Biology subjects

Kraxner, J.

Publications and source records attributed to Kraxner, J..

7 recordsLinked to original sources

bioRxiv2026

Tunable Laminar Perfusion Coordinates Endothelial and Perivascular Remodeling in Angiogenic Vasculature-on-Chip

Perfusable vascular microphysiological systems are increasingly used to model angiogenesis, tissue crosstalk, and disease. However, many platforms still rely on oscillatory, discontinuous, or poorly controlled perfusion regimes, limiting the study of sustained flow-dependent vascular remodeling. Here, we establish a tunable, unidirectional laminar flow workflow for long-term perfusion of angiogenic vasculature-on-chip cultures and use it to investigate endothelial, perivascular, and immune cell responses to sustained flow. Using an AIM Biotech microfluidic platform containing 14-day-old human umbilical vein endothelial cell-derived angiogenic sprouts and pericytes, continuous perfusion enabled intraluminal transport of 1 m tracer beads through vessels, demonstrating stable flow across the vascular bed. Sustained laminar flow induced endothelial remodeling at both the mother vessel and sprout levels, with cellular alignment evident in both compartments. Quantitative analysis of the mother vessel further revealed Golgi polarization against the direction of flow. Sustained perfusion also increased pericyte recruitment to angiogenic sprouts and reduced endothelial proliferation within the mother vessel, consistent with flow-driven vascular maturation and quiescence. Live-cell imaging further captured directional endothelial migration against the flow, lumen remodeling, and dynamic pericyte behavior under continuous perfusion. In immune-cell assays performed under continuous-flow conditions, interactions with untreated endothelium were limited, whereas inflammatory activation increased immune-cell adhesion and crawling. These observations suggest that sustained flow supports a quiescent endothelial phenotype and demonstrate the suitability of the platform for studying inflammatory activation and immune-vascular communication under controlled hemodynamic conditions. Beyond its biological relevance, the workflow provides practical advantages for live-cell imaging, low medium consumption, and downstream perturbation studies. Moreover, the modular design of the platform makes it well suited for vascular-organ crosstalk applications. Collectively, these results establish laminar flow angiogenic vasculature-on-chip as an experimentally tractable model for studying vascular mechanobiology, vascular maturation, and dynamic cell interactions under defined hemodynamic conditions.

bioengineering↗

bioRxiv2024

Contractility of cardiac and skeletal muscle tissue increases with environmental stiffness

The mechanical interplay between contractility and mechanosensing in striated muscles is of fundamental importance for tissue morphogenesis, load adaptation, and disease progression, but remains poorly understood. In this study, we investigate the dependence of contractile force generation of cardiac and skeletal muscle on environmental stiffness. Using in vitro engineered muscle micro-tissues that are attached to flexible elastic pillars, we vary the stiffness of the microenvironment over three orders of magnitude and study its effect on contractility. We find that the active contractile force upon electrical stimulation of both cardiac and skeletal micro-tissues increases with environmental stiffness according to a strong power-law relationship. To explore the role of adhesion-mediated mechanotransduction processes, we deplete the focal adhesion protein {beta}-parvin in skeletal micro-tissues. This reduces the absolute contractile force but leaves the mechanoresponsiveness unaffected. Our findings highlight the influence of external stiffness on the adaptive behavior of muscle tissue and shed light on the complex mechanoadaptation processes in striated muscle.

biophysics↗

bioRxiv2024

Polarity-JaM: An image analysis toolbox for cell polarity, junction and morphology quantification

Cell polarity involves the asymmetric distribution of cellular components such as signaling molecules and organelles within a cell, asymmetries of a cell"s shape as well as contacts with neighbouring cells. Gradients and mechanical forces often act as global cues that bias cell polarity and orientation, and polarity is coordinated by communication between adjacent cells. Advances in fluorescence microscopy combined with deep learning algorithms for image segmentation open up a wealth of possibilities to understand cell polarity behaviour in health and disease. We have therefore developed the open-source package Polarity-JaM, which offers versatile methods for performing reproducible exploratory image analysis. Multi-channel single cell segmentation is performed using a flexible and userfriendly interface to state-of-the-art deep learning algorithms. Interpretable single-cell features are automatically extracted, including cell and organelle orientation, cell-cell contact morphology, signaling molecule gradients, as well as collective orientation, tissue-wide size and shape variation. Circular statistics of cell polarity, including polarity indices, confidence intervals, and circular correlation analysis, can be computed using our web application. We have developed data graphs for comprehensive visualisation of key statistical measures and suggest the use of an adapted polarity index when the expected polarisation direction or the direction of a global cue is known a priori. The focus of our analysis is on fluorescence image data from endothelial cells (ECs) and their polarisation behaviour. ECs line the inside of blood vessels and are essential for vessel formation and repair, as well as for various cardiovascular diseases, cancer, and inflammation. However, the general architecture of the software will allow it to be applied to other cell types and image modalities. The package is built in in Python, allowing researchers to seamlessly integrate Polarity-JaM into their image and data analysis workflows, see https://polarityjam. readthedocs.io. In addition, a web application for statistical analysis, available at www.polarityjam.com, and a Napari plugin are available, each with a graphical user interface to facilitate exploratory analysis.

bioinformatics↗

bioRxiv2023

Vimentin filaments integrate low complexity domains in a highly complex helical structure

Intermediate filaments (IFs) are integral components of the cytoskeleton. They provide cells with tissue-specific mechanical properties and are involved in numerous cellular processes. Due to their intricate architecture, a 3D structure of IFs has remained elusive. Here we use cryo-focused ion beam milling, cryo-electron microscopy and tomography, to obtain a 3D structure of vimentin IFs (VIFs). VIFs assemble into a modular, densely-packed and highly-ordered helical symmetric structure of 40 -helices in cross-section, organized into 5 protofibrils. Surprisingly, the intrinsically disordered head domains form an amyloid-like fiber in the center of VIFs, while the intrinsically disordered tails form lateral connections between the protofibrils. Our findings demonstrate how protein domains of low sequence complexity can complement well-folded protein domains to construct a biopolymer with striking strength and stretchability.

biochemistry↗

bioRxiv2020

Post-Translational Modifications Soften Intermediate Filaments

The mechanical properties of biological cells are determined by the cytoskeleton, a composite biopolymer network consisting of microtubules, actin filaments and intermediate filaments (IFs). By differential expression of cytoskeletal proteins, modulation of the network architecture and interactions between the filaments, cell mechanics may be adapted to varying requirements on the cell. Here, we focus on the intermediate filament protein vimentin and introduce post-translational modifications as an additional, much faster mechanism for mechanical modulation. We study the impact of phosphorylation on filament mechanics by recording force-strain curves using optical traps. Partial phosphorylation softens the filaments. We show that binding of the protein 14-3-3 to phosphorylated vimentin IFs further enhances this effect and speculate that in the cell 14-3-3 may serve to preserve the softening and thereby the altered cell mechanics. We explain our observation by the additional charges introduced during phosphorylation.

biophysics↗

bioRxiv2019

Lateral Subunit Coupling Determines Intermediate Filament Mechanics

The cytoskeleton is a composite network of three types of protein filaments, among which in-termediate filaments (IFs) are the most extensible ones. Two very important IFs are keratin and vimentin, which have similar molecular architectures, but different mechanical behaviors. Here we compare the mechanical response of single keratin and vimentin filaments using optical tweezers. We show that the mechanics of vimentin strongly depends on the ionic strength of the buffer and that its force-strain curve suggests a high degree of cooperativity between subunits. Indeed, a computational model indicates that in contrast to keratin, vimentin is characterized by strong lateral subunit coupling of its charged monomers during unfolding of -helices. We conclude that cells can tune their mechanics by differential use of keratin versus vimentin.

biophysics↗

bioRxiv2019

Vimentin intermediate filaments undergo irreversible conformational changes during cyclic loading

Intermediate filaments (IFs) are part of the cytoskeleton of eukaryotic cells and are thus largely responsible for the cells mechanical properties. IFs are characterized by a pronounced extensibility and remarkable resilience that enable them to support cells in extreme situations. Previous experiments showed that under strain, -helices in vimentin IFs might unfold to {beta}-sheets. Upon repeated stretching, the filaments soften, however, the remaining plastic strain is negligible. Here we observe that vimentin IFs do not recover their original stiffness on reasonable time scales, and we explain these seemingly contradicting results by introducing a third, less well-defined conformational state. Reversibility on the nanoscale can be fully rescued by introducing crosslinkers that prevent transition to the {beta}-sheet. Our results classify IFs as a nano-material with intriguing mechanical properties, which is likely to play a major role for the cells local adaption to external stimuli.

biophysics↗