Verbruggen Gust, Wang Jun, Wang Lai, Elewaut Dirk, Veys Eric M
Department of Rhumatology, Ghent University Hospital, Ghent University, Ghent, Belgium.
Methods Mol Med. 2004;100:183-208. doi: 10.1385/1-59259-810-2:183.
Flow cytometry has been used as a procedure to characterize the phenotype and function of human articular cartilage cells cultured as monolayers or in gelled artificial matrices. Procedures allowing intact cells with their cell-associated matrix, to be obtained have been described. Appropriate monoclonal antibodies have allowed plasma membrane-associated proteins, e.g., growth factors and cytokine receptors, as well as the cell-associated extracellular matrix macromolecules, to be studied. Intracellular compounds have been traced in permeabilized cells after blocking of their intracellular transport and secretion mechanisms. We report the use of fluorescent dye-labeled monoclonal antibodies or specific binding proteins against extracellular matrix compounds such as hyaluronan, aggrecan, types I and II collagen, and fibronectin. The autocrine and paracrine growth factor and cytokine pathways considered include the insulin-like growth factor-1 (IGF-1)/IGF receptor I (IGFRI), and the transforming growth factor-beta1 (TGF-beta1)/TGF-beta receptor II (TGF-betaRII) cascades, as well as the interleukin-1alpha/beta (IL-1alpha/beta)/interleukin-1 receptors I and II (IL-1RI and II) systems. Catabolic enzymes that mediate extracellular matrix turnover, e.g., some matrix metalloproteinases and their natural inhibitors, were also studied. Finally, flow cytometry was used to assess the results of some pharmacological interventions on the aforementioned variables in cultured chondrocytes.
