Biology subjects

June, R.

Publications and source records attributed to June, R..

8 recordsLinked to original sources

bioRxiv2026

Expanded stoichiometric model of chondrocyte metabolism: response to cyclical shear and compressive loading

Cartilage deterioration is a hallmark osteoarthritis, and there is substantial interest in developing strategies for cartilage repair. Cyclical mechanical stimulation has been known for decades to drive synthesis of cartilage matrix proteins. Matrix synthesis requires activation of central metabolism for producing precursors to non-essential amino acids required for protein translation. However, there are gaps in knowledge regarding how mechanical stimuli affect chondrocyte central metabolism. Here, we find that cyclical shear and compression drive differences in chondrocyte central metabolism in a sex-dependent manner. Based on established biochemistry, we developed and tested a stoichiometric model containing 139 metabolites and 172 reactions from central metabolism that includes production of key cartilage matrix proteins. We then used experimental metabolomics data from shear and compressive stimulation of osteoarthritic chondrocytes to constrain this model and ran multiple simulations examining the potential for producing matrix proteins and ATP. Our results show that both shear and compression can stimulate osteoarthritic chondrocyte metabolism in a manner consistent with production of cartilage matrix proteins, with notable differences between male and female chondrocytes. Additionally, and importantly, our simulation results suggest that nitrogen availability is a key limitation to chondrocyte synthesis of matrix proteins. These results are a starting point for using central metabolism of chondrocytes to optimize synthesis of matrix proteins for cartilage repair. For example, increasing glutamine levels in the presence of cyclical compression has potential to increase production of both types II and VI collagen. These strategies have potential for improving cartilage tissue engineering and repair.

bioengineering↗

bioRxiv2025

Targeted analysis of chondrocyte central metabolites in response to cyclical compression and shear deformations

Osteoarthritis results in deterioration of articular cartilage, the soft tissue covering articulating surfaces of bones in joints like the knee and hip. Cyclical mechanical stimulation of articular cartilage results in synthesis of cartilage matrix, suggesting that therapeutic mechanical stimulation might be beneficial for cartilage repair in osteoarthritis. Several prior studies identify ion channels and cytoskeletal molecules as components of chondrocyte mechanotransduction. The pathways of central metabolism, glycolysis, the pentose phosphate pathway, and the tricarboxylic acid cycle are necessary for producing non-essential amino acids that are needed for synthesizing matrix proteins for cartilage repair. However, it is currently unknown if and how levels of central metabolites change with applied mechanical stimulation of chondrocytes. Here, we show that applied cyclical shear and compressive deformations drive changes in multiple central metabolites in primary chondrocytes. These results add to a rich picture of chondrocyte mechanotransduction, including calcium signaling, ion channels, integrins, and cytoskeletal components. By finding compression- and shear-induced changes in central metabolites, these data support the potential for therapeutic mechanotransduction toward cartilage repair. Future studies may build on these results to understand the relationships between mechanical stimulation and chondrocyte central metabolism.

bioengineering↗

bioRxiv2024

Metabolomic Profiling and Characterization of a Novel 3D Culture System for Studying Chondrocyte Mechanotransduction

Background/ObjectiveArticular chondrocytes synthesize and maintain the avascular and aneural articular cartilage. In vivo these cells are surrounded by a 3D pericellular matrix (PCM) containing predominantly collagen VI. The PCM protects chondrocytes and facilitates mechanotransduction. PCM stiffness is critical in transmitting biomechanical signals to chondrocytes. Various culture systems with different hydrogels are used to encapsulate chondrocytes for 3D culture, but many lack either the PCM or the in vivo stiffness of the cartilage matrix. This study aimed at establishing a culture system to investigate a) if chondrocytes cultured in alginate will develop a PCM and b) study mechanotransduction via metabolic changes induced in 3D agarose-embedded chondrocytes upon mechanical stimulation. MethodsWe cultured primary human and bovine chondrocytes in monolayers or as alginate encapsulated cells in media containing sodium L-ascorbate. PCM expression was analyzed by immunofluorescence and western blots. We further characterized the response of chondrocytes embedded in physiologically stiff agarose to dynamic compression through metabolomic profiling. ResultsWe found that primary human and bovine chondrocytes, when cultured in alginate beads with addition of sodium L-ascorbate for 7 days, had a pronounced PCM, retained their phenotype, and synthesized both collagens VI and II. This novel culture system enables alginate-encapsulated chondrocytes to develop a robust PCM thereby creating a model system to study mechanotransduction in the presence of an endogenous PCM. We also observed distinct compression-induced changes in metabolomic profiles between the monolayer-agarose and alginate-released agarose-embedded chondrocytes indicating physiological changes in cell metabolism. Conclusion/SignificanceThese data show that 3D preculture of chondrocytes in alginate before encapsulation in physiologically stiff agarose leads to pronounced development of pericellular matrix that is sustained in the presence of ascorbate. This model can be useful in studying the mechanism by which chondrocytes respond to cyclical compression and other types of loading simulating in vivo physiological conditions.

bioengineering↗

bioRxiv2024

The metabolome of individuals with knee osteoarthritis is influenced by 18-months of an exercise and weight loss intervention and sex: the IDEA trial

ObjectiveThe Intensive Diet and Exercise for Arthritis (IDEA) trial was conducted to evaluate the effects of diet and exercise on osteoarthritis (OA), the most prevalent form of arthritis. Various risk factors, such as obesity and sex, contribute to the debilitating nature of OA. While diet and exercise are known to improve OA symptoms, cellular and molecular mechanisms underlying these interventions, as well as effects of participant sex, remain elusive. MethodsSerum was obtained at three timepoints from IDEA participants assigned to groups of diet, exercise, or combined diet and exercise (n=10 per group). All serum metabolites were extracted and analyzed via liquid chromatography-mass spectrometry combined with metabolomic profiling. Extracted serum was pooled and fragmentation patterns were analyzed to identify metabolites that statistically differentially regulated between groups. ResultsChanges in metabolism across male and female IDEA participants after 18-months of diet, exercise, and combined diet and excise intervention mapped to lipid, amino acid, carbohydrate, vitamin, and matrix metabolism. The diverse metabolic landscape detected across IDEA participants shows that intervention type impacts the serum metabolome of individuals with OA in distinct patterns. Moreover, differences in the serum metabolome corresponded with participant sex. ConclusionsThese findings suggest that intensive weight loss among male and female subjects offers potential metabolic benefits for individuals with knee OA. This provides a deeper understanding of dysregulation occurring during OA development that may pave the way for improved interventions, treatments, and quality of life of those impacted by this disease.

biochemistry↗

bioRxiv2022

The gut microbiome has sexually dimorphic effects on bone tissue energy metabolism and multiscale bone quality in C57BL/6J mice

The gut microbiome impacts bone mass, implying a disruption to bone homeostasis, yet significant uncertainty remains regarding the impacts of the gut microbiome on remodeling bone cells. The gut microbiome is thought to be essential for normal biomineralization, but the specific consequences of the absent gut microbiome on tissue mineralization and multiscale bone quality are not determined. We hypothesized that bone homeostasis and tissue-scale metabolism, tissue mineralization, and whole-bone biomechanics are altered in germ-free (GF) C57BL/6J mice. Further, because many characteristics of the gut microbiome are sexually dimorphic, we hypothesized that the gut microbiome would show important sex differences with regards to its impact on bone quality. Differences between GF and conventional mouse bone extended from bone tissue metabolism to whole bone biomechanics. Cortical bone tissue from male mice had a greater signature of amino acid metabolism whereas female cells had a greater signature of lipid metabolism. These sex differences were also present in GF mice and were indeed even more stark. GF increased cortical femur bone formation for both sexes and decreased bone resorption and osteoclast density only in females. GF similarly increased cortical femur tissue mineralization and altered collagen structure for both sexes but led to greater gains in distal femur trabecular microarchitecture for males. Whole femur strength was similarly increased with GF for both sexes, but males had a greater increase in modulus. GF did not impact fracture toughness for either sex. The altered bone quality with GF is multifactorial and is likely contributed to by differences in tissue-scale composition as well as lower cortical porosity. Together, these data demonstrate that the gut microbiome influences bone cells and multiscale bone quality, but that the specific relationships that underlie these effects to bone are different for females and males.

microbiology↗

bioRxiv2019

Primary human chondrocytes respond to compression with plasma membrane receptors and by microtubule activation: a phosphoproteomic study

Chondrocytes are responsible for maintaining the cartilage that helps joints like the knee and hip bear load and move smoothly. These cells typically respond to physiological compression with pathways consistent with matrix synthesis, and chondrocyte mechanotransduction is essential for tissue and joint homeostasis. In osteoarthritis (OA), chondrocyte mechanotransduction appears to be dysregulated, yet many pathways and mechanisms of osteoarthritic chondrocyte mechanotransduction remain poorly understood. The objective of this study is to document the phosphoproteomic responses of primary osteoarthritic chondrocytes to physiological sinusoidal compression. Here we show that OA chondrocytes respond to physiological compression by first activating proteins consistent with cytoskeletal remodeling and decreased transcription, and then later activating proteins for transcription. These results show that several microtubule-related proteins respond to compression, as well as proteins related to calcium signaling, which has previously been extensively shown in chondrocytes. Our results demonstrate that compression is a relevant physiological stimulus for osteoarthritic chondrocytes. We anticipate these data to be a starting point for more sophisticated analysis of both normal and osteoarthritic chondrocyte mechanotransduction. For example, finding differences in compression-induced phosphoproteins between normal and OA cells may lead to druggable targets to restore homeostasis to diseased joints.

bioengineering↗

bioRxiv2018

Characterization of osteoarthritis phenotypes by global metabolomic profiling of human synovial fluid

ObjectiveOsteoarthritis (OA) is a multifactorial disease with etiological heterogeneity. The objective of this study was to classify OA subgroups by generating metabolic phenotypes of OA from human synovial fluid.\n\nDesignPost mortem synovial fluids (n=75) were analyzed by high performance-liquid chromatography mass spectrometry (HPLC-MS) to measure changes in the global metabolome. Comparisons of healthy (grade 0), early OA (grades I-II), and late OA (grades III-IV) donor populations were considered to reveal phenotypes throughout disease progression.\n\nResultsGlobal metabolomic profiles in synovial fluid were distinct between healthy, early OA, and late OA donors. Pathways differentially activated among these groups included structural deterioration, glycerophospholipid metabolism, inflammation, central energy metabolism, oxidative stress, and vitamin metabolism. Within disease states (early and late OA), subgroups of donors revealed distinct phenotypes. Phenotypes of OA exhibited increased inflammation (early and late OA), oxidative stress (late OA), or structural deterioration (early and late OA) in the synovial fluid.\n\nConclusionThese results revealed distinct metabolic phenotypes of OA in human synovial fluid, provide insight into pathogenesis, represent novel biomarkers and assist in developing personalized interventions for subgroups of OA patients.

pathology↗

bioRxiv2018

Inhibition of Early Response Genes Prevents Changes in Global Joint Metabolomic Profiles in Mouse Post-Traumatic Osteoarthritis

Osteoarthritis (OA) is the most common degenerative joint disease, and joint injury increases the risk of OA by 10-fold. Although the injury event itself damages joint tissues, a substantial amount of secondary damage is mediated by the cellular responses to the injury. Cellular responses include the production and activation of proteases (MMPs, ADAMTSs, Cathepsins), the production of inflammatory cytokines, and we hypothesize, changes to the joint metabolome. The trajectory of cellular responses is driven by the transcriptional activation of early response genes, which requires Cdk9-dependent RNA Polymerase II phosphorylation. Flavopiridol is a potent and selective inhibitor of Cdk9 kinase activity, which prevents the transcriptional activation of early response genes. To model post-traumatic osteoarthritis, we subjected mice to non-invasive ACL-rupture joint injury. Following injury, mice were treated with flavopiridol to inhibit Cdk9-dependent transcriptional activation, or vehicle control. Global joint metabolomics were analyzed 1 hour after injury. We found that injury induced metabolomic changes, including increases in Vitamin D3 metabolism and others. Importantly, we found that inhibition of primary response gene activation at the time of injury largely prevented the global changes in the metabolomics profiles. Cluster analysis of joint metabolomes identified groups of injury-induced and drug-responsive metabolites, which may offer novel targets for cell-mediated secondary joint damage. Metabolomic profiling provides an instantaneous snapshot of biochemical activity representing cellular responses, and these data demonstrate the potential for inhibition of early response genes to alter the trajectory of cell-mediated degenerative changes following joint injury.\n\nSignificance StatementJoint injury is an excellent predictor of future osteoarthritis. It is increasingly apparent that the acute cellular responses to injury contribute to the initiation and pathogenesis of OA. Although changes to the joint transcriptome have been extensively studied in the context of joint injury, little is known about changes to small-molecule metabolites. Here we use a non-invasive ACL rupture model of joint injury in mice to identify injury-induced changes to the global metabolomic profiles. In one experimental group we prevented the activation of primary response gene transcription using the Cdk9 inhibitor flavopiridol. Through this comparison, we identified two sets of metabolites that change acutely after joint injury: those that require transcription of primary response genes, and those that do not.

pathology↗