Expression of receptor for ecotropic murine leukemia virus on hematopoietic cells

A fusion protein (F-SU/GFP) which is comprised of the surface (SU) subunit of the Friend MuLV envelop glycoprotein and the green fluorescence protein (GFP) was generated by a baculovirus expression system. The F-SU/GFP specifically bound to mammalian tissue cultured cells expressing the mCAT-1, the receptor for ecotropic murine leukemia virus (Eco-MuLV). The expression level of mCAT-1 on hematopoietic cells as measured based on the capacity of cells to absorb the F-SU/GFP. In BALB/c mice susceptible to Eco-MuLV infection, all hematopoietic cell subpopulations absorbed the F-SU/GFP ith the most prominent absorption observed in the TER119+ erythroblasts. Hematopoietic cells from C4W and AKR mice did not absorb the F-SU/GFP as readily as the BALB/c cells, probably due to expression of the envelope glycoproteins of endogeneous Eco-MuLVs in these mice. Although AKR mice are susceptible to Eco-MuLV infection hile C4W mice are resistant, these mice showed no apparent difference in the F-SU/GFP-absorbing capacity

Early detection of p53-/- thymocytes appeared during radiation-induced thymic lymphomagenesis in p53 heterozygous (+/-) B10 mice

Radiation exposure is know to be highly leukemmogenic, but the mechanism action is not yet understood. Studies with the p53 heterozygous mouse have shown that a single dose of radiation can induce a high incidence of thymic lymphomas, most of which manifest the loss of wild-type p53 allele. The loss of heterozygosity (LOH) in the p53 gene is thus thought to be an early event in radiation-induced thymic lymphomagenesis. Thus, the p53 heterozygous mice might be a useful model for the study of consequences of radiation-exposure. However, sensitive detection of p53-/- thymocytes appeared in p53+/- thymocyte population was needed to examine appearance of p53-/- thymocyte in p53 heterozygous (+/-) B10 mice in the early stage of radiation-induced thymic lymphomagenesis. In this study, we characterized highly radioresistant thymocytes as a candidate of p53-/- thymocytes, because the enrichment of p53-/- thymocyte by a high dose of irradiation could be expected from the facts that p53+/- thymocytes are sensitive to radiation-induced apoptosis but p53-/- thymocytes highly resistant. At various times post irradiation (6Gy) as lymphoma-inducing treatment, p53 heterozygous B10 mice were treated with a high dose (30Gy)of whole-body irradiation to enrich p53-/- thymocytes and, 24 hours later, the remaining thymocytes were assayed for surface marker and p53 genotype. In a significant fraction of the mice 7wks after 6Gy-irradiation, we observe an abnormal increase of relative cell number of CD4+CD8+ thymocytes remaining in the thymus of highly irradiated mice and such abnormal thymocytes could be detected even 5 weeks after the irradiation. The abnormal CD4+CD8+ thymocytes werer shown by PCR analysis of sorted cells to contain cells ith p53-/- genotype. The data obtained so far suggest that p53-/- thymocytes may appear as prelymphoma cells within several weeks post irradiation in CD4+CD8+ thymocyte population of irradiated p53 heterozygous mice.第１２回国際放射線研究会

Direct detection of p53-/- thymocyte appearing at an early stage of radiation-induced thymic lymphomagenesis in p53+/- heterozygous B10 mice

Purpose: The appearance of tumor suppressor protein 53 (p53) -/- thymocytes at an early stage of radiation-induced lymphomagenesis was investigated in the p53 heterozygous (+/-) B10 mice following a single dose of irradiation, since most thymic lymphomas manifested the loss of the wild-type p53 allele and the loss of heterozygosity was thought to be an early event critical for radiation-induced thymic lymphomagenesis in p53+/- mice. \nMaterials and methods: The mice were exposed to a single dose (6 Gy) of irradiation to induce thymic lymphomas and, at various times after irradiation, treated with an extremely high dose (30 Gy) of whole-body irradiation to enrich p53-/- thymocytes and, 24 h later, the remaining thymocytes were assayed for cell surface markers and p53 genotype. \nResults: In a significant fraction of the p53+/- mice 5 weeks after 6 Gy irradiation, there was a relative increase in the number of cluster of differentiation (CD) 4+CD8+ thymocyte subpopulation among thymocytes remaining after 30 Gy irradiation. The CD4+CD8+ double-positive (DP) thymocytes were shown to contain p53-/- cells, and the number of p53-/- thymocytes was more than 105 in those individuals. \nConclusions: The results clearly indicated that an extremely high dose (30 Gy) of whole-body irradiation enabled us to directly detect p53-/- thymocytes in an abundant p53+/- thymocyte population and that proliferative p53-/- thymocytes develop in a CD4+CD8+ DP thymocyte subpopulation within a few weeks after a single dose (6 Gy) of irradiation

Radiation-Induced Apoptosis in Peritoneal Resident Macrophages of C3H Mice

Gamma ray-radiation induced significant apoptosis in peritoneal resident macrophages (PRMs) of C3H/HeJ (C3H) mice, but not other strains of mice. In order to investigate the role of DNA damage in the apoptosis, DNA damage was quantified in PRMs using the alkaline single cell gel electrophoresis (Comet) assay. No significant difference was found between C3H and C57Black/6 mice in either radiation-induced DNA damage or repair. Radiation induced apoptosis at the same level in PRMs of p53 knockout mice and atm knockout mice as in those of wild type C3H mice, however radiation-induced apoptosis was significantly less extensive in thymocytes of these mutant mice than those of wild type mice. Apoptosis was also induced at the same level by irradiation in PRMs of C3H scid mice as in those of wild type C3H mice. Therefore, it was suggested that radiation-induced DNA damage and TP53, ATM or DNA-PK mediated cellular responses occurring downstream thereof were not involved in the radiation-induced apoptotic cell death in C3H mouse PRMs

Stem-cell leukemia: p53 deficiency mediated suppresion of leukemic differentiation in C3H/He myeloid leukemia

C3H/He mice produce myeloid leukemias after whole body irradiation of 1-3Gy as compared with non-irradiated controls that produce fewer than 1% of leukemia [Radiation Research 127 (1991) 146]. Thus, p53-deficient C57BL/6 strain, a malignant lymphoma prone, was crossed back into C3H/He strain. Lethally irradiated wild-type mice to which p53-deficient bone marrow cells were transplanted (transplantation assay) showed dramatic change in the propensity of leukemia of myeloid lineages, the cells lacking CD3, Thy1.2, sIgM, B220, Mac-1, Gr-1, but being positive for c-Kit and CD44. Furthermore, transplanted mice subjected to 3Gy irradiation gave rise to a faster development of leukemia and a higher frequency of double-lineage leukemias than the non-irradiated control