A Microactuator Array Based on Ionic Electroactive Artificial Muscles for Cell Mechanical Stimulation.

Mechanical stimulation is prevalent within organisms, and appropriate regulation of such stimulation can significantly enhance cellular functions. Consequently, the in vitro construction and simulation of mechanical stimulation have emerged as a research hotspot in biomechanics. In recent years, a class of artificial muscles named electroactive polymers (EAPs), especially ionic EAPs, have shown promising applications in biomechanics. While several techniques utilizing ionic EAPs for cell mechanical stimulation have been reported, further research is needed to advance and enhance their practical applications. Here, we prepared a microactuator array based on ionic EAP artificial muscles for cell mechanical stimulation. As a preliminary effort, we created a 5 × 5 microactuator array on a supporting membrane by employing laser cutting. We evaluated the electro-actuation performance of the microactuators through experimental testing and numerical simulations, affirming the potential use of the microactuator array for cell mechanical stimulation. The devised approach could inspire innovative design concepts in the development of miniaturized intelligent electronic devices, not only in biomechanics and biomimetics but also in other related fields.

Engineering Soft Spring Gauges for In Situ Biomaterial and Tissue Weighing.

Hydrogels have gained great attention and broad applications in tissue engineering, regenerative medicine, and drug delivery due to their excellent biocompatibility and degradability. However, accurately and noninvasively characterizing the degradation process of hydrogels remains a challenge. To address this, we have developed a method using soft spring gauges (SSGs) for the in situ weighing of hydrogels. Our approach uses a simple hydrogel-based sacrificial template method to fabricate polydimethylsiloxane (PDMS) SSGs. The SSGs used in this study can characterize hydrogels with a minimum wet weight of approximately 30 mg. Through theoretical derivations, numerical simulations, and experimental characterization, we confirmed that the length change of the SSGs in a buffer solution correlates linearly with the applied hanging weights. This allows us to track and assess the solid mass change of hydrogels during degradation with high feasibility and accuracy. Additionally, we have demonstrated the potential application of SSGs for the in situ characterization of engineered tissue growth. This method represents an advanced approach for in situ hydrogel weighing, holding great promise for advancing the development of hydrogels and other biomaterials in biomedical applications.

Mechanotherapy in oncology: Targeting nuclear mechanics and mechanotransduction.

Mechanotherapy is proposed as a new option for cancer treatment. Increasing evidence suggests that characteristic differences are present in the nuclear mechanics and mechanotransduction of cancer cells compared with those of normal cells. Recent advances in understanding nuclear mechanics and mechanotransduction provide not only further insights into the process of malignant transformation but also useful references for developing new therapeutic approaches. Herein, we present an overview of the alterations of nuclear mechanics and mechanotransduction in cancer cells and highlight their implications in cancer mechanotherapy.

Differential Responses of Primary Astrocytes Under Hyperthermic Temperatures.

Astrocyte is the most abundant cells in brain and plays critical roles in brain homeostasis and functions. Although hyperthermia (or fever) is a common symptom in patients, its influence on astrocyte viability, morphology, and functions remains elusive. Here we developed an in vitro astrocyte culture system capable of precisely controlling culture temperature to study astrocyte responses under clinically-relevant hyperthermic temperatures (38 ∼ 41 °C). We found that hyperthermia in this temperature range does not alter cell morphology, but significantly affects cell viability, activation and functions. Specifically, high-hyperthermia (40 °C and 41 °C) causes irreversible and permanent damages to astrocytes and compromises their normal viability and functionalities repairing damaged neural tissue, recycling neurotransmitters, and promoting brain development, while mild-hyperthermia (38 °C and 39 °C) induces astrocyte activation and cytokine secretion without significant decreases in cell viability. This study sheds new insights into our understanding of various fever-associated symptoms, enabling the future development of astrocyte-targeted therapy to treat brain diseases via hyperthermia.

An ECM-Mimicking, Injectable, Viscoelastic Hydrogel for Treatment of Brain Lesions.

Brain lesions can arise from traumatic brain injury, infection, and craniotomy. Although injectable hydrogels show promise for promoting healing of lesions and health of surrounding tissue, enabling cellular ingrowth and restoring neural tissue continue to be challenging. It is hypothesized that these challenges arise in part from the mismatch of composition, stiffness, and viscoelasticity between the hydrogel and the brain parenchyma, and this hypothesis is tested by developing and evaluating a self-healing hydrogel that not only mimics the composition, but also the stiffness and viscoelasticity of native brain parenchyma. The hydrogel is crosslinked by dynamic boronate ester bonds between phenylboronic acid grafted hyaluronic acid (HA-PBA) and dopamine grafted gelatin (Gel-Dopa). This HA-PBA/Gel-Dopa hydrogel could be injected into a lesion cavity in a shear-thinning manner with rapid hemostasis, high tissue adhesion, and efficient self-healing. In an in vivo mouse model of brain lesions, the multi-functional injectable hydrogel is found to support neural cell infiltration, decrease astrogliosis and glial scars, and close the lesions. The results suggest a role for extracellular matrix-mimicking viscoelasticity in brain lesion healing, and motivate additional experimentation in larger animals as the technology progresses toward potential application in humans.

Dynamic and static biomechanical traits of cardiac fibrosis.

Cardiac fibrosis is a common pathology in cardiovascular diseases which are reported as the leading cause of death globally. In recent decades, accumulating evidence has shown that the biomechanical traits of fibrosis play important roles in cardiac fibrosis initiation, progression and treatment. In this review, we summarize the four main distinct biomechanical traits (i.e., stretch, fluid shear stress, ECM microarchitecture, and ECM stiffness) and categorize them into two different types (i.e., static and dynamic), mainly consulting the unique characteristic of the heart. Moreover, we also provide a comprehensive overview of the effect of different biomechanical traits on cardiac fibrosis, their transduction mechanisms, and in-vitro engineered models targeting biomechanical traits that will aid the identification and prediction of mechano-based therapeutic targets to ameliorate cardiac fibrosis.

A positive mechanobiological feedback loop controls bistable switching of cardiac fibroblast phenotype.

Cardiac fibrosis is associated with activation of cardiac fibroblasts (CFs), a pathological, phenotypic transition that is widely believed to be irreversible in the late stages of disease development. Sensing of a stiffened mechanical environment through regulation of integrin-based adhesion plaques and activation of the Piezo1 mechanosensitive ion channel is known to factor into this transition. Here, using integrated in vitro and in silico models, we discovered a mutually reinforcing, mechanical positive feedback loop between integrin ß1 and Piezo1 activation that forms a bistable switch. The bistable switch is initiated by perturbations in matrix elastic modulus that amplify to trigger downstream signaling involving Ca2+ and YAP that, recursively, leads fibroblasts to further stiffen their environment. By simultaneously interfering with the newly identified mechanical positive feedback loop and modulating matrix elastic modulus, we reversed markers of phenotypical transition of CF, suggesting new therapeutic targets for fibrotic disease.

Plasmonic Ag Nanoparticle-Loaded n-p Bi2O2CO3/α-Bi2O3 Heterojunction Microtubes with Enhanced Visible-Light-Driven Photocatalytic Activity.

In this study, n-p Bi2O2CO3/α-Bi2O3 heterojunction microtubes were prepared via a one-step solvothermal route in an H2O-ethylenediamine mixed solvent for the first time. Then, Ag nanoparticles were loaded onto the microtubes using a photo-deposition process. It was found that a Bi2O2CO3/α-Bi2O3 heterostructure was formed as a result of the in situ carbonatization of α-Bi2O3microtubes on the surface. The photocatalytic activities of α-Bi2O3 microtubes, Bi2O2CO3/α-Bi2O3 microtubes, and Ag nanoparticle-loaded Bi2O2CO3/α-Bi2O3 microtubes were evaluated based on their degradation of methyl orange under visible-light irradiation (λ > 420 nm). The results indicated that Bi2O2CO3/α-Bi2O3 with a Bi2O2CO3 mass fraction of 6.1% exhibited higher photocatalytic activity than α-Bi2O3. Loading the microtubes with Ag nanoparticles significantly improved the photocatalytic activity of Bi2O2CO3/α-Bi2O3. This should be ascribed to the internal static electric field built at the heterojunction interface of Bi2O2CO3 and α-Bi2O3 resulting in superior electron conductivity due to the Ag nanoparticles; additionally, the heterojunction at the interfaces between two semiconductors and Ag nanoparticles and the local electromagnetic field induced by the surface plasmon resonance effect of Ag nanoparticles effectively facilitate the photoinduced charge carrier transfer and separation of α-Bi2O3. Furthermore, loading of Ag nanoparticles leads to the formation of new reactive sites, and a new reactive species ·O2− for photocatalysis, compared with Bi2O2CO3/α-Bi2O3.

Bioinspired Microstructure Platform for Modular Cell-Laden Microgel Fabrication.

Cell-laden microgels have attracted increasing interest in various biomedical fields, as living building blocks to construct spatially organized multicellular structures or complex tissue features (e.g., cell spheroids and aligned cells/fibers). Although numerous approaches have been developed to tailor cell-laden microgels, there is still an unmet need for modular, versatile, convenient, and high-throughput methods. In this study, as inspired by the phenomena of water droplet manipulation from natural microstructures, a novel platform is developed to manipulate microscale hydrogel droplets and fabricate modular cell-laden microgels. First, taking antenna-like trichome as a template, catcher-like bioinspired microstructures are fabricated and hydrogel droplets are manipulated modularly in a versatile, convenient, and high-throughput manner, which is compatible with various types of hydrogels (e.g., photo-cross-linking, thermal-cross-linking, and ion-cross-linking). It is demonstrated that this platform can manipulate cell-laden microgels as modular units, such as two or more cell-laden microgels on one single catcher-like structure and different structures on one single chip. The authors also demonstrate the application of this platform on constructing complex tissue features like myocardial fibrosis tissue models to study cardiac fibrosis. The developed platform will be a powerful tool for engineering various in vitro tissue models for widespread biomedical applications.

A new model of myofibroblast-cardiomyocyte interactions and their differences across species.

Although coupling between cardiomyocytes and myofibroblasts is well known to affect the physiology and pathophysiology of cardiac tissues across species, relating these observations to humans is challenging because the effect of this coupling varies across species and because the sources of these effects are not known. To identify the sources of cross-species variation, we built upon previous mathematical models of myofibroblast electrophysiology and developed a mechanoelectrical model of cardiomyocyte-myofibroblast interactions as mediated by electrotonic coupling and transforming growth factor-ß1. The model, as verified by experimental data from the literature, predicted that both electrotonic coupling and transforming growth factor-ß1 interaction between myocytes and myofibroblast prolonged action potential in rat myocytes but shortened action potential in human myocytes. This variance could be explained by differences in the transient outward K+ current associated with differential Kv4.2 gene expression across species. Results are useful for efforts to extrapolate the results of animal models to the predicted effects in humans and point to potential therapeutic targets for fibrotic cardiomyopathy.

Effect of gene mutation of plants on their mechano-sensibility: the mutant of EXO70H4 influences the buckling of Arabidopsis trichomes.

With the development of molecular biology, more and more mutants of plants have been constructed, where gene mutants have been found to influence not only the biological processes but also biophysical behaviors of plant cells. Trichomes are an important appendage, which has been found to act as an active mechanosensory switch transducing mechanical signals into physiology changes, where the mechanical property of trichomes is vital for such functions. Up to now, over 40 different genes have been found with the function of regulating trichome cell morphogenesis; however, the effect of gene mutants on trichome mechanosensory function remains elusive. In this study, we found that EXO70H4, one of the most up-regulated genes in the mature trichome, not only affects the thickness of the trichome cell wall but also the mechanical property (i.e., the Young's modulus) of trichomes. Finite element method simulation results show that the buckling instability and stress concentration (e.g., exerted by insects) cannot occur on the base of the mutant exo70H4 trichome, which might further interrupt the mechanical signal transduction from branches to the base of trichomes. These results indicated that the mutant exo70H4 trichome might lack the ability to act as an active mechanosensory switch against chewing insect herbivores. Our findings provide new information about the effect of gene mutation (like crop mutants) on the mechano-sensibility and capability to resist the agricultural pests or lodging, which could be of great significance to the development of agriculture.

Investigating the Effect of Substrate Stiffness on the Redox State of Cardiac Fibroblasts Using Scanning Electrochemical Microscopy.

Cardiac fibrosis, in which cardiac fibroblasts differentiate into myofibroblasts, leads to oversecretion of the extracellular matrix, results in increased stiffness, and facilitates disequilibrium of cellular redox state, further leading to oxidative stress and various degrees of cell death. However, the relationship between the matrix stiffness and the redox status of cardiac fibroblasts remains unclear. In this work, we constructed an in vitro cardiac fibrosis model by culturing cardiac fibroblasts on polyacrylamide gels with tunable stiffness and characterized the differentiation of cardiac fibroblasts to myofibroblasts by immunofluorescence staining of α-smooth muscle actin. We then applied scanning electrochemical microscopy (SECM) with a depth scan mode to in situ and quantitatively assess the redox status by monitoring the glutathione (GSH) efflux rate (k) through the redox reaction between GSH (a typical indicator of cellular redox level) released from cardiac fibroblasts and SECM probe-oxidized ferrocenecarboxylic acid ([FcCOOH]+). The SECM results demonstrate that the GSH efflux from the cardiac fibroblasts decreased with increasing substrate stiffness (i.e., mimicking the increased fibrosis degree), indicating that a more oxidizing microenvironment facilitates the cell differentiation and GSH may serve as a biomarker to predict the degree of cardiac fibrosis. This work provides an SECM approach to quantify the redox state of cardiac fibroblasts by recording the GSH efflux rate. In addition, the newly established relationship between the redox balance and the substrate stiffness would help to better understand the redox state of cardiac fibroblasts during cardiac fibrosis.

The Plasticity of Nanofibrous Matrix Regulates Fibroblast Activation in Fibrosis.

Natural extracellular matrix (ECM) mostly has a fibrous structure that supports and mechanically interacts with local residing cells to guide their behaviors. The effect of ECM elasticity on cell behaviors has been extensively investigated, while less attention has been paid to the effect of matrix fiber-network plasticity at microscale, although plastic remodeling of fibrous matrix is a common phenomenon in fibrosis. Here, a significant decrease is found in plasticity of native fibrotic tissues, which is associated with an increase in matrix crosslinking. To explore the role of plasticity in fibrosis development, a set of 3D collagen nanofibrous matrix with constant modulus but tunable plasticity is constructed by adjusting the crosslinking degree. Using plasticity-controlled 3D culture models, it is demonstrated that the decrease of matrix plasticity promotes fibroblast activation and spreading. Further, a coarse-grained molecular dynamic model is developed to simulate the cell-matrix interaction at microscale. Combining with molecular experiments, it is revealed that the enhanced fibroblast activation is mediated through cytoskeletal tension and nuclear translocation of Yes-associated protein. Taken together, the results clarify the effects of crosslinking-induced plasticity changes of nanofibrous matrix on the development of fibrotic diseases and highlight plasticity as an important mechanical cue in understanding cell-matrix interactions.

Engineering Biomaterials and Approaches for Mechanical Stretching of Cells in Three Dimensions.

Mechanical stretch is widely experienced by cells of different tissues in the human body and plays critical roles in regulating their behaviors. Numerous studies have been devoted to investigating the responses of cells to mechanical stretch, providing us with fruitful findings. However, these findings have been mostly observed from two-dimensional studies and increasing evidence suggests that cells in three dimensions may behave more closely to their in vivo behaviors. While significant efforts and progresses have been made in the engineering of biomaterials and approaches for mechanical stretching of cells in three dimensions, much work remains to be done. Here, we briefly review the state-of-the-art researches in this area, with focus on discussing biomaterial considerations and stretching approaches. We envision that with the development of advanced biomaterials, actuators and microengineering technologies, more versatile and predictive three-dimensional cell stretching models would be available soon for extensive applications in such fields as mechanobiology, tissue engineering, and drug screening.

Fluorescent conjugated polymer nanovector for in vivo tracking and regulating the fate of stem cells for restoring infarcted myocardium.

Stem cell therapy holds great promise for cardiac regeneration. However, the lack of ability to control stem cell fate after in vivo transplantation greatly restricts its therapeutic outcomes. MicroRNA delivery has emerged as a powerful tool to control stem cell fate for enhanced cardiac regeneration. However, the clinical translation of therapy based on gene-transfected stem cells remains challenging, due to the unknown in vivo behaviors of stem cells. Here, we developed a nano-platform (i.e., PFBT@miR-1-Tat NPs) that can achieve triggered release of microRNA-1 to promote cardiac differentiation of mesenchymal stem cells (MSCs), and long-term tracking of transplanted MSCs through bright and ultra-stable fluorescence of conjugated polymer poly(9,9-dioctylfluorene-alt-benzothiadiazole) (PFBT). We found that PFBT@miR-1-Tat NP-treated MSCs significantly restored the infarcted myocardium by promoting stem cell cardiac differentiation and integration with the in situ cardiac tissues. Meanwhile, MSCs without gene delivery improved the infarcted heart functions mainly through a paracrine effect and blood vessel formation. The developed conjugated polymer nanovector should be a powerful tool for manipulating as well as revealing the fate of therapeutic cells in vivo, which is critical for optimizing the therapeutic route of gene and cell combined therapy and therefore for accelerating clinical translation. STATEMENT OF SIGNIFICANCE: The lack of controllability in stem cell fate and the unclear in vivo cellular behaviors restrict the therapeutic outcomes of stem cell therapy. Herein, we engineered fluorescent conjugated polymer nanoparticles as gene delivery nanovectors with controlled release and high intracellular delivery capability to harness the fate of mesenchymal stem cells (MSCs) in vivo, meanwhile to reveal the cellular mechanism of gene-treated stem cell therapy. As compared with only MSC treatment that improves infarcted myocardium functions through paracrine effect, treatment with conjugated polymer nanovector-treated MSCs significantly restored infarcted myocardium through enhancing MSC cardiac differentiation and integration with the in-situ cardiac tissues. These findings demonstrate that the conjugated polymer nanovector would be a powerful tool in optimizing gene and cell combined therapy.

Matrix stiffness controls cardiac fibroblast activation through regulating YAP via AT1 R.

Cardiac fibrosis is a common pathway leading to heart failure and involves continued activation of cardiac fibroblasts (CFs) into myofibroblasts during myocardium damage, causing excessive deposition of the extracellular matrix (ECM) and thus increases matrix stiffness. Increasing evidence has shown that stiffened matrix plays an important role in promoting CF activation and cardiac fibrosis, and several signaling factors mediating CF mechanotransduction have been identified. However, the key molecules that perceive matrix stiffness to regulate CF activation remain to be further explored. Here, we detected significantly increased expression and nuclear localization of Yes-associated protein (YAP) in native fibrotic cardiac tissues. By using mechanically regulated in vitro cell culture models, we found that a stiff matrix-induced high expression and nuclear localization of YAP in CFs, accompanied by enhanced cell activation. We also demonstrated that YAP knockdown decreased fibrogenic response of CFs and that YAP overexpression promoted CF activation, indicating that YAP plays an important role in mediating matrix stiffness-induced CF activation. Further mechanistic studies revealed that the YAP pathway is an important signaling branch downstream of angiotensin II type 1 receptor in CF mechanotransduction. The findings help elucidate the mechanism of fibrotic mechanotransduction and may contribute to the development of new approaches for treating fibrotic diseases.

Nanoscale integrin cluster dynamics controls cellular mechanosensing via FAKY397 phosphorylation.

Transduction of extracellular matrix mechanics affects cell migration, proliferation, and differentiation. While this mechanotransduction is known to depend on the regulation of focal adhesion kinase phosphorylation on Y397 (FAKpY397), the mechanism remains elusive. To address this, we developed a mathematical model to test the hypothesis that FAKpY397-based mechanosensing arises from the dynamics of nanoscale integrin clustering, stiffness-dependent disassembly of integrin clusters, and FAKY397 phosphorylation within integrin clusters. Modeling results predicted that integrin clustering dynamics governs how cells convert substrate stiffness to FAKpY397, and hence governs how different cell types transduce mechanical signals. Existing experiments on MDCK cells and HT1080 cells, as well as our new experiments on 3T3 fibroblasts, confirmed our predictions and supported our model. Our results suggest a new pathway by which integrin clusters enable cells to calibrate responses to their mechanical microenvironment.

Solvent-Free Fabrication of Carbon Nanotube/Silk Fibroin Electrospun Matrices for Enhancing Cardiomyocyte Functionalities.

Cardiac tissue engineering holds great potential in regenerating functional cardiac tissues for various applications. The major strategy is to design scaffolds recapitulating the native cardiac microenvironment to enhance cell and tissue functionalities. Among various biomaterial systems, nanofibrous matrices with aligned morphologies and enhanced conductivity incline to induce the formation of oriented engineered cardiac tissues with enhanced functionalities. The challenge is to functionalize the scaffolds with conductive additives without influencing their biocompatibility. In this study, we developed a fully aqueous process for the fabrication of conductive carbon nanotube/silk fibroin (CNT/silk) electrospun scaffolds. The carbon nanotubes are well dispersed within the nanofibers, providing the scaffolds with enhanced conductivity and excellent biocompatibility for the culture of neonatal rat cardiomyocytes with improved cell spreading and enhanced expression of cardiac-specific proteins. Moreover, the aligned CNT/silk fibroin composite scaffolds exhibit abilities to guide the oriented organization of cardiac tissues and the biomimicking distribution of sarcomeres and gap junctions. The findings demonstrate the great potential of the CNT/silk scaffolds prepared through this aqueous processing method in supporting the formation of cardiac tissues with enhanced functionalities.

Microchannel Stiffness and Confinement Jointly Induce the Mesenchymal-Amoeboid Transition of Cancer Cell Migration.

The physical confinement of cell microenvironment could enhance the invasive capability and drug resistance of cancer cells. However, due to the lack of in vitro experimental platform to mimic both stiffness and confinement of the tumor microenvironment, the underlying mechanism remains elusive. Here, we developed a hydrogel-based microchannel platform with independently tunable channel stiffness and width in a physiological range. We found that the migration speed of the cancer cell is influenced by the synergistic effect of channel stiffness and width. In addition, the mesenchymal-amoeboid transition has a strong correlation with the channel stiffness. Besides, with a developed computational model, the role of nuclear stiffness on cancer migration speed and thus the mesenchymal-amoeboid transition in microchannels was also revealed. This platform is capable of mimicking the native physical microenvironment during metastasis, providing a powerful tool for high-throughput screening applications and investigating the interaction between cancer migration and biophysical microenvironment.