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2. A Scalable Method to Fabricate 2D Hydrogel Substrates for Mechanobiology Studies with Independent Tuning of Adhesiveness and Stiffness.

Author

Gandin, Alessandro, Torresan, Veronica, Panciera, Tito, and Brusatin, Giovanna

Subjects
MATERIALS science, CYTOLOGY, BIOLOGICAL laboratories, POLYETHYLENE glycol, PEPTIDES
Abstract

Mechanical signals from the extracellular matrix are crucial in guiding cellular behavior. Two-dimensional hydrogel substrates for cell cultures serve as exceptional tools for mechanobiology studies because they mimic the biomechanical and adhesive characteristics of natural environments. However, the interdisciplinary knowledge required to synthetize and manipulate these biomaterials typically restricts their widespread use in biological laboratories, which may not have the material science expertise or specialized instrumentation. To address this, we propose a scalable method that requires minimal setup to produce 2D hydrogel substrates with independent modulation of the rigidity and adhesiveness within the range typical of natural tissues. In this method, norbornene-terminated 8-arm polyethylene glycol is stoichiometrically functionalized with RGD peptides and crosslinked with a di-cysteine terminated peptide via a thiol–ene click reaction. Since the synthesis process significantly influences the final properties of the hydrogels, we provide a detailed description of the chemical procedure to ensure reproducibility and high throughput results. We demonstrate examples of cell mechanosignaling by monitoring the activation state of the mechanoeffector proteins YAP/TAZ. This method effectively dissects the influence of biophysical and adhesive cues on cell behavior. We believe that our procedure will be easily adopted by other cell biology laboratories, improving its accessibility and practical application. [ABSTRACT FROM AUTHOR]

Published
2024
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5. Broadly Applicable Hydrogel Fabrication Procedures Guided by YAP/TAZ‐Activity Reveal Stiffness, Adhesiveness, and Nuclear Projected Area as Checkpoints for Mechanosensing (Adv. Healthcare Mater. 3/2022)

Author

Gandin, Alessandro, primary, Torresan, Veronica, additional, Ulliana, Lorenzo, additional, Panciera, Tito, additional, Contessotto, Paolo, additional, Citron, Anna, additional, Zanconato, Francesca, additional, Cordenonsi, Michelangelo, additional, Piccolo, Stefano, additional, and Brusatin, Giovanna, additional

Published
2022
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6. Broadly Applicable Hydrogel Fabrication Procedures Guided by YAP/TAZ‐Activity Reveal Stiffness, Adhesiveness, and Nuclear Projected Area as Checkpoints for Mechanosensing

Author

Gandin, Alessandro, primary, Torresan, Veronica, additional, Ulliana, Lorenzo, additional, Panciera, Tito, additional, Contessotto, Paolo, additional, Citron, Anna, additional, Zanconato, Francesca, additional, Cordenonsi, Michelangelo, additional, Piccolo, Stefano, additional, and Brusatin, Giovanna, additional

Published
2021
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7. Biocompatible Iron–Boron Nanoparticles Designed for Neutron Capture Therapy Guided by Magnetic Resonance Imaging

Author

Torresan, Veronica, primary, Guadagnini, Andrea, additional, Badocco, Denis, additional, Pastore, Paolo, additional, Muñoz Medina, Guillermo Arturo, additional, Fernàndez van Raap, Marcela B., additional, Postuma, Ian, additional, Bortolussi, Silva, additional, Bekić, Marina, additional, Čolić, Miodrag, additional, Gerosa, Marco, additional, Busato, Alice, additional, Marzola, Pasquina, additional, and Amendola, Vincenzo, additional

Published
2020
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8. 4D Multimodal Nanomedicines Made of Nonequilibrium Au–Fe Alloy Nanoparticles

Author

Torresan, Veronica, primary, Forrer, Daniel, additional, Guadagnini, Andrea, additional, Badocco, Denis, additional, Pastore, Paolo, additional, Casarin, Maurizio, additional, Selloni, Annabella, additional, Coral, Diego, additional, Ceolin, Marcelo, additional, Fernández van Raap, Marcela B., additional, Busato, Alice, additional, Marzola, Pasquina, additional, Spinelli, Antonello E., additional, and Amendola, Vincenzo, additional

Published
2020
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10. Simple yet effective methods to probe hydrogel stiffness for mechanobiology

Author

Alessandro Gandin, Yaswanth Murugesan, Maurizio Ventre, Anna Citron, Stefano Piccolo, Giovanna Brusatin, Tito Panciera, Paolo Contessotto, Lucia Nicola, Giusy Battilana, Lorenzo Ulliana, A. Paolo Netti, Veronica Torresan, Gandin, Alessandro, Murugesan, Yaswanth, Torresan, Veronica, Ulliana, Lorenzo, Citron, Anna, Contessotto, Paolo, Battilana, Giusy, Panciera, Tito, Ventre, Maurizio, Netti, A. Paolo, Nicola, Lucia, Piccolo, Stefano, and Brusatin, Giovanna

Subjects
0303 health sciences, Multidisciplinary, Computer science, Atomic force microscopy, Science, Uniaxial compression, Stiffness, Nanotechnology, 02 engineering and technology, 021001 nanoscience & nanotechnology, Characterization and analytical techniques, Article, 03 medical and health sciences, Mechanobiology, Self-healing hydrogels, medicine, Medicine, Biomaterials - cells, medicine.symptom, Gels and hydrogels, 0210 nano-technology, 030304 developmental biology
Abstract

In spite of tremendous advances made in the comprehension of mechanotransduction, implementation of mechanobiology assays remains challenging for the broad community of cell biologists. Hydrogel substrates with tunable stiffness are essential tool in mechanobiology, allowing to investigate the effects of mechanical signals on cell behavior. A bottleneck that slows down the popularization of hydrogel formulations for mechanobiology is the assessment of their stiffness, typically requiring expensive and sophisticated methodologies in the domain of material science. Here we overcome such barriers offering the reader protocols to set-up and interpret two straightforward, low cost and high-throughput tools to measure hydrogel stiffness: static macroindentation and micropipette aspiration. We advanced on how to build up these tools and on the underlying theoretical modeling. Specifically, we validated our tools by comparing them with leading techniques used for measuring hydrogel stiffness (atomic force microscopy, uniaxial compression and rheometric analysis) with consistent results on PAA hydrogels or their modification. In so doing, we also took advantage of YAP/TAZ nuclear localization as biologically validated and sensitive readers of mechanosensing, all in all presenting a suite of biologically and theoretically proven protocols to be implemented in most biological laboratories to approach mechanobiology.

Published
2021

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