**3. Tissue regeneration**

TEHV is not the sole approach investigated to obtain new viable valve devices: tissue-guided regeneration has been proposed as an alternative method for *in vivo* direct tissue reconstruc‐ tion, by exploiting ECM instructive abilities. Once eliminated the allogeneic or xenogeneic cell component through decellularizing treatment, the fibre mesh still maintains biomechanical proficiency assuring in vivo prompt restoration of hydrodynamic performance. Furthermore, in the body, conceived as physiological bioreactor, naked natural scaffolds recruit recipient's cells thanks to their chemo-attractant properties. Positive aspects associated with this option should be identified in the possibility to construct autologous-like tissues, by skipping difficultly controllable procedures of cell seeding and chemo-mechanical stimulation *in vitro*.

Among the first experimental evidences, biomaterials, as patches of pure type I collagen, have been successfully introduced in the therapy of ischemic myocardium: once applied to the diseased tissue, the collagen sponge attracts progenitors and less undifferentiated cells, which in turn or alone are able to fully colonize it and start a cardiovascular transdifferentiation [85]. It is noteworthy to remember that these patches, either synthetic or cell-purified from biolog‐ ical tissues, have found FDA-approved applications as haemostatics or for skin reconstruction with excellent results [86].

A further surprising element for a positive consideration of this method has been given by Campbell and colleagues, who were able to obtain a tubular cell construct by implanting a polymeric tube in the animal peritoneal cavity. The newly formed tissue, pulled from the tube, had anatomical and histological resemblance to a quite mature blood vessel and it could be hence considered an optimal vascular substitute [87].

Elastomeric poly(glycerol sebacate) scaffolds treated with multiple coating strategies based on

In respect to the application of bioabsorbable polymers, the other TEHV modality, founded on natural ECMs, was experimented some years later. After the development of various decellularizing treatments, the combination with differentiated cells, as ECs and VICs, was able to generate directly *in vitro* by static conditioning surrogates of early heart valve tissues [24, 81]. As well as polymeric TEHVs, cell-repopulated decellularized ECMs were positively remodelled after dynamic stimulation with proper mechanical signals. In this case, actually,

The usage of stem cells as cell source for the engineering of plain ECMs led to even better *in vitro* outcomes. Multipotent differentiation potential of human bone marrow MSCs can represent the ideal characteristic for complete repopulation of natural valve matrices. MSC engrafting ability was evaluated on decellularized porcine and human scaffolds. In both considered interactions, stem cells were able to adhere, spread within the ECM and transdif‐ ferentiate towards typical valve phenotypes (ECs, VICs). Collagen, GAG and elastin synthesis was indeed activated in engrafted cells, which tend to distribute similarly to the original valve cell topography. It was, however, the homotypic combination to better favour MSC-to-SMC

TEHV is not the sole approach investigated to obtain new viable valve devices: tissue-guided regeneration has been proposed as an alternative method for *in vivo* direct tissue reconstruc‐ tion, by exploiting ECM instructive abilities. Once eliminated the allogeneic or xenogeneic cell component through decellularizing treatment, the fibre mesh still maintains biomechanical proficiency assuring in vivo prompt restoration of hydrodynamic performance. Furthermore, in the body, conceived as physiological bioreactor, naked natural scaffolds recruit recipient's cells thanks to their chemo-attractant properties. Positive aspects associated with this option should be identified in the possibility to construct autologous-like tissues, by skipping difficultly controllable procedures of cell seeding and chemo-mechanical stimulation *in vitro*.

Among the first experimental evidences, biomaterials, as patches of pure type I collagen, have been successfully introduced in the therapy of ischemic myocardium: once applied to the diseased tissue, the collagen sponge attracts progenitors and less undifferentiated cells, which in turn or alone are able to fully colonize it and start a cardiovascular transdifferentiation [85]. It is noteworthy to remember that these patches, either synthetic or cell-purified from biolog‐ ical tissues, have found FDA-approved applications as haemostatics or for skin reconstruction

A further surprising element for a positive consideration of this method has been given by Campbell and colleagues, who were able to obtain a tubular cell construct by implanting a polymeric tube in the animal peritoneal cavity. The newly formed tissue, pulled from the tube,

ECM-derived proteins allowed adhesion and transdifferentiation of EPCs [61, 80].

also elastin content was demonstrated to increase [82-84].

conversion in the *ventricularis* layer [25].

**3. Tissue regeneration**

254 Calcific Aortic Valve Disease

with excellent results [86].

Also offering the opportunity to create tissue banks for ready-to-use devices at the moment of clinical need, the investigation on tissue-guided regenerated heart valves (TGRHVs) has particularly increased in recent years.
