Spatiotemporal Assessment of (Patho)physiological Tissue Remodeling and Mechanics in In Situ Tissue-Engineered Pulmonary Valves in Sheep

The use of acellular synthetic scaffolds to replace diseased heart valves represents a translationally attractive strategy, with proof-of-concept demonstrated in preclinical and clinical studies. Nevertheless, in vivo remodeling remains poorly understood, particularly regarding spatial heterogeneity in tissue formation and scaffold resorption. We hypothesized that these variations are, at least partly, driven by differences in local mechanical loading. This study aimed to map the spatiotemporal expression of matrix and immune cell phenotypes in tissue-engineered heart valves (TEHV) implants and correlate these with mechanical stresses and strains. A retrospective analysis was performed on explanted valves from a 12-month preclinical ovine study, using Raman microspectroscopy and immunohistochemistry. Valve-specific geometry and mechanical properties were incorporated into finite element models to compute local principal stresses and strains. Raman microspectroscopy revealed progressive tissue development in the fibrosa and spongiosa, while the ventricularis region exhibited limited elastic fiber formation. Scaffold degradation was associated with inflammatory cell infiltration in localized areas of foreign body response. One malfunctioning valve, previously diagnosed with regurgitation, displayed distinct matrix composition and strong expression of fibrotic markers, confirmed by immunohistochemistry. Regional stresses and strains exhibited increasing heterogeneity over time, reflecting dynamic remodeling process. Correlation analysis showed that strain (but not stress) positively associated with markers of matrix remodeling and inflammation (e.g. IL-10, CD163, TNF-a), suggesting strain as a key driver of local tissue responses. These findings highlight the regulatory role of mechanical strain in in situ regeneration and support the development of scaffold designs that promote favorable mechanical environments for functional TEHV remodeling.