Abstract
Cells respond to the mechanical properties of the extracellular matrix (ECM) through formation of focal adhesions (FAs), re-organization of the actin cytoskeleton and adjustment of cell contractility. These are energy-demanding processes, but a potential causality between mechanical cues (matrix stiffness) and cellular (energy) metabolism remains largely unexplored. Here, we cultured human mesenchymal stem cells (hMSCs) on stiff (20 kPa) or soft (1 kPa) substrate and demonstrate that cytoskeletal reorganization and FA formation spreading on stiff substrates lead to a drop in intracellular ATP levels, correlating with activation of AMP-activated protein kinase (AMPK). The resulting increase in ATP levels further facilitates cell spreading and reinforces cell tension of the steady state, and coincides with nuclear localization of YAP/TAZ and Runx2. While on soft substrates (1 kPa), lowered ATP levels limit these cellular mechanoresponses. Furthermore, genetic ablation of AMPK lowered cellular ATP levels on stiff substrate and strongly reduced responses to substrate stiffness. Together, these findings reveal a hitherto unidentified relationship between energy expenditure and the cellular mechanoresponse, and point to AMPK as a key mediator of stem cell fate in response to ECM mechanics.
Highlights
The physical properties of the extracellular matrix (ECM) have a profound impact on cell behavior and stem cell fate, and a direct rela tionship between matrix stiffness, cell spreading and lineage selection has been demonstrated [1,2,3,4,5,6]
We first determined the influence of substrate stiffness on indicators of energy expenditure such as intracellular ATP levels and glucose up take rates
We cultured human mesenchymal stem cells (hMSCs) on 1 kPa or 20 kPa poly acrylamide (PAAm) gels functionalized with collagen (50 μg/mL), and determined intracellular ATP levels at different time points, from initial adhesion and spreading of cells to steady state (Fig. 1a and Supplementary Fig. 1a and b)
Summary
The physical properties of the extracellular matrix (ECM) have a profound impact on cell behavior and stem cell fate, and a direct rela tionship between matrix stiffness, cell spreading and lineage selection has been demonstrated [1,2,3,4,5,6]. Forces generated within the actin cyto skeleton and transmitted through FAs, play a major role in the cellular response to biophysical cues [7,8,9,10]. Much attention has been given to how cells respond to mechanical forces. In order to understand how mechanical forces shape the cellular phenotype and regulate cell fate, it is necessary to consider the highly dynamic nature of the cellular response, which involves a hitherto overlooked large energy expenditure. The cellular response is a well-tuned process that must balance energy supply with energy demand to allow proper actin polymerization, FA formation and cellular contractility buildup. It remains unclear how cells respond to this energy expenditure, tune energy supply and demand, and maintain energy homeostasis for mechanotransduction
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