Browsing by Author "Brahmachary, Priyanka P."

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Correlations between metabolites in the synovial fluid and serum: A mouse injury study
(Wiley, 2022-03) Wallace, Cameron W.; Hislop, Brady; Hahn, Alyssa K.; Erdogan, Ayten E.; Brahmachary, Priyanka P.; June, Ronald K.
Osteoarthritis occurs frequently after joint injury. Currently, osteoarthritis is diagnosed by radiographic changes that are typically found after the disease has progressed to multiple tissues. The primary objective was to compare potential metabolomic biomarkers of joint injury between synovial fluid and serum in a mouse model of posttraumatic osteoarthritis. The secondary objective was to gain insight into the pathophysiology of osteoarthritis by examining metabolomic profiles after joint injury. Twelve-week-old adult female C57BL/6 mice (n = 12) were randomly assigned to control, Day 1, or Day 8 postinjury groups. Randomly selected stifle joints were subjected to a single rapid compression. At Days 1 and 8 postinjury, serum was extracted before mice were euthanized for synovial fluid collection. Metabolomic profiling detected ~2500 metabolites across serum and synovial fluid. Of these, 179 were positively correlated and 51 were negatively correlated between synovial fluid and serum, indicating the potential for the development of metabolomic biomarkers. Synovial fluid captured injury-induced differences in metabolomic profiles at both Days 1 and 8 after injury whereas serum did not. However, synovial fluid and serum were distinct at both time points after injury. In synovial fluid, pathways of interest mapped to amino acid synthesis and degradation, bupropion degradation, and transfer RNA (tRNA) charging. In serum, pathways were amino acid synthesis and degradation, the phospholipase pathway, and nicotine degradation. These results provide a rich picture of the injury response at early time points after joint injury. Furthermore, the correlations between synovial fluid and serum metabolites suggest the potential to gain insight into intra-articular pathophysiology through analysis of serum metabolites.
Metabolomic Profiling and Mechanotransduction of Single Chondrocytes Encapsulated in Alginate Microgels
(MDPI AG, 2022-03) Fredrikson, Jacob P.; Brahmachary, Priyanka P.; Erdoğan, Ayten E.; Archambault, Zachary K.; Wilking, James N.; June, Ronald K.; Chang, Connie B.
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 matrix synthesis and maintenance, it is important to understand how mechanical loads affect the cellular responses of chondrocytes. Many studies have examined chondrocyte responses to in vitro mechanical loading by embedding chondrocytes in 3-D hydrogels. However, these experiments are mostly performed in the absence of PCM, which may obscure important responses to mechanotransduction. Here, drop-based microfluidics is used to culture single chondrocytes in alginate microgels for cell-directed PCM synthesis that closely mimics the in vivo microenvironment. Chondrocytes formed PCM over 10 days in these single-cell 3-D microenvironments. Mechanotransduction studies were performed, in which single-cell microgels mimicking the cartilage PCM were embedded in high-stiffness agarose. 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.
Pericellular Matrix Formation and Atomic Force Microscopy of Single Primary Human Chondrocytes Cultured in Alginate Microgels
(Wiley, 2023-09) Fredrikson, Jacob P.; Brahmachary, Priyanka P.; June, Ronald K.; Cox, Lewis M.; Chang, Connie B.
One of the main components of articular cartilage is the chondrocyte's pericellular matrix (PCM), which is critical for regulating mechanotransduction, biochemical cues, and healthy cartilage development. Here, individual primary human chondrocytes (PHC) are encapsulated and cultured in 50 µm diameter alginate microgels using drop-based microfluidics. This unique culturing method enables PCM formation and manipulation of individual cells. Over ten days, matrix formation is observed using autofluorescence imaging, and the elastic moduli of isolated cells are measured using AFM. Matrix production and elastic modulus increase are observed for the chondrons cultured in microgels. Furthermore, the elastic modulus of cells grown in microgels increases ≈ten-fold over ten days, nearly reaching the elastic modulus of in vivo PCM. The AFM data is further analyzed using a Gaussian mixture model and shows that the population of PHCs grown in microgels exhibit two distinct populations with elastic moduli averaging 9.0 and 38.0 kPa. Overall, this work shows that microgels provide an excellent culture platform for the growth and isolation of PHCs, enabling PCM formation that is mechanically similar to native PCM. The microgel culture platform presented here has the potential to revolutionize cartilage regeneration procedures through the inclusion of in vitro developed PCM.