Transcriptomic Profiling of Murine Chondrocytes Following APOE Knockout: Insights into Gene Expression Alterations
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE296343 | GEO | 2025/08/20
REPOSITORIES: GEO
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Project description:Osteoarthritis (OA) is a degenerative joint disease with limited effective therapies. Cold weathers have been shown to affect joint pain in OA patients. However, the impact of cold climate on OA progression is debated, with the underlying mechanisms not fully understood. To investigate the mechanisms by which low temperatures exacerbate OA, RNA sequencing on human chondrocytes cultured at 33 °C and 37 °C was performed. The gene expression in non-OA region chondrocytes and OA region chondrocytes at 33 °C was compared to that at 37 °C. The Apoe gene was identified as a temperature-relevant gene associated with exacerbation of osteoarthritis. Apoe expression was downregulated in both OA and non-OA chondrocytes cultured at 33°C compared to their 37°C counterparts.
2025-08-20 | GSE296344 | GEO
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Project description:The resting zone (RZ) in mammalian growth plates is critical for maintaining and regulating chondrocyte turnover during longitudinal bone growth as a control tower and stem cell reservoir. Although recent lineage tracing studies have identified several markers for stem cells in the RZ, these markers only partially label chondrocytes in the RZ, suggesting that the resting chondrocytes (RCs) are a heterogeneous population with different types of stem cells. Since a comprehensive marker for RCs is still lacking, the RZ is generally determined based on ambiguous histological criteria, such as small and round chondrocytes without columnar formation, which may lead to inconsistencies among researchers. Therefore, in this study, we used single-cell RNA sequencing (scRNAseq) of growth plate chondrocytes followed by validation by fluorescence in situ hybridization (FISH) to precisely annotate cell clusters in scRNAseq and search for a marker of RCs. The scRNAseq analysis revealed that apolipoprotein E (Apoe) was the top-hit gene, which was ubiquitously expressed in the RC cluster. FISH confirmed that Apoe was exclusively localized to the histologically defined RZ. In newly generated ApoemCherry knock-in mice, we further confirmed that mCherry expression mirrored the distribution of Apoe-expressing chondrocytes in the RZ particularly after the formation of the secondary ossification center. These mCherry+ RCs were slow cycling in vivo and exhibited stem cell properties in vitro. Moreover, APOE was detected in human growth plate RCs. These findings suggest that apolipoprotein E is a novel pan-RC marker in both mouse and human growth plates.
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Project description:As the unique cell type in articular cartilage, chondrocyte senescence is a crucial cellular event contributing to osteoarthritis development. Here we show that clathrin-mediated endocytosis and activation of Notch signaling promotes chondrocyte senescence and osteoarthritis development, which is negatively regulated by myosin light chain 3. Myosin light chain 3 (MYL3) protein levels decline sharply in senescent chondrocytes of cartilages from model mice and osteoarthritis (OA) patients. Conditional deletion of Myl3 in chondrocytes significantly promoted, whereas intra-articular injection of adeno-associated virus overexpressing MYL3 delayed, OA progression in male mice. MYL3 deficiency led to enhanced clathrin-mediated endocytosis by promoting the interaction between myosin VI and clathrin, further inducing the internalization of Notch and resulting in activation of Notch signaling in chondrocytes. Pharmacologic blockade of clathrin-mediated endocytosis-Notch signaling prevented MYL3 loss-induced chondrocyte senescence and alleviated OA progression in mice. Our results establish a previously unknown mechanism essential for cellular senescence and provide a potential therapeutic direction for OA.
2023-09-07 | PXD044684 | Pride
Project description:Osteoarthritis (OA) is an aging-associated disease of diarthrodial joints that involves changes in the bone, cartilage and soft tissue. Chondrocytes are the only cell type in cartilage and play a pivotal role in tissue homeostasis and cartilage remodeling in OA. Age- or OA-dependent metabolic changes affecting extracelluar matrix composition as well as variations in the response to cytokines and growth factors have been investigated in human chondrocytes. However, the molecular mechanisms causing age-dependent variation of the chondrocyte phenotype are poorly characterized. The proinflammatory mediator nitric oxide (NO) is produced when chondrocytes are exposed to cytokines such as IL-1. NO affects the energy metabolism of cells through interfering with mitochondrial respiration and glycolysis and has been implicated in extracellular matrix synthesis and degradation and chondrocyte death. NO may contribute to the aging process by compromising the energy balance ! in chondrocytes through the generation of an oxidizing environment and this may in turn affect matrix gene expression. In human chondrocytes NO effects on the expression of extacelluar matrix genes or genes related to matrix structure have not been addressed comprehensively. In chondrocytes NO has profound effects on extracellular matrix: Interleukin-1 induced the production of nitric oxide in human articular chondrocytes in alginate culture and simultaneously inhibited the biosynthesis of proteoglycans. NG-monomethyl-L-arginine (L-NMMA) inhibited nitric oxide formation and the suppression of proteoglycan synthesis. The nitric oxide donor, S-nitrosyl-acetyl-D,L- penicillamine (SNAP) also inhibited proteoglycan biosynthesis suggesting that interleukin-1 suppresses matrix synthesis through mechanisms involving the production of NO. Slices of rabbit articular cartilage synthesized large quantities of nitric oxide (NO) following exposure to IL-1 and strongly suppressed the incorporation of 35SO4 into the glycosaminoglycans (GAGs) of cartilage proteoglycans. Simultaneous treatment of cartilage fragments with IL-1 and L-NMMA inhibited NO synthesis in response to IL-1 and restored proteoglycan synthesis. SNAP reversibly mimicked the effect! of IL-1 on proteoglycan synthesis. These data suggest that endogenously synthesized NO is the mediator which reduces cartilage proteoglycan synthesis in response to cytokines such as IL-1. Aggrecan and type II collagen synthesis are also affected by NO. Our hypothesis entails that in human chondrocytes endogenous NO modulates the expression of matrix genes as well as enzymes involved in extracellular matrix remodelling. Experimental approach: In this experiment cultured chondrocytes from 3 donors was left unstimulated (control) or stimulated with IL-1, L-NMMA or IL-1 + L-NMMA for a period of 8 days in order to achieve sufficiently high levels of intracellular NO and stable expression of a subset of genes. Total RNA was isolated using the Trizol procedure. Gene expression will be probed using the Glycov2 chip. In this study cultured chondrocytes from three donors will either be left unstimulated (control) or stimulated with IL-1, L-NMMA or IL-1 L-NMMA for a period of 8 days in order to achieve sufficiently high levels of intracellular NO and stable expression of a subset of genes. 12 Samples were hybridized and analyzed using the GLYCOv2 array, three from each class.
2011-02-17 | E-GEOD-27357 | biostudies-arrayexpress
Project description:Examination of the genome-wide distribution of 5hmC in osteoarthritic chondrocytes compared to normal chondrocytes in order to elucidate the effect on OA-specific gene expression. 5hmC-sequencing was performed and data was compared with microarray gene expression data to identify genes with differential expression between normal and OA chondrocytes that are potentially under epigenetic regulation. High-throughput sequencing of 5hmC in 4 normal and 4 OA chondrocyte samples.
2015-06-30 | E-GEOD-64393 | biostudies-arrayexpress
Project description:Bovine chondrocyte-seeded and mesenchymal stem cell (MSC)-seeded agarose were cultured for 28 days in chemically defined media containing 10 ng/mL TGF-beta3. Chondrogenic differentiated MSCs were compared to chondrocytes at this timepoint and to undifferentiated MSCs harvested at day 0. Donor-matched sets of chondrocytes and MSCs were used, with three donors total.
2010-10-02 | E-GEOD-18394 | biostudies-arrayexpress
Project description:To survey altered gene expreesion in round, colmnar and hypertrophic chondrocytes of Tric-b-KO mice, total RNA preparations from round chondrocytes and colmnar/hypertrophic chondrocytes were subjected to gene microarray analysis. Results provide insight into the transcriptional profile of each Tric-b-KO chondrocyte and provide further insight into their functions.
2023-01-31 | GSE223776 | GEO
Project description:Background: Chondrocyte homeostasis is vital for maintaining the extracellular matrix (ECM) and overall cartilage health. In osteoarthritis (OA), oxidative stress, often caused by redox imbalances, can disrupt chondrocyte homeostasis, leading to cartilage degradation. Hydrogen peroxide (H2O2), a reactive oxygen species (ROS), is a key mediator of oxidative stress and contributes to chondrocyte apoptosis and ECM degradation. Previous studies have explored individual protein responses to oxidative stress; however, a comprehensive proteomic analysis in chondrocytes has not been conducted. In this study, we aimed to assess the global proteomic alterations in chondrocytes exposed to H2O2 using a shotgun proteomics approach, which enables the detection of a broader spectrum of proteomic changes. Methods: Chondrocytes were treated with H2O2 for 1, 4, and 16 h followed by protein extraction and processing, including denaturation, alkylation, and trypsin digestion. The peptides were then acidified, desalted, dried, and resuspended for LC-MS/MS. Proteomics data were analyzed using MaxQuant software to identify and quantify proteins. Secretome analysis was performed to examine protein secretion changes under oxidative stress. The statistical significance of all proteomics and secretome data was assessed using a two-tailed Student’s t-test in the Perseus software. Complementary methods, including quantitative PCR, western blotting, and immunofluorescence, were employed to validate the proteomic analysis. Results: Our findings revealed that oxidative stress primarily affected the endoplasmic reticulum (ER), causing notable alterations in the expression of ER-associated proteins, redox-responsive enzymes, chaperones, and sialyltransferases. These changes increased intracellular accumulation of ECM proteins and decreased secretion into the extracellular environment, indicating impaired protein trafficking and secretion. Additionally, immune-related pathways were activated in the long term, with a short-term upregulation of inflammatory markers, such as interleukin (IL)-6) and IL-18, although the levels of matrix metalloproteinases (MMPs) remained stable, indicating a complex inflammatory response and stress responses that disrupt ECM homeostasis. Conclusions: Our study demonstrates a detailed proteomic view of the stress response of H2O2-treated chondrocytes, highlighting the significant changes in ER function, cytoskeletal remodeling, protein secretion, and immune responses. These changes suggest that oxidative stress impacts ECM balance and contributes to OA progression, offering new insights into the molecular mechanisms underlying oxidative stress in OA.
2025-06-09 | PXD056759 | Pride
Project description:Osteoarthritis (OA) of the hand is a common disease resulting in pain and impaired function. The pathogenesis of hand OA (HOA) is elusive and models to study it have not been described so far. Culture of chondrocytes is a model to study the development of cartilage degeneration, which is a hallmark of OA and well established in OA of the knee and hip. In the current study we investigated the feasibility human chondrocyte culture derived from proximal interphalangeal (PIP) finger joints of dissecting room cadavers. Index and middle fingers without signs of osteoarthritis were obtained from 30 cadavers using two different protocols. Hyaline cartilage from both articulating surfaces of the proximal interphalangeal (PIP) joint was harvested and digested in collagenase. Cultured chondrocytes were monitored for contamination, viability, and expression of chondrocyte specific genes. Chondrocytes derived from knee joints of the cadavers were cultured under identical conditions. Gene expression comparing chondrocytes from PIP and knee joints was carried out using Affymetrix GeneChip Human 2.0 ST arrays. The resulting differentially expressed genes were validated by real-time PCR and immunohistochemistry.Chondrocytes harvested up to 101 hours after death of the donors were viable. mRNA expression of collagen 2A1, aggrecan and Sox9 was significantly higher in chondrocytes as compared to cultured fibroblasts. Comparison of gene expression by chondrocytes from PIP and knee joints yielded 528 differentially expressed genes. Chondrocytes from the same joint region had a higher grade of similarity than chondrocytes of the same individual. These results were validated using real-time PCR and immunohistochemistry.We demonstrate for the first time a reliable method for culture of chondrocytes derived from PIP joints. PIP chondrocytes show a specific gene expression pattern and could be used as tool to study cartilage degeneration in HOA. Three samples of cultured chondrocytes from knee and proximal interphalangeal finger joints were compared. Gene expression of the four most differentially regulated genes was confirmed by real-time PCR in 10 independent samples.
2016-07-03 | E-GEOD-68038 | biostudies-arrayexpress