The intricate structure of the brain at various scales is linked to its functional and biomechanical properties. Since understanding the biomechanical properties is vital for comprehending brain diseases, development, and injuries, measurements at different scales are necessary. Here we introduce methods to measure the biomechanical properties at both tissue and cellular levels using a mouse model. A specially designed magnetic resonance elastography system is introduced for imaging the mouse brain, enabling in vivo mapping of its shear modulus. Additionally, protocols for isolating and culturing primary neurons and astrocytes from the hippocampus and cerebral cortex of the mouse brain are presented. The nanoindentation technique using atomic force microscopy is employed to measure the biomechanical properties of individual cells. The results indicate that the storage/loss modulus of the mouse cerebral cortex and hippocampus are 8.07 ± 1.28 kPa / 3.20 ± 0.66 kPa and 6.60 ± 0.52 kPa / 2.52 ± 0.33 kPa, respectively. Meanwhile, the Young’s modulus for neurons and astrocytes is 470.88 ± 17.67 Pa and 681.13 ± 14.15 Pa, respectively. These findings demonstrate that the brain exhibits distinct biomechanical properties at different scales. The proposed methods offer general techniques for investigating the multiscale biomechanical properties of the brain.

The care of patients with cholestatic liver diseases such as primary sclerosing cholangitis (PSC) is challenging, partly due to the lack of knowledge of disease pathogenesis and suitable in vitro models for disease modeling and drug screening. Although the pathogenesis of cholestatic liver diseases like PSC remains unknown, the importance of the vascular-biliary interface is clear. Cholangiocyte injury not only impairs barrier function such that bile leaks and damages periductal tissue, but also activates cholangiocytes to secret pro-inflammatory and pro-fibrogenic mediators to stimulate immune cells and mesenchymal cells, ultimately causing damage to the liver. Here we describe a detailed protocol for fabricating a human vascularized bile duct-on-a-chip (VBDOC) that consists of a vascular channel, biliary channel, and neighboring mesenchymal cells in a collagen gel that models the vascular-biliary interface structurally and functionally in three dimensions. This device is notable in maintaining cholangiocyte polarity and barrier function, recapitulating physiological functions and responses of the large bile ducts, and enabling manipulation of components of the mechanical microenvironment such as matrix stiffness and shear flow in the lumens. This practical workflow could help researchers manufacture the VBDOC in their own labs and apply it to studies of various cholestatic liver diseases.

α-Synuclein (α-Syn) is a presynaptic protein primarily associated with Parkinson’s disease and other neurodegenerative diseases. The cholesterol content in SV membranes regulates α-Syn binding to synaptic vesicles, changing its function and modifying its aggregation. Using single-vesicle imaging, we show that low concentrations of cholesterol reduce vesicle clustering, and high concentrations enhance vesicle clustering mediated by α-Syn. Furthermore, using all-atom molecular dynamics simulation, we investigate the role of cholesterol in synaptic-like vesicle clustering mediated by α-Syn. In particular, we found cholesterol reduces hydrogen bonds and interaction energies in low concentrations, while high concentrations of cholesterol increase hydrogen bonds and interaction energies. Moreover, cholesterol also regulates lipid packing defects, and the condensation of cholesterol leads to the suppression of shallow packing defects, and enhancement of large defects with increasing cholesterol concentration. We revealed that cholesterol promoted vesicle clustering is due to the electrostatic interaction between cholesterol in the membrane and the N-terminal region of α-Syn. Moreover, this increased electrostatic interaction arises from a change in packing defect distribution of the protein–membrane interface induced by cholesterol condensation. This work highlights the complex interplay between α-Syn and cholesterol, emphasizing the importance of cholesterol levels in membranes and their impact on α-Syn function.

Increased extracellular matrix (ECM) stiffness, a hallmark of risk in cardiovascular disease (CVD), is closely associated with inflammation triggered by immune cell infiltration in the vessel wall. While numerous immunotherapies targeting inflammation in CVD are being developed, the specific immune components and key factors that respond to arterial stiffness remain unclear. In this work, we analyzed single-cell transcriptomics to identify immune cell populations sensitive to mechanical stress in stiffened carotid plaques. We utilized an in vitro model of polyacrylamide gels with varying stiffness and an in vivo mouse model of acute calcification to replicate arterial stiffening. An imaging flow cytometry panel was employed to determine specific cell populations and gene expression in response to ECM stiffening. The scRNA-seq analysis revealed that SPP1^high macrophages constitute a prominent myeloid population influencing extracellular matrix composition. We uncovered that macrophages exhibit elevated SPP1 protein levels when cultured on a stiffer matrix. Additionally, the percentage of SPP1^high macrophages increased in the stiffened arterial wall in the mouse model of vascular calcification. Collectively, we combined single-cell transcriptomics analysis with in vitro imaging flow cytometry studies to identify SPP1^high macrophages as a population sensitive to ECM stiffness. Our findings suggest that macrophage SPP1 could serve as a potential biomarker for patients experiencing arterial stiffening.

Deep tissue injury (DTI) is a serious condition primarily triggered by prolonged mechanical loading rather than short-term excessive force. Cyclic force stimulation has been shown to enhance cellular protection; however, the biomechanical mechanisms underlying tissue and cellular responses to such stimulation remain poorly understood. This study investigates the biomechanical effects of cyclic force transmission from viscoelastic muscle tissue to muscle fibers using a multiscale finite element model with varying cyclic force parameters. Finite element analysis was used to examine the impact of different frequencies and amplitudes of force stimulation, considering the viscoelastic properties of tissues and cells. Results indicated that, on each one of the three model scales, the average maximal Von Mises stress during stress relaxation increased with cyclic force amplitude at a constant frequency. At a fixed amplitude, frequency variations did not influence the average maximal Von Mises stress in the macroscopic and mesoscopic models. However, in the microscopic model, higher frequencies resulted in lower average maximal Von Mises stress at low amplitudes and higher average maximal Von Mises stress at high amplitudes. Notably, a high-frequency, low-amplitude cyclic force mode reduced the average maximal minimum principal stress by 3.6% in the first four seconds and 5.8% in the last 16 seconds of the 20-second stress relaxation period. These findings suggested that such a cyclic force mode may mitigate or delay mechanical damage caused by prolonged mechanical loading, offering insights into potential strategies for preventing DTI.

Circulating tumor cells (CTCs) are cancerous cells that break away from the primary tumor, enter the bloodstream, and travel to another part of the body. Research into CTCs, particularly their biological phenotypes and molecular mechanisms, has provided critical insights into metastasis and potential therapeutic targets. From a biophysical or mechanobiological perspective, CTCs must undergo biomechanical adaptations to navigate the processes of intravasation, circulation, arrest, and extravasation. These adaptations enable them to interact with blood components and survive in the circulatory system for hours or even days, ultimately facilitating metastatic progression. As research on metastasis within the bloodstream advances, this review explores the mechanobiology of CTCs, emphasizing the cellular and molecular mechanisms that regulate their suspension and adhesion states. Understanding these dynamic behaviors will offer deeper insights into CTC biology and the metastatic cascade.