old succumbed to myelogenous leukemia by 15 months after phlebotomy, the majority of them dying between 7 and 10 months after this treatment. These results suggest that (i) anemia is an effective stress for triggering myelogenous leukemia in animals that are predisposed to the disease, (ii) the RF mouse is "naturally" prone to the development of myelogenous leukemia, and (iii) the concept of two-step de novo induction of myelogenous leukemia appears to be applicable in this animal.

Anemic stress as a trigger of myelogenous leukemia in rats rendered leukemia-prone by x-ray.

All of the 128 Sprague-Dawley female rats bled two-thirds of the blood volume at 1, 2, or 3 months after irradiation (50, 170, or 350 roentgens) succumbed to leukemia by 16 months after bleeding. Some nonbled irradiated rats developed leukemia possibly as a result of triggering by a radiation-induced anemia. The threshold leukemogenic x-ray dose is probably below 25 roentgens. Elevated levels of basophils, neutrophils containing the "pseudo-Pelger" type nuclear anomaly, and myeloblasts exhibiting Auer rods seen in these animals were generally not found in earlier models of rat myelogenous leukemia.