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  • Title: Effects of low dose rate irradiation on plateau phase bone marrow stromal cells in vitro: demonstration of a new form of non-lethal, physiologic damage to support of hematopoietic stem cells.
    Author: Greenberger JS, Klassen V, Kase K, Shadduck RK, Sakakeeny MA.
    Journal: Int J Radiat Oncol Biol Phys; 1984 Jul; 10(7):1027-37. PubMed ID: 6611329.
    Abstract:
    The clinical use of low dose rate (LDR) (5-25 rad/min) total body irradiation in bone marrow transplantation patients is well established. We have developed an in vitro system for study of the effects of LDR irradiation on bone marrow stromal cells. Purified mouse bone marrow stromal cell cultures in plateau phase with no detectable hematopoiesis were prepared and were then "engrafted" in vitro by addition of purified nonadherent hematopoietic cells from continuous bone marrow cultures. Hematopoietic cells were added in liquid medium or suspended in an overlay of semisolid 0.4% agar-containing medium. Other agar overlays contained Interleukin-3-dependent cloned multipotential hematopoietic stem cell line B6SUtA. In parallel experiments, a cloned permanent bone marrow stromal cell line D2XRII was used in place of purified stromal cell cultures. Stromal cultures were irradiated at 5 rad/min, 20 rad/min, or 200 rad/min, 24 hours or 3 weeks prior to "engraftment." Two classes of irradiation damage were demonstrated following 1000 rad irradiation at 200 rad/min: 1) Decreased clonagenic survival of trypsinized replated marrow stromal cells (lethal effect); and 2) decreased production by marrow stromal cells or D2XRII cells of colony stimulating factors (CSF)s for granulocyte-macrophage progenitor cells and B6SUtA cells (physiologic effect). Holding the cultures in plateau phase for 3 weeks after irradiation was associated with significantly more repair of the lethal effect compared to the physiologic effect. Cultures irradiated at 5 rad/min or 20 rad/min to doses producing significantly less lethal effect showed a complex alteration of production of growth factors. Cumulative cell production by hemopoietic stem cells added in liquid culture was comparably decreased for all three dose rates. These data demonstrate a distinct physiologic expression of irradiation damage to bone marrow stromal cells that affects cell to cell interaction, responds differently to changes in dose rate, and is repaired with kinetics different from those of the lethal effect of irradiation. The present system should prove valuable for investigation of cellular interactions in hematopoietic stem cell engraftment that are altered by total body irradiation.
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