Binding of type II collagen fragments to the surface of human articular chondrocytes

Type II collagen binds to chondrocytes through integrins and Annexin V. While the potential integrin binding sites have been identified, it is unclear which domains bind to Annexin V. Proteolytic fragments of collagen are known to modulate cell signaling pathways resulting in degradation of articular cartilage; it is unknown whether Annexin V binds the fragments. One of the goals of this study was to determine the binding of type II collagen and its fragments to chondrocytes using flow cytometry and fluorescence microscopy. Our data suggest that the binding of the N-telopeptide of type II collagen is through Annexin V, while binding of the C-telopeptide and the triple helical peptide to the surface of chondrocytes are potentially facilitated through other collagen receptors, such as integrins and cell-associated matrix proteins.; One of the characteristics of Annexin V is that it is a calcium ion receptor. Most physiological processes utilize intracellular and/or extracellular calcium ions. Calcium ions within the cytosol act as a key intracellular second messenger and as an enzymatic cofactor. Therefore, intracellular calcium flux was measured upon N-telopeptide binding to Annexin V. Flow cytometry data demonstrated that upon N-telopeptide binding to the cell surface, calcium is rapidly released from intracellular stores. Calcium release was also observed with C-telopeptide and helical peptide, but to a lesser extent. In addition, chondrocyte cytosolic baseline calcium levels decreased after prolonged exposure to all three peptides. Further investigation with bradykinin, a known inflammatory agent, demonstrated a rapid release of stored calcium. The cells pre-treated with all three peptides seem to be more sensative to bradykinin than the cells treated with type II collagen. It may be hypothesized that in the diseased tissue cells that are exposed to fragments of type II collagen may have increased sensitivity to inflammatory stimuli that may be present in the matrix such as various cytokines.