Lichtresponsive Gele zum Verständnis der Mechanotransduktion bei Herzinfarkten und Muskeldystrophie
Zellbiologie
Zusammenfassung der Projektergebnisse
In a first study, we first used a 2D light-responsive hydrogel system to mechanically actuate C2C12 mouse myoblasts in the absence of any additional differentiation factors. We found that actuation enhances cell proliferation and migration, accompanied by nuclear translocation of myogenic proteins MyoD and Yes-associated protein (YAP). Additionally, deposition of collagen type I and fibronectin is reduced, suggesting an antifibrotic effect. Myoblast proliferation and mechanotransduction was dependent on the actuation time (training schedule), underlining that mechanical stimulation can support myogenesis in vitro. In a second study, we investigated how mechanical actuation with our hydrogel system affects mesenchymal stem cells (MSCs) without the presence of differentiation factors. Both continuous and intermittent actuation of MSCs for 3 days led to a significantly enhanced cell spreading and nuclear translocation of Runt-related transcription factor 2 (Runx2) and YAP, indicating that cyclic mechanical stimulation on a soft, ridged substrate modulates the MSC fate commitment in the direction of osteogenesis and not into myoblasts. Therefore, in followup studies, we continued our studies with myoblasts. In a third study, we observed myotube formation on our hydrogel system by increasing the number of myoblasts, but when we started actuation on these confluent cell layers, we soon identified that the setup was not stable enough to perform the required long-term actuation experiments (>5 days). After a thorough optimization process of both the acrylsilanization and protein functionalization, it was concluded that the setup would not be reliable enough to perform further experiments. Therefore, we decided to develop a 3D light-actuatable hydrogel system, using similar chemistry as the 2D hydrogel film. We hypothesized that a 3D microgel-assembly could both provide sufficient space for the myoblasts to grow, differentiate and interact with each other to form myotubes, while the microgels can be locally actuated with light. This approach was first tested with thermoresponsive microgels produced via an adapted particle replication in non-wetting templates (PRINT) technique we developed in our group. While initial results demonstrated efficient myotube formation in the porous structure in between microgels, we were not able to actuate the microgels efficiently due to the low amount of gold nanorods (AuNRs) we could incorporate inside the microgels. Due to this limitation, we decided to produce the microgels via microfluidics and developed a new method called thermally-assisted microfluidics to produce a large library of thermoresponsive microgels that can be actuated under physiological conditions with different structures, morphologies, stiffness, and steepness of their thermal transition. One of these microgels was used to form a mature monolayer-microgel construct designed to mechanically stimulate cells on a substrate mimicking native tissue properties, such as stiffness and curvature. Currently, we are producing rod-shaped microgels with the same chemical compositions using microfluidics as above and realized that myoblasts can assemble the microgels into a 3D construct without the need to chemically interlink the microgels, as we demonstrated with fibroblasts and spherical microgels before. We are now continuing these studies further to develop a 3D light actuating in vitro gym for muscle cells in health and disease. In conclusion, including cyclic signals to in vitro models reveals the effect of cyclic deformation on both myoblasts and MSCs, and our translation of the chemistry to 3D in the form of microgel spheres and rods provides a new way to incorporate these signals in 3D culture systems.
Projektbezogene Publikationen (Auswahl)
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Cells feel the beat – temporal effect of cyclic mechanical actuation on muscle cells. Applied Materials Today, 27, 101492.
Chandorkar, Yashoda; Bastard, Céline; Di Russo, Jacopo; Haraszti, Tamás & De Laporte, Laura
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Poster: Gordon Research Conference: Multiscale Mechanochemistry and Mechanobiology. July 31-August 5, 2022, Ventura, United States.
Laura De Laporte
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Actuation of Soft Thermoresponsive Hydrogels Mechanically Stimulates Osteogenesis in Human Mesenchymal Stem Cells without Biochemical Factors. ACS Applied Materials & Interfaces, 16(1), 30-43.
Castro Nava, Arturo; Doolaar, Iris C.; Labude-Weber, Norina; Malyaran, Hanna; Babu, Susan; Chandorkar, Yashoda; Di Russo, Jacopo; Neuss, Sabine & De Laporte, Laura
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Thermally Assisted Microfluidics to Produce Chemically Equivalent Microgels with Tunable Network Morphologies. Angewandte Chemie International Edition, 64(1).
Rommel, Dirk; Häßel, Bernhard; Pietryszek, Philip; Mork, Matthias; Jung, Oliver; Emondts, Meike; Norkin, Nikita; Doolaar, Iris Christine; Kittel, Yonca; Yazdani, Ghazaleh; Omidinia‐Anarkoli, Abdolrahman; Schweizerhof, Sjören; Kim, Kyoohyun; Mourran, Ahmed; Möller, Martin; Guck, Jochen & De Laporte, Laura
