Kaoru Tanaka and Eiichi Kojima
- Keywords:megakaryocyte progenitor cells (CFU-Meg), fractionated irradiation, mice
Hematopoietic systems are the most sensitive systems in the body to radiation. Only a few studies have attempted to examine the effects of fractionated irradiation on hematopoietic progenitor cells in mice. We investigated the effects of in vitro fractionated irradiation with two equal split-doses on megakaryocyte progenitor cells (CFU-Meg) in the bone marrow and spleen of mice . ¡¡Female BALB¡¿c mice, 10-20 weeks old, were used for this study. CFU-Meg cultures were prepared according to the fibrin clot culture system of Kuriya et al. Femoral bone marrow (5¡ß10£µcells¡¿ml) or spleen cells (50¡ß10£µcells¡¿ml) were suspended in culture medium containing 10¡ó spleen lymphocyte-conditioned medium (as a source of colony-stimulating factor) and fibrinogen solution and then were plated. The fibrin clots were first exposed to X-rays (200kVp, 20mA, 0.7Gy¡¿min) at graded doses (0.1-1.9Gy) and were kept at 37¡î for various times (1, 2, 3, 4, 5, and 24h). Then, they were irradiated a second time at the same doses as the first ones. Four days after irradiation and incubation, the cultures were fixed with 5¡ó glutaraldehyde and were stained for acetylcholinest-erase activity. Colonies consisting of 4 or more acetylcholinesterase-positive cells were scored as CFU-Meg. ¡¡The results were evaluated by comparison of the dose-survival relationships of CFU-Meg irradiated with a single dose and split doses of X-rays. The radiation-dose response curves for CFU-Meg in both bone marrow and spleen after a single irradiation with X-rays exhibited no shoulder with a single slope. A similar profile was found in the dose response curves after split doses of X-irradiation. The Do values of the dose response curves were measured at various time intervals between two equal split doses of X-irradiation. The data indicated that there was an apparent absence of recovery in the survival of femoral CFU-Meg after the fractionated doses of irradiation (Fig.1) and the small recovery of fractionated irradiation was found at 3 and 24h intervals for splenic CFU-Meg. In relation to the concentration of cultured cells, the dose response curves for CFU-Meg in the bone marrow slowed with the increase of the plated cells. The present studies also showed that the slope of the spleen dose-survival curves was slow in contrast with that of bone mallow, but the concentration of cultured spleen cells was usually ten times that of cultured bone marrow cells. When the sizes of individual colonies were examined by counting the number of acetylcholinesterase-positive cells per colony, there was little difference in the distribution of colony sizes between groups having single and fractionated doses of irradiation.
Fig.1 Changes in the Do values at different time intervals for single and two equal split doses of irradiation.
a Higher Contents
Contents
Next
a Higher Next
2¡¥Pre-T Cell Activity in the Bone Marrow during Early Stages after Irradiation in Mice with Different Susceptibilities to Radiation-Induced Lymphomagenesis
Masahiro Muto, Ying Chen and Eiko Kubo (£±China Institute for Radiation Protection)
- Keywords :pre T cell, strain difference, radiation-induced T cell lymphomas
¡¡It is known that exposure of B10. Thy 1.1 mice to fractionated X-irradiation induces a high incidence (¡ä95¡ó) of thymic lymphomas, whereas the incidence for STS¡¿A mice is very low (¡ã8-9¡ó). ¡¡Under the influence of an irradiated host environment, radiation-induced thymic lymphomas can originate from cells present in nonirradiated thymus grafts (Muto et al. Cancer Res., 43, 3822, 1983). Injection of the syngenic normal bone marrow cells after irradiation has been shown to enhance the progressive regeneration of the radiation-injured thymus and effectively prevent the development of thymic lymphomas. Thus, the cellular processes leading to the development of thymic prelymphoma cells seem to have involved a complex process in which the interactions between target cells for neoplastic transformation and thymic microenvironment and also bone marrow were of critical importance. ¡¡To investigate the cellular mechanism of strain difference with regard to radiation-induced lymphomagenesis, we established the STSA.Thy 1.1 strain by introducing Thy 1.1 marker from the B10.Thy 1.1 strain into STS¡¿A strain by 12 repeated backcrosses followed by brother-sister matings. ¡¡Using B10.Thy 1 congenic mice and STS¡¿A.Thy 1 congenic mice, we first investigated the radiosensitivity of pre-T cells in the bone marrow from B10.Thy 1.1 and STS¡¿A. Thy 1.1 mice. Bone marrow cells were taken from individual B10.Thy 1.1 and STS¡¿A.Thy 1.1 mice at various times after leukemogenic irradiation and a limited number of these cells (6 X 10£³-1 X 10£µ) was injected intrathymically into 3.78 Gy-irradiated B10.Thy 1.2 and STS¡¿A.Thy 1.2 mice, respectively. One month after transplantation, the thymocytes were stained with FITC-labeled anti-Thy 1.1 and biotin-labeled anti-Thy 1.2 followed by avidin-PE, and the percentage of donor-type (Thy 1.1¡Ü) cells was analyzed. The titration curves were drawn by limiting dilution analysis according to a Poisson distribution, and the number of pre-T cells in the bone marrow from B10.Thy 1.1 and STS¡¿A.Thy 1.1 mice after irradiation were evaluated. The results indicated that thenumber of pre-T cells in the bone marrow from B10.Thy 1.1 mice was severely depressed during the time up to 70 days after irradiation, while that from STS¡¿A.Thy 1.1 mice recovered to 22.4¡ó of that of non-irradiationed mice. ¡¡Moreover, leukemogenic irradiation resulted in a sustained depression of the level of mixed lymphocyte reactivity (MLR) of the thymus from B10.Thy 1.1 mice; in contrast, MLR of the thymus from STS¡¿A mice after irradiation was higher than that from the control group. TL-2 positive cells which were not expressed on normal thymocytes of B10.Thy 1.1 mice appeared significantly in the thymocytes of irradiated B10. Thy 1.1 mice, but the proportion of TL-2 positive thymocytes from irradiated STS¡¿A mice did not change when compared with normal thymocytes of STS¡¿A mice. Abnormal expressions of IL-2Ron thymocytes of irradiatedB10. Thy 1.1 mice were observed, but not on those of irradiated STS¡¿A mice. ¡¡Thus, in the B10. Thy 1.1 mice, irradiation (1.61 Gy X 4) causes atrophy of the thymus as well as depletion of pre T cells in the bone marrow. Under these conditions, a differentiation arrest and¡¿or abnormal T cell differentiation might occur among regenerating immature lymphoid cells, and some of these regenerating cells may then undergo preneoplastic change or transformation due to induced abnormal gene expression followed by genomic instability and chromosome aberrations. On the other hand, radioresistant T cell precursors from STS¡¿A mice appear to undergo normal differentiation. This might lead to prevention of the appearance of prelymphoma cells in irradiated STS¡¿A mice.
[Publications]1)Muto, M., Chen, Y., Kubo, E. and Mita, K.: Jpn. J. Cancer Res., 87, 247-257, 1996. 2)Muto, M., Utsuyama, M., Horiguchi, T., Kubo, E., Sado, T. and Hirokawa, K.:Cell Prolif., 28, 645-657, 1995.
a Higher ContentsContentsNexta Higher Next
3¡¥Possible Induction of GVL Effect against a Leukemia Refractory to Anti-leukemia Response in Usual MHC-compatible, Allogeneic Bone Marrow Transplantation
Shiro Aizawa, Hitoko Kamisaku and Toshihiko Sado
- Keywords:CD8¡Ü, T cells, GVL effect, GVHD, leukemia
¡¡Our previous results for a murine model indicated that GVL effect against leukemia LE750 could not be induced in MHC-compatible allogeneic BMT of leukemia-bearing C3H mice, whereas a significant GVL effect was observed against leukemia 8313. Furthermore, we showed that CD8¡Ü T cells contaminating donor BM were consistently critical for the induction of GVL effect in allogeneic BMT of leukemia-bearing C3H mice and, furthermore, CD8¡Ü T cells of MHC-compatible, allogeneic AKR donor mice could preferentially induce the GVL effect without significant lethal GVHD. ¡¡To elucidate cellular factors which influence sensitivity of leukemias to anti-leukemic response induced in allogeneic BMT of leukemia-bearing host, we examined why a significant GVL effect can not be induced against a leukemia (LE750) in ordinary MHC-compatible, allogeneic BMT. The resistance of LE750 leukemic cells to the induction of GVL effect was attributed to neither less sensitivity to lysis by minor H antigen-specific, cytotoxic T cells nor an immuno-suppressive activity of LE750 leukemic cells in leukemia-bearing mice. To investigate the significance of dose effect of cells responsible for the induction of GVL effect, we used CD8¡Ü T cells of AKR donor mice which have been shown to preferentially induce GVL effect without lethal GVHD and therefore allowed us to increase the cell number of CD8¡Ü T cells in the donor bone marrow inoculum. Although the addition of unprimed CD8¡Ü T cells into the donor inoculum could not induce a significant GVL effect against LE750 leukemia, CD8¡Ü T cells of donor mice which were primed with host cells manifested improvement in survival and the effect of the inoculation of primed CD8¡Ü T cells was dose-dependent. The results indicate that the outcome of anti-leukemic response in allogeneic BMT of leukemic recipients may be determined, at least in part, by the balance between the malignancy of leukemias and the number of anti-leukemic effector cells.
[Publications]
a Higher ContentsContentsNexta Higher Next
4¡¥Effect of Fractionated Radiation Exposure on Hematopoietic Potentials in Radiation-Induced Thymic Lymphoma-Susceptible B10 Mice and -Refractory C3H Mice
Hitoko Kamisaku, Shiro Aizawa, Kumie Nemoto, Masanobu Kitagawa¡ö, Yoshinori Ikarashi, and Toshihiko Sado (¡öTokyo Medical and Dental University, Tokyo)
- Keywords:fractionated radiation, hematopoiesis, thymic lymphoma
We previously reported from dose-response analysis of thymocyte precursors by intrathymic injection that the number of thymocyte precursors present in the bone marrow of fractionated whole-body X-irradiation (FX) -treated B10 mice was greatly reduced as compared to that present in the normal bone marrow. The results suggested that the depletion of pre T cells in the bone marrow following FX-treatment was an important cellular events that led to the development of thymic lymphomas. ¡¡In order to explore the significance of the shortage in supply of pre T cells from the bone marrow during FX-induced thymic lymphomagenesis, thymocyte precursor potentials in bone marrow of FX-treated B10 mice was compared to that of lymphomagenesis-refractory C3H mice. Furthermore, the recovery of other hematopoietic activities following FX-treatment was also examined. The results showed that the frequency of pre T cells in the bone marrow of FX-treated mice was greatly reduced to a few percent 4 weeks after irradiation in not only lymphoma-prone B10 but also -refractory C3H mice. On the other hand, the activities of hematopoietic stem cells (CFU-s and GM CFU-c) in the bone marrow of FX-treated mice were reduced to a few percent of those in normal bone marrow 3 days after irradiation but progressively recovered to about 20 to 50 ¡ó 4 weeks later . The kinetics for the numbers of pre B , mature T and B cells in FX-treated mice also showed that the regeneration of all lymphoid lineage cells was severely delayed in both strains of mice as compared to that of Gr-1¡Ü polymorphonuclear cells. These results indicate that fractionated irradiation severely injured overall lymphopoiesis more than hematopoiesis including CFU-s, GM CFU-c and polymorphonuclear cells.
[Publications]Kitagawa, M., Aizawa, S., Kamisaku, H., Ikeda, H., Hirokawa, K. and Sado, T.: Blood 86, 603-609, 1995.
a Higher ContentsContentsa Higher Next