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• W45361012 abstract "This doctoral thesis focuses on the approaches currently in use to develop new vital valve devices: heart valve tissue engineering and heart valve tissue guided regeneration. Indeed, it investigates some aspects able to modify the preservation of the implanted construct, i.e. the immunogenic properties of the extracellular matrix and the biological entity of stem cell populations in heart valve leaflets.The porcine semilunar valves, as the bovine pericardial derived ones, find large clinical usage for the substitution of dysfunctional valves. The rational of their employ is the high morphological and functional analogies with the native human ones and hence they can provide a valid extracellular matrix (ECM) for bioengineered prosthesis. Once eliminated the xenogeneic cell component through a decellularizingtreatment, it is possible to benefit from the extracellular fiber mesh as a template for repopulation with human stem cells in order to develop autologous-like replacements. So far the influence on cellular attachment exerted by the anisotropic ECM distribution in fibrosa and ventricularis has never beeninvestigated. Porcine pulmonary leaflets were decellularized with a Triton X110/Sodium Cholate-based protocol. The absence of alpha-Gal epitopes, highly immunogenic for the human species, has been demonstrated through a double fluorescence technique using an isolectin and a specific monoclonal antibody. After treatment with fetal bovine serum and fibronectin to increase adhesion, human bone marrow mesenchymal stem cells have been seeded either onto the ventricularis or fibrosa and statically maintained in culture for 30 days. Not only ventricularis is able to consent a higher attachment, but also increased spreading and early cell differentiation in the cusp stroma. The interaction between the same cells and decellularized human leaflets has been further evaluated by ventricularis seeding. The homologous combination favors proliferation, with decrease of apoptotic events and enhanced cell maturation level, so that it is possible to appreciate the expression of typical mature smooth muscle markers in ventricularis. ECM performs consequently an essential role for cell integration by furnishing specific signals for the acquisition of the correct valvular phenotype.A further hypothesis for the achievement of new vital substitutes is likely represented by tissue guided regeneration operated in vivo by ECM in respect to the recipient's cells. Decellularized aortic roots were used to mimic a reconstruction of the RVOT in minipigs and performances were followed echocardiographically for 12-14 months, revealing a progressive function improvement. A continuous engraftment is observed in the explanted specimens, even if the layers less exposed to blood circulation are still devoid of cells. The observations on primary cultures of the vessel and leaflet tissues lead to hypothesize the contribution of two main phenotypes to repopulation: pulmonary artery smooth musclecells and mobilized mesenchymal stem cells.Particularly important is the preservation of the bioconstruct once the function has been rescued, but at the same time it is essential to understand the key cell effectors participating to valve dysfunction in order to prevent the phenomenon. Scarcely studied, possible stem cell populations in the valves could be involved in the homeostatic tissue remodeling or in adverse pathophysiological events. Aortic roots (n=27) with related mitral leaflets (n=27) have been classified in 5 groups depending on allograft donor's age (10- 60 years). In addition, aortic and mitral pathological leaflets (n=10) were harvested during valve replacements. Cusp cryosections have been analysed through classical histology, Mallory's trichrome, VonKossa, Oil Red O and immunohistochemistry for differentiated, inflammatory, calcifying, different lineagederived stem cell markers. With aging the typical observed feature is the accumulation of lipids, both as small droplets in the subventricularis layer or as cholesterol crystals in the fibrosa. Calcifications appear rare. Stem cell epitopes are highly expressed in the leaflet to reveal a specific spatial distribution. Primary cultures obtained from these specimens showed a phenotype and a differentiation potentiality, suggestiveof mesenchymal stem progenitors without apparent osteogenic induction. Further studies are needed to better understand the contribution of these cells in heart valve calcification.In conclusion, both proposed modalities for the achievement of new valve substitutes appear valid instruments for the aim, because relying on fully decellularized alpha-Gal negative matrices permissive of cell-repopulation. Future efforts and application of new biomimetic strategies would surely implement the encouraging results here demonstrated. New insights on stem cell valvular biology have been proposed here for the first time with important confirmations on the continuous cell remodeling interesting the cusp tissue." @default.
• W45361012 created "2016-06-24" @default.
• W45361012 creator A5072593015 @default.
• W45361012 date "2010-03-12" @default.
• W45361012 modified "2023-09-27" @default.
• W45361012 title "From the anatomical study to the application of different bioengineering techniques for the creation of new vital heart valve substitutes" @default.
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