title: The effects of variation of cryopreservation protocols on the immunogenicity of allogeneic skin grafts.
abstract: The use of allografts to effect wound closure on full thickness skin injuries is limited by unpredictable graft rejection times. If the period of graft take could be extended, the use of allografts would reduce the morbidity and mortality associated with these wounds. This study evaluated the effects of variation in the cryopreservation protocol on the viability and immunogenicity of skin using a murine model system. Immunogenicity was assessed by the stimulatory activity of C3H (H-2K) skin-derived epidermal cells (EC) in primary one-way EC/lymphocyte reactions with BALB/c (H-2d) and CBA (H-2K) responder lymphocytes. Viability was determined by measuring tetrazolium reductase activity. The following cryopreservation protocols were assessed: freezing at 1, 30, 64, and > 100 degrees C/min in 10 and 15% (v/v) Me2SO and freezing at 30 degrees C/min in 5 to 20% (v/v) Me2SO or glycerol. A cryopreservation protocol of 30 degrees C/min in 15% (v/v) Me2SO proved optimal for murine skin allograft storage and immunomodulation. The viability of skin treated by this protocol was maintained (78% of fresh skin viability, no significant difference analysis of variance). The stimulatory capacity of treated EC for H-2K and H-2d lymphocytes was 5 +/- 4 and 5 +/- 9% (+/- 95% confidence limits) of fresh EC (100%) activity. Langerhans cell numbers in epidermal sheets and EC suspensions did not correlate with the stimulatory capacity of fresh and treated EC for allogeneic lymphocytes. A functional impairment of Langerhans cell immunostimulatory capacity was implied.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Microbiology, University of Leeds, United Kingdom.
authors.forename: E
authors.initials: E
authors.lastname: Ingham
authors.forename: J B
authors.initials: JB
authors.lastname: Matthews
authors.forename: J N
authors.initials: JN
authors.lastname: Kearney
authors.forename: G
authors.initials: G
authors.lastname: Gowland
mesh_terms: D000818:Animals; D003080:Cold Temperature; D015925:Cryopreservation / Q000379:methods*; D004121:Dimethyl Sulfoxide; D005069:Evaluation Studies as Topic; D005260:Female; D005990:Glycerol; D006085:Graft Survival; D066298:In Vitro Techniques; D007959:Lymphocyte Culture Test, Mixed; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008808:Mice, Inbred CBA; D012867:Skin / Q000276:immunology; D016038:Skin Transplantation / Q000276:immunology*; D013997:Time Factors; D014021:Tissue Preservation / Q000379:methods*; D014184:Transplantation, Homologous
chemical_list: D005990:Glycerol; D004121:Dimethyl Sulfoxide