Project Title: 14. Scope: Molecular and Cellular Radiobiology Effects of Radiation and Chemicals on Control of Hemopoiesis RX-03-02-(b) (Conte'd.) thymidine and P-32 labeled diisopropylfluorophosphate (DFP-32); depletion by extracorporeal irradiation of the blood (ECIB); Fe-55 suicide of erythropoietic precursors; and production of humoral regulators of hemopoiesis in vivo in renal glomerular tissue culture systems. _ Fe-55 suicide, recently developed here, allows the near-total obliteration of that segment of erythropoiesis which is incorporating iron into hemoglobin, The short path length of Fe-55 Auger electrons (about I um) results in deposition of almost all the energy within the cell boundaries of the red cell precursors. Thus with a sufficient amount of Fe-55, these cells are killed--eliminating new production of red cells while the HSC con- tinues to flow down the erythropoietic pathway. If there is an intramedullary feedback loop from the differentiated red cell compartment to the pluripotent HSC which senses diminished red cell production before there is anemia, an increased flow ot the HSC into erythropoiesis would be expected with an initial dimtnution in size of the HSC pool, measureable in mouse by reduction in number of colony forming units in the spleen (CFUs). The relative proportion of pluripotent HSC's that are dormant or actively in cycle is measured by killing the cells in cycle either by hydroxyurea or by suicide through incorporation of tritiated thymidine. The quantitative effect of these treatments is measur- ed by CFUs or a limiting dilution analysis of the number of stem cells growing in diffusion chambers. A combination of these techniques applied serially to mice receiving continuous Fe-55 suicide will be used to determine the rate at which the HSC is diverted down the erythropoietic pathway and whether the HSC has a limited capacity for mitosis as stated in the Hayflick hypothesis or an infinite capacity, a question in chemotherapy of human malignant diseases, Erythropoiesis is regulated in part (EP), and a, glycoprotein. by the hormone erythropoietin To date EP has been obtainable only by extrac- tion from the blood plasma or urine of severely hypoxic or anemic human beings or animals. The development of renal glomerular cultures from goat and human kidneys synthesizing EP, assayable in the supernate, will allow biochemical study of the factors that may control production of EP at the cellular level (pO, pCO, pH, androgens, etc.) and will permit purification and characterization of human EP, Studies will then be directed to the production of antibodies against EP and development of me assays for clinical application. If purified radiolabeled weed by adding amino acids labeled with carbon-14 or tritium to the es has specific activity high enough for autoradiography, attempts will B& made to identify the site of genetic control of erythropoiesis by combining autoradiography and karyotyping. Granulopoiesis is studied in the intact mammal utilizing labeling with tritiated thymidine, determining the DNA content d the cells in various stages of granulopoiesis, and determining the changes in the (See Continuation Sheet) bi 39286 RX-226