Enzymatic extracellular matrix remodeling in ex vivo slices of aged myocardium

Cardiac aging is accompanied by extracellular matrix (ECM) remodeling and increased myocardial stiffness, which may contribute to age-associated functional decline. However, whether direct remodeling of the aged cardiac ECM can modify myocardial mechanics and function remains poorly understood. Here, we established an ex vivo living myocardial slice platform to investigate whether enzymatic remodeling of the aged ECM can alter myocardial structure, mechanics, and function. Left ventricular myocardial slices (LMS) from young and aged mice were maintained under static stretch for 48 hours using a custom 3D-printed culture setup. Aged slices were treated with collagenase and subsequently characterized using quantitative imaging, tensile testing, contractility measurements, and transcriptomic analysis. Aged myocardium exhibited increased collagen and fibronectin abundance, altered collagen network architecture, and increased stiffness compared with young tissue. Collagenase treatment reduced collagen and network complexity and reduced myocardial stiffness toward young tissue values without compromising ultimate tensile stress. Despite these structural and biomechanical changes, age-associated differences in myocardial contractility remained comparable across groups. Transcriptomic analysis indicated that treated aged tissue largely retained its age-associated molecular phenotype, despite pathway-level shifts involving metabolism, cardiac function, and inflammatory responses. These findings demonstrate that enzymatic ECM remodeling can modify the aged myocardial mechanical environment while selected functional and transcriptional features of aging persist over 48 hours. The LMS platform provides a controllable ex vivo tissue-level approach for investigating ECM contributions during cardiac aging and evaluating strategies for ECM engineering. Statement of significance Cardiac aging is accompanied by progressive extracellular matrix (ECM) remodeling and tissue stiffening, which are associated with impaired cardiac function. However, studying how these age-related changes influence living myocardium requires models that balance physiological complexity and experimental control. This study combines living myocardial slice culture with controlled enzymatic ECM remodeling to examine aged tissue responses within the native myocardial environment. Collagenase treatment reduced tensile modulus and altered collagen network organization, while selected functional and transcriptional features of aging persisted over 48 hours. These findings demonstrate the utility of living myocardial slices for evaluating matrix-targeted interventions and highlight the importance of assessing biological responses alongside changes in ECM structure and tissue mechanics.