Immune and non-immune factors in cryopreserved tissues

Background. Although cryopreserved tissues are used clinically, the effects of cryopreservation on antigenicity leading to immune and non-immune responses are not well known. Methods. We investigated the change of inflammatory effects of cryopreserved tissue by using spleen and aortic allografts from Class I antigen-disparate B6.C-H-2(bm1) (bm 1; K-bm1, IA(b), D-b), Class II antigen-disparate B6.C-H-2(bm12) (bm12; K-b, IA(bm12), D-b) and Class I and Class II antigen-disparate (bm1 x bm12)F1 (K-bm1 (x) (b), IA(b) (x) (bm12), D-b) mice against C57BL/6 Cr Slc (B6; H-2(b)) mice. Cryopreservation was done in a programmed freezer and cryopreserved tissues were kept in the vapor phase of liquid nitrogen for 2 weeks and thawed at room temperature. Results. Cryopreserved B6 spleen cells expressed, almost the same levels of Class I (K-b and D-b) and Class II (IA(b)) antigens as observed in fresh B6 spleen cells. Cryopreserved bm1 and bm12 spleen cells had the same stimulator activities in mixed-lymphocyte reaction (MLR) and cytotoxic T-lymphocyte (CTL) assays compared with fresh bm1 and bm12 spleen cells, respectively. To elucidate the effects of cryopreserved tissues on immune response of recipients, descending aortas of (bm1 x bm12)F1 mice were implanted into the right common carotid artery of B6 (H-2(b)) mice with the cuff technique and the reactivities of recipient B6 mice against Class I antigen-disparate bm1 antigens and Class II antigen-disparate bm12 antigens were examined 4 weeks after implantation. In both MLR and CTL assays against bm1 or bm12 antigens, anti-donor reactivities were augmented and there was no significant difference between B6 mice grafted with fresh aortic allografts and those grafted with cryopreserved ones. Histologic analysis showed that mild infiltration of mononuclear cells into the adventitia was observed in both fresh and cryopreserved aortic allografts. The fibrous change was observed more strongly in cryopreserved aortic allografts compared with fresh aortic allografts. Conclusions. Cryopreservation has no effect on eliciting immune responses to Class I or Class II alloantigens, but has some effect on promoting fibrous change.