PhD Scientific Days 2023

Budapest, 22-23 June 2023

Translational Medicine - Posters P

Promotion of Fibroblast-Myofibroblast Transition Through Modulation of Polyacrilamide Hydrogel Stiffness

F. Palmieri1, M. Parigi2, R. Garella1, A. Tani2, F. Chellini2, A. Cappitti3, M. Salzano4,
D. Martella5, R. Squecco1, C. Sassoli2

1 Department of Experimental and Clinical Medicine, Section of Physiological
Sciences, University of Florence, 50134 Florence, Italy
2 Department of Experimental and Clinical Medicine, Section of Anatomy and Histology, Imaging Platform, University of Florence, 50134 Florence, Italy
3 Department of Chemistry “Ugo Schiff” , University of Florence, 50019 Sesto Fiorentino, Italy
4 Department of Chemical, Materials and Industrial Production Engineering, University of Naples, 80125 Napoli, Italy
5 European Laboratory for Non-Linear Spectroscopy (LENS), 50019 Sesto Fiorentino,
Italy
Istituto Nazionale di Ricerca Metrologica (INRiM), 10135 Torino, Italy

Text of the abstract

Introduction: Mechano-mimetic materials are particularly attractive for providing in vitro models for fibroblast to myofibroblast (Myof) transition, a key process in the physiological repair of damaged tissue and regarded as the “core cellular mechanism” of pathological fibrosis. In vivo mechanical and chemical stimuli from extracellular matrix (ECM) and profibrotic factors such as transforming growth factor (TGF)-β1 respectively, are determinant in promoting fibroblast differentiation.
Aim: In this in vitro study we explore the use of polymeric hydrogels as mechano-mimetic scaffold in comparison to conventional glass supports for fibroblast cell culture, that do not account for the tissue specific ECM mechanics.
Methods: Different mechano-mimetic substrates of polyacrylamide (PA) gels have been prepared by free radical polymerization, with an accurate tailoring of water content and crosslinking degree, to modulate the mechanical properties. The resulting stiffness ranged from 29-1 kPa. Glass coverslips coated with PA hydrogels were used to culture murine NIH-3T3 fibroblasts in the absence or presence of TGF‐β1. After 48h of culture, the cells were subjected to morphological and functional analyses, to estimate the acquisition of a differentiated Myof phenotype. In particular, cell shape and area, stress fiber assembly, α-smooth muscle actin expression were assessed by confocal laser scanning microscopy whereas cell membrane passive properties and ion currents by whole cell patch-clamp.
Results: A major finding obtained from our experiments is that soft materials (Young modulus<1 kPa) triggered an efficient mechanical cue to induce fibroblast differentiation, better than stiffer ones, even in the absence of TGF-β1, potentially mimicking the mechanical characteristics of ECM soon after wounding in vivo.
Conclusion: These findings may open the way to high-throughput in vitro Myof models with the great advantage of using biomaterials reproducible on a large scale. In addition, since the acrylamide monomers are commercially available and the technical approach used to create the scaffolds is easy, these hydrogels could be suitable to a wide range of biomedical applications.
Funding: Supported by the annual financing fund by MIUR (Ministry of Education, University and Research, Italy), University of Florence ROBERTASQUECCORICATEN23.