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Macrophages in the vascular wall ingest and clear lipids, but abundant lipid accumulation leads to foam cell formation and atherosclerosis, a pathological condition often characterized by tissue stiffening. While the role of biochemical stimuli in the modulation of macrophage function is well studied, the role of biophysical cues and the molecules involved in mechanosensation are less well understood. Here, we use genetic and pharmacological tools to show extracellular oxidized low-density lipoproteins (oxLDLs) stimulate Ca2+ signaling through activation of the mechanically gated ion channel Piezo1. Moreover, macrophage Piezo1 expression is critical in the transduction of environmental stiffness and channel deletion suppresses, whereas a gain-of-function mutation exacerbates oxLDL uptake. Additionally, we find that depletion of myeloid Piezo1 protects from atherosclerotic plaque formation in vivo. Together, our study highlights an important role for Piezo1 and its respective mutations in macrophage mechanosensing, lipid uptake, and cardiovascular disease.

We propose a model for the feedback control processes that underlie the robustness and high sensitivity of mechanosensory hair cells. Our model encompasses self-tuning active processes intrinsic to these cells, which drive the amplification of mechanical stimuli by consuming metabolic energy, and a neural input process that protects these cells from damage caused by powerful stimuli. We explore the effects of these two feedback mechanisms on mechanical self-oscillations of the sense cells and their response to external forcing.

Cells intricately sense mechanical forces from their surroundings, driving biophysical and biochemical activities. This mechanosensing phenomenon occurs at the cell-matrix interface, where mechanical forces resulting from cellular motion, such as migration or matrix stretching, are exchanged through surface receptors, primarily integrins, and their corresponding matrix ligands. A pivotal player in this interaction is the α5β1 integrin and fibronectin (FN) bond, known for its role in establishing cell adhesion sites for migration. However, upregulation of the α5β1-FN bond is associated with uncontrolled cell metastasis. This bond operates through catch bond dynamics, wherein the bond lifetime paradoxically increases with greater force. The mechanism sustaining the characteristic catch bond dynamics of α5β1-FN remains unclear. Leveraging molecular dynamics simulations, our approach unveils a pivot-clip mechanism. Two key binding sites on FN, namely the synergy site and the RGD (Arg-Gly-Asp) motif, act as active points for structural changes in α5β1 integrin. Conformational adaptations at these sites are induced by a series of hydrogen bond formations and breaks at the synergy site. We disrupt these adaptations through a double mutation on FN, known to reduce cell adhesion. A whole-cell finite-element model is employed to elucidate how the synergy site may promote dynamic α5β1-FN binding, resisting cell contraction. In summary, our study integrates molecular- and cellular-level modeling to propose that FNs synergy site reinforces cell adhesion through enhanced binding dynamics and a mechanosensitive pivot-clip mechanism. This work sheds light on the interplay between mechanical forces and cell-matrix interactions, contributing to our understanding of cellular behaviors in physiological and pathological contexts.

While extracellular matrix (ECM) stress relaxation is increasingly appreciated to regulate stem cell fate commitment and other behaviors, much remains unknown about how cells process stress-relaxation cues in tissue-like three-dimensional (3D) geometries versus traditional 2D cell culture. Here, we develop an oligonucleotide-crosslinked hyaluronic acid-based ECM platform with tunable stress relaxation properties capable of use in either 2D or 3D. Strikingly, stress relaxation favors neural stem cell (NSC) neurogenesis in 3D but suppresses it in 2D. RNA sequencing and functional studies implicate the membrane-associated protein spectrin as a key 3D-specific transducer of stress-relaxation cues. Confining stress drives spectrins recruitment to the F-actin cytoskeleton, where it mechanically reinforces the cortex and potentiates mechanotransductive signaling. Increased spectrin expression is also accompanied by increased expression of the transcription factor EGR1, which we previously showed mediates NSC stiffness-dependent lineage commitment in 3D. Our work highlights spectrin as an important molecular sensor and transducer of 3D stress-relaxation cues.

Adult neural stem cells (NSCs) reside in the dentate gyrus of the hippocampus, and their capacity to generate neurons and glia plays a role in learning and memory. In addition, neurodegenerative diseases are known to be caused by a loss of neurons and glial cells, resulting in a need to better understand stem cell fate commitment processes. We previously showed that NSC fate commitment toward a neuronal or glial lineage is strongly influenced by extracellular matrix stiffness, a property of elastic materials. However, tissues in vivo are not purely elastic and have varying degrees of viscous character. Relatively little is known about how the viscoelastic properties of the substrate impact NSC fate commitment. Here, we introduce a polyacrylamide-based cell culture platform that incorporates mismatched DNA oligonucleotide-based cross-links as well as covalent cross-links. This platform allows for tunable viscous stress relaxation properties via variation in the number of mismatched base pairs. We find that NSCs exhibit increased astrocytic differentiation as the degree of stress relaxation is increased. Furthermore, culturing NSCs on increasingly stress-relaxing substrates impacts cytoskeletal dynamics by decreasing intracellular actin flow rates and stimulating cyclic activation of the mechanosensitive protein RhoA. Additionally, inhibition of motor-clutch model components such as myosin II and focal adhesion kinase partially or completely reverts cells to lineage distributions observed on elastic substrates. Collectively, our results introduce a unique system for controlling matrix stress relaxation properties and offer insight into how NSCs integrate viscoelastic cues to direct fate commitment.

After traumatic brain injury, the brain extracellular matrix undergoes structural rearrangement due to changes in matrix composition, activation of proteases, and deposition of chondroitin sulfate proteoglycans by reactive astrocytes to produce the glial scar. These changes lead to a softening of the tissue, where the stiffness of the contusion core and peripheral pericontusional regions becomes softer than that of healthy tissue. Pioneering mechanotransduction studies have shown that soft substrates upregulate intermediate filament proteins in reactive astrocytes; however, many other aspects of astrocyte biology remain unclear. Here, we developed a platform for the culture of cortical astrocytes using polyacrylamide (PA) gels of varying stiffness (measured in Pascal; Pa) to mimic injury-related regions in order to investigate the effects of tissue stiffness on astrocyte reactivity and morphology. Our results show that substrate stiffness influences astrocyte phenotype; soft 300 Pa substrates led to increased GFAP immunoreactivity, proliferation, and complexity of processes. Intermediate 800 Pa substrates increased Aggrecan+, Brevican+, and Neurocan+ astrocytes. The stiffest 1 kPa substrates led to astrocytes with basal morphologies, similar to a physiological state. These results advance our understanding of astrocyte mechanotransduction processes and provide evidence of how substrates with engineered stiffness can mimic the injury microenvironment.

The cytoskeleton of the cell is constantly exposed to physical forces that regulate cellular functions. Selected members of the LIM (Lin-11, Isl-1, and Mec-3) domain-containing protein family accumulate along force-bearing actin fibers, with evidence supporting that the LIM domain is solely responsible for this force-induced interaction. However, LIM domain's force-induced interactions are not limited to actin. LIMK1 and LMO1, both containing only two tandem LIM domains, are recruited to force-bearing keratin fibers in epithelial cells. This unique recruitment is mediated by their LIM domains and regulated by the sequences outside the LIM domains. Based on in vitro reconstitution of this interaction, LIMK1 and LMO1 directly interact with stretched keratin 8/18 fibers. These results show that LIM domain's mechano-sensing abilities extend to the keratin cytoskeleton, highlighting the diverse role of LIM proteins in force-regulated signaling.

The gastrointestinal system is a hollow organ affected by fibrostenotic diseases that cause volumetric compromise of the lumen via smooth muscle hypertrophy and fibrosis. Many of the driving mechanisms remain unclear. Yes-associated protein-1 (YAP) is a critical mechanosensory transcriptional regulator that mediates cell hypertrophy in response to elevated extracellular rigidity. In the type 2 inflammatory disorder, eosinophilic esophagitis (EoE), phospholamban (PLN) can induce smooth muscle cell hypertrophy. We used EoE as a disease model for understanding a mechanistic pathway in which PLN and YAP interact in response to rigid extracellular substrate to induce smooth muscle cell hypertrophy. PLN-induced YAP nuclear sequestration in a feed-forward loop caused increased cell size in response to a rigid substrate. This mechanism of rigidity sensing may have previously unappreciated clinical implications for PLN-expressing hollow systems such as the esophagus and heart.