Mechanotransduction from the ECM to the genome: Are the pieces now in place?

A multitude of biochemical signaling processes have been characterized that affect gene expression and cellular activity. However, living cells often need to integrate biochemical signals with mechanical information from their microenvironment as they respond. In fact, the signals received by shape alone can dictate cell fate. This mechanotrasduction of information is powerful, eliciting proliferation, differentiation, or apoptosis in a manner dependent upon the extent of physical deformation. The cells internal "prestressed" structure and its "hardwired" interaction with the extra-cellular matrix (ECM) appear to confer this ability to filter biochemical signals and decide between divergent cell functions influenced by the nature of signals from the mechanical environment. In some instances mechanical signaling through the tissue microenvironment has been shown to be dominant over genomic defects, imparting a normal phenotype on cells that otherwise have transforming genetic lesions. This mechanical control of phenotype is postulated to have a central role in embryogenesis, tissue physiology as well as the pathology of a wide variety of diseases, including cancer. We will briefly review studies showing physical continuity between the external cellular microenvironment and the interior of the cell nucleus. Newly characterized structures, termed nuclear envelope lamina spanning complexes (NELSC), and their interactions will be described as part of a model for mechanical transduction of extracellular cues from the ECM to the genome.     

The cardiac nanoenvironment: form and function at the nanoscale

Mechanical forces in the cardiovascular system occur over a wide range of length scales. At the whole organ level, large scale forces drive the beating heart as a synergistic unit. On the microscale, individual cells and their surrounding extracellular matrix (ECM) exhibit dynamic reciprocity, with mechanical feedback moving bidirectionally. Finally, in the nanometer regime, molecular features of cells and the ECM show remarkable sensitivity to mechanical cues. While small, these nanoscale properties are in many cases directly responsible for the mechanosensitive signaling processes that elicit cellular outcomes. Given the inherent challenges in observing, quantifying, and reconstituting this nanoscale environment, it is not surprising that this landscape has been understudied compared to larger length scales. Here, we aim to shine light upon the cardiac nanoenvironment, which plays a crucial role in maintaining physiological homeostasis while also underlying pathological processes. Thus, we will highlight strategies aimed at (1) elucidating the nanoscale components of the cardiac matrix, and (2) designing new materials and biosystems capable of mimicking these features in vitro.     

Interaction between cell and extracellular matrix in heart disease: multiple roles of tenascin-C in tissue remodeling

The heart remodels myocardial tissue in physiological and pathological response. The cell-extracellular matrix (ECM) interaction provides not only structural and mechanical support but also important biological signaling during tissue remodeling. Among various ECM molecules, tenascin-C (TNC) is well known as a regulator of multiple cellular functions during embryogenesis, wound healing or cancer progression. In the heart, TNC appears in several important steps of embryonic development such as the initial differentiation of cardiomyocytes or coronary vasculo/angiogenesis, but it is not detected in a normal adult myocardium. However, TNC is found to re-express after myocardial injury and may regulate cellular behavior during tissue remodeling by modulating the attachment of cardiomyocytes to connective tissue, by enhancing migration and differentiation of myofibroblasts, and by inducing matrix metallo-proteinases. TNC also interacts with other ECM molecules and may modulate progression of fibrosis. Furthermore, transient and site specific expression of TNC closely associated with myocardial injury and inflammation suggests not only its key roles during tissue remodeling but also that TNC can be a marker for myocardial disease activity.     

Controlling BMP growth factor bioavailability: The extracellular matrix as multi skilled platform

Bone morphogenetic proteins (BMPs) belong to the TGF-β superfamily of signaling ligands which comprise a family of pluripotent cytokines regulating a multitude of cellular events. Although BMPs were originally discovered as potent factors extractable from bone matrix that are capable to induce ectopic bone formation in soft tissues, their mode of action has been mostly studied as soluble ligands in absence of the physiologically relevant cellular microenvironment. This micro milieu is defined by supramolecular networks of extracellular matrix (ECM) proteins that specifically target BMP ligands, present them to their cellular receptors, and allow their controlled release. Here we focus on functional interactions and mechanisms that were described to control BMP bioavailability in a spatio-temporal manner within the respective tissue context. Structural disturbance of the ECM architecture due to mutations in ECM proteins leads to dysregulated BMP signaling as underlying cause for connective tissue disease pathways. We will provide an overview about current mechanistic concepts of how aberrant BMP signaling drives connective tissue destruction in inherited and chronic diseases.     

活性氧、钙和心力衰竭

活性氧信号和钙信号广泛存在于机体内,两者相互作用,共同参与调节机体多种生理功能及病理过程。本文对这两个信号系统之间的相互作用及相关机制、在心力衰竭过程中的作用及可能的临床应用前景进行了综述。     

Geometric Dependence of 3D Collective Cancer Invasion

Metastasis of mesenchymal tumor cells is traditionally considered as a single-cell process. Here, we report an emergent collective phenomenon in which the dissemination rate of mesenchymal breast cancer cells from three-dimensional tumors depends on the tumor geometry. Combining experimental measurements and computational modeling, we demonstrate that the collective dynamics is coordinated by the mechanical feedback between individual cells and their surrounding extracellular matrix (ECM). We find the tissue-like fibrous ECM supports long-range physical interactions between cells, which turn geometric cues into regulated cell dissemination dynamics. Our results suggest that migrating cells in three-dimensional ECM represent a distinct class of an active particle system in which the collective dynamics is governed by the remodeling of the environment rather than direct particle-particle interactions.     

Extracellular matrix-cell interactions: Focus on therapeutic applications

Extracellular matrix (ECM) macromolecules together with a multitude of different molecules residing in the extracellular space play a vital role in the regulation of cellular phenotype and behavior. This is achieved via constant reciprocal interactions between the molecules of the ECM and the cells. The ECM-cell interactions are mediated via cell surface receptors either directly or indirectly with co-operative molecules. The ECM is also under perpetual remodeling process influencing cell-signaling pathways on its part. The fragmentation of ECM macromolecules provides even further complexity for the intricate environment of the cells. However, as long as the interactions between the ECM and the cells are in balance, the health of the body is retained. Alternatively, any dysregulation in these interactions can lead to pathological processes and finally to various diseases. Thus, therapeutic applications that are based on retaining normal ECM-cell interactions are highly rationale. Moreover, in the light of the current knowledge, also concurrent multi-targeting of the complex ECM-cell interactions is required for potent pharmacotherapies to be developed in the future.     

A Hybrid Model of Tumor–Stromal Interactions in Breast Cancer

Ductal carcinoma in situ (DCIS) is an early stage noninvasive breast cancer that originates in the epithelial lining of the milk ducts, but it can evolve into comedo DCIS and ultimately, into the most common type of breast cancer, invasive ductal carcinoma. Understanding the progression and how to effectively intervene in it presents a major scientific challenge. The extracellular matrix (ECM) surrounding a duct contains several types of cells and several types of growth factors that are known to individually affect tumor growth, but at present the complex biochemical and mechanical interactions of these stromal cells and growth factors with tumor cells is poorly understood. Here we develop a mathematical model that incorporates the cross-talk between stromal and tumor cells, which can predict how perturbations of the local biochemical and mechanical state influence tumor evolution. We focus on the EGF and TGF-β signaling pathways and show how up- or down-regulation of components in these pathways affects cell growth and proliferation. We then study a hybrid model for the interaction of cells with the tumor microenvironment (TME), in which epithelial cells (ECs) are modeled individually while the ECM is treated as a continuum, and show how these interactions affect the early development of tumors. Finally, we incorporate breakdown of the epithelium into the model and predict the early stages of tumor invasion into the stroma. Our results shed light on the interactions between growth factors, mechanical properties of the ECM, and feedback signaling loops between stromal and tumor cells, and suggest how epigenetic changes in transformed cells affect tumor progression.     

Dynamic Model for Characterizing Contractile Behaviors and Mechanical Properties of a Cardiomyocyte

Studies on the contractile dynamics of heart cells have attracted broad attention for the development of both heart disease therapies and cardiomyocyte-actuated micro-robotics. In this study, a linear dynamic model of a single cardiomyocyte cell was proposed at the subcellular scale to characterize the contractile behaviors of heart cells, with system parameters representing the mechanical properties of the subcellular components of living cardiomyocytes. The system parameters of the dynamic model were identified with the cellular beating pattern measured by a scanning ion conductance microscope. The experiments were implemented with cardiomyocytes in one control group and two experimental groups with the drugs cytochalasin-D or nocodazole, to identify the system parameters of the model based on scanning ion conductance microscope measurements, measurement of the cellular Young’s modulus with atomic force microscopy indentation, measurement of cellular contraction forces using the micro-pillar technique, and immunofluorescence staining and imaging of the cytoskeleton. The proposed mathematical model was both indirectly and qualitatively verified by the variation in cytoskeleton, beating amplitude, and contractility of cardiomyocytes among the control and the experimental groups, as well as directly and quantitatively validated by the simulation and the significant consistency of 90.5% in the comparison between the ratios of the Young’s modulus and the equivalent comprehensive cellular elasticities of cells in the experimental groups to those in the control group. Apart from mechanical properties (mass, elasticity, and viscosity) of subcellular structures, other properties of cardiomyocytes have also been studied, such as the properties of the relative action potential pattern and cellular beating frequency. This work has potential implications for research on cytobiology, drug screening, mechanisms of the heart, and cardiomyocyte-based bio-syncretic robotics.     

A genetic strategy for the dynamic and graded control of cell mechanics, motility, and matrix remodeling

Cellular mechanical properties have emerged as central regulators of many critical cell behaviors, including proliferation, motility, and differentiation. Although investigators have developed numerous techniques to influence these properties indirectly by engineering the extracellular matrix (ECM), relatively few tools are available to directly engineer the cells themselves. Here we present a genetic strategy for obtaining graded, dynamic control over cellular mechanical properties by regulating the expression of mutant mechanotransductive proteins from a single copy of a gene placed under a repressible promoter. With the use of constitutively active mutants of RhoA GTPase and myosin light chain kinase, we show that varying the expression level of either protein produces graded changes in stress fiber assembly, traction force generation, cellular stiffness, and migration speed. Using this approach, we demonstrate that soft ECMs render cells maximally sensitive to changes in RhoA activity, and that by modulating the ability of cells to engage and contract soft ECMs, we can dynamically control cell spreading, migration, and matrix remodeling. Thus, in addition to providing quantitative relationships between mechanotransductive signaling, cellular mechanical properties, and dynamic cell behaviors, this strategy enables us to control the physical interactions between cells and the ECM and thereby dictate how cells respond to matrix properties.     

Biomechanics of the lung parenchyma: critical roles of collagen and mechanical forces.

The biomechanical properties of connective tissues play fundamental roles in how mechanical interactions of the body with its environment produce physical forces at the cellular level. It is now recognized that mechanical interactions between cells and the extracellular matrix (ECM) have major regulatory effects on cellular physiology and cell-cycle kinetics that can lead to the reorganization and remodeling of the ECM. The connective tissues are composed of cells and the ECM, which includes water and a variety of biological macromolecules. The macromolecules that are most important in determining the mechanical properties of these tissues are collagen, elastin, and proteoglycans. Among these macromolecules, the most abundant and perhaps most critical for structural integrity is collagen. In this review, we examine how mechanical forces affect the physiological functioning of the lung parenchyma, with special emphasis on the role of collagen. First, we overview the composition of the connective tissue of the lung and their complex structural organization. We then describe how mechanical properties of the parenchyma arise from its composition as well as from the architectural organization of the connective tissue. We argue that, because collagen is the most important load-bearing component of the parenchymal connective tissue, it is also critical in determining the homeostasis and cellular responses to injury. Finally, we overview the interactions between the parenchymal collagen network and cellular remodeling and speculate how mechanotransduction might contribute to disease propagation and the development of small- and large-scale heterogeneities with implications to impaired lung function in emphysema.     

HSP90 and eNOS partially co-localize and change cellular localization in relation to different ECM components in 2D and 3D cultures of adult rat cardiomyocytes

Background information. Cultivation techniques promoting three‐dimensional organization of mammalian cells are of increasing interest, since they confer key functionalities of the native ECM (extracellular matrix) with a power for regenerative medicine applications. Since ECM compliance influences a number of cell functions, Matrigel‐based gels have become attractive tools, because of the ease with which their mechanical properties can be controlled. In the present study, we took advantage of the chemical and mechanical tunability of commonly used cell culture substrates, and co‐cultures to evaluate, on both two‐ and three‐dimensional cultivated adult rat cardiomyocytes, the impact of ECM chemistry and mechanics on the cellular localization of two interacting signalling proteins: HSP90 (heat‐shock protein of 90 kDa) and eNOS (endothelial nitric oxide synthase). Results. Freshly isolated rat cardiomyocytes were cultured on fibronectin, Matrigel gel or laminin, or in co‐culture with cardiac fibroblasts, and tested for both integrity and viability. As validation criteria, integrity of both plasma membrane and mitochondria was evaluated by transmission electron microscopy. Cell sensitivity to microenvironmental stimuli was monitored by immunofluorescence and confocal microscopy. We found that HSP90 and eNOS expression and localization are affected by changes in ECM composition. Elaboration of the images revealed, on Matrigel‐cultured cardiomyocytes, areas of high co‐localization between HSP90 and eNOS and co‐localization coefficients, which indicated the highest correlation with respect to the other substrates. Conclusions. Our three‐dimensional adult cardiomyocyte cultures are suitable for both analysing cell—ECM interactions at electron and confocal microscopy levels and monitoring micro‐environment impact on cardiomyocyte phenotype.     

Preparation of Hydroxy-PAAm Hydrogels for Decoupling the Effects of Mechanotransduction Cues

It is now well established that many cellular functions are regulated by interactions of cells with physicochemical and mechanical cues of their extracellular matrix (ECM) environment. Eukaryotic cells constantly sense their local microenvironment through surface mechanosensors to transduce physical changes of ECM into biochemical signals, and integrate these signals to achieve specific changes in gene expression. Interestingly, physicochemical and mechanical parameters of the ECM can couple with each other to regulate cell fate. Therefore, a key to understanding mechanotransduction is to decouple the relative contribution of ECM cues on cellular functions.Here we present a detailed experimental protocol to rapidly and easily generate biologically relevant hydrogels for the independent tuning of mechanotransduction cues in vitro. We chemically modified polyacrylamide hydrogels (PAAm) to surmount their intrinsically non-adhesive properties by incorporating hydroxyl-functionalized acrylamide monomers during the polymerization. We obtained a novel PAAm hydrogel, called hydroxy-PAAm, which permits immobilization of any desired nature of ECM proteins. The combination of hydroxy-PAAm hydrogels with microcontact printing allows to independently control the morphology of single-cells, the matrix stiffness, the nature and the density of ECM proteins. We provide a simple and rapid method that can be set up in every biology lab to study in vitro cell mechanotransduction processes. We validate this novel two-dimensional platform by conducting experiments on endothelial cells that demonstrate a mechanical coupling between ECM stiffness and the nucleus.     

The extracellular matrix in development and morphogenesis: A dynamic view

The extracellular matrix (ECM) is synthesized and secreted by embryonic cells beginning at the earliest stages of development. Our understanding of ECM composition, structure and function has grown considerably in the last several decades and this knowledge has revealed that the extracellular microenvironment is critically important for cell growth, survival, differentiation and morphogenesis. ECM and the cellular receptors that interact with it mediate both physical linkages with the cytoskeleton and the bidirectional flow of information between the extracellular and intracellular compartments. This review considers the range of cell and tissue functions attributed to ECM molecules and summarizes recent findings specific to key developmental processes. The importance of ECM as a dynamic repository for growth factors is highlighted along with more recent studies implicating the 3-dimensional organization and physical properties of the ECM as it relates to cell signaling and the regulation of morphogenetic cell behaviors. Embryonic cell and tissue generated forces and mechanical signals arising from ECM adhesion represent emerging areas of interest in this field.     

Strain Transfer in Ventricular Cardiomyocytes to Their Transverse Tubular System Revealed by Scanning Confocal Microscopy

The transverse tubular system (t-system) is a major site for signaling in mammalian ventricular cardiomyocytes including electrical signaling and excitation-contraction coupling. It consists of membrane invaginations, which are decorated with various proteins including mechanosensitive ion channels. Here, we investigated mechanical modulation of the t-system. By applying fluorescent markers, three-dimensional scanning confocal microscopy, and methods of digital image analysis, we studied isolated ventricular cardiomyocytes under different strains. We demonstrate that strain at the cellular level is transmitted to the t-system, reducing the length and volume of tubules and altering their cross-sectional shape. Our data suggest that a cellular strain of as little as 5% affects the shape of transverse tubules, which has important implications for the function of mechanosensitive ion channels found in them. Furthermore, our study supports a prior hypothesis that strain can cause fluid exchange between the t-system and extracellular space.     

Brushes,cables, and anchors: Recent insights into multiscale assembly and mechanics of cellular structural networks

The remarkable ability of living cells to sense, process, and respond to mechanical stimuli in their environment depends on the rapid and efficient interconversion of mechanical and chemical energy at specific times and places within the cell. For example, application of force to cells leads to conformational changes in specific mechanosensitive molecules which then trigger cellular signaling cascades that may alter cellular structure, mechanics, and migration and profoundly influence gene expression. Similarly, the sensitivity of cells to mechanical stresses is governed by the composition, architecture, and mechanics of the cellular cytoskeleton and extracellular matrix (ECM), which are in turn driven by molecular-scale forces between the constituent biopolymers. Understanding how these mechanochemical systems coordinate over multiple length and time scales to produce orchestrated cell behaviors represents a fundamental challenge in cell biology. Here, we review recent advances in our understanding of these complex processes in three experimental systems: the assembly of axonal neurofilaments, generation of tensile forces by actomyosin stress fiber bundles, and mechanical control of adhesion assembly.     

Mechanical Cell-Matrix Feedback Explains Pairwise and Collective Endothelial Cell Behavior In Vitro

In vitro cultures of endothelial cells are a widely used model system of the collective behavior of endothelial cells during vasculogenesis and angiogenesis. When seeded in an extracellular matrix, endothelial cells can form blood vessel-like structures, including vascular networks and sprouts. Endothelial morphogenesis depends on a large number of chemical and mechanical factors, including the compliancy of the extracellular matrix, the available growth factors, the adhesion of cells to the extracellular matrix, cell-cell signaling, etc. Although various computational models have been proposed to explain the role of each of these biochemical and biomechanical effects, the understanding of the mechanisms underlying in vitro angiogenesis is still incomplete. Most explanations focus on predicting the whole vascular network or sprout from the underlying cell behavior, and do not check if the same model also correctly captures the intermediate scale: the pairwise cell-cell interactions or single cell responses to ECM mechanics. Here we show, using a hybrid cellular Potts and finite element computational model, that a single set of biologically plausible rules describing (a) the contractile forces that endothelial cells exert on the ECM, (b) the resulting strains in the extracellular matrix, and (c) the cellular response to the strains, suffices for reproducing the behavior of individual endothelial cells and the interactions of endothelial cell pairs in compliant matrices. With the same set of rules, the model also reproduces network formation from scattered cells, and sprouting from endothelial spheroids. Combining the present mechanical model with aspects of previously proposed mechanical and chemical models may lead to a more complete understanding of in vitro angiogenesis.     

Biophysical interactions between components of the tumor microenvironment promote metastasis

During metastasis, tumor cells need to adapt to their dynamic microenvironment and modify their mechanical properties in response to both chemical and mechanical stimulation. Physical interactions occur between cancer cells and the surrounding matrix including cell movements and cell shape alterations through the process of mechanotransduction. The latter describes the translation of external mechanical cues into intracellular biochemical signaling. Reorganization of both the cytoskeleton and the extracellular matrix (ECM) plays a critical role in these spreading steps. Migrating tumor cells show increased motility in order to cross the tumor microenvironment, migrate through ECM and reach the bloodstream to the metastatic site. There are specific factors affecting these processes, as well as the survival of circulating tumor cells (CTC) in the blood flow until they finally invade the secondary tissue to form metastasis. This review aims to study the mechanisms of metastasis from a biomechanical perspective and investigate cell migration, with a focus on the alterations in the cytoskeleton through this journey and the effect of biologic fluids on metastasis. Understanding of the biophysical mechanisms that promote tumor metastasis may contribute successful therapeutic approaches in the fight against cancer.