Maxillary expansion is a common orthodontic procedure for treating maxillary transverse deficiency. However, the cell responses to mechanical force may vary across different age groups, suggesting the need for age-specific treatment protocols. To compare the age-related responses to the mechanical force, we examined the 6-week- and 12-month-old mice undergoing maxillary expansion with 0.012-in. stainless steel orthodontic wire bonded to the maxillary first and second molars (25 g force). Mice were euthanized on days 0, 3, 7, and 14 for analysis. MicroCT analysis, tartrate-resistant acid phosphatase (TRAP) stain, and immunofluorescence/immunohistochemistry stain using antibodies to RUNX2, alkaline phosphatase (ALP), Gli1 and Ki67 along with the TUNEL assay, were conducted to evaluate suture width, osteoclast activity, new bone formation and mesenchymal stem cell (MSC) proliferation and apoptosis. Both 6-week- and 12-month-old mice exhibited successful midpalatal suture opening, but young mice demonstrated earlier and more intense osteoclast activity, along with higher expression of RUNX2 and ALP. Young mice also exhibited a higher percentage of Gli1+Ki67+ immunopositive cells, while middle-aged mice showed a higher percentage of Gli1+TUNEL+ positive cells on day 3 after maxillary expansion. Our findings suggest that aging negatively impacts mechanical force-induced bone remodeling by reducing osteoclastogenesis, osteogenesis, and MSC proliferation while increasing MSC apoptosis.

Esophageal squamous cell carcinoma (ESCC) is one of the most prevalent cancers worldwide, characterized by a dismal prognosis and elusive therapeutic targets. Dysregulated cholesterol metabolism is a critical hallmark of cancer cells, facilitating tumor progression. Here, we used whole genome sequencing data from several ESCC cohorts to identify the important role of squalene epoxidase (SQLE) in promoting ESCC tumorigenesis and metastasis. Specifically, our findings highlight the significance of 2,3-oxidosqualene, an intermediate metabolite of cholesterol biosynthesis, synthesized by SQLE and metabolized by lanosterol synthase (LSS), as a key regulator of ESCC progression. Mechanistically, the interaction between 2,3-oxidosqualene and vinculin enhances the nuclear accumulation of Yes-associated protein 1 (YAP), thereby increasing YAP/TEAD-dependent gene expression and accelerating both tumor growth and metastasis. In a 4-nitroquinoline 1-oxide (4-NQO)-induced ESCC mouse model, overexpression of Sqle resulted in accelerated tumorigenesis compared to wild-type controls, highlighting the pivotal role of SQLE in vivo. Furthermore, elevated SQLE expression in ESCC patients correlates with a poorer prognoses, suggesting potential therapeutic avenues for treatment. In conclusion, our study elucidates the oncogenic function of 2,3-oxidosqualene as a naturally occurring metabolite and proposes modulation of its levels as a promising therapeutic strategy for ESCC.

Tendon injuries are often exacerbated by persistent inflammation, which hampers tissue regeneration. In this study, we developed a noninvasive, wirelessly controlled, and self-powered piezoelectric nanofilm fabricated by coaxial electrospinning of polycaprolactone (PCL) and tetragonal barium titanate nanoparticles (BTO), and investigated its roles in modulating inflammation and repairing Achilles tendon defects as well as the mechanism in a rat model. In vitro study and in vivo study upon subcutaneous implantation showed that the piezoelectric PCL/BTO nanofilms could inhibit M1 macrophage polarization and reduce the secretion of inflammatory factors. Moreover, when bridging an Achilles tendon defect, the nanofilms could promote tenogenic gene expression including collagen deposition, and collagen remodeling, facilitate functional tendon recovery and significantly reduce tissue inflammation by suppressing M1 macrophage polarization and promoting M2 polarization. Moreover, the piezoelectric stimulation could also enhance tendon regeneration by inhibiting angiogenesis, reducing lipid deposition, and decreasing ectopic ossification. Mechanistically, the piezoelectric nanofilms reduced tissue inflammation mainly via inhibiting the nuclear factor (NF)-κB signaling pathway that is mediated by interleukin (IL)-17A secreted from CD3+ T cells, and thus to reduce proinflammatory factors, such as IL-1β and IL-6, inducible nitric oxide synthase, monocyte chemoattractant protein-1, and tumor necrosis factor-α. These findings indicate the potential of piezoelectric stimulation in immunomodulation, and in promoting tendon regeneration via IL-17A/NF-κB-mediated pathway.

Microspheres (MPs) and porous microspheres (PMPs) are the two most widely used microparticles in tissue engineering and stem cell therapy. However, how stem cells perceive the topological differences between them to regulate cell function remains to be unclear. Here, we systematically studied the changes in stem cell function under the action of MPs and PMPs and elucidated the related mechanisms. Our findings show that the porous structure of PMPs can be sensed by focal adhesions (FAs), which triggers the synthesis of F-actin to inhibit the phosphorylation and degradation of Yes-associated protein (YAP), while also transmitting stress to the nucleus through the contraction of F-actin, thereby enhancing the nuclear translocation of YAP protein. The activation of YAP significantly enhances the proliferation, osteogenesis, paracrine and glucose metabolism of BMSCs, making them exhibit stronger bone repair ability in both in vivo and in vitro experiments. In summary, this study provides a comprehensive and reliable understanding of the behavior of BMSCs in response to MPs and PMPs. It also deepens our understanding of the association between microparticles' topological cues and biological functions, which will provide valuable guidance for the construction of bone tissue engineering (BTE) scaffolds.

Renal fibrosis is a fundamental pathological change in the progression of various chronic kidney diseases to the end stage of renal disease. The Hippo signaling pathway is an evolutionary highly conserved signaling pathway that is involved in the regulation of organ size, tissue regeneration, and human reproduction and development. Currently, many studies have shown that it is closely associated with renal diseases, such as, renal fibrosis, diabetic nephropathy, and renal cancer. Here, we review the current researches on the effect of Hippo signaling pathway on renal fibrosis, which provides new ideas and theoretical basis for clinical therapeutics of renal fibrosis.

Soft micropillar arrays enable detailed studies of cellular mechanotransduction and biomechanics using traditional beam-bending models. However, they often rely on simplified assumptions, leading to significant errors in force estimation. We present MechanoBioCAD (MBC), a finite element method (FEM)-based tool designed specifically for micropillar research and error estimation. Unlike traditional methods, MBC leverages the principle of minimizing total potential energy, avoiding errors associated with beam bending assumptions. MBC automates FEM model generation, analysis, and post-processing, providing accurate force quantification based on deflection input. The tool addresses critical issues such as substrate deformation, interpillar interactions, improper load application heights, and nonlinear effects. Applied to fibroblast cell traction and Caenorhabditis elegans (C. elegans) thrashing cases, MBC recorded 23% and 34% errors in the estimated forces, respectively, compared to traditional methods. As an open-access tool with the Abaqus Student Edition, MBC democratizes rational design, analysis, and error estimation for researchers who are not subject matter experts in FEM.

The Hippo/YAP signaling pathway is closely related to the occurrence and development of cardiovascular diseases. However, it's still unclear whether this pathway plays a certain role in hypertension. In this study, aortic morphology and function in spontaneously hypertensive rats (SHR) were comprehensively evaluated using Wistar-Kyoto rats (WKY) as controls. Results indicated that the aorta of SHRs have distinct changes in pathological structure. Furthermore, the proliferative activity of vascular smooth muscle cells (VSMCs) was enhanced, with vascular fibrosis being aggravated. Immunohistochemical analysis revealed that SHRs exhibited high expression of Yes-Associated Protein (YAP). Western Blot analysis showed that cytoplasmic YAP and TAZ expression decreased in hypertensive rats, indicating that YAP/TAZ nuclear transfer increased and Hippo/YAP signaling pathway had been activated. The cell function experiments of VSMCs extracted from rat aorta showed that the cell viability and proliferation ability of VSMCs in SHRs were enhanced, the expression of Fibronectin and collagen I was increased, and vascular fibrosis was aggravated. siRNA-YAP (si-YAP) can reverse the above phenomenon in VSMCs. Knockdown of YAP can inhibit Foxm1 expression. As an inhibitor of large tumor suppressor kinases LAST1/2, GA-107 can inhibit the phosphorylation level of YAP, increase blood pressure, aggravate aortic pathomorphological changes, promote VSMCs proliferation and vascular fibrosis, and thus aggravate hypertension symptoms in SHRs. However, these effects of GA-107 can be antagonized by inhibiting Foxm1 with thiosulfathiazole (Thio). Conclusively, Hippo/YAP signaling pathway promotes vascular remodeling through the regulation of Foxm1 and causes hypertension.

Cells sense physical cues from their environment and convert them into biochemical responses through mechanotransduction. Unlike solid tumours, the role of such forces in haematological cancers is underexplored. In this context, immune cells experience dynamic mechanical stimuli as they migrate, extravasate and home to specific tissues. Understanding how these forces shape B-cell function and malignancy represents a groundbreaking area of research. This review examines the key mechanosensory pathways and molecules involved in lymphocyte mechanotransduction, beginning with mechanosensory proteins at the plasma membrane, followed by intracellular signal propagation through the cytoskeleton, eventually highlighting the nucleus as a 'signal actuator'. Subsequently, we cover some measurement approaches and advanced systems to investigate tumour biomechanics, highlighting their application in the context of B cells. Finally, we focus on the implications of mechanobiology in leukaemia, identifying molecules involved in B-cell malignancies that could serve as potential 'mechano-targets' for personalised therapies. This review emphasises the need to understand how lymphocytes generate, sense and respond to mechanical stimuli, which could open avenues for future biomedical innovations. Impact statement Our review is particularly valuable in highlighting the underexplored role of mechanobiology in B cell function and malignancies, while also discussing emerging techniques that can advance this research area. It bridges mechanotransduction, immunology, and cancer biology in a way that will be of interest to researchers across these three main fields.

Synphilin-1 has been studied extensively in the context of Parkinson's disease pathology. However, the biophysical functions of synphilin-1 remain unexplored. To investigate its novel functionalities herein, cellular traction force and rigidity sensing ability are analyzed based on synphilin-1 overexpression using elastomeric pillar arrays and substrates of varying stiffness. Molecular changes are analyzed using RNA sequencing-based transcriptomic and liquid chromatography-tandem mass spectrometry-based proteomic analyses.

Synphilin-1 overexpression reduces cell area, with a decline of local contraction on elastomeric pillar arrays. Cells overexpressing synphilin-1 exhibit an impaired ability to respond to substrate rigidity; however, synphilin-1 knockdown restores rigidity sensing abilities. Integrated omics analysis and in silico prediction corroborate the phenotypic alterations induced by synphilin-1 overexpression at a biophysical level. Zyxin emerges as a novel synphilin-1 binding protein, and synphilin-1 overexpression reduces the nuclear translocation of yes-associated protein.

These findings provide novel insights into the biophysical functions of synphilin-1, suggesting a potential protective role to the altered extracellular matrix, which may be relevant to neurodegenerative conditions such as Parkinson's disease.

All biological surfaces possess distinct dynamic surface topographies. Due to their versatility, these topographies play a crucial role in modulating cell behavior and, when intentionally designed, can precisely guide cellular responses. So far, biomechanical responses have predominantly been studied on static surfaces, overlooking the dynamic environment in the body, where cells constantly interact with shifting biomechanical cues. In this work, we designed and fabricated a light-responsive liquid crystal polymer film to study the effect of micrometer-scale, dynamic surface topographies on cells under physiologically relevant conditions. The light-responsive liquid crystal polymers enable on-demand surface topographical changes, reaching pillar heights of 800 nm and grooved topographies with 700 nm height differences at 37 °C in water. The light-induced surface topographies increased mechanosensitive cell signaling by up to 2-fold higher yes-associated protein (YAP) translocation to the nucleus, as well as up to 3-fold more heterogeneity in distribution of focal adhesions, in a topography-related manner. The pillared topography was seen to cause a lower cellular response, while the grooved topography caused an increased mechanical activation, as well as cell alignment due to a more continuous and aligned physical cue that enhances cell organization. Excitingly, we observed that subsequent surface topography changes induced a 3-fold higher YAP nuclear translocation in fibroblast cells, as well as a 5-fold higher vinculin heterogeneity distribution, indicating that multiple cycles of topography exposure ampliated the cell response. Our work emphasizes the potential of light-responsive liquid crystal polymer films generating dynamic biomechanical cues that allow us to modulate and steer cells in vitro.

The behavior of cells is governed by signals originating from their local environment, including mechanical forces exerted on the cells. Forces are transduced by mechanosensitive proteins, which can impinge on signaling cascades that are also activated by growth factors. We investigated the cross-talk between mechanical and biochemical signals in the regulation of intracellular signaling networks in epithelial monolayers. Phosphoproteomic and transcriptomic analyses on epithelial monolayers subjected to mechanical strain revealed the activation of extracellular signal-regulated kinase (ERK) downstream of the epidermal growth factor receptor (EGFR) as a predominant strain-induced signaling event. Strain-induced EGFR-ERK signaling depended on mechanosensitive E-cadherin adhesions. Proximity labeling showed that the metalloproteinase ADAM17, an enzyme that mediates shedding of soluble EGFR ligands, was closely associated with E-cadherin. A probe that we developed to monitor ADAM-mediated shedding demonstrated that mechanical strain induced ADAM activation. Mechanically induced ADAM activation was essential for mechanosensitive, E-cadherin-dependent EGFR-ERK signaling. Together, our data demonstrate that mechanical strain transduced by E-cadherin adhesion triggers the shedding of EGFR ligands that stimulate downstream ERK activity. Our findings illustrate how mechanical signals and biochemical ligands can operate within a linear signaling cascade.

Lung cancer, a major cause of cancer-related mortality, has limited therapeutic options, especially for advanced cases. Ferroptosis, an iron-dependent form of cell death, is a potential therapeutic strategy for this disease; however, resistance mechanisms in the tumor microenvironment impede its effectiveness. Therefore, in this study, we aimed to investigate the efficacy of sulfasalazine (SAS), a ferroptosis inducer, and auranofin (AUR), a Food and Drug Administration-approved anti-inflammatory agent, combination to counteract ferroptosis resistance in lung cancer. SAS induced ferroptosis in vitro; however, its efficacy in vivo was limited, possibly because of factors, such as nutrient deprivation and high cell density, in the microenvironment that suppressed the activities of Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), key regulators of ferroptosis resistance. Screening of 2483 drugs revealed AUR as a compound resensitizing the YAP/TAZ-deficient lung cancer cells to ferroptosis. Moreover, SAS and AUR combination significantly enhanced lipid peroxidation and reactive oxygen species accumulation, further driving ferroptosis in cells. This combination effectively inhibited tumor growth and enhanced survival in a murine lung cancer model. Overall, our findings suggest that AUR potentiates ferroptosis-based therapies, serving as an effective candidate to overcome ferroptosis resistance in lung cancer.

During the first cell fate decision in mammalian embryos, the inner cell mass cells, which will give rise to the embryo proper and other extraembryonic tissues, segregate from the trophectoderm cells, the precursors of the placenta. Cell fate segregation proceeds in a gradual manner encompassing two rounds of cell division, as well as cell positional and morphological changes. While it is known that the activity of the Hippo signaling pathway and the subcellular localization of its downstream effector YAP dictate lineage specific gene expression, the response of YAP to these dynamic cellular changes remains incompletely understood. Here we address these questions by quantitative live imaging of endogenously tagged YAP while simultaneously monitoring geometric cellular features and cell cycle progression throughout cell fate segregation. We apply a probabilistic model to our dynamic data, providing a quantitative characterization of the mutual effects of YAP and cellular relative exposed area, which has previously been shown to correlate with subcellular YAP localization in fixed samples. Additionally, we study how nuclear YAP levels are influenced by other factors, such as the decreasing pool of maternally provided YAP that is partitioned to daughter cells through cleavage divisions, cell cycle-associated nuclear volume changes, and a delay after divisions in adjusting YAP levels to new cell positions. Interestingly, we find that establishing low nuclear YAP levels required for the inner cell mass fate is largely achieved by passive cell cycle-associated mechanisms. Moreover, contrary to expectations, we find that mechanical perturbations that result in cell and nuclear shape changes do not influence YAP localization in the embryo. Together our work identifies how various inputs are integrated over a dynamic developmental time course to shape the levels of a key molecular determinant of the first cell fate choice.

Vascularization has been considered an essential strategy for bone regeneration and can be promoted by magnesium ions (Mg2+). During angiogenesis, the differentiation of endothelial cells (ECs) into tip cell is a critical step since it controls the growth direction and pattern of new vascular sprouts. While several studies have noted the pro-angiogenic effects of Mg2+, however, their specific influence on tip cell formation is unclear. Therefore, this research seeks to examine the impact of Mg2+ on tip cells and elucidate the potential mechanisms involved. The results reveal that Mg2+ shows good compatibility and stimulates ECs to migrate and invade in vitro. Moreover, Mg2+ enhances EC spheroids sprouting and elevates the expression of genes linked to tip cells. The underlying mechanisms are that Mg2+ facilitates tip cell differentiation via the VEGFA-VEGFR2/Notch1 signaling pathway crosstalk and promotes migration and filopodia formation of tip cells and proliferation of stalk cells by inducing YAP nuclear translocation, culminating in the maturation of vascular networks. Furthermore, EC spheroids stimulated by Mg2+ load in hydrogel enhance vascularized bone regeneration in vivo. These findings enrich the understanding of how Mg2+ influence blood vessel formation and provide practical strategies for the development and design of magnesium-based biomaterials.

The limited regenerative ability of "permanent" cells is a major barrier to treating conditions like spinal cord injury (SCI) and myocardial infarction (MI). The delivery of stem cells, which can generate various cell types, offer potential for personalized therapy with reduced immunoreaction and recovery time. However, restoring function to these tissues also requires new or replacement cells to align properly. Neurons, for example, must organize and extend parallel axons, mimicking their natural structure for directional signal propagation. Current stem cell differentiation methods lack guidance, resulting in randomly distributed axons and limited repair effectiveness. Advancing methods and materials to guide stem cell differentiation into functional, aligned nerve bundles is crucial for improving SCI treatment outcomes. This study aimed to develop an in vitro system to promote aligned neural differentiation by applying cyclic uniaxial tension to PC-12 stem cells adhered to 3D-printed elastic scaffolds. We created a simple loading device which can apply cyclic and controllable stretching force to a scaffold, which in turn transmits uniaxial tension to cells adhered to the scaffold during their differentiation. An elastomer ink for 3D printing scaffolds was formulated and surface treatment processes were investigated to enhance the cell-scaffold adhesion to support the dynamic loading. It was revealed that a corona discharge treatment while the scaffold is soaked with type I collagen can significantly enhance cell adhesion. A range of strain magnitudes and frequencies were revealed to enhance the differentiation of neural tissue derived PC-12 cells to neuron cells and increase the length of their neurites up to 76%. The combination of 3% maximum strain and 1 Hz loading frequency maximized differentiation and neurite extension. These findings demonstrate that dynamic mechanical stimulation enhances neural differentiation and organization, offering an alternative approach for regenerative therapies targeting SCI and similar conditions.

Cells perceive external and internally generated forces of different kinds, significantly impacting their cellular biology. In the relatively nascent field of mechanobiology, the impact of such forces is studied and further utilized to broaden the knowledge of cellular developmental pathways, disease progression, tissue engineering, and developing novel regenerative strategies. However, extensive considerations of mechanotransduction pathways for biomedical applications are still broadly limited due to a lack of affordable technologies in terms of devices and simple magnetic actuatable materials. Herein, synthesizing a monophasic, macroporous, in situ-fabricated gelatin-based ferrogel is reported using polyethylene glycol (PEG) coated-iron oxide (magnetite) particles with high magnetization. Developing a 3D printed, compact, and wireless device capable of providing a wide range of magneto-mechanical actuation using magnetic field susceptible materials in a noncontact manner is reported. Using the device and ferrogel, C2C12 myoblast differentiation is studied under magnetic field actuation, and significant differences in the myogenin, a differentiation marker, expression behavior are observed. Due to careful design considerations, robust component selection, and easy availability of low-cost precursor for magnetic responsive material fabrication, the device-ferrogel combination can be easily adapted to routine biological studies, thereby helping mechanobiology to be utilized for developing exciting new biomedical strategies.

The viscoelastic properties of tissues influence their morphology and cellular behavior, yet little is known about changes in these properties during brain malformations. Lissencephaly, a severe cortical malformation caused by LIS1 mutations, results in a smooth cortex. Here, we show that human-derived brain organoids with LIS1 mutation exhibit increased stiffness compared to controls at multiple developmental stages. This stiffening correlates with abnormal extracellular matrix (ECM) expression and organization, as well as elevated water content, measured by diffusion-weighted MRI. Short-term MMP9 treatment reduces both stiffness and water diffusion levels to control values. Additionally, a computational microstructure mechanical model predicts mechanical changes based on ECM organization. These findings suggest that LIS1 plays a critical role in ECM regulation during brain development and that its mutation leads to significant viscoelastic alterations.

Cellular mechanotransduction is a complex physiological process that integrates alterations in the external environment with cellular behaviours. In recent years, the role of the nucleus in mechanotransduction has gathered increased attention. Our research investigated the involvement of lamin A/C, a component of the nuclear envelope, in the mechanotransduction of macrophages under compressive force. We discovered that hydrostatic compressive force induces heterochromatin formation, decreases SUN1/SUN2 levels, and transiently downregulates lamin A/C. Notably, downregulated lamin A/C increased nuclear permeability to yes-associated protein 1 (YAP1), thereby amplifying certain effects of force, such as inflammation induction and proliferation inhibition. Additionally, lamin A/C deficiency detached the linker of nucleoskeleton and cytoskeleton (LINC) complex from nuclear envelope, consequently reducing force-induced DNA damage and IRF4 expression. In summary, lamin A/C exerted dual effects on macrophage responses to mechanical compression, promoting certain outcomes while inhibiting others. It operated through two distinct mechanisms: enhancing nuclear permeability and impairing intracellular mechanotransmission. The results of this study support the understanding of the mechanisms of intracellular mechanotransduction and may assist in identifying potential therapeutic targets for mechanotransduction-related diseases.

Parkinson's disease (PD) is exacerbated by dysfunction of inter-organelle contact, which depends on cellular responses to the mechanical microenvironment and can be regulated by external mechanical forces. Delivering dynamic mechanical forces to neural cells proves challenging due to the skull. Inspired by the effects of massage; here PEGylated black phosphorus nanosheets (PEG-BPNS), known for their excellent biocompatibility, biodegradability, specific surface area, mechanical strength, and flexibility, are introduced, which are capable of adhering to neural cell membrane and generating mechanical stimulation with their lateral size of 200 nm, exhibiting therapeutic potential in a 1-methyl-4-phenyl-1,2,3,6-te-trahydropyridine-induced PD mouse model by regulating inter-organelle contacts. Specifically, it is found that 200 nm PEG-BPNS, acting as "NanoMassage," significantly increase  plasma membrane tension, as evidenced by fluorescent lipid tension reporter fluorescence lifetime analysis. This mechanical force modulates actin reorganization, subsequently regulating the contacts between actin, mitochondria, and endoplasmic reticulum, further controlling mitochondrial fission and mitigating mitochondrial dysfunction in PD, exhibiting therapeutic efficacy via intranasal administration. These findings provide a noninvasive strategy for applying mechanical stimulation to deep brain areas and elucidate the mechanism of NanoMassage mediating inter-organelle contacts, suggesting the rational design of "NanoMassage" to remodel inter-organelle communications in neurodegenerative disease treatment.

The components of the extracellular matrix (ECM) are dynamic, and they mediate mechanical signals that modulate cellular behaviors. Disruption of the ECM can induce the migration and invasion of cancer cells via specific signaling pathways and cytokines. Metastasis is a leading cause of high mortality in malignancies, and early intervention can improve survival rates. However, breast cancer is frequently diagnosed subsequent to metastasis, resulting in poor prognosis and distant metastasis poses substantial hurdles in therapy. In breast cancer, there is notable tissue remodeling of ECM proteins, with several identified as essential components for metastasis. Moreover, specific ECM molecules, receptors, enzymes, and various signaling pathways play crucial roles in breast cancer metastasis, drug treatment, and resistance. The in-depth consideration of these elements could provide potential therapeutic targets to enhance the survival rates and quality of life for breast cancer patients. This review explores the mechanisms by which alterations in the ECM contribute to breast cancer metastasis and discusses current clinical applications targeting ECM in breast cancer treatment, offering valuable perspectives for future ECM-based therapies.

The mechanical properties of the extracellular matrix (ECM), particularly stiffness, regulate endothelial progenitor responses during vascular development, yet their behavior in physiologically compliant matrices (<1 kPa) remains underexplored. Using norbornene-modified hyaluronic acid (NorHA) hydrogels with tunable stiffness (190-884 Pa), we investigated how hydrogel stiffness influences cell morphology, endothelial maturation, mechanotransduction, and microvascular network formation in human induced pluripotent stem cell-derived endothelial progenitors (hiPSC-EPs). Our findings reveal a stiffness-dependent tradeoff between mechanotransduction and vascular network formation. At intermediate stiffness (551 Pa), cells exhibited the greatest increase in endothelial marker CD31 expression and Yes-associated protein (YAP)/ transcriptional coactivator with PDZ-binding motif (TAZ) nuclear translocation, indicating enhanced mechanotransduction and endothelial maturation. However, this did not translate to superior plexus formation. Instead, the most compliant matrix (190 Pa) supported greater vascular connectivity, characterized by longer branches (∼0.03/volume vs. 0.015 at 551 Pa) and enhanced actin remodeling. 3D cell contraction measurements revealed a 15.6-fold higher basal displacement in compliant hydrogels, suggesting that cell-generated forces and matrix deformability collectively drive vascular morphogenesis. Unlike prior studies focusing on pathological stiffness ranges (>10 kPa), our results emphasize that vascularization is not solely driven by the most mechanotransductive environment but rather by a balance of compliance, contractility, and cell-induced remodeling. These findings underscore the need to design hydrogels that provide sufficient mechanotransduction for endothelial maturation while maintaining compliance to support dynamic vascular morphogenesis. This work provides a mechanically tuned framework for optimizing microenvironments to balance endothelial differentiation and vascular network formation in tissue engineering and regenerative medicine.

The field of mechanobiology has grown in the past decade, but limited studies investigate how the extracellular matrix affects the cell metabolome. The methionine cycle involves the catabolism and regeneration of methionine through the donation and recovery of a single methyl group; this methyl group can methylate DNA, RNA, and proteins to alter gene expression and protein-protein interactions. Through studying cells cultured on fibrous (mimicking healthy extracellular matrice (ECM)) and flat (mimicking severely fibrotic ECM) substrates, we observed an increase in methionine cycle enzyme expression in cells on the flat substrate. We also present how the methionine cycle is modulated by the ECM through transmembrane protein integrin β1. By inhibiting integrin activation through the ligand-mimicking peptide RGD, we observed that the methionine cycle was protected from alteration. The results presented provide insight into possible therapeutic targets for fibrotic diseases and knowledge of mechanisms by which the ECM alters cell processes.

Multiple mechanisms were proposed to mediate the nuclear import of TAZ/YAP, transcriptional co-activators regulating organ growth and regeneration. Our earlier observations showed that TAZ/YAP harbor a C-terminal, unconventional nuclear localization signal (NLS). Here, we show that this sequence, necessary and sufficient for basal, ATP-independent nuclear import, contains an indispensable central methionine flanked by negatively charged residues. Based on these features, we define the M-motif and propose that it is a new class of NLS, also present and import-competent in other cellular (STAT1 and cyclin B1) and viral (ORF6 of SARS-CoV2, VSV-M) proteins. Accordingly, ORF6 SARS-Cov2 competitively inhibits TAZ/YAP uptake, while TAZ abrogates STAT1 import. Similar to viral M-motif proteins, TAZ binds RAE1 and inhibits classic nuclear protein import, including that of antiviral factors (IRF3 and NF-κB). However, RAE1 is dispensable for TAZ import itself. Thus, the TAZ/YAP NLS has a dual function: it mediates unconventional nuclear import and inhibits classic import, contributing to the suppression of antiviral responses.

Culture platforms that closely mimic the spatial architecture, cellular diversity, and extracellular matrix composition of native tissues can serve as invaluable tools for a range of scientific discovery and biomedical applications. Organoids have emerged as a promising alternative to both traditional 2D cell culture and animal models, offering a physiologically relevant 3D culture system for studying human cell biology. Organoids provide a manipulable platform to investigate organ development and function as well as to model patient-specific phenotypes. This mini review examines various methods used for culturing organoids to model normal and disease conditions in gastrointestinal tissues. We focus on how the matrix composition and media formulations can impact cell signaling, altering the baseline cellular phenotypes as well as response to perturbations. We discuss future directions for optimizing organoid culture conditions to improve basic and translational potential.

Brain arteriovenous malformations (bAVMs) are complex vascular lesions with significant clinical risks. The Hippo signaling pathway, particularly its downstream effector YAP, plays a crucial role in angiogenesis and vascular remodeling. This study investigates the role of YAP and related molecular markers in bAVMs, focusing on the effects of embolization. Immunohistochemical analysis was conducted on tissue samples from bAVM patients (n = 127), as well as on healthy blood vessels (n = 17). YAP, HIF-1α, FGFR1, CTGF, and CYR61 expression were quantified and correlated with clinical parameters. Results: In healthy vessels, YAP exhibited nuclear localization in (sub)endothelial cells and the tunica media, while CTGF and CYR61 were detected in the cytoplasm and extracellular matrix. The expression of YAP, CTGF, and CYR61 was significantly lower in bAVM tissues. Embolized bAVMs exhibited significantly higher expression of YAP, CTGF, and CYR61 compared to non-embolized tissues, suggesting a link between embolization and pro-angiogenic signaling. Additionally, FGFR1 was upregulated in embolized tissues. These results suggest that upregulation of YAP expression via the Hippo pathway might play a key role in bAVM pathophysiology. Embolization may further promote vascular remodeling. Dysregulation of YAP and related molecules in bAVMs warrants further studies to explore potential therapeutic strategies targeting the Hippo pathway.

Atherosclerosis is characterized by the accumulation of fatty and fibrotic plaques, which preferentially develop at curvatures and branches along the arterial trees that are exposed to disturbed flow. However, the mechanisms by which endothelial cells sense disturbed flow are still unclear.

The partial carotid ligation mouse model was used to investigate disturbed flow-induced atherogenesis. In vitro experiments were performed using the ibidi system to generate oscillatory shear stress and laminar shear stress. ApoE-/- mice with endothelium-specific knockout or overexpression of 5-HT1B (serotonin receptor 1B) were used to investigate the role of endothelial 5-HT1B in atherosclerosis. RNA sequencing analysis, immunofluorescence analysis, and molecular biological techniques were used to explore the role of 5-HT1B in mechanotransduction and endothelial activation.

The data showed that human endothelial cells express a high level of 5-HT1B, which is a serotonin receptor subtype. Endothelial 5-HT1B is upregulated in atherosclerotic areas of both humans and rodents and is increased by disturbed flow both in vivo and in vitro. Endothelium-specific overexpression of 5-HT1B exacerbates, whereas knockout or knockdown of 5-HT1B in endothelium inhibits disturbed flow-induced endothelial inflammation and atherogenesis in both male and female ApoE-/- mice. We reveal a previously unknown role of 5-HT1B as a mechanosensor in endothelial cells in response to mechanical stimuli. Upon activation by oscillatory shear stress, 5-HT1B recruits β-arrestin, orchestrates RhoA (ras homolog family member A), and then activates mechanosensitive YAP (yes-associated protein), thereby enhancing endothelial inflammation and monocyte infiltration. Pharmacological blockade of 5-HT1B suppresses endothelial activation and atherogenesis via inhibition of YAP.

Taken together, these results uncover that endothelial 5-HT1B acts as a mechanosensor for disturbed flow and contributes to atherogenesis. Inhibition of 5-HT1B could be a promising therapeutic strategy for atherosclerosis.

There is an unmet need for point-of-care therapies to prevent scarring and promote corneal clarity after injury, which is essential for maintaining vision. Verteporfin, an inhibitor of Yes-associated protein (YAP), has been shown to prevent fibrosis in several organs. Visudyne (VP) is an FDA-approved liposomal formulation of verteporfin used to treat abnormal blood vessels in the eye. Here, we showed that VP reduces myofibroblast formation in corneal stromal fibroblasts. To prolong the residence time of verteporfin on the ocular surface, the cohesive viscoelastic ProVisc® hyaluronic acid (HA) gel was hybridized to VP. This formulation is readily translatable because both VP and ProVisc® HA gel are FDA-approved agents. The ProVisc® HA gel increased the residence of subconjunctivally injected verteporfin 12-fold at 24 h after injection compared with pure VP. A single subconjunctival administration of VP hybridized within ProVisc® HA gel (VP/HA hydrogel) significantly reduced YAP activation, corneal fibrosis, neovascularization, and inflammation, leading to reduced opacity without compromising epithelial wound healing in mechanically injured rat corneas. This work demonstrated that VP hybridized with a viscoelastic HA gel can be readily repurposed to promote scar-less healing in the cornea.

Tumor microenvironment (TME) is a complex ecosystem composed of both cellular and non-cellular components that surround tumor tissue. The extracellular matrix (ECM) is a key component of the TME, performing multiple essential functions by providing mechanical support, shaping the TME, regulating metabolism and signaling, and modulating immune responses, all of which profoundly influence cell behavior. The quantity and cross-linking status of stromal components are primary determinants of tissue stiffness. During tumor development, ECM stiffness not only serves as a barrier to hinder drug delivery but also promotes cancer progression by inducing mechanical stimulation that activates cell membrane receptors and mechanical sensors. Thus, a comprehensive understanding of how ECM stiffness regulates tumor progression is crucial for identifying potential therapeutic targets for cancer. This review examines the effects of ECM stiffness on tumor progression, encompassing proliferation, migration, metastasis, drug resistance, angiogenesis, epithelial-mesenchymal transition (EMT), immune evasion, stemness, metabolic reprogramming, and genomic stability. Finally, we explore therapeutic strategies that target ECM stiffness and their implications for tumor progression.

Idiopathic pulmonary fibrosis (IPF) is an irreversible and fatal interstitial lung disease, characterized by excessive extracellular matrix (ECM) secretion that disrupts normal alveolar structure. This study aims to explore the potential molecular mechanisms underlying the promotion of IPF development.

Firstly, we compared the transcriptome and single-cell sequencing data from lung tissue samples of patients with IPF and healthy individuals. Subsequently, we conducted Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses on the differentially expressed genes (DEGs). Furthermore, we employed sodium alginate hydrogels with varying degrees of crosslinking to provide differential mechanical stress, mimicking the mechanical microenvironment in vivo during lung fibrosis. On this basis, we examined cytoskeletal remodeling in fibroblasts MRC-5, mRNA expression of multiple related genes, immunofluorescence localization, and cellular proliferation capacity.

Bioinformatics analysis revealed a series of DEGs associated with IPF. Further functional and pathway enrichment analyses indicated that these DEGs were primarily enriched in ECM-related biological processes. Single-cell sequencing data revealed that fibroblasts and myofibroblasts are the main contributors to excessive ECM secretion and suggested activation of mechanotransduction and the Hippo/YAP signaling pathway in myofibroblasts. Cellular experiments demonstrated that sodium alginate hydrogels with different stiffness can simulate different mechanical stress environments, thereby affecting cytoskeletal rearrangement and Hippo/YAP pathway activity in MRC-5 lung fibroblasts. Notably, high levels of mechanical stress promoted YAP nuclear translocation, increased expression of type I collagen and α-SMA, and enhanced proliferative capacity. Additionally, we also found that fibroblasts primarily participate in mechanotransduction through the Rho/ROCK and Integrin/FAK pathways under high mechanical stress conditions, ultimately upregulating the gene expression of CCNE1/2, CTGF, and FGF1.

Our study uncovers the crucial role of cytoskeletal mechanotransduction in myofibroblast transformation and IPF development through activation of the Hippo/YAP pathway, providing new insights into understanding the pathogenesis of IPF.

Physical interactions between cells, such as cell-cell junctions, can profoundly impact cell fate. A vital cell fate for normal development and homeostasis is programmed cell death. Cells fated to die must be efficiently cleared away via phagocytosis, and defects are associated with a variety of diseased states. Whether cell-cell physical associations affect programmed cell elimination has not been well-explored. Here we describe, in vivo, a cell-cell adhesion-driven signaling pathway that ensures compartment-specific cell clearance during development. We previously described the specialized cell death program "Compartmentalized Cell Elimination" (CCE) in the C. elegans embryo. During CCE, the tail-spike cell (TSC), a polarized epithelial cell, undergoes a tripartite, ordered, and organized death sequence, allowing for the study of three distinct death modalities in a single cell setting. Prior to its demise, the TSC serves as a scaffold for the tail tip, formed by the hyp10 epithelial cell which develops along the TSC process. The hyp10 cell in turn also serves as the phagocyte for the dying TSC process. Here we present data suggesting that the physical association between the dying TSC and hyp10 phagocyte via CDH-3/cadherin mediates function of the mechanosensitive transcriptional coactivator YAP-1/YAP and its partner EGL-44/TEAD in the hyp10 phagocyte to promote localization of hyp10 SYX-2/Syntaxin around the dying TSC remnant. This pathway facilitates the phagocytic function of EFF-1/fusogen, which we have previously shown to be required for phagosome sealing during CCE. Our work sheds additional light on a poorly understood step of phagocytosis and implicates adhesive forces and signaling between cells as important in cell uptake.

The human brain is a highly complex organ characterized by intricate neural networks, biochemical signaling, and unique mechanical properties. The soft and dynamic viscoelastic extracellular matrix (ECM) plays a crucial role in supporting different types of brain cells and influencing their behavior. Understanding how brain cells respond to mechanical stimuli within this complex environment is essential for unraveling fundamental mechanisms of healthy, unhealthy, and regenerative functions within the central nervous system. This requires the development of advanced materials and techniques to study the interplay between mechanical cues and cell responses. Hydrogels have become essential in this research, mimicking the brain's ECM in both chemical composition and mechanical behavior. Conventional hydrogels, while helpful, are static and lack dynamic stimulation. On the other hand, dynamic hydrogels provide reversible, dynamic stimulation, closely replicating the brain's ECM properties. This review discusses current hydrogel platforms used to investigate brain function in health and disease, focusing on traumatic brain injury (TBI)-like conditions and brain tumors. These dynamic materials offer sophisticated tools for understanding brain cell mechanobiology and developing new therapeutic approaches.

MST2 (STK3) is a major upstream kinase in the Hippo signalling pathway, an evolutionary conserved pathway in regulation of organ size, self-renewal and tissue homeostasis. Its downstream effectors are the transcriptional regulators YAP and TAZ. This pathway is regulated by a variety of factors, such as substrate stiffness or cell-cell contacts. Using a yeast two-hybrid screen, we detected a novel interaction between the kinases MST2 and CDK5, which we further confirmed by co-immunoprecipitation experiments. Cyclin-dependent kinase 5 (CDK5) is an unusual member of the family of cyclin-dependent kinases, involved in tumour growth and angiogenesis. Although a link between CDK5 and Hippo has been previously postulated, the mode of action is still elusive. Here, we show that knockdown of CDK5 causes reduced transcriptional activity of YAP and that CDK5 influences the phosphorylation levels of the Hippo upstream kinase LATS1. Moreover, a phosphoproteomics approach revealed that CDK5 interferes with the phosphorylation of DLG5, another upstream kinase, which regulates the Hippo pathway. Hence, CDK5 seems to act as a signalling hub for integrating the Hippo pathway and other signalling cascades. These interactions might have important implications for the use of CDK5 inhibitors, which are already in clinical use for tumour diseases.

During lung development, the embryonic airway originates as a wishbone-shaped epithelial tube, which undergoes a series of branching events to build the bronchial tree. This process depends crucially on cell proliferation and is thought to involve distinct branching modes: lateral branching, wherein daughter branches emerge along the length of a parent branch, and bifurcations, wherein the tip of a parent branch splits to form two new daughter branches. The developing airway is fluid-filled, and previous studies have shown that altered luminal pressure can influence rates of branching morphogenesis. However, it is not clear if altered tissue mechanics influence patterns of proliferation along the embryonic airway epithelium nor if individual branching modes are affected differently by changes in luminal pressure. Here, we focused on mechanisms of lateral branching and used as a model system the embryonic avian lung, which forms exclusively via this branching mode during early development. We used microinjected fluid droplets or pharmacological modulators of fluid secretion to alter luminal fluid pressure either locally or globally within cultured embryonic lungs. Somewhat surprisingly, we found both local and global increases in luminal pressure to suppress the formation of new lateral branches while also promoting increased epithelial proliferation. In a consistent manner, decreased luminal pressure led to an increase in lateral branching morphogenesis. Morphometric analysis of airway branching patterns revealed that altered luminal pressure shifts the overall branching program, rather than simply changing rates of morphogenesis. Taken together, these results highlight the importance of mechanical forces during airway branching and suggest that different branching modes may be affected differently by luminal fluid pressure.

Contact inhibition of proliferation is a critical cell density control mechanism governed by the Hippo signalling pathway. The biochemical signalling underlying cell density-dependent cues regulating Hippo signalling and its downstream effectors, YAP, remains poorly understood. Here, we reveal that the tight junction protein ZO-2 is required for the contact-mediated inhibition of proliferation. We additionally determined that the well-established molecular players of this process, namely Hippo kinase LATS1 and YAP, are regulated by ZO-2 and that the scaffolding function of ZO-2 promotes the interaction with and phosphorylation of YAP by LATS1. Mechanistically, YAP is phosphorylated when ZO-2 brings LATS1 and YAP together via its SH3 and PDZ domains, respectively, subsequently leading to the cytoplasmic retention and inactivation of YAP. In conclusion, we demonstrate that ZO-2 maintains Hippo signalling pathway activation by promoting the stability of LATS1 to inactivate YAP.

Biomaterial scaffold engineering presents great potential in promoting axonal regrowth after spinal cord injury (SCI), yet persistent challenges remain, including the surrounding host foreign body reaction and improper host-implant integration. Recent advances in mechanobiology spark interest in optimizing the mechanical properties of biomaterial scaffolds to alleviate the foreign body reaction and facilitate seamless integration. The impact of scaffold stiffness on injured spinal cords has not been thoroughly investigated. Herein, we introduce stiffness-varied alginate anisotropic capillary hydrogel scaffolds implanted into adult rat C5 spinal cords post-lateral hemisection. Four weeks post-implantation, scaffolds with a stiffness approaching that of the spinal cord effectively minimize the host foreign body reaction via yes-associated protein (YAP) nuclear translocation. Concurrently, the softest scaffolds maximize cell infiltration and angiogenesis, fostering significant axonal regrowth but limiting the rostral-caudal linear growth. Furthermore, as measured by atomic force microscopy (AFM), the surrounding spinal cord softens when in contact with the stiffest scaffold while maintaining a physiological level in contact with the softest one. In conclusion, our findings underscore the pivotal role of stiffness in scaffold engineering for SCI in vivo, paving the way for the optimal development of efficacious biomaterial scaffolds for tissue engineering in the central nervous system.

Age is a critical determinant of arterial disease, including aneurysm formation. Here, to understand the impact of aging on the arterial transcriptome, we leveraged RNA-sequencing data to define transcripts that change with advancing age in human arteries. Among the most repressed transcripts in aged individuals were those that are relevant for actomyosin structure and organization, including both myosin light chain kinase (MYLK) and smooth muscle γ-actin (ACTG2). This was associated with a reduction of serum response factor (SRF), which controls these transcripts via defined promoter elements. To determine the consequences of isolated Srf depletion, we conditionally deleted Srf in vascular smooth muscle of young mice (i8-SRF-KO mice). This led to a reduction of the SRF regulon, including Mylk and Actg2, and impaired arterial contractility, but left endothelial-dependent dilatation unaffected. Srf-depletion also increased aortic diameter and Alcian blue staining of the aortic media, which are cardinal features of aortopathy, such as aortic aneurysmal disease. Despite this, i8-SRF-KO mice were protected from aortic lesions elicited by angiotensin II (AngII). Proteomics demonstrated that Srf-depletion mimicked a protein signature of AngII treatment involving increases of the mechanoresponsive transcriptional coactivators YAP and TAZ and reduction of the Hippo kinase Lats2. Protection from aortopathy could be overcome by changing the order of KO induction and AngII administration resulting in advanced aneurysms in both i8-SRF-KO and control mice. Our work provides important insights into the molecular underpinnings of age-dependent changes in aortic function and mechanisms of adaptation in hypertension.

Scar formation is characterized by dynamic alterations in collagen secretion, which critically determine scar morphology and pathological progression. In fibroblasts, collagen secretion is initiated through the activation of cytokine- and integrin-mediated signaling pathways, which promote collagen gene transcription. The procollagen polypeptide α chains undergo extensive post-translational modifications, including hydroxylation and glycosylation, within the endoplasmic reticulum (ER), followed by folding and assembly into triple-helical procollagen. Subsequent intracellular trafficking involves the sequential transport of procollagen through the ER, Golgi apparatus, and plasma membrane, accompanied by further structural refinements prior to extracellular secretion. Once secreted, procollagen is enzymatically processed to form mature collagen fibrils, which drive scar tissue remodeling. Recent advances in elucidating regulation of collagen secretion have identified pivotal molecular targets, such as transforming growth factor-beta 1 (TGF-β1), prolyl 4-hydroxylase (P4H), heat shock protein 47 (HSP47), and transport and Golgi organization protein 1 (TANGO1), providing novel therapeutic strategies to mitigate pathological scar hyperplasia and improve regenerative outcomes. This review provides a comprehensive analysis of the molecular mechanisms governing collagen secretion during scar formation, with emphasis on signaling cascades, procollagen biosynthesis, intracellular transport dynamics, and post-translational modifications, thereby offering a framework for developing targeted anti-scar therapies.

Mechanical unloading leads to profound musculoskeletal degeneration, muscle wasting, and weakness. Understanding the specific signaling pathways involved is essential for uncovering effective interventions. This review provides new perspectives on mechanotransduction pathways, focusing on the critical roles of focal adhesions (FAs) and oxidative stress in skeletal muscle atrophy under mechanical unloading. As pivotal mechanosensors, FAs integrate mechanical and biochemical signals to sustain muscle structural integrity. When disrupted, these complexes impair force transmission, activating proteolytic pathways (e.g., ubiquitin-proteasome system) that accelerate atrophy. Oxidative stress, driven by mitochondrial dysfunction and NADPH oxidase-2 (NOX2) hyperactivation, exacerbates muscle degeneration through excessive reactive oxygen species (ROS) production, impaired repair mechanisms, and dysregulated redox signaling. The interplay between FA dysfunction and oxidative stress underscores the complexity of muscle atrophy pathogenesis: FA destabilization heightens oxidative damage, while ROS overproduction further disrupts FA integrity, creating a self-amplifying vicious cycle. Therapeutic strategies, such as NOX2 inhibitors, mitochondrial-targeted antioxidants, and FAK-activating compounds, promise to mitigate muscle atrophy by preserving mechanotransduction signaling and restoring redox balance. By elucidating these pathways, this review advances the understanding of muscle degeneration during unloading and identifies promising synergistic therapeutic targets, emphasizing the need for combinatorial approaches to disrupt the FA-ROS feedback loop.

Cells sense and respond to the matrix by exerting traction force through binding of integrins to an integrin-specific ligand. Here, Arg-Gly-Asp (RGD) peptide is covalently conjugated to the double-stranded DNA (dsDNA) and stem-loop DNA (slDNA) tethers with a tension tolerance of 43pN and immobilized on a PEG substrate. Unlike dsDNA, which is ruptured under high tension, leading to the removal of RGD, slDNA remains bound even when ruptured. Our results suggest that cells adapt their adhesion state by modulating actin filament polymerization and cofilin phosphorylation, effectively balancing the talin conformation to prevent dsDNA rupture and maintain normal adhesion. This phenomenon, termed integrin-ligand coupling strength, mediated cellular adaptive mechanosensing. Furthermore, we demonstrate that positive durotaxis can shift to negative durotaxis, depending on the integrin-ligand coupling strength. This study highlights the significance of the coupling strength in cell-extracellular matrix (ECM) interactions and offers new insights into designing biomaterials with tunable adhesive properties for cell-based applications.

In conditions of microgravity the human body undergoes extensive alterations in physiological function. However, it has proven challenging to determine how these changes are mediated at the molecular and cellular level. Here, we investigated whether ELKIN1, a mechanically activated ion channel, regulates changes in cellular and molecular structures in conditions of simulated microgravity. Deletion of ELKIN1 inhibited the simulated microgravity-induced alterations of cellular structure and attachment. In addition, cells lacking ELKIN1 did not exhibit changes in focal adhesion structures and redistribution of the YAP1 transcription factor in response to simulated microgravity, consistent with wild type cells. Finally, melanoma cell invasion of a collagen gel, from organotypic spheroids, was reduced in simulated microgravity, in an ELKIN1 dependent manner. Thus, the force sensing molecule, ELKIN1, modulates the impact of microgravity at both the molecular and cellular levels, revealing one of the molecular mechanisms that underpins cellular adaptations to conditions of microgravity.

Cytoplasmic β- and γ-actin isoforms, along with non-muscle myosin 2 isoforms, are tightly regulated in epithelial cells and compose the actomyosin cytoskeleton at the apical junctional complex. However, their specific role in regulating the mechanics of the membrane cortex and the organization of junctions, and which biomechanical circuitries modulate their expression remain poorly understood. Here, we show that γ-actin depletion in MDCK and other epithelial cells results in increased expression and junctional accumulation of β-actin and increased tight junction membrane tortuosity, both dependent on nonmuscle myosin-2A upregulation. The knock-out of γ-actin also decreases apical membrane stiffness and increases dynamic exchange of the cytoplasmic tight junction proteins like ZO-1 and cingulin, without affecting tight junction organization and barrier function. In summary, our findings uncover a biomechanical circuitry linking γ-actin to β-actin expression through nonmuscle myosin-2A and reveal γ-actin as a key regulator of tight junction and apical membrane cortex mechanics, and the dynamics of cytoskeleton-associated tight junction proteins in epithelial cells.