Publications

Cardiac fibrosis occurs following insults to the myocardium and is characterized by the abnormal accumulation of non-compliant extracellular matrix (ECM), which compromises cardiomyocyte contractile activity and eventually leads to heart failure. This phenomenon is driven by the activation of cardiac fibroblasts (cFbs) to myofibroblasts and results in changes in ECM biochemical, structural and mechanical properties. The lack of predictive in vitro models of heart fibrosis has so far hampered the search for innovative treatments, as most of the cellular-based in vitro reductionist models do not take into account the leading role of ECM cues in driving the progression of the pathology. Here, we devised a single-step decellularization protocol to obtain and thoroughly characterize the biochemical and micro-mechanical properties of the ECM secreted by activated cFbs differentiated from human induced pluripotent stem cells (iPSCs). We activated iPSC-derived cFbs to the myofibroblast phenotype by tuning basic fibroblast growth factor (bFGF) and transforming growth factor beta 1 (TGF-beta 1) signalling and confirmed that activated cells acquired key features of myofibroblast phenotype, like SMAD2/3 nuclear shuttling, the formation of aligned alpha-smooth muscle actin (alpha- SMA)-rich stress fibres and increased focal adhesions (FAs) assembly. Next, we used Mass Spectrometry, nanoindentation, scanning electron and confocal microscopy to unveil the characteristic composition and the visco-elastic properties of the abundant, collagen-rich ECM deposited by cardiac myofibroblasts in vitro. Finally, we demonstrated that the fibrotic ECM activates mechanosensitive pathways in iPSC-derived cardiomyocytes, impacting on their shape, sarcomere assembly, phenotype, and calcium handling properties. We thus propose human bio-inspired decellularized matrices as animal-free, isogenic cardiomyocyte culture substrates recapitulating key pathophysiological changes occurring at the cellular level during cardiac fibrosis.

JTD Keywords: Adhesio, Cardiac fibrosis modelling, Decellularized extracellular matrix, Differentiation, Expression, Fibroblast activation, Fibronectin, Heart, Induced pluripotent stem cells, Ipsc-derived-cardiac fibroblasts, Ipsc-derived-cardiomyocyte, Myocardial-infarction, Neonatal cardiomyocytes, Smooth muscle actin, Substrate stiffness

This work proposes a microfibers-hydrogel assembled composite as delivery vehicle able to combine into a single system both burst and prolonged release of lactate. The prolonged release of lactate has been achieved by electrospinning a mixture of polylactic acid and proteinase K (26.0 mg of proteinase K and 0.99 g of PLA dissolved in 6 mL of 2:1 chloroform:acetone in the optimal case), which is a protease that catalyzes the degradation of polylactic acid into lactate. The degradation of microfibers into lactate reflects that proteinase K preserves its enzymatic activity even after the electrospinning process because of the mild operational conditions used. Besides, burst release is obtained from the lactate-loaded alginate hydrogel. The successful assembly between the lactate-loaded hydrogel and the polylactic acid/proteinase K fibers has been favored by applying a low-pressure (0.3 mbar at 300 W) oxygen plasma treatment, which transforms hydrophobic fibers into hydrophilic while the enzymatic activity is still maintained. The composite displays both fast (< 24 h) and sustained (> 10 days) lactate release, and allows the modulation of the release by adjusting either the amount of loaded lactate or the amount of active enzyme.Copyright © 2023. Published by Elsevier B.V.

JTD Keywords: biodegradable fibers, proteases, regeneration, repair, Alginate, Alginates, Biodegradable fibers, Endopeptidase k, Hydrogels, Lactic acid, Myocardial-infarction, Polymers, Proteases

The term 'mechanosensation' describes the capacity of cells to translate mechanical stimuli into the coordinated regulation of intracellular signals, cellular function, gene expression and epigenetic programming. This capacity is related not only to the sensitivity of the cells to tissue motion, but also to the decryption of tissue geometric arrangement and mechanical properties. The cardiac stroma, composed of fibroblasts, has been historically considered a mechanically passive component of the heart. However, the latest research suggests that the mechanical functions of these cells are an active and necessary component of the developmental biology programme of the heart that is involved in myocardial growth and homeostasis, and a crucial determinant of cardiac repair and disease. In this Review, we discuss the general concept of cell mechanosensation and force generation as potent regulators in heart development and pathology, and describe the integration of mechanical and biohumoral pathways predisposing the heart to fibrosis and failure. Next, we address the use of 3D culture systems to integrate tissue mechanics to mimic cardiac remodelling. Finally, we highlight the potential of mechanotherapeutic strategies, including pharmacological treatment and device-mediated left ventricular unloading, to reverse remodelling in the failing heart.© 2022. Springer Nature Limited.

JTD Keywords: cardiomyocyte proliferation, cross-linking, extracellular-matrix, focal adhesions, gene-expression, mechanical regulation, myocardial-infarction, substrate stiffness affects, t-cells, Ventricular assist device