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Selective killing of leucocytes by freezing: potential for reducing the immunogenicity of pancreatic islets
M J Taylor1, H L Bank, M J Benton
1Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston 29425.
Recent developments in transplantation immunobiology, concerning the mechanism of tissue rejection, clearly indicate that antigen recognition alone is not sufficient for lymphocyte activation. "Passenger" leucocytes (antigen presenting cells) carried in the donor tissue are now recognized as the major immunogenic stimulus, such that removal of these contaminating leucocytes, using a variety of procedures, has enabled the immunogenicity of allografts to be reduced and the survival of the graft to be significantly extended. Remarkable advances have been made in recent years in preventing rejection of islet allografts, and even xenografts, in experimental animals by using procedures which do not involve continuous immunosuppressive therapy. Cryopreservation offers not only the means by which donor tissue can be stored effectively during such procedures but also the possibility that, under appropriate conditions, the freezing and thawing process itself could modulate tissue immunogenicity by allowing the selective killing of immunocompetent leucocytes whilst preserving the function of parenchymal cells in the graft. In this preliminary study we have characterized the survival of leucocytes and islets from the same species (rat) after cryopreservation by the same technique using dimethyl sulphoxide (Me2SO) as the cryoprotectant. Optimum survival of rat lymphocytes and macrophages was found at cooling rates in the range of 0.3-5 degrees C/min, after cooling at rates greater than 75 degrees C/min, survival was reduced to a negligible level. On the other hand, recovery of islets was 73 +/- 9% at 75 degrees C/min, indicating that depletion of lymphoid cells, with satisfactory preservation of endocrine cells, should be obtainable at this cooling rate.
Recent developments in transplantation immunobiology, concerning the mechanism of tissue rejection, clearly indicate that antigen recognition alone is not sufficient for lymphocyte activation. "Passenger" leucocytes (antigen presenting cells) carried in the donor tissue are now recognized as the major immunogenic stimulus, such that removal of these contaminating leucocytes, using a variety of procedures, has enabled the immunogenicity of allografts to be reduced and the survival of the graft to be significantly extended. Remarkable advances have been made in recent years in preventing rejection of islet allografts, and even xenografts, in experimental animals by using procedures which do not involve continuous immunosuppressive therapy. Cryopreservation offers not only the means by which donor tissue can be stored effectively during such procedures but also the possibility that, under appropriate conditions, the freezing and thawing process itself could modulate tissue immunogenicity by allowing the selective killing of immunocompetent leucocytes whilst preserving the function of parenchymal cells in the graft. In this preliminary study we have characterized the survival of leucocytes and islets from the same species (rat) after cryopreservation by the same technique using dimethyl sulphoxide (Me2SO) as the cryoprotectant. Optimum survival of rat lymphocytes and macrophages was found at cooling rates in the range of 0.3-5 degrees C/min, after cooling at rates greater than 75 degrees C/min, survival was reduced to a negligible level. On the other hand, recovery of islets was 73 +/- 9% at 75 degrees C/min, indicating that depletion of lymphoid cells, with satisfactory preservation of endocrine cells, should be obtainable at this cooling rate.