The LINC Complex Regulates Tendon Elastic Modulus, Collagen Crimp, and Lateral Expansion During Early Postnatal Development.

There is limited understanding of how mechanical signals regulate tendon development. The nucleus has emerged as a major regulator of cellular mechanosensation via the linker of nucleoskeleton and cytoskeleton (LINC) protein complex. Specific roles of LINC in tenogenesis have not been explored.

Depletion of SUN1/2 induces heterochromatin accrual in mesenchymal stem cells during adipogenesis.

Critical to the mechano-regulation of mesenchymal stem cells (MSC), Linker of the Nucleoskeleton and Cytoskeleton (LINC) complex transduces cytoskeletal forces to the nuclei. The LINC complex contains outer nuclear membrane Nesprin proteins that associate with the cytoskeleton and their inner nuclear membrane couplers, SUN proteins. Here we tested the hypothesis that severing of the LINC complex-mediated cytoskeletal connections may have different effects on chromatin organization and MSC differentiation than those due to ablation of SUN proteins.

Low intensity mechanical signals promote proliferation in a cell-specific manner: Tailoring a non-drug strategy to enhance biomanufacturing yields.

Biomanufacturing relies on living cells to produce biotechnology-based therapeutics, tissue engineering constructs, vaccines, and a vast range of agricultural and industrial products. With the escalating demand for these bio-based products, any process that could improve yields and shorten outcome timelines by accelerating cell proliferation would have a significant impact across the discipline. While these goals are primarily achieved using or strategies, harnessing cell mechanosensitivity represents a promising - albeit less studied - pathway to promote bioprocessing endpoints, yet identifying which mechanical parameters influence cell activities has remained elusive.

Development and characterization of a low intensity vibrational system for microgravity studies.

Extended-duration human spaceflight necessitates a better understanding of the physiological impacts of microgravity. While the ground-based microgravity simulations identified low intensity vibration (LIV) as a possible countermeasure, how cells may respond to LIV under real microgravity remain unexplored. In this way, adaptation of LIV bioreactors for space remains limited, resulting in a significant gap in microgravity research.

Prrx1-driven LINC complex disruption in vivo reduces osteoid deposition but not bone quality after voluntary wheel running.

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex serves to connect the nuclear envelope and the cytoskeleton, influencing cellular processes such as nuclear arrangement, architecture, and mechanotransduction. The role LINC plays in mechanotransduction pathways in bone progenitor cells has been well studied; however, the mechanisms by which LINC complexes govern in vivo bone formation remain less clear. To bridge this knowledge gap, we established a murine model disrupting LINC using transgenic Prx-Cre mice and floxed Tg(CAG-LacZ/EGFP-KASH2) mice.

Data-Driven and Cell-Specific Determination of Nuclei-Associated Actin Structure.

Quantitative and volumetric assessment of filamentous actin fibers (F-actin) remains challenging due to their interconnected nature, leading researchers to utilize threshold based or qualitative measurement methods with poor reproducibility. Here we introduce a novel machine learning based methodology for accurate quantification and reconstruction of nuclei-associated F-actin. Utilizing a Convolutional Neural Network (CNN), we segment actin filaments and nuclei from 3D confocal microscopy images and then reconstruct each fiber by connecting intersecting contours on cross-sectional slices.

Nuclear actin structure regulates chromatin accessibility.

Polymerized β-actin may provide a structural basis for chromatin accessibility and actin transport into the nucleus can guide mesenchymal stem cell (MSC) differentiation. Using MSC, we show that using CK666 to inhibit Arp2/3 directed secondary actin branching results in decreased nuclear actin structure, and significantly alters chromatin access measured with ATACseq at 24 h. The ATAC-seq results due to CK666 are distinct from those caused by cytochalasin D (CytoD), which enhances nuclear actin structure.

Porcine computational modeling to investigate developmental dysplasia of the hip.

While it is well-established that early detection and initiation of treatment of developmental dysplasia of the hip (DDH) is crucial to successful clinical outcomes, research on the mechanics of the hip joint during healthy and pathological hip development in infants is limited. Quantification of mechanical behavior in both the healthy and dysplastic developing joints may provide insight into the causes of DDH and facilitate innovation in treatment options. In this study, subject-specific three-dimensional finite element models of two pigs were developed: one healthy pig and one pig with induced dysplasia in the right hindlimb.

Osterix-driven LINC complex disruption in vivo diminishes osteogenesis at 8 weeks but not at 15 weeks.

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex is a crucial connective component between the nuclear envelope and the cytoskeleton involving various cellular processes including nuclear positioning, nuclear architecture, and mechanotransduction. How LINC complexes regulate bone formation in vivo, however, is not well understood. To start bridging this gap, here we created a LINC disruption murine model using transgenic mice expressing Cre recombinase enzyme under the control of the Osterix (Osx-Cre) which is primarily active in pre-osteoblasts and floxed Tg(CAG-LacZ/EGFP-KASH2) mice.

Diet-Stimulated Marrow Adiposity Fails to Worsen Early, Age-Related Bone Loss.

Introduction: Longitudinal effect of diet-induced obesity on bone is uncertain. Prior work showed both no effect and a decrement in bone density or quality when obesity begins prior to skeletal maturity. We aimed to quantify long-term effects of obesity on bone and bone marrow adipose tissue (BMAT) in adulthood.

Development and Characterization of a low intensity vibrational system for microgravity studies.

The advent of extended-duration human spaceflight demands a better comprehension of the physiological impacts of microgravity. One primary concern is the adverse impact on the musculoskeletal system, including muscle atrophy and bone density reduction. Ground-based microgravity simulations have provided insights, with vibrational bioreactors emerging as potential mitigators of these negative effects.

The LINC complex regulates Achilles tendon elastic modulus, Achilles and tail tendon collagen crimp, and Achilles and tail tendon lateral expansion during early postnatal development.

There is limited understanding of how mechanical signals regulate tendon development. The nucleus has emerged as a major regulator of cellular mechanosensation, via the linker of nucleoskeleton and cytoskeleton (LINC) protein complex. Specific roles of LINC in tenogenesis have not been explored.

Increased deformations are dispensable for cell mechanoresponse in engineered bone analogs mimicking aging bone marrow.

Aged individuals and astronauts experience bone loss despite rigorous physical activity. Bone mechanoresponse is in-part regulated by mesenchymal stem cells (MSCs) that respond to mechanical stimuli. Direct delivery of low intensity vibration (LIV) recovers MSC proliferation in senescence and simulated microgravity models, indicating that age-related reductions in mechanical signal delivery within bone marrow may contribute to declining bone mechanoresponse.

Prrx1-driven LINC complex disruption in vivo reduces osteoid deposition but not bone quality after voluntary wheel running.

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex serves to connect the nuclear envelope and the cytoskeleton, influencing cellular processes such as nuclear arrangement, architecture, and mechanotransduction. The role LINC plays in mechanotransduction pathways in bone progenitor cells has been well studied; however, the mechanisms by which LINC complexes govern bone formation remain less clear. To bridge this knowledge gap, we established a murine model disrupting LINC using transgenic Prx-Cre mice and floxed Tg(CAG-LacZ/EGFP-KASH2) mice.

Osterix-driven LINC complex disruption diminishes bone microarchitecture in 8-week male mice but not after 6-week voluntary wheel running.

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex is a crucial connective component between the nuclear envelope and the cytoskeleton involving various cellular processes including nuclear positioning, nuclear architecture, and mechanotransduction. How LINC complexes regulate bone formation , however, is not well understood. To start bridging this gap, here we created a LINC disruption murine model using transgenic mice expressing Cre recombinase enzyme under the control of the Osterix (Osx-Cre) which is primarily active in pre-osteoblasts and floxed Tg(CAG-LacZ/EGFP-KASH2) mice.

Low intensity mechanical signals promote proliferation in a cell-specific manner: Tailoring a non-drug strategy to enhance biomanufacturing yields.

Biomanufacturing relies on living cells to produce biotechnology-based therapeutics, tissue engineering constructs, vaccines, and a vast range of agricultural and industrial products. With the escalating demand for these bio-based products, any process that could improve yields and shorten outcome timelines by accelerating cell proliferation would have a significant impact across the discipline. While these goals are primarily achieved using or strategies, harnessing cell mechanosensitivity represents a promising - albeit less studied - pathway to promote bioprocessing endpoints, yet identifying which mechanical parameters influence cell activities has remained elusive.

Data driven and cell specific determination of nuclei-associated actin structure.

Quantitative and volumetric assessment of filamentous actin fibers (F-actin) remains challenging due to their interconnected nature, leading researchers to utilize threshold based or qualitative measurement methods with poor reproducibility. Here we introduce a novel machine learning based methodology for accurate quantification and reconstruction of nuclei-associated F-actin. Utilizing a Convolutional Neural Network (CNN), we segment actin filaments and nuclei from 3D confocal microscopy images and then reconstruct each fiber by connecting intersecting contours on cross-sectional slices.

Atomic Force Microscopy Cantilever-Based Nanoindentation: Mechanical Property Measurements at the Nanoscale in Air and Fluid.

An atomic force microscope (AFM) fundamentally measures the interaction between a nanoscale AFM probe tip and the sample surface. If the force applied by the probe tip and its contact area with the sample can be quantified, it is possible to determine the nanoscale mechanical properties (e.g.

Mechanically Induced Nuclear Shuttling of β-Catenin Requires Co-transfer of Actin.

Mesenchymal stem cells (MSCs) respond to environmental forces with both cytoskeletal re-structuring and activation of protein chaperones of mechanical information, β-catenin, and yes-associated protein 1 (YAP1). To function, MSCs must differentiate between dynamic forces such as cyclic strains of extracellular matrix due to physical activity and static strains due to ECM stiffening. To delineate how MSCs recognize and respond differently to both force types, we compared effects of dynamic (200 cycles × 2%) and static (1 × 2% hold) strain on nuclear translocation of β-catenin and YAP1 at 3 hours after force application.

Architectural control of mesenchymal stem cell phenotype through nuclear actin.

There is growing appreciation that architectural components of the nucleus regulate gene accessibility by altering chromatin organization. While nuclear membrane connector proteins link the mechanosensitive actin cytoskeleton to the nucleoskeleton, actin's contribution to the inner architecture of the nucleus remains enigmatic. Control of actin transport into the nucleus, plus the presence of proteins that control actin structure (the actin tool-box) within the nucleus, suggests that nuclear actin may support biomechanical regulation of gene expression.

Nuclear envelope mechanobiology: linking the nuclear structure and function.

The nucleus, central to cellular activity, relies on both direct mechanical input as well as its molecular transducers to sense external stimuli and respond by regulating intra-nuclear chromatin organization that determines cell function and fate. In mesenchymal stem cells of musculoskeletal tissues, changes in nuclear structures are emerging as a key modulator of their differentiation and proliferation programs. In this review we will first introduce the structural elements of the nucleoskeleton and discuss the current literature on how nuclear structure and signaling are altered in relation to environmental and tissue level mechanical cues.

Modeling stem cell nucleus mechanics using confocal microscopy.

Nuclear mechanics is emerging as a key component of stem cell function and differentiation. While changes in nuclear structure can be visually imaged with confocal microscopy, mechanical characterization of the nucleus and its sub-cellular components require specialized testing equipment. A computational model permitting cell-specific mechanical information directly from confocal and atomic force microscopy of cell nuclei would be of great value.

Lamin A/C Is Dispensable to Mechanical Repression of Adipogenesis.

Mesenchymal stem cells (MSCs) maintain the musculoskeletal system by differentiating into multiple lineages, including osteoblasts and adipocytes. Mechanical signals, including strain and low-intensity vibration (LIV), are important regulators of MSC differentiation via control exerted through the cell structure. Lamin A/C is a protein vital to the nuclear architecture that supports chromatin organization and differentiation and contributes to the mechanical integrity of the nucleus.

At the nuclear envelope of bone mechanobiology.

The nuclear envelope and nucleoskeleton are emerging as signaling centers that regulate how physical information from the extracellular matrix is biochemically transduced into the nucleus, affecting chromatin and controlling cell function. Bone is a mechanically driven tissue that relies on physical information to maintain its physiological function and structure. Disorder that present with musculoskeletal and cardiac symptoms, such as Emery-Dreifuss muscular dystrophies and progeria, correlate with mutations in nuclear envelope proteins including Linker of Nucleoskeleton and Cytoskeleton (LINC) complex, Lamin A/C, and emerin.

Low-intensity vibration restores nuclear YAP levels and acute YAP nuclear shuttling in mesenchymal stem cells subjected to simulated microgravity.

Reducing the musculoskeletal deterioration that astronauts experience in microgravity requires countermeasures that can improve the effectiveness of otherwise rigorous and time-expensive exercise regimens in space. The ability of low-intensity vibrations (LIV) to activate force-responsive signaling pathways in cells suggests LIV as a potential countermeasure to improve cell responsiveness to subsequent mechanical challenge. Mechanoresponse of mesenchymal stem cells (MSC), which maintain bone-making osteoblasts, is in part controlled by the "mechanotransducer" protein YAP (Yes-associated protein), which is shuttled into the nucleus in response to cyto-mechanical forces.