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Bone Marrow Cryopreservation: Improved Recovery Due to Bioantioxidant Additives in the Freezing Solution

National Centre for Cell Science, Pune, India

Key Words. Mouse bone marrow • Human bone marrow • DMSO • Antioxidants • GM • GEMM

Dr. Lalita S. Limaye, National Centre for Cell Science, Ganeshkhind, Pune 411007, India.

One hypothesis of cryoinjury is the damaging effect of oxygen-free radicals formed during freezing and thawing. Addition of physiologically acceptable antioxidants into the preservation solution improved the cryoprotection of bone marrow cells. Bone marrow nucleated cells were frozen using rate-controlled freezing devices. Antioxidants used in combination with 10% dimethylsulfoxide were tocopheryl acetate, catalase, ascorbic acid, superoxide dismutase and reduced glutathione. The parameters used to assess the efficacy of cryopreservation were viability, nucleated cell recovery, and colony-forming unit assays: granulocyte-macrophage and granulocyte-erythroid-macrophage-megakaryocyte. Results obtained indicate that the first three antioxidants increase the post-thaw recovery of cells, particularly in terms of early and late progenitors. Superoxide dismutase and reduced glutathione, however, have no beneficial effect on the preservation. The response of cryopreserved cells to suboptimal concentrations of colony-stimulating factors in in vitro assays was also restored to some extent when the cells were frozen with antioxidants.