title: The "in situ" expression of human leukocyte antigen class I antigens is not altered by cryopreservation in human arterial allografts.
abstract: This study was aimed to establish whether the cryopreservation procedure we currently use in clinics can modify arterial homograft antigenicity. To this purpose, we performed an immunohistochemical study on fresh and cryopreserved human arterial homografts to visualize the expression of HLA class I heavy and light chains "in situ" by using the HC-10 and Namb-1 monoclonal antibodies. Human femoral arteries and thoracic aortas were harvested from 18 heart-beating donors and sampled before and after cryopreservation. Arterial segments were frozen in liquid nitrogen vapors in a controlled rate freezing system. After thawing, samples were processed for routine immunohistochemistry. To standardize immunostaining, flow-cytometry indirect immunofluorescence analysis was performed on HUVEC; immunohistochemistry of human ovarian cortical vessels was performed as an additional positive control. Negative controls were performed by omitting tissue incubation with primary antibodies. HLA-class I antigens were markedly expressed by endothelial cells lining surface intima and adventitial vasa vasorum; a moderate expression was found in medial smooth muscle cells. Except for the surface unreactivity caused by loss of endothelium, results from cryopreserved arterial allografts were strictly comparable to those observed in fresh, unfrozen tissues. These results support the view that cryopreserved arterial allografts are immunogenic as their fresh counterparts; apart from smooth muscle cells which retained a moderate expression of HLA class I antigens following cryopreservation, our study suggests that the highly HC-10 positive endothelial cells we found to line the rich adventitial network of vasa vasorum are expected to be one of the major targets of the serological response in the recipient.
authors.affiliation: Clinical Pathology Unit, Department of Experimental Pathology, Policlinico S. Orsola, University of Bologna, Blg 11, via Massarenti 9, 40138, Bologna, Italy. gianandr.pasquinelli@unibo.it
authors.forename: G
authors.initials: G
authors.lastname: Pasquinelli
authors.forename: M P
authors.initials: MP
authors.lastname: Pistillo
authors.forename: F
authors.initials: F
authors.lastname: Ricci
authors.forename: M
authors.initials: M
authors.lastname: Buzzi
authors.forename: P L
authors.initials: PL
authors.lastname: Tazzari
authors.forename: L
authors.initials: L
authors.lastname: Foroni
authors.forename: C
authors.initials: C
authors.lastname: Manferdini
authors.forename: C
authors.initials: C
authors.lastname: Ceccarelli
authors.forename: A
authors.initials: A
authors.lastname: Stella
authors.forename: R
authors.initials: R
authors.lastname: Conte
mesh_terms: D000328:Adult; D000911:Antibodies, Monoclonal / Q000276:immunology; D001158:Arteries / Q000276:immunology* / Q000637:transplantation*; D015925:Cryopreservation; D005260:Female; D005434:Flow Cytometry; D006680:HLA Antigens / Q000276:immunology*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D014184:Transplantation, Homologous
chemical_list: D000911:Antibodies, Monoclonal; D006680:HLA Antigens