Biology subjects

Neu, C.

Publications and source records attributed to Neu, C..

15 recordsLinked to original sources

bioRxiv2026

Human Osteochondral Granular Extracellular Matrix (gECM) Hydrogels Drive Tissue-Specific Composition and Mechanics

Osteochondral defects remain a major clinical challenge due to the limited regenerative capacity of cartilage and the complexity of the osteochondral interface. Here, we present a human-derived granular extracellular matrix (gECM) hydrogel platform designed for translational osteochondral repair. Using otherwise discarded human donor tissues, we developed cartilage and bone gECM hydrogels under current good manufacturing practice workflows. These materials are shear-thinning, immediately hold their form, and crosslink under physiological conditions to form stable constructs. Proteomic analysis confirmed that cartilage and bone gECM retain distinct tissue-specific biochemical signatures, while mechanical characterization demonstrated tissue-relevant stiffness, with bone gECM hydrogels exhibiting greater stiffness than cartilage gECM hydrogel. Particle packing density primarily governed viscosity, whereas tissue type contributed strongly to bulk stiffness. Together, these findings establish a scalable, human-derived gECM platform that integrates tissue-specific structural and mechanical cues, and advances a clinically translatable strategy for osteochondral repair.

bioengineering↗

bioRxiv2026

Lamin A/C coordinates nuclear mechanics, chromatin architecture, and transcriptional homeostasis in skeletal muscle in vivo

The nuclear lamina provides mechanical integrity to the eukaryotic nucleus and organizes lamina-associated chromatin domains that are important for chromatin architecture and gene-expression regulation. Lamin A/C, a major component of the nuclear lamina, is disrupted in hereditary laminopathies and has also been implicated in aging-associated nuclear dysfunction. Although the mechanical role of Lamin A/C has been extensively studied in vitro, particularly in monolayer cell culture and isolated nuclei, its role in maintaining nuclear mechanics and chromatin organization in intact tissues remains incompletely understood. Here, we investigated how partial and complete Lamin A/C disruption affects nuclear shape, chromatin architecture, intranuclear mechanics, and gene expression in vivo. Across multiple murine tissues, Lamin A/C deficiency did not cause a generalized collapse of nuclear shape or gross tissue architecture. Instead, Lamin A/C disruption preferentially altered chromatin architecture in mechanically stiff tissues, including skeletal muscle and heart. Using live in vivo deformation microscopy during controlled hindlimb muscle stimulation, we quantified real-time multiscale deformation of skeletal muscle tissue and nuclei. These measurements revealed reduced effective nuclear stiffness and altered load sharing between euchromatin-rich and heterochromatin-rich domains after Lamin A/C loss. Super-resolution imaging further showed that partial and complete Lamin A/C disruption uncoupled H3K9me3 from DAPI-dense heterochromatin, indicating a spatial disruption of repressive chromatin organization. Exploratory ATAC-seq suggested increased chromatin accessibility in heterozygous muscle, whereas RNA-seq showed that complete Lamin A/C loss caused broad myopathic transcriptional dysregulation while partial loss preserved a near-wild-type transcriptomic state. Integrated analysis identified HDAC2 as a candidate mechanosensitive compensatory node that may help buffer gene expression after partial Lamin A/C disruption. Together, these results establish Lamin A/C as an in vivo coordinator of nuclear mechanics, heterochromatin organization, and transcriptional homeostasis in skeletal muscle.

cell biology↗

bioRxiv2026

Granular Extracellular Matrix (gECM) Hydrogel Wafers as Shelf-Stable 2.5D Substrates for Microphysiological Modeling

Understanding disease pathology and evaluating emerging therapeutics require in vitro models that accurately recapitulate human tissue environments. However, existing microphysiological systems often compromise either biomimicry or ease-of-use, limiting widespread adoption and scalability. Here, we present lyophilized granular extracellular matrix (gECM) hydrogel wafers as shelf-stable, humanized 2.5D substrates that enable physiologically relevant modeling while simplifying integration into experimental workflows. Derived from decellularized human cartilage and bone, gECM hydrogel wafers retain tissue-specific architecture while introducing microporosity and surface topography through lyophilization. These wafers maintain swelling behavior, structural integrity, and mechanical properties over three months of room-temperature storage, allowing pre-fabrication and on-demand use without loss of function. gECM hydrogel wafers support direct cell seeding without encapsulation and sustain viability and proliferation of human adipose-derived mesenchymal stromal cells over 21 days, with gene expression trends comparable to 3D gECM hydrogels. Furthermore, wafers can be readily integrated into microfluidic systems with in situ hydration and transport of large biomolecules. Together, this platform bridges the gap between conventional 2D culture ease-of-use and 3D biomaterial biomimicry, providing a scalable and physiologically relevant in vitro model approach for high-throughput disease studies and therapeutic screening.

bioengineering↗

bioRxiv2026

Multi-step femtosecond laser-fabricated membranes for regulated migration of biomolecules and cells

Organ-on-chip (OoC) systems enable the recapitulation of key structural and functional characteristics of human tissues within controlled micro-engineered environments. In mechanically active tissues such as musculoskeletal, cardiac, and vascular systems, the incorporation of dynamic physical forces is essential for replicating the biomechanical cues governing cellular morphology and functional responses in-vivo. Without such stimuli, OoC models may fail to capture physiologically relevant tissue behaviors. Porous and semi-permeable membranes are critical components of OoCs, facilitating selective transport of nutrients, gases, and signaling molecules between cellular compartments to support biologically accurate barrier replication. Hence, fabrication strategies that permit precise modulation of membrane permeability are desirable to accommodate for the varying needs in pore size and porosity across organ systems. This study presents a two-stage fabrication process for stretchable, microporous polydimethylsiloxane (PDMS) membranes using femtosecond (fs-) pulse laser drilling. The laser-ablated pores exhibit a characteristic conical morphology, with diameters tapering from the laser entry to exit point. By modulating laser power and number of pulses, 6-15 m exit-end pore diameters were achieved in 50 m thick PDMS films. The membranes demonstrated strong mechanical resilience, with a 5-12% reduction in Youngs modulus after 500 cycles of strain loading. Furthermore, membranes fabricated at lower laser powers exhibited superior retention of elasticity, highlighting the influence of processing parameters on mechanical behavior. Cytocompatibility and permeability assessments confirmed that the membranes supported sustained cell viability and proliferation over at least three days. In size-restricted membrane pore geometries, cellular migration was constrained without any inhibition of biomolecular transport. This selective permeability is critical in multilayer OoC architectures, where a balance between biomolecular diffusion and cellular compartmentalization is necessary to preserve distinct tissue interfaces and functional organization. This work presents fs-laser micro-drilling as a robust and tunable fabrication strategy for producing mechanically resilient, selectively permeable PDMS membranes for physiologically relevant OoC applications.

bioengineering↗

bioRxiv2025

Perturbed cyclic strain results in a mechanically-induced, premature senescent phenotype in cardiac fibroblasts

The cardiovascular system operates under continuous cyclic mechanical stretch, and changes in mechanics and biochemical responses observed in cardiovascular disorders activate the cardiac fibroblasts (CFs) and increase cellular senescence in the tissue. However, it is unclear if cellular senescence is initiated from biomechanical stimulus alone. Here, we subjected murine CFs to uniaxial stretch and perturbed the mechanical stimulation to examine the impact on induction of a senescent phenotype. Loss of stretch magnitude and increase in frequency, mimicking an injurious hypertrophic or fibrotic response, led to a senescence-like phenotype, including cell cycle, lamin B expression, and DNA damage. Mechanical induction of CF senescence relied on p53/p21, whereas CF senescence induction triggered by reactive oxygen species or involving a mutation in lamin A/C gene occurred through p16. Moreover, mechanical induction of premature senescence was associated with decreases in the nuclear envelope protein emerin. These results demonstrate perturbed mechanical stimulation can initiate a senescent state and altered nuclear integrity may initiate this phenotype. TeaserPerturbed mechanics initiates a premature senescent-like phenotype in cardiac fibroblasts.

biophysics↗

bioRxiv2022

Increasing information gain in animal research by improving statistical model accuracy

Reduction of the numbers of laboratory animals is one of the three pillars of ethical animal research. Equivalently, information gain per animal should be maximized. A road towards this goal that is barely taken in current animal research is the more accurate statistical modeling of experiments. Here we show for a typical experiment ("open field test") with outcomes that are non-normally distributed count data, how this can be implemented and what information gain is achieved. We contrast the state of the art - the use of confidence intervals based on null-hypothesis significance testing (NHST) -, with a Bayesian approach with the same underlying normal model, and a Bayesian approach with a more accurate negative binomial model. We find that the more accurate model leads to a marked improvement of knowledge gained with the experiment, especially for small sample sizes. As experimental data that violate assumptions of simple, conventional models are frequent, our findings have wider implications.

bioinformatics↗

bioRxiv2022

High-frame-rate analysis of knee cartilage deformation by spiral dualMRI and relaxation mapping

PurposeDaily activities including walking impose high frequency cyclic forces on cartilage and repetitive compressive deformation. Analyzing cartilage deformation during walking would provide spatial maps of displacement, strain, and enable viscoelastic characterization, which may serve as imaging biomarkers for early cartilage degeneration when the damage is still reversible. However, the time-dependent biomechanics of cartilage is not well described, and how defects in the joint impact the viscoelastic response is unclear. MethodsWe used spiral acquisition with displacement encoding MRI to quantify displacement and strain maps at a high frame rate (40 ms; 25 frames/sec) in tibiofemoral joints. We also employed relaxometry methods (T1, T1{rho}, T2, T2*) on the cartilage. ResultsNormal and shear strains were concentrated on the tibiofemoral contact area during loading, and the defected joint exhibited larger compressive strains. We also determined a positive correlation between the change of T1{rho} in cartilage after cyclic loading and increased compressive strain on the defected joint. Viscoelastic behavior was quantified by the time-dependent displacement, where the damaged joint showed increased creep behavior compared to the intact. ConclusionsOur results indicate that spiral scanning with displacement encoding can quantitatively differentiate the damaged from intact joint using the strain and creep response. The viscoelastic response identified with this methodology could serve as biomarkers to detect defects in joints in vivo and facilitate the early diagnosis of joint diseases such as osteoarthritis.

bioengineering↗

bioRxiv2021

Epigenetic remodeling during monolayer cell expansion reduces therapeutic potential

Understanding how cells remember previous mechanical environments to influence their fate, or mechanical memory, informs the design of biomaterials and therapies in medicine. Current regeneration therapies require two-dimensional (2D) cell expansion processes to achieve large cell populations critical for the repair of damaged (e.g. connective and musculoskeletal) tissues. However, the influence of mechanical memory on cell fate following expansion is unknown, and mechanisms defining how physical environments influence the therapeutic potential of cells remain poorly understood. Here, we show that the organization of histone H3 trimethylated at lysine 9 (H3K9me3) and expression of tissue-identifying genes in primary cartilage cells (chondrocytes) transferred to three-dimensional (3D) hydrogels depends on the number of previous population doublings on tissue culture plastic during 2D cell expansion. Decreased levels of H3K9me3 occupying promoters of dedifferentiation genes after the 2D culture were also retained in 3D culture. Suppression of H3K9me3 during expansion of cells isolated from a murine model similarly resulted in the loss of the chondrocyte phenotype and global remodeling of nuclear architecture. In contrast, increasing levels of H3K9me3 through inhibiting H3K9 demethylases partially rescued the chondrogenic nuclear architecture and gene expression, which has important implications for tissue repair therapies, where expansion of large numbers of phenotypically-suitable cells is required. Overall, our findings indicate mechanical memory in primary cells is encoded in the chromatin architecture, which impacts cell fate and the phenotype of expanded cells. SIGNIFICANCE STATEMENTTissue regeneration procedures, such as cartilage defect repair (e.g. Matrix-induced Autologous Chondrocyte Implantation) often require cell expansion processes to achieve sufficient cells to transplant into an in vivo environment. However, the chondrocyte cell expansion on 2D stiff substrates induces epigenetic changes that persist even when the chondrocytes are transferred to a different (e.g. 3D) or in vivo environment. Treatments to alter epigenetic gene regulation may be a viable strategy to improve existing cartilage defect repair procedures and other tissue engineering procedures that involve cell expansion.

bioengineering↗

bioRxiv2021

Microinductor-Fused Atomic Force Microscopy Cantilevers for Dynamic Imaging and Multimodal Manipulation

Recent advances in atomic force microscopy (AFM) imaging and force spectroscopy have demonstrated improvements in rapid acquisition of quantitative data for applications in materials science, surface characterization, and biology. However, conventional AFM technology is limited in detection sensitivity and the ability to excite at off-resonance frequencies restricting broad utility of the technology. Here we demonstrate new AFM cantilevers fabricated with a planar microcoil at the tip region, which can be used to generate or sense highly-localized magnetic fields. Torsion/bending actuation of the cantilevers is accomplished with simple experimental configurations, enabling quantitative and simultaneous mapping of both stiffness and friction at the sample surface with more than one order of magnitude improvement in compositional contrast. Our method is compatible with commercial AFM systems, allowing us to clearly resolve small stiffness and friction variations in copolymer and biological samples that were difficult to detect by conventional AFM methods. In combination with fluorescence microscopy, we also generated localized fields to selectively stimulate and monitor structural changes in viable cells with nm-scale detail. Hybrid AFM cantilevers may be useful to characterize a broad range of complex material surfaces, in addition to combined physical and chemical analyses of single cells and biological microenvironments.

biophysics↗

bioRxiv2020

Epigenetic mechanisms mediating cell state transitions in chondrocytes.

Epigenetic modifications play critical roles in regulating cell lineage differentiation, but the epigenetic mechanisms guiding specific differentiation steps within a cell lineage have rarely been investigated. To decipher such mechanisms, we used the defined transition from proliferating (PC) into hypertrophic chondrocytes (HC) during endochondral ossification as a model. We established a map of activating and repressive histone modifications for each cell type. ChromHMM state transition analysis and Pareto-based integration of differential levels of mRNA and epigenetic marks revealed that differentiation associated gene repression is initiated by the addition of H3K27me3 to promoters still carrying substantial levels of activating marks. Moreover, the integrative analysis identified genes specifically expressed in cells undergoing the transition into hypertrophy. Investigation of enhancer profiles detected surprising differences in enhancer number, location, and transcription factor binding sites between the two closely related cell types. Furthermore, cell type-specific upregulation of gene expression was associated with a shift from low to high H3K27ac decoration. Pathway analysis identified PC-specific enhancers associated with chondrogenic genes, while HC-specific enhancers mainly control metabolic pathways linking epigenetic signature to biological functions.

developmental biology↗

bioRxiv2020

Nuclear stiffness decreases with disruption of the extracellular matrix in living tissues

Reciprocal interactions between the cell nucleus and the extracellular matrix lead to macroscale tissue phenotype changes. The extracellular environment is physically linked to the nuclear envelope and provides cues to maintain nuclear structure and cellular homeostasis regulated in part by mechanotransduction mechanisms. However, little is known about how structure and properties of the extracellular matrix in living tissues impacts nuclear mechanics, and current experimental challenges limit the ability to detect and directly measure nuclear mechanics while cells are within the native tissue environment. Here, we hypothesized that enzymatic disruption of the tissue matrix results in a softer tissue, affecting the stiffness of embedded cell and nuclear structures. We aimed to directly measure nuclear mechanics without perturbing the native tissue structure to better understand nuclear interplay with the cell and tissue microenvironments. To accomplish this, we expanded an atomic force microscopy needle-tip probe technique that probes nuclear stiffness in cultured cells to measure the nuclear envelope and cell membrane stiffness within native tissue. We validated this technique by imaging needle penetration and subsequent repair of the plasma and nuclear membranes of HeLa cells stably expressing the membrane repair protein CHMP4B-GFP. In the native tissue environment ex vivo, we found that while enzymatic degradation of viable cartilage tissues with collagenase 3 (MMP-13) and aggrecanase-1 (ADAMTS-4) decreased tissue matrix stiffness, cell and nuclear membrane stiffness is also decreased. Finally, we demonstrated the capability for cell and nucleus elastography using the AFM needle-tip technique. These results demonstrate disruption of the native tissue environment that propagates to the plasma membrane and interior nuclear envelope structures of viable cells.

biophysics↗

bioRxiv2020

Percolation of Microparticle Matrix Promotes Cell Migration and Integration while Supporting Native Tissue Architecture

Cells embedded in the extracellular matrix of tissues play a critical role in maintaining homeostasis while promoting integration and regeneration following damage or disease. Emerging engineered biomaterials utilize decellularized extracellular matrix as a tissue-specific support structure; however, many dense, structured biomaterials unfortunately demonstrate limited formability, fail to promote cell migration, and result in limited tissue repair. Here, we developed a reinforced composite material of densely packed acellular extracellular matrix microparticles in a hydrogel, termed tissue clay, that can be molded and crosslinked to mimic native tissue architecture. We utilized hyaluronic acid-based hydrogels, amorphously packed with acellular articular cartilage tissue particulated to ~125-250 microns in diameter and defined a percolation threshold of 0.57 (v/v) beyond which the compressive modulus exceeded 300kPa. Remarkably, primary chondrocytes recellularized particles within 48 hours, a process driven by chemotaxis, exhibited distributed cellularity in large engineered composites, and expressed genes consistent with native cartilage repair. We additionally demonstrated broad utility of tissue clays through recellularization and persistence of muscle, skin, and cartilage composites in a subcutaneous in vivo mouse model. Our findings suggest optimal strategies and material architectures to balance concurrent demands for large-scale mechanical properties while also supporting integration of dense musculoskeletal and connective tissues.

bioengineering↗

bioRxiv2020

Stretched-Exponential Modeling of Anomalous T1{rho} and T2 Relaxation in the Intervertebral Disc In Vivo

PurposeIntervertebral disc degeneration (IVDD), resulting in the depletion of hydrophilic glycosaminoglycans (GAGs) located in the nucleus pulposus (NP), can lead to debilitating neck and back pain. Magnetic Resonance Imaging (MRI) is a promising means of IVD assessment due to the sensitivity of MRI tissue relaxation properties to matrix composition. Furthermore, anomalous (i.e. non-monoexponential) relaxation models have shown higher sensitivity to specific matrix components compared to conventional monoexponential models. Here, we extend the use of the stretched exponential model, an anomalous relaxation model, to IVD relaxometry data. Theory and MethodsT1{rho} and T2 relaxation data were measured in the cervical IVDs of healthy volunteers and IVDs adjacent to cervical fusion, and analyzed using both conventional and stretched-exponential (SE) models. Model differences were evaluated via goodness of fit in the healthy data. Normalized histograms of the resultant quantitative MRI (qMRI) maps were described using stable distributions, and data were compared across adjacent disc segments. ResultsIn the healthy IVDs, we found lower mean squared error in the SE relaxation model fitting behavior compared to monoexponential models, supporting anomalous relaxation behavior in healthy IVDs. SE model parameter T1{rho} increased level-wise in the caudal direction, especially in the nucleus pulposus, while conventional T1{rho} and T2 monoexponential measures did not vary. For IVDs adjacent to cervical fusion, SE parameters deviated near the fusion site compared with those in the healthy population. ConclusionSE modeling of T1{rho} relaxation provides greater sensitivity to level-wise variation in IVD matrix properties compared with conventional relaxation modeling, and could provide improved sensitivity to early stages of IVD degeneration. The improved model fit and correlation between the SE T1{rho} parameter with IVD level suggests SE modeling may be a more sensitive method for detection of GAG content variation.

biophysics↗

bioRxiv2020

Image-based elastography of heterochromatin and euchromatin domains in the deforming cell nucleus

Chromatin of the eukaryotic cell nucleus comprises of microscopically dense heterochromatin and loosely packed euchromatin domains, each with distinct transcriptional ability and roles in cellular mechanotransduction. While recent methods have been developed to characterize the nucleus, measurement of intranuclear mechanics remains largely unknown. Here, we describe the development of nuclear elastography, which combines microscopic imaging and computational modeling to quantify the relative elasticity of the heterochromatin and euchromatin domains. Using contracting murine embryonic cardiomyocytes, nuclear elastography reveals that the heterochromatin is almost four times stiffer than the euchromatin at peak deformation. The relative elasticity between the two domains changes rapidly during the active deformation of the cardiomyocyte in the normal physiological condition but progresses more slowly in cells cultured in a mechanically stiff environment, although the relative stiffness at peak deformation does not change. Further, we found that the disruption of the LINC complex in cardiomyocytes compromises the intranuclear elasticity distribution resulting in elastically similar heterochromatin and euchromatin. These results provide insight into the elastography dynamics of heterochromatin and euchromatin domains, and provide a non-invasive framework to further investigate the mechanobiological function of subcellular and subnuclear domains limited only by the spatiotemporal resolution of the image acquisition method.

cell biology↗

bioRxiv2019

MACS: A Novel Concept for High-Magnification Imaging of Stretch-Activated Cells Reveals Divergent Nuclear Behavior to Different Levels of Strain

Mechanical cues from the environment influence cell behavior. Mechanisms of cellular mechanosensation are unclear, partially due to a lack of methods that can reveal dynamic processes. Here, we present a new concept for a low-cost, 3D-printed TENSCell (TENSion in Cells) device, that enables high-magnification imaging of cells during stretch. Using this device, we observed that nuclei of mouse embryonic skin fibroblasts underwent rapid and divergent responses, characterized by nuclear area expansion during 5% strain, but nuclear area shrinkage during 20% strain. Only responses to low strain were dependent on calcium signaling, while actin inhibition abrogated all nuclear responses and increased nuclear strain transfer and DNA damage. Imaging of actin dynamics during stretch revealed similar divergent trends, with F-actin shifting away from (5% strain) or towards (20% strain) the nuclear periphery. Our findings emphasize the importance of simultaneous stimulation and data acquisition to capture rapid mechanosensitive processes and suggest that mechanical confinement of nuclei through actin may be a protective mechanism during high strain loads.\n\nSTATEMENT OF SIGNIFICANCECells can sense and respond to mechanical cues in their environment. These responses can be rapid, on the time scale of seconds, and new methods are required for their acquisition and study. We introduce a new concept for a 3D-printed cell-stretch device that allows for simultaneous high-resolution imaging, while also being low-cost and easy to assemble to enable broad applicability. Using this device, we further demonstrated to importance of simultaneous stimulation and data acquisition to elicit mechanosensitive cell behavior as we observed rapid changes in nuclear size and reorganization of actin filaments around the nuclear border in skin cells. Overall, our results suggest that the rapid reorganization of actin during high loads might protect the genome from strain-induced damage.

cell biology↗