Altered extracellular matrix and mechanotransduction gene expression in rat bone tissue following long-term estrogen deficiency
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PROVIDER: S-EPMC11347883 | biostudies-literature | 2024 Jul
REPOSITORIES: biostudies-literature
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Project description:Mechanical properties of extracellular matrices (ECMs) regulate essential cell behaviours, including differentiation, migration and proliferation, through mechanotransduction. Studies of cell-ECM mechanotransduction have largely focused on cells cultured in 2D, on top of elastic substrates with a range of stiffnesses. However, cells often interact with ECMs in vivo in a 3D context, and cell-ECM interactions and mechanisms of mechanotransduction in 3D can differ from those in 2D. The ECM exhibits various structural features as well as complex mechanical properties. In 3D, mechanical confinement by the surrounding ECM restricts changes in cell volume and cell shape but allows cells to generate force on the matrix by extending protrusions and regulating cell volume as well as through actomyosin-based contractility. Furthermore, cell-matrix interactions are dynamic owing to matrix remodelling. Accordingly, ECM stiffness, viscoelasticity and degradability often play a critical role in regulating cell behaviours in 3D. Mechanisms of 3D mechanotransduction include traditional integrin-mediated pathways that sense mechanical properties and more recently described mechanosensitive ion channel-mediated pathways that sense 3D confinement, with both converging on the nucleus for downstream control of transcription and phenotype. Mechanotransduction is involved in tissues from development to cancer and is being increasingly harnessed towards mechanotherapy. Here we discuss recent progress in our understanding of cell-ECM mechanotransduction in 3D.
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Project description:The loss of TGFbeta or its downstream mediator, Smad3, key players in tissue repair, accelerates closure of incisional wounds in mice. In contrast, we now report that excisional ear wounds in mice lacking Smad3 enlarge compared with wild-type controls resulting from changes in extracellular matrix molecules, which alter the mechanotransduction properties of these wounds. Specifically, levels of elastin and glycosoaminoglycans are increased, collagen fibers are more compactly organized, and matrix modulators like integrins, TGFbeta1, and matrix metalloproteinases (MMPs) are altered both basally and after wounding in Smad3 knockout mice. Mechanical testing of dorsal skin correlates these changes in matrix composition with functional parameters, specifically an increased elastic modulus, suggesting an imbalance of tissue forces. We propose that the altered mechanical elastic properties translate into a persistent retractile force that is opposed by decreased wound contractile forces contributing to the enlarging ear wound in Smad3 knockout mice. These studies highlight a previously undescribed role for Smad3 in the mechanotransduction of matrix unsupported ear wound closure.
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Project description:Extracellular matrix (ECM) is comprised of different types of proteins, which change in composition and ratios during morphogenesis and disease progression. ECM proteins provide cell adhesion and impart mechanical cues to the cells. Increasing substrate stiffness has been shown to induce Yes-associated protein (YAP) translocation from the cytoplasm to the nucleus, yet these mechanistic studies used fibronectin only as the biochemical cue. How varying the types of ECM modulates mechanotransduction of stem cells remains largely unknown. Using polyacrylamide hydrogels with tunable stiffness as substrates, here we conjugated four major ECM proteins commonly used for cell adhesion: fibronectin, collagen I, collagen IV and laminin, and assessed the effects of varying ECM type and density on YAP translocation in human mesenchymal stem cells (hMSCs). For all four ECM types, increasing ECM ligand density alone can induce YAP nuclear translocation without changing substrate stiffness. The ligand threshold for such biochemical ligand-induced YAP translocation differs across ECM types. While stiffness-dependent YAP translocation can be induced by all four ECM types, each ECM requires a different optimized ligand density for this to occur. Using antibody blocking, we further identified integrin subunits specifically involved in mechanotransduction of different ECM types. Finally, we demonstrated that altering ECM type further modulates hMSC osteogenesis without changing substrate stiffness. These findings highlight the important role of ECM type in modulating mechanotransduction and differentiation of stem cells, and call for future mechanistic studies to further elucidate the role of changes in ECM compositions in mediating mechanotransduction during morphogenesis and disease progression. STATEMENT OF SIGNIFICANCE: Our study addresses a critical gap of knowledge in mechanobiology. Increasing substrate stiffness has been shown to induce nuclear YAP translocation, yet only on fibronectin-coated substrates. However, extracellular matrix (ECM) is comprised of different protein types. How varying the type of ECM modulates stem cell mechanotransduction remains largely unknown. We here reveal that the choice of ECM type can directly modulate stem cell mechanotransduction, filling this critical gap. This work has broad impacts in mechanobiology and biomaterials, as it provides the first evidence that varying ECM type can impact YAP translocation independent of substrate stiffness, opening doors for a more rational biomaterials design tuning ECM properties to control cell fate for promoting normal development and for preventing disease progression.
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Project description:Tooth eruption is a multifactorial process involving movement of existing tissues and formation of new tissues coordinated by a complex set of genetic events. We have used the model of the unopposed rodent molar to study morphological and genetic mechanisms involved in axial movement of teeth. Following extraction of opposing upper molars, lower molars supererupted by 0.13 mm. Labeled tissue sections revealed significant amounts of new bone and cementum apposition at the root apex of the unopposed side following supereruption for 12 days. Newly apposited cementum and alveolar bone layers were approximately 3-fold thicker in the experimental vs the control group, whereas periodontal ligament width was maintained. Tartrate-resistant acid phosphatase staining indicated bone resorption at the mesial alveolar walls of unopposed molars and provided in tandem with new bone formation at the distal alveolar walls an explanation for the distal drift of molars in this model. Microarray analysis and semiquantitative RT-PCR demonstrated a significant increase in collagen I, integrin beta5, and SPARC gene expression as revealed by comparison between the unopposed molar group and the control group. Immunohistochemical verification revealed increased levels of integrin beta5 and SPARC labeling in the periodontal ligament of the unopposed molar. Together our findings suggest that posteruptive axial movement of teeth was accomplished by significant formation of new root cementum and alveolar bone at the root apex in tandem with upregulation of collagen I, integrin beta5, and SPARC gene expression.
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Project description:Understanding how biomineralization occurs in the extracellular matrix (ECM) of bone cells is crucial to the understanding of bone formation and the development of a successfully engineered bone tissue scaffold. It is still unclear how ECM mechanical properties affect protein-mineral interactions in early stages of bone mineralization. We investigated the longitudinal mineralization properties of MC3T3-E1 cells and the elastic modulus of their ECM using shear modulation force microscopy, synchrotron grazing incidence X-ray diffraction (GIXD), scanning electron microscopy, energy dispersive X-ray spectroscopy, and confocal laser scanning microscopy (CLSM). The elastic modulus of the ECM fibers underwent significant changes for the mineralizing cells, which were not observed in the nonmineralizing cells. On substrates conducive to ECM network production, the elastic modulus of mineralizing cells increased at time points corresponding to mineral production, whereas that of the nonmineralizing cells did not vary over time. The presence of hydroxyapatite in mineralizing cells and the absence thereof in the nonmineralizing ones were confirmed by GIXD, and CLSM showed that a restructuring of actin occurred only for mineral-producing cells. These results show that the correct and complete development of the ECM network is required for osteoblasts to mineralize. This in turn requires a suitably prepared synthetic substrate for bone development to succeed in vitro.
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Project description:Extracellular biophysical properties have particular implications for a wide spectrum of cellular behaviors and functions, including growth, motility, differentiation, apoptosis, gene expression, cell-matrix and cell-cell adhesion, and signal transduction including mechanotransduction. Cells not only react to unambiguously mechanical cues from the extracellular matrix (ECM), but can occasionally manipulate the mechanical features of the matrix in parallel with biological characteristics, thus interfering with downstream matrix-based cues in both physiological and pathological processes. Bidirectional interactions between cells and (bio)materials in vitro can alter cell phenotype and mechanotransduction, as well as ECM structure, intentionally or unintentionally. Interactions between cell and matrix mechanics in vivo are of particular importance in a variety of diseases, including primarily cancer. Stiffness values between normal and cancerous tissue can range between 500 Pa (soft) and 48 kPa (stiff), respectively. Even the shear flow can increase from 0.1-1 dyn/cm2 (normal tissue) to 1-10 dyn/cm2 (cancerous tissue). There are currently many new areas of activity in tumor research on various biological length scales, which are highlighted in this review. Moreover, the complexity of interactions between ECM and cancer cells is reduced to common features of different tumors and the characteristics are highlighted to identify the main pathways of interaction. This all contributes to the standardization of mechanotransduction models and approaches, which, ultimately, increases the understanding of the complex interaction. Finally, both the in vitro and in vivo effects of this mechanics-biology pairing have key insights and implications for clinical practice in tumor treatment and, consequently, clinical translation.
| S-EPMC10777970 | biostudies-literature
Project description:ObjectiveTranscriptomic changes in joint tissues during the development of osteoarthritis (OA) are of interest for the discovery of biomarkers and mechanisms of disease. The objective of this study was to use the rat medial meniscus transection (MMT) model to discover stage and tissue-specific transcriptomic changes.DesignSham or MMT surgeries were performed in mature rats. Cartilage, menisci and synovium were scored for histopathological changes at 2, 4 and 6 weeks post-surgery and processed for RNA-sequencing. Differentially expressed genes (DEG) were used to identify pathways and mechanisms. Published transcriptomic datasets from animal models and human OA were used to confirm and extend present findings.ResultsThe total number of DEGs was already high at 2 weeks (723 in meniscus), followed by cartilage (259) and synovium (42) and declined to varying degrees in meniscus and synovium but increased in cartilage at 6 weeks. The most upregulated genes included tenascins. The 'response to mechanical stimulus' and extracellular matrix-related pathways were enriched in both cartilage and meniscus. Pathways that were enriched in synovium at 4 weeks indicate processes related to synovial hyperplasia and fibrosis. Synovium also showed upregulation of IL-11 and several MMPs. The mechanical stimulus pathway included upregulation of the mechanoreceptors PIEZO1, PIEZO2 and TRPV4 and nerve growth factor. Analysis of data from prior RNA-sequencing studies of animal models and human OA support these findings.ConclusionThese results indicate several shared pathways that are affected during OA in cartilage and meniscus and support the role of mechanotransduction and other pathways in OA pathogenesis.
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Project description:BackgroundPost-menopausal osteoporosis is a common health problem worldwide, most commonly caused by estrogen deficiency. Most of the information regarding the skeletal effects of this disease relates to trabecular bone, while cortical bone is less studied. The purpose of this study was to evaluate the influence of estrogen deficiency on the structure and mechanical properties of cortical bone.MethodsEight ovariectomized (OVH) and eight intact (control) Sprague Dawley rats were used.Structural features of femoral cortical bone were studied by light microscopy, scanning electron microscopy and synchrotron-based microcomputer-tomography and their mechanical properties determined by nano-indentation.ResultsCortical bone of both study groups contains two distinct regions: organized circumferential lamellae and disordered bone with highly mineralized cartilaginous islands. Lacunar volume was lower in the OVH group both in the lamellar and disorganized regions (182 ± 75 µm3 vs 232 ± 106 µm3, P < 0.001 and 195 ± 86 µm3 vs. 247 ± 106 µm3, P < 0.001, respectively). Lacunar density was also lower in both bone regions of the OVH group (40 ± 18 ×103 lacunae/mm3 vs. 47 ± 9×103 lacunae/mm3 in the lamellar region, P = 0.003 and 63 ± 18×103lacunae/mm3 vs. 75 ± 13×103 lacunae/mm3 in the disorganized region, P < 0.001). Vascular canal volume was lower in the disorganized region of the bone in the OVH group compared to the same region in the control group (P < 0.001). Indentation moduli were not different between the study groups in both bone regions.DiscussionChanges to cortical bone associated with estrogen deficiency in rats require high-resolution methods for detection. Caution is required in the application of these results to humans due to major structural differences between human and rat bone.
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Project description:E26 transformation specific (ETS) family transcription factors are expressed during embryogenesis and are involved in various cellular processes such as proliferation, migration, differentiation, angiogenesis, apoptosis, and survival of cellular lineages to ensure appropriate development. Dysregulated expression of many of the ETS family members is detected in different cancers. The human ELF3, a member of the ETS family of transcription factors, plays a role in the induction and progression of human cancers is well studied. However, little is known about the role of ELF3 in early development. Here, the zebrafish elf3 was cloned, and its expression was analyzed during zebrafish development. Zebrafish elf3 is maternally deposited. At different developmental stages, elf3 expression was detected in different tissue, mainly neural tissues, endoderm-derived tissues, cartilage, heart, pronephric duct, blood vessels, and notochord. The expression levels were high at the tissue boundaries. Elf3 loss-of-function consequences were examined by using translation blocking antisense morpholino oligonucleotides, and effects were validated using CRISPR/Cas9 knockdown. Elf3-knockdown produced short and bent larvae with notochord, craniofacial cartilage, and fin defects. The extracellular matrix (ECM) in the fin and notochord was disorganized. Neural defects were also observed. Optic nerve fasciculation (bundling) and arborization in the optic tectum were defective in Elf3-morphants, and fragmentation of spinal motor neurons were evident. Dysregulation of genes encoding ECM proteins and matrix metalloprotease (MMP) and disorganization of ECM may play a role in the observed defects in Elf3 morphants. We conclude that zebrafish Elf3 is required for epidermal, mesenchymal, and neural tissue development.
| S-EPMC9668168 | biostudies-literature
Project description:Dysfunctional extracellular matrices (ECM) contribute to aging and disease. Repairing dysfunctional ECM could potentially prevent age-related pathologies. Interventions promoting longevity also impact ECM gene expression. However, the role of ECM composition changes in healthy aging remains unclear. Here we perform proteomics and in-vivo monitoring to systematically investigate ECM composition (matreotype) during aging in C. elegans revealing three distinct collagen dynamics. Longevity interventions slow age-related collagen stiffening and prolong the expression of collagens that are turned over. These prolonged collagen dynamics are mediated by a mechanical feedback loop of hemidesmosome-containing structures that span from the exoskeletal ECM through the hypodermis, basement membrane ECM, to the muscles, coupling mechanical forces to adjust ECM gene expression and longevity via the transcriptional co-activator YAP-1 across tissues. Our results provide in-vivo evidence that coordinated ECM remodeling through mechanotransduction is required and sufficient to promote longevity, offering potential avenues for interventions targeting ECM dynamics.
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