Periodic Reporting for period 2 - MECHANO FIBROSIS (Regulation of mechanotransduction through motor-molecules activation of focal adhesion kinase in progressive fibrosis)
Okres sprawozdawczy: 2023-01-01 do 2023-12-31
Mechanical forces arising from cell-ECM interactions are key regulators of tissue development, health, and fibrotic disease progression.
Tissue fibrosis contributes to 45% of all deaths in the developed world and is characterized by progressive extracellular matrix (ECM) stiffening and altered patterns of cell adhesion signalling (i.e. focal adhesion kinase, FAK). Yet, studying dynamic ECM-Cell biophysical interactions with static structures, biochemically centric methodologies, and incomplete ECM-Cell descriptions presents an unprecedented challenge in tissue engineering and regenerative medicine (TERM). Focal adhesions (FAs), nanoscale complexes of structural and signalling molecules that link the extracellular matrix (ECM) to the cytoskeleton through integrin receptors, function as principal sites of mechanotransduction. Studies with contractility inhibitors, deformable substrates, and laser tweezers have established that force regulates FA assembly and signalling and identified key molecules in these mechanoresponses, yet very little is known about how forces are integrated into biochemical signalling. Most of our understanding of cell-ECM interactions comes from either cell population-based or ‘whole’ cell assays on static substrates of varying stiffness where the cell is viewed as in a ‘uniform’ stress state. These analyses provide only ‘averaged’ metrics that do not capture/reveal important relationships at the cell-ECM interface due to the heterogeneity of individual subcellular components. With this project, the fellow will decisively overcome the technical limitations of studying cell-ECM dynamics in fibrotic microenvironments by developing engineering tools and investigative in vivo models to examine the role of focal adhesion mechanobiology in tissue homeostasis and fibrosis disease progression.
Conclusions of the Action:
The MECHANO FIBROSIS project successfully developed and implemented a novel engineering tool, capable of applying forces to molecular bonds at the cell-ECM interface, demonstrating its utility in studying mechanotransduction at the molecular level.
The project's findings have significantly advanced the understanding of cell-ECM interactions in healthy (soft) and fibrotic (stiff) environments, highlighting the importance of mechanical forces in cellular signaling and disease progression.
The outcomes of this research have potential implications for developing new diagnostic and therapeutic tools for fibrotic diseases, thereby addressing a critical societal health issue.
Tissue fibrosis contributes to 45% of all deaths in the developed world and is characterized by progressive extracellular matrix (ECM) stiffening and altered patterns of cell adhesion signalling (i.e. focal adhesion kinase, FAK). Yet, studying dynamic ECM-Cell biophysical interactions with static structures, biochemically centric methodologies, and incomplete ECM-Cell descriptions presents an unprecedented challenge in tissue engineering and regenerative medicine (TERM). Focal adhesions (FAs), nanoscale complexes of structural and signalling molecules that link the extracellular matrix (ECM) to the cytoskeleton through integrin receptors, function as principal sites of mechanotransduction. Studies with contractility inhibitors, deformable substrates, and laser tweezers have established that force regulates FA assembly and signalling and identified key molecules in these mechanoresponses, yet very little is known about how forces are integrated into biochemical signalling. Most of our understanding of cell-ECM interactions comes from either cell population-based or ‘whole’ cell assays on static substrates of varying stiffness where the cell is viewed as in a ‘uniform’ stress state. These analyses provide only ‘averaged’ metrics that do not capture/reveal important relationships at the cell-ECM interface due to the heterogeneity of individual subcellular components. With this project, the fellow will decisively overcome the technical limitations of studying cell-ECM dynamics in fibrotic microenvironments by developing engineering tools and investigative in vivo models to examine the role of focal adhesion mechanobiology in tissue homeostasis and fibrosis disease progression.
Conclusions of the Action:
The MECHANO FIBROSIS project successfully developed and implemented a novel engineering tool, capable of applying forces to molecular bonds at the cell-ECM interface, demonstrating its utility in studying mechanotransduction at the molecular level.
The project's findings have significantly advanced the understanding of cell-ECM interactions in healthy (soft) and fibrotic (stiff) environments, highlighting the importance of mechanical forces in cellular signaling and disease progression.
The outcomes of this research have potential implications for developing new diagnostic and therapeutic tools for fibrotic diseases, thereby addressing a critical societal health issue.
The MECHANO FIBROSIS project culminated with significant insights into the mechanobiology of fibrosis. The project's results, both in vitro and in vivo, have been critical in unraveling the complex cellular responses in fibrotic environments.
During its outgoing phase, the project developed an in vitro model revealing the regulatory coupling of force and focal adhesion kinase (FAK) signaling at focal adhesions (FAs). This breakthrough demonstrated the linear relationship between traction force and FAK phosphorylation at individual FAs in fibrotic/stiff environments. These findings were crucial in understanding the coordinated cellular responses to external forces, such as cell migration and tissue-scale force coordination, critical in tissue stiffness and fibrosis progression. Finally, an in vivo model further illuminated how cell adhesive forces regulate immune cell function during tissue repair processes. Impaired tissue revascularization and aberrant immune cell migration are hallmarks of tissue fibrosis progression. By using the dorsal skinfold window chamber model and intravital imaging, we assessed the impact of cell adhesive forces on immune cell migration and tissue revascularization, underscoring the complexity of fibrotic tissue dynamics.
In the final phase, the project optimized a disease model that more accurately replicates the fibrotic environment. The novel molecular tools and biosensors developed enabled a deeper understanding of the cellular mechanotransduction mechanisms involved in fibrotic disease progression.
Exploitation, and Dissemination:
The project's achievements were widely disseminated through high-impact journals(i.e. Nat Comm, Advanced materials and Science Advances), international conferences (Society for Biomaterials (SFB), European Society for Biomaterials (ESB) and GRC Fibronectin, integrins and other related molecules), and biomaterials workshops (INM, Germany and NCSU, USA), ensuring knowledge transfer within the scientific community to extend the international network of the MECHANO FIBROSIS project. Particularly, in the final phase the integration of FRET biosensor technology and cell engineering techniques in collaborating labs at the host institution underscores the project's significant impact on the broader research field.
Finally, the findings have opened avenues for novel therapeutic strategies targeting mechanotransduction pathways in fibrotic diseases. The developed tools can be incorporated into a biomaterial (hydrogel) offering unique properties and potential for use as anti-fibrotic therapeutic device or diagnostic tools.
In conclusion, the MECHANO FIBROSIS project not only achieved its scientific objectives but also laid down a foundation for future therapeutic developments and opened new horizons in the field of mechanobiology and tissue engineering
During its outgoing phase, the project developed an in vitro model revealing the regulatory coupling of force and focal adhesion kinase (FAK) signaling at focal adhesions (FAs). This breakthrough demonstrated the linear relationship between traction force and FAK phosphorylation at individual FAs in fibrotic/stiff environments. These findings were crucial in understanding the coordinated cellular responses to external forces, such as cell migration and tissue-scale force coordination, critical in tissue stiffness and fibrosis progression. Finally, an in vivo model further illuminated how cell adhesive forces regulate immune cell function during tissue repair processes. Impaired tissue revascularization and aberrant immune cell migration are hallmarks of tissue fibrosis progression. By using the dorsal skinfold window chamber model and intravital imaging, we assessed the impact of cell adhesive forces on immune cell migration and tissue revascularization, underscoring the complexity of fibrotic tissue dynamics.
In the final phase, the project optimized a disease model that more accurately replicates the fibrotic environment. The novel molecular tools and biosensors developed enabled a deeper understanding of the cellular mechanotransduction mechanisms involved in fibrotic disease progression.
Exploitation, and Dissemination:
The project's achievements were widely disseminated through high-impact journals(i.e. Nat Comm, Advanced materials and Science Advances), international conferences (Society for Biomaterials (SFB), European Society for Biomaterials (ESB) and GRC Fibronectin, integrins and other related molecules), and biomaterials workshops (INM, Germany and NCSU, USA), ensuring knowledge transfer within the scientific community to extend the international network of the MECHANO FIBROSIS project. Particularly, in the final phase the integration of FRET biosensor technology and cell engineering techniques in collaborating labs at the host institution underscores the project's significant impact on the broader research field.
Finally, the findings have opened avenues for novel therapeutic strategies targeting mechanotransduction pathways in fibrotic diseases. The developed tools can be incorporated into a biomaterial (hydrogel) offering unique properties and potential for use as anti-fibrotic therapeutic device or diagnostic tools.
In conclusion, the MECHANO FIBROSIS project not only achieved its scientific objectives but also laid down a foundation for future therapeutic developments and opened new horizons in the field of mechanobiology and tissue engineering
The project has made substantial progress in understanding the complex dynamics of cell-biomaterial interactions, especially in the context of fibrotic diseases.
Outgoing phase: Provided vital insights into how focal adhesions (FAs) convert mechanical forces into focal adhesion kinase (FAK) signaling events that regulate cell migration and influence tissue outcomes.
The project explored the complex dynamics between biomaterials, immune cells, and fibrogenic cells. Advanced techniques like intravital imaging and computational flow cytometry revealed a previously unreported heterogeneity in biomaterial-instructed immune cells, challenging traditional views of their homogeneity. These findings are critical for developing new biomaterial-based strategies in regenerative medicine and for treating inflammatory diseases.
Final phase: Optimized a mechanically active platform to apply mechanical forces at the cell-ECM molecular interface.
A platform was optimised to precisely apply mechanical forces at the molecular cell-ECM interface. This led to a deeper understanding of mechanotransduction, particularly how molecular motors can manipulate forces at a molecular scale, affecting key mechanoresponsive molecules like Talin. This represents a significant advancement over current methodologies.
Overall the project's outcomes have considerable implications for the field of regenerative medicine and the development of therapeutic strategies for fibrotic diseases. The advancements in understanding the mechanobiology at the molecular/cellular level, coupled with the development of innovative in vitro and in vivo models, pave the way for more effective treatments for fibrosis. These results have socio-economic impacts by potentially reducing the burden of fibrotic diseases and improving patient outcomes, and they have broader societal implications by enhancing our fundamental understanding of cellular behavior in response to mechanical stimuli.
Outgoing phase: Provided vital insights into how focal adhesions (FAs) convert mechanical forces into focal adhesion kinase (FAK) signaling events that regulate cell migration and influence tissue outcomes.
The project explored the complex dynamics between biomaterials, immune cells, and fibrogenic cells. Advanced techniques like intravital imaging and computational flow cytometry revealed a previously unreported heterogeneity in biomaterial-instructed immune cells, challenging traditional views of their homogeneity. These findings are critical for developing new biomaterial-based strategies in regenerative medicine and for treating inflammatory diseases.
Final phase: Optimized a mechanically active platform to apply mechanical forces at the cell-ECM molecular interface.
A platform was optimised to precisely apply mechanical forces at the molecular cell-ECM interface. This led to a deeper understanding of mechanotransduction, particularly how molecular motors can manipulate forces at a molecular scale, affecting key mechanoresponsive molecules like Talin. This represents a significant advancement over current methodologies.
Overall the project's outcomes have considerable implications for the field of regenerative medicine and the development of therapeutic strategies for fibrotic diseases. The advancements in understanding the mechanobiology at the molecular/cellular level, coupled with the development of innovative in vitro and in vivo models, pave the way for more effective treatments for fibrosis. These results have socio-economic impacts by potentially reducing the burden of fibrotic diseases and improving patient outcomes, and they have broader societal implications by enhancing our fundamental understanding of cellular behavior in response to mechanical stimuli.