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In Vitro Modulation Of Human Immune Responses Of Cryopreserved Aorta And Heart Valves By Vitrification Treatment
Benjamin A. Hoegerle1, Martina Seifert2, Karin Sudrow2, Isabella Werner3, Nadija Souidi2, Meagan Stolk2, Kelvin GM Brockbank4, Andres Beiras1, Anton Moritz1,
Ulrich A. Stock1.
1University Hospital Frankfurt, Frankfurt am Main, Germany, 2Charité Universitätsmedizin Berlin, Berlin, Germany, 3University Hospital Frankfurt, Frankfurt, Germany, 4Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC, USA.

Introduction: Current challenge of allogenic heart valves is minimizing inflammation and rejection. The present in vitro study analyzed whether treatment of cryopreserved human aortic valves by vitrification solution modulates immune responses. Methods: 5 human aortic valves were frozen and thawed according to protocols of conventional cryopreservation (CFC). Additionally tissue was thawed and treated by vitrification solution (CFC+V). Tissue punches (n=6) were co-cultured for 7 days with human peripheral blood mononuclear cells (PBMC) to quantify induced proliferation by flow cytometry. Co-cultures with sorted CD14+ monocytes were analyzed for activation, differentiation and polarization into macrophages. Release of cytokines by cytometric bead array was determined. Results: In all matrix/PBMC settings we found neither induction of immune cell proliferation nor release of all tested pro-inflammatory cytokines compared to controls with mitogen-activate cells. Co-cultures with CD14+ monocytes showed marginal increase in proportion of CD206+ cells for both groups, however no increase regarding fraction of CCR7 expressing cells. Percentage was higher only in co-cultures with CFC+V aorta but not in valves. In all co-cultures IL-6 was exclusively detected, showing reduced levels only in aortic co-cultures of CFC+V aortic tissue compared to CFC. Conclusion: Our data emphasize potential of vitrification to modify immunogenic properties of allogenic aortic and heart valve tissue. Increase of proportion of macrophages with a more anti-inflammatory phenotype and decrease of IL-6 levels for aortic tissue by vitrification treatment might offer modulation of immune responses in general. Vitrification is easy, fast, and safe preserving biological tissues and promoting better clinical graft performance.

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