Background/Purpose: Huntingtin-interacting protein 1 (Hip1) is an arthritis severity and joint damage gene recently discovered in rodent models of arthritis. Hip1 regulates fibroblast-like synoviocytes (FLS) invasiveness, and FLS invasiveness strongly correlates with joint damage. Previous studies suggested that Hip1 regulates PDGFR signaling in rodent cells. In the present study we aimed at determining whether Hip1 mediates PDGFβ and EGF-induced invasion in RA-derived FLS.

Methods: FLS cell lines from RA patients were transfected with siRNA targeting Hip1 or controls. Knock-down was confirmed with qPCR and western blot. Invasion was studied in a well-established two-chamber assay through Matrigel, where lower chamber contained either PDGFβ (50ng/ml), EGF (1ug/ml) in SFM. Cell were also treated with PDGFβ for 5, 15 and 30 minutes and analyzed by immunofluorescence for pFAK localization and actin cytoskeleton. Total cell lysates from RA FLS stimulated with PDGF were used to quantify matrix metalloproteases (MMP1-3, 8-10, 12, 13), as well as total and phospho-FAK.

Results: siRNA Hip1 knock down in RA FLS (n=6) significantly reduced invasiveness induced by PDGFβ (40% reduction; p=0.03) and EGF (50%; p=0.005) compared with siRNA control cells. Hip1 knock-down cells had fewer thick actin filaments and developed an unusual star-like morphology unable to take the polarized shape typically required for movement and invasion, with impaired formation of lamellipodia. Total levels of phospho-FAK did not differ between Hip1 knock-down and control FLS. However, in Hip1 knock-down FLS phospho-FAK distribution was significantly different from controls and distant from the cell periphery. Levels of MMPs were not affected by siRNA Hip1.

Conclusion: We describe a new role for Hip1 in the regulation of EGF and PDGF-induced invasiveness of RA FLS, and show that in the absence of Hip1 the cells takes an unusual and non-invading morphology without lamellipodia. These new findings provide new understanding of events regulating FLS behavior in RA and have the potential to generate a new prognostic biomarkers and a new intra-cellular target for therapies aimed at reducing joint damage.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/huntingtin-interactin-protein-1-hip1-regulates-invasiveness-actin-filament-and-lamellipodia-formation-in-rheumatoid-arthritis-fibroblast-like-synoviocytes-fls/