Background and Objectives Lasp-1 is an important actin-crosslinking protein. It’s localises at focal adhesions, along stress fibres and leading edges and is involved in cellular migration and metastatic dissemination of different cancers. Rheumatoid arthritis synovial fibroblasts (RASF) are also able to enter the bloodstream from sites of cartilage destruction to new locations where they re-initiate the destructive processes. The underlying mechanisms of this process are of special interest, but currently unclear. Therefore we have investigated the role of Lasp-1 in synovial fibroblast migration during RA.
Materials and Methods Lasp-1 expression and its subcellular location was analysed using Western blotting and immunofluorescence microscopy of cells seeded on various coatings, including fibronectin, laminin and a self-purified collagen matrix. Furthermore, Lasp-1 deficient mice were generated, which were interbred with hTNFtg mice, to form a model of inflammatory arthritis. Mice of all genotypes were analysed using a clinical score, measuring weight, grip strength and paw swelling over a time course of 14 weeks. Hind paws from each genotype were isolated, paraffin-embedded and toluidine blue stained in order to assess synovial inflammation, cartilage degradation and SF attachment. Cell migration properties of SFs derived from WT, Lasp-1-/-, hTNFtg and Lasp1-/-/hTNFtg were measured using a modified scratch assay and by live cell imaging.
Results Lasp-1 expression was up regulated in RASF and in SF from hTNFtg mice compared to healthy controls. In immunofluorescence, Lasp-1 was co-localised with cortactin, a marker for structures of cell adhesion and invasion, particularly when cultured on a purified collagen matrix. Lasp1-/-/hTNFtg mice showed milder clinical symptoms in comparison to hTNFtg mice. Notably, histopathological analyses revealed less cartilage destruction (0.4 mm vs. 1.6 mm, p<0.05) and less attachment of synovial tissue to the cartilage surface (5% vs. 35%, p<0.05) in Lasp1-/-/hTNFtg mice at 14 weeks of age. Lasp-1-/- SFs and Lasp1-/-/hTNFtg SFs exhibited a significantly reduced migration rate in comparison to WT and hTNFtg SFs, with significant differences observed in cell migration speed, cell spreading and in the formation of stress fibres and filopodia.
Conclusion Lasp-1 deficiency leads to significantly reduced migration rates of RASF and hTNFtg SF in vitro and results in a decreased cartilage degradation and destruction and less SF attachment to articular cartilage in hTNFtg mice in vivo. Therefore, Lasp-1 is an important target for therapeutic strategies aimed to reduce the invasive and migratory behaviour of synovial fibroblast in rheumatoid arthritis.
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