Deep-learning-based 3D cellular force reconstruction directly from volumetric images.

The forces exerted by single cells in the three-dimensional (3D) environments play a crucial role in modulating cellular functions and behaviors closely related to physiological and pathological processes. Cellular force microscopy (CFM) provides a feasible solution for quantifying mechanical interactions, which usually regains cellular forces from deformation information of extracellular matrices embedded with fluorescent beads. Owing to computational complexity, traditional 3D-CFM is usually extremely time consuming, which makes it challenging for efficient force recovery and large-scale sample analysis. With the aid of deep neural networks, this study puts forward a novel, data-driven 3D-CFM to reconstruct 3D cellular force fields directly from volumetric images with random fluorescence patterns. The deep-learning-based network is established through stacking deep convolutional neural networks (DCNN) and specific function layers. Some necessary physical information associated with constitutive relation of extracellular matrix material is coupled to the data-driven network. The mini-batch stochastic-gradient-descent and back-propagation algorithms are introduced to ensure its convergence and training efficiency. The networks not only have good generalization ability and robustness but also can recover 3D cellular forces directly from the input fluorescence image pairs. Particularly, the computational efficiency of the deep-learning-based network is at least one to two orders of magnitude higher than that of traditional 3D-CFM. This study provides a novel scheme for developing high-performance 3D-CFM to quantitatively characterize mechanical interactions between single cells and surrounding extracellular matrices, which is of vital importance for quantitative investigations in biomechanics and mechanobiology.

Catch bond-inspired hydrogels with repeatable and loading rate-sensitive specific adhesion.

Biological receptor-ligand adhesion governed by mammalian cells involves a series of mechanochemical processes that can realize reversible, loading rate-dependent specific interfacial bonding, and even exhibit a counterintuitive behavior called catch bonds that tend to have much longer lifetimes when larger pulling forces are applied. Inspired by these catch bonds, we designed a hydrogen bonding-meditated hydrogel made from acrylic acid-N-acryloyl glycinamide (AA-NAGA) copolymers and tannic acids (TA), which formed repeatable specific adhesion to polar surfaces in an ultra-fast and robust way, but hardly adhered to nonpolar materials. It demonstrated up to five-fold increase in shear adhesive strength and interfacial adhesive toughness with external loading rates varying from 5 to 500 mm min-1. With a mechanochemical coupling model based on Monte Carlo simulations, we quantitatively revealed the nonlinear dependence of rate-sensitive interfacial adhesion on external loading, which was in good agreement with the experimental data. Likewise, the developed hydrogels were biocompatible, possessed antioxidant and antibacterial properties and promoted wound healing. This work not only reports a stimuli-responsive hydrogel adhesive suitable for multiple biomedical applications, but also offers an innovative strategy for bionic designs of smart hydrogels with loading rate-sensitive specific adhesion for various emerging areas including flexible electronics and soft robotics.

High-Throughput Electromechanical Coupling Chip Systems for Real-Time 3D Invasion/Migration Assay of Cells.

Cell invasion/migration through three-dimensional (3D) tissues is not only essential for physiological/pathological processes, but a hallmark of cancer malignancy. However, how to quantify spatiotemporal dynamics of 3D cell migration/invasion is challenging. Here, this work reports a 3D cell invasion/migration assay (3D-CIMA) based on electromechanical coupling chip systems, which can monitor spatiotemporal dynamics of 3D cell invasion/migration in a real-time, label-free, nondestructive, and high-throughput way. In combination with 3D topological networks and complex impedance detection technology, this work shows that 3D-CIMA can quantitively characterize collective invasion/migration dynamics of cancer cells in 3D extracellular matrix (ECM) with controllable biophysical/biomechanical properties. More importantly, this work further reveals that it has the capability to not only carry out quantitative evaluation of anti-tumor drugs in 3D microenvironments that minimize the impact of cell culture dimensions, but also grade clinical cancer specimens. The proposed 3D-CIMA offers a new quantitative methodology for investigating cell interactions with 3D extracellular microenvironments, which has potential applications in various fields like mechanobiology, drug screening, and even precision medicine.

Geometric constraint-triggered collagen expression mediates bacterial-host adhesion.

Cells living in geometrically confined microenvironments are ubiquitous in various physiological processes, e.g., wound closure. However, it remains unclear whether and how spatially geometric constraints on host cells regulate bacteria-host interactions. Here, we reveal that interactions between bacteria and spatially constrained cell monolayers exhibit strong spatial heterogeneity, and that bacteria tend to adhere to these cells near the outer edges of confined monolayers. The bacterial adhesion force near the edges of the micropatterned monolayers is up to 75 nN, which is ~3 times higher than that at the centers, depending on the underlying substrate rigidities. Single-cell RNA sequencing experiments indicate that spatially heterogeneous expression of collagen IV with significant edge effects is responsible for the location-dependent bacterial adhesion. Finally, we show that collagen IV inhibitors can potentially be utilized as adjuvants to reduce bacterial adhesion and thus markedly enhance the efficacy of antibiotics, as demonstrated in animal experiments.

Spontaneous formation and spatial self-organization of mechanically induced mesenchymal-like cells within geometrically confined cancer cell monolayers.

There is spatiotemporal heterogeneity in cell phenotypes and mechanical properties in tumor tissues, which is associated with cancer invasion and metastasis. It is well-known that exogenous growth factors like transforming growth factor (TGF)-ß, can induce epithelial-mesenchymal transition (EMT)-based phenotypic transformation and the formation of EMT patterning on geometrically confined monolayers with mechanics heterogeneity. In the absence of exogenous TGF-ß stimulation, however, whether geometric confinement-caused mechanics heterogeneity of cancer cell monolayers alone can trigger the EMT-based phenotypic heterogeneity still remains mysterious. Here, we develop a micropattern-based cell monolayer model to investigate the regulation of mechanics heterogeneity on the cell phenotypic switch. We reveal that mechanics heterogeneity itself is enough to spontaneously induce the emergence of mesenchymal-like phenotype and asymmetrical activation of TGF-ß-SMAD signaling. Spatiotemporal dynamics of patterned cell monolayers with mesenchymal-like phenotypes is essentially regulated by tissue-scale cell behaviors like proliferation, migration as well as heterogeneous cytoskeletal contraction. The inhibition of cell contraction abrogates the asymmetrical TGF-ß-SMAD signaling activation level and the emergence of mesenchymal-like phenotype. Our work not only sheds light on the key regulation of mechanics heterogeneity caused by spatially geometric confinement on regional mesenchymal-like phenotype of cancer cell monolayers, but highlights the key role of biophysical/mechanical cues in triggering phenotypic switch.

Extracellular matrix stiffness modulates host-bacteria interactions and antibiotic therapy of bacterial internalization.

Pathogenic bacteria evolve multiple strategies to hijack host cells for intracellular survival and persistent infections. Previous studies have revealed the intricate interactions between bacteria and host cells at genetic, biochemical and even single molecular levels. Mechanical interactions and mechanotransduction exert a crucial impact on the behaviors and functions of pathogenic bacteria and host cells, owing to the ubiquitous mechanical microenvironments like extracellular matrix (ECM) stiffness. Nevertheless, it remains unclear whether and how ECM stiffness modulates bacterial infections and the sequential outcome of antibacterial therapy. Here we show that bacteria tend to adhere to and invade epithelial cells located on the regions with relatively high traction forces. ECM stiffness regulates spatial distributions of bacteria during the invasion through arrangements of F-actin cytoskeletons in host cells. Depolymerization of cytoskeletons in the host cells induced by bacterial infection decreases intracellular accumulation of antibiotics, thus preventing the eradication of invaded bacterial pathogens. These findings not only reveal the key regulatory role of ECM stiffness, but suggest that the coordination of cytoskeletons may provide alternative approaches to improve antibiotic therapy against multidrug resistant bacteria in clinic.

A Solvent Co-cross-linked Organogel with Fast Self-Healing Capability and Reversible Adhesiveness at Extreme Temperatures.

Antifreezing gels are promising in diverse engineering applications such as structural soft matters, sensors, and wearable devices. However, the capability of fast self-healing and reversible adhesiveness still remain a huge challenge for gels at extreme temperatures. Here, we proposed a solvent-involved cross-linking system composed of polyacrylic acid, polyvinyl alcohol, borax, ethylene glycol, and water, capable of antifreezing below -90 °C. It was not only antifreezing, anticrystalline, and abundant in dynamic bonds but also highly transparent, stretchable (over 800%), and conductive over the scope of temperature from -60 to 60 °C. Moreover, this gel could self-heal within 1 min and repeatedly adhere to multiple substrates including glass, metal, and rubber with an adhesive strength greater than 18 kPa. These key functions of the gel could be mostly preserved after 5 days of storage at 70% relative humidity. It is anticipated that our research opens a new scope for high-performance extreme environment-tolerant adhesives or wearable devices.