In vitro and in vivo systems mechanobiology of osteoarthritic chondrocytes
Zignego, Donald Lee
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All cells are subjected to and respond to mechanical forces, but the underlying processes linking the mechanical stimuli to biological responses are poorly understood. In the joints of the body (e.g. the knee, hip, etc...) articular cartilage serves as a low friction, load bearing material and is subjected to near-constant mechanical loading. Through excessive loading of the joint, usually caused by obesity or injury, the protective articular cartilage begins to diminish, leading to the progression of osteoarthritis (OA). Osteoarthritis is the most common joint disorder in the world and is characterized by the deterioration of articular cartilage. Determining the link between cartilage deterioration and mechanical loading is one motivation that drove this research. Articular cartilage is composed of a dense extracellular matrix (ECM), a less-stiff pericelluar matrix (PCM), and highly specialized cells called chondrocytes. As the sole cell type in cartilage, chondrocytes are responsible for the healthy turnover of the ECM by creating, maintaining, and repairing the matrix. Multiple lines of evidence suggest chondrocytes can transduce mechanical stimuli into biological signals. The hypothesis for this research is that physiologically pertinent loading of chondrocytes results in a specific set of bio-signals resulting in matrix synthesis. To test this hypothesis, two unbiased, large-scale metabolomic and phosphoproteomic datasets were generated by modeling physiological compressive loading on 3D-embedded chondrocytes. To assess loading-induced changes in metabolites (e.g. small molecules representing the functional state of the cell) and proteome-wide patterns of post-translational modifications (i.e. phosphorylation), chondrocytes were encapsulated in physiologically stiff agarose, compressively loaded in tissue culture, and analyzed via liquid chromatography -- mass spectrometry (LC-MS). The results helped identify global and local biological patterns in the chondrocytes which are a direct result from mechanical loading. In addition, a novel mouse model that expresses cartilage specific bioluminescence was used to assess loading induced changes in vivo. The results from the mouse model allowed for in vivo validation and integration of the in vitro results from the metabolomic and phosphoproteomic results. To my knowledge, such research has never been done, and considerably expands the scientific knowledge of chondrocyte mechanotransduction.