Biology subjects

Brahmachary, P.

Publications and source records attributed to Brahmachary, P..

5 recordsLinked to original sources

bioRxiv2026

Viscoelastic recovery time of chondrocytes from monolayer and alginate cultures

This study uses a previously reported 3D-printed variable-height flow cell to investigate the viscoelastic properties of chondrocytes from 2D monolayer and 3D alginate cultures. It was hypothesized that chondrocytes could be distinguished by phenotype associated with their culture environment using viscoelastic recovery time, owing to variation in the pericellular matrix (PCM) produced by chondrocytes from different culture methods. The PCM surrounding the chondrocytes was imaged with confocal microscopy during applied deformation and subsequent recovery. The projected cell area was fitted with a Burgers mechanical model to extract the viscoelastic recovery time. No difference between bovine and primary OA cells from monolayer cultures was observed. However, a statistically significant difference in recovery time was observed between cells from monolayer and alginate cultures in both the bovine (31 s vs. 13 s) and primary OA (34 s vs. 13 s) groups. This work shows that viscoelastic recovery time is influenced by the culture method used for chondrocytes and further demonstrates the role of the PCM as a mechanical protector of chondrocytes.

cell biology↗

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

Metabolomic Profiles and Pathways in Osteoarthritic Human Cartilage: A Comparative Analysis with Healthy Cartilage

ObjectiveOsteoarthritis (OA) is a chronic joint disease with heterogenous metabolic pathology. To gain insight into OA-related metabolism, healthy and end-stage osteoarthritic cartilage were compared metabolically to uncover disease-associated profiles, classify OA-specific metabolic endotypes, and identify targets for intervention for the diverse populations of individuals affected by OA. DesignFemoral head cartilage (n=35) from osteoarthritis patients were collected post-total joint arthroplasty. Healthy cartilage (n=11) was obtained from a tissue bank. Metabolites from all cartilage samples were extracted and analyzed using liquid chromatography-mass spectrometry metabolomic profiling. Additionally, cartilage extracts were pooled and underwent fragmentation analysis for biochemical identification of metabolites. ResultsSpecific metabolites and metabolic pathways, including lipid- and amino acid pathways, were differentially regulated between osteoarthritis-derived and healthy cartilage. The detected alterations of amino acids and lipids highlight key differences in bioenergetic resources, matrix homeostasis, and mitochondrial alterations in osteoarthritis-derived cartilage compared to healthy. Moreover, metabolomic profiles of osteoarthritic cartilage separated into four distinct endotypes highlighting the heterogenous nature of OA metabolism and diverse landscape within the joint between patients. ConclusionsThe results of this study demonstrate that human cartilage has distinct metabolomic profiles between healthy and end-stage osteoarthritis patients. By taking a comprehensive approach to assess metabolic differences between healthy and osteoarthritic cartilage, and within osteoarthritic cartilage alone, several metabolic pathways with distinct regulation patterns were detected. Additional investigation may lead to the identification of metabolites that may serve as valuable indicators of disease status or potential therapeutic targets.

biochemistry↗

bioRxiv2020

Metabolomic Profiling and Mechanotransduction of Single Chondrocytes Encapsulated in Alginate Microgels

Human articular cartilage is comprised of two main components, the extracellular matrix (ECM) and the pericellular matrix (PCM). The PCM helps to protect chondrocytes in the cartilage from mechanical loads, but in patients with osteoarthritis, the PCM is weakened resulting in increased chondrocyte stress. As chondrocytes are responsible for cartilage synthesis and maintenance, it is important to understand how mechanical loads affect cellular responses of chondrocytes. Many studies have examined the chondrocyte response to in vitro mechanical loading by embedding in stiff agarose. However, these experiments are mostly performed in the absence of PCM which may obscure important responses to mechanotransduction. Here, we demonstrate that drop-based microfluidics allows culture of single chondrocytes in alginate microgels for cell-directed PCM synthesis that closely mimics the in vivo microenvironment. Chondrocytes form PCM over 10 days in these single cell microenvironments. Single cell microgels and monolayer controls were encapsulated in high stiffness agarose to mimic the cartilage PCM. After physiological dynamic compression in a custom-built bioreactor, microgels exhibited distinct metabolomic profiles from both uncompressed and monolayer controls. These results demonstrate the potential of single cell encapsulation in alginate microgels to advance cartilage tissue engineering and basic chondrocyte mechanobiology.

bioengineering↗