and objectives The production of ELR+ CXC chemokines is widely studied in arthritis and is postulated to contribute to the infl ammatory phenomena that lead to cartilage breakdown. Healthy articular chondrocytes however, also express their own chemokine receptors and ligands. The function of CXC chemokine receptors in these cells is puzzling because chondrocytes are encased in a dense extracellular matrix and are not known to migrate in vitro. This study aims to identify the function of this

more »

... naling mechanism in articular cartilage. Materials and methods Adult human articular chondrocytes (AHAC) were expanded in monolayer culture under standard conditions. Receptor expression was confi rmed using semiquantitative RT-PCR, western blot and immunohistochemistry. CXCR1/2 combined and individual functionality was tested using an in vitro calcium mobilisation assay. CXCR1/2 signalling was blocked at specifi c receptor level using validated blocking antibodies and siRNA, or at the downstream level using Pertussis toxin, PI3K inhibitors and intracellular calcium chelators. Chondrocyte phenotypic gene expression was assessed using real time RT-PCR. The content of highly sulphated proteoglycans in chondrocyte micromasses was analysed using Alcian blue staining followed by guanidine HCl extraction and spectrophotometric quantifi cation. CXCL6 and CXCL8 were detected in heparitinase digested, chondroitinase ABC digested and un-digested paraffi n sections from on 24 July 2018 by guest. Protected by copyright.