More Like this

1. Variable fluid flow regimes alter human brain microvascular endothelial c ell–cell junctions and cytoskeletal structure

   https://doi.org/10.1002/cm.21687  

   Ranadewa, Dilshan ; Wu, Jingwen ; Subramanianbalachandar, Vignesh A. ; Steward, Jr, Robert L. ( September 2021 , Cytoskeleton)

   The human brain microvasculature is constantly exposed to variable fluid flow regimes and their influence on the endothelium depends in part on the synchronous cooperative behavior between cell–cell junctions and the cytoskeleton. In this study, we exposed human cerebral microvascular endothelial cells to a low laminar flow (1 dyne⋅cm⁻²), high laminar flow (10 dyne⋅cm⁻²), low oscillatory flow (±1 dyne⋅cm⁻²), or high oscillatory flow (±10 dyne⋅cm⁻²) for 24 hr. After this time, endothelial cell–cell junction and cytoskeletal structural response was characterized through observation of zonula occludens‐1 (ZO‐1), claudin‐5, junctional adhesion molecule‐A (JAM‐A), vascular endothelial cadherin (VE‐Cad), and F‐actin. In addition, we also characterized cell morphology through measurement of cell area and cell eccentricity. Our results revealed the greatest change in junctional structure reorganization for ZO‐1 and JAM‐A to be observed under low laminar flow conditions while claudin‐5 exhibited the greatest change in structural reorganization under both low and high laminar flow conditions. However, VE‐Cad displayed the greatest structural response under a high laminar flow, reflecting the unique responses each cell–cell junction protein had to each fluid flow regime. In addition, cell area and cell eccentricity displayed most significant changes under the high laminar flow and low oscillatory flow, respectively. We believe this study will be useful to the field of cell mechanics and mechanobiology.

    

   more » « less
2. Characterization of the strain-rate–dependent mechanical response of single cell–cell junctions

   https://doi.org/10.1073/pnas.2019347118  

   Esfahani, Amir Monemian ; Rosenbohm, Jordan ; Safa, Bahareh Tajvidi ; Lavrik, Nickolay V. ; Minnick, Grayson ; Zhou, Quan ; Kong, Fang ; Jin, Xiaowei ; Kim, Eunju ; Liu, Ying ; et al ( February 2021 , Proceedings of the National Academy of Sciences)

   Cell–cell adhesions are often subjected to mechanical strains of different rates and magnitudes in normal tissue function. However, the rate-dependent mechanical behavior of individual cell–cell adhesions has not been fully characterized due to the lack of proper experimental techniques and therefore remains elusive. This is particularly true under large strain conditions, which may potentially lead to cell–cell adhesion dissociation and ultimately tissue fracture. In this study, we designed and fabricated a single-cell adhesion micro tensile tester (SCAµTT) using two-photon polymerization and performed displacement-controlled tensile tests of individual pairs of adherent epithelial cells with a mature cell–cell adhesion. Straining the cytoskeleton–cell adhesion complex system reveals a passive shear-thinning viscoelastic behavior and a rate-dependent active stress-relaxation mechanism mediated by cytoskeleton growth. Under low strain rates, stress relaxation mediated by the cytoskeleton can effectively relax junctional stress buildup and prevent adhesion bond rupture. Cadherin bond dissociation also exhibits rate-dependent strengthening, in which increased strain rate results in elevated stress levels at which cadherin bonds fail. This bond dissociation becomes a synchronized catastrophic event that leads to junction fracture at high strain rates. Even at high strain rates, a single cell–cell junction displays a remarkable tensile strength to sustain a strain as much as 200% before complete junction rupture. Collectively, the platform and the biophysical understandings in this study are expected to build a foundation for the mechanistic investigation of the adaptive viscoelasticity of the cell–cell junction.

    

   more » « less
3. Mapping Tyrosine Kinase Receptor Dimerization to Receptor Expression and Ligand Affinities

   https://doi.org/10.3390/pr7050288  

   Mamer, Spencer B. ; Palasz, Alexandra A. ; Imoukhuede, P. I. ( May 2019 , Processes)

   Tyrosine kinase receptor (RTK) ligation and dimerization is a key mechanism for translating external cell stimuli into internal signaling events. This process is critical to several key cell and physiological processes, such as in angiogenesis and embryogenesis, among others. While modulating RTK activation is a promising therapeutic target, RTK signaling axes have been shown to involve complicated interactions between ligands and receptors both within and across different protein families. In angiogenesis, for example, several signaling protein families, including vascular endothelial growth factors and platelet-derived growth factors, exhibit significant cross-family interactions that can influence pathway activation. Computational approaches can provide key insight to detangle these signaling pathways but have been limited by the sparse knowledge of these cross-family interactions. Here, we present a framework for studying known and potential non-canonical interactions. We constructed generalized models of RTK ligation and dimerization for systems of two, three and four receptor types and different degrees of cross-family ligation. Across each model, we developed parameter-space maps that fully determine relative pathway activation for any set of ligand-receptor binding constants, ligand concentrations and receptor concentrations. Therefore, our generalized models serve as a powerful reference tool for predicting not only known ligand: Receptor axes but also how unknown interactions could alter signaling dimerization patterns. Accordingly, it will drive the exploration of cross-family interactions and help guide therapeutic developments across processes like cancer and cardiovascular diseases, which depend on RTK-mediated signaling. 

   more » « less
4. Probing Single-Cell Mechanical Allostasis Using Ultrasound Tweezers

   https://doi.org/10.1007/s12195-019-00578-z  

   Qian, Weiyi ; Chen, Weiqiang ( January 2019 , Cellular and Molecular Bioengineering)

   Introduction— In response to external stress, cells alter their morphology, metabolic activity, and functions to mechanically adapt to the dynamic, local environment through cell allostasis. To explore mechanotransduction in cellular allostasis, we applied an integrated micromechanical system that combines an ‘ultrasound tweezers’-based mechanical stressor and a Förster resonance energy transfer (FRET)-based molecular force biosensor, termed “actinin-sstFRET,” to monitor in situ single-cell allostasis in response to transient stimulation in real time. Methods— The ultrasound tweezers utilize 1 Hz, 10-second transient ultrasound pulses to acoustically excite a lipid-encapsulated microbubble, which is bound to the cell membrane, and apply a pico- to nano-Newton range of forces to cells through an RGD-integrin linkage. The actinin-sstFRET molecular sensor, which engages the actin stress fibers in live cells, is used to map real-time actomyosin force dynamics over time. Then, the mechanosensitive behaviors were examined by profiling the dynamics in Ca2+ influx, actomyosin cytoskeleton (CSK) activity, and GTPase RhoA signaling to define a single-cell mechanical allostasis. Results—By subjecting a 1 Hz, 10-second physical stress, single vascular smooth muscle cells (VSMCs) were observed to remodeled themselves in a biphasic mechanical allostatic manner within 30 minutes that caused them to adjust their contractility and actomyosin activities. The cellular machinery that underscores the vital role of CSK equilibrium in cellular mechanical allostasis, includes Ca2+ influx, remodeling of actomyosin CSK and contraction, and GTPase RhoA signaling. Mechanical allostasis was observed to be compromised in VSMCs from patients with type II diabetes mellitus (T2DM), which could potentiate an allostatic maladaptation. Conclusions— By integrating tools that simultaneously permit localized mechanical perturbation and map actomyosin forces, we revealed distinct cellular mechanical allostasis profiles in our micromechanical system. Our findings of cell mechanical allostasis and maladaptation provide the potential for mechanophenotyping cells to reveal their pathogenic contexts and their biophysical mediators that underlie multi-etiological diseases such as diabetes, hypertension, or aging. 

   more » « less
5. Magnesium Regulates Endothelial Barrier Functions through TRPM7, MagT1, and S1P1

   https://doi.org/10.1002/advs.201901166  

   Zhu, Donghui ; You, Jing ; Zhao, Nan ; Xu, Huaxi ( July 2019 , Advanced Science)

   Mg²⁺‐deficiency is linked to hypertension, Alzheimer's disease, stroke, migraine headaches, cardiovascular diseases, and diabetes, etc., but its exact role in these pathophysiological conditions remains elusive. Mg²⁺can regulate vascular functions, yet the mechanistic insight remains ill‐defined. Data show that extracellular Mg²⁺enters endothelium mainly through the TRPM7 channel and MagT1 transporter. Mg²⁺can act as an antagonist to reduce Ca²⁺signaling in endothelium. Mg²⁺also reduces the intracellular reactive oxygen species (ROS) level and inflammation. In addition, Mg²⁺‐signaling increases endothelial survival and growth, adhesion, and migration. Endothelial barrier integrity is significantly enhanced with Mg²⁺‐treatment through S1P1‐Rac1 pathways and barrier‐stabilizing mediators including cAMP, FGF1/2, and eNOS. Mg²⁺also promotes cytoskeletal reorganization and junction proteins to tighten up the barrier. Moreover, Mg²⁺‐deficiency enhances endothelial barrier permeability in mice, and Mg²⁺‐treatment rescues histamine‐induced transient vessel hyper‐permeability in vivo. In summary, Mg²⁺‐deficiency can cause deleterious effects in endothelium integrity, and Mg²⁺‐treatment may be effective in the prevention or treatment of vascular dysfunction.

    

   more » « less