title: Xeno-immunogenicity of ice-free cryopreserved porcine leaflets.
abstract: BACKGROUND
  Undesirable processes of inflammation, calcification, or immune-mediated reactions are limiting factors in long-term survival of heart valves in patients. In this study, we target the modulatory effects of ice-free cryopreservation (IFC) of xenogeneic heart valve leaflet matrices, without decellularization, on the adaptive human immune responses in vitro.
  
  
  METHODS
  We tested porcine leaflet matrices from fresh untreated, conventionally cryopreserved (CFC), and IFC pulmonary valves by culturing them with human blood mononuclear cells for 5 d in vitro. No other tissue treatment protocols to modify possible immune responses were used. Matrices alone or in addition with a low-dose second stimulus were analyzed for induction of proliferation and cytokine release by flow cytometry-based techniques. Evaluation of the α-Gal epitope expression was performed by immunohistochemistry with fluorochrome-labeled B4 isolectin.
  
  
  RESULTS
  None of the tested leaflet treatment groups directly triggered the proliferation of immune cells. But when tested in combination with a second trigger by anti-CD3, IFC valves showed significantly reduced proliferation of T cells, especially effector memory T cells, in comparison with fresh or CFC tissue. Moreover, the cytokine levels for interferon-γ (IFNγ), tumor necrosis factor α, and interleukin-10 were reduced for the IFC-treated group being significantly different compared with the CFC group. However, no difference between treatment groups in the expression of the α-Gal antigen was observed.
  
  
  CONCLUSIONS
  IFC of xenogeneic tissue might be an appropriate treatment method or processing step to prevent responses of the adaptive immune system.
authors.affiliation: Institute of Medical Immunology, Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany. Electronic address: martina.seifert@charite.de.
authors.forename: Martina
authors.initials: M
authors.lastname: Seifert
authors.affiliation: Institute of Medical Immunology, Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Bayrak
authors.affiliation: Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany.
authors.forename: Meaghan
authors.initials: M
authors.lastname: Stolk
authors.affiliation: Institute of Medical Immunology, Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany.
authors.forename: Naima
authors.initials: N
authors.lastname: Souidi
authors.affiliation: Institute of Medical Immunology, Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Schneider
authors.affiliation: Department for Thoracic and Cardiovascular Surgery, Johann Wolfgang Goethe University, Frankfurt am Main, Germany.
authors.forename: Ulrich A
authors.initials: UA
authors.lastname: Stock
authors.affiliation: Cell and Tissue Systems, North Charleston, South Carolina; Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, South Carolina; Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, Georgia.
authors.forename: Kelvin G M
authors.initials: KG
authors.lastname: Brockbank
mesh_terms: D000818:Animals; D016207:Cytokines / Q000378:metabolism; D000939:Epitopes / Q000378:metabolism; D006351:Heart Valves / Q000276:immunology / Q000637:transplantation*; D064593:Heterografts / Q000276:immunology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000502:physiology; D011897:Random Allocation; D013552:Swine; D014181:Transplantation Immunology; D014183:Transplantation, Heterologous
chemical_list: D016207:Cytokines; D000939:Epitopes
keywords: Cryopreservation; Cytokine; Heart valve; Immune response; Inflammation