Il ruolo delle HDL nel controllo dell'infiammazione sinoviale

Recent reports suggest that apolipoproteins (apo) exert an important role in controlling inflammation. It has been demonstrated that high density lipoprotein (HDL) are able to block the contact-mediated activation of monocytes-macrophages by stimulated T lymphocytes and inhibit the production of IL-1ß and TNF?. Aim of the thesis: To investigate the effects of HDL on MCP-1 release from monosodium urate crystals-stimulated synoviocytes and IL-1ß, TNF? and IL-1Ra release from microparticles-stimulated monocytes. Methods: Human synoviocytes were obtained by synovial tissue explants from patients with osteoarthritis and stimulated with monosodium urate (MSU) crystals (0.01-0.25 mg/ml) in the presence or absence of human HDL (50 e 100 ?g/ml). Microparticles (MP) were isolated by ultracentrifugation from T lymphocytes and HUT-78 cultures stimulated with phorbol myristate acetate (PMA) (5 ng/ml) and phytohemagglutinin (PHA) (1 ?g/ml) for 48 and 6 h respectively. The cellular origin of MP was determined by flow cytometry. Human monocytes were activated for 48 h by MP from T lymphocytes at concentrations of 13.3 ?g/ml and 26.6 ?g/ml. HUT-78-derived MP were used at a concentration of 1.5-6 ?g/ml in the presence or absence of human HDL (0.2 mg/ml). HDL were isolated from peripheral blood of healthy volunteers by ultracentrifugation. MCP-1 was determined in cultured cells by western blotting and confocal microscopy, while the production of IL-1ß, TNF? and IL-1Ra was measured in culture supernatants by ELISA. Results: Confocal microscopy and western blotting analysis revealed that MCP-1 resides in small cytoplasmatic granules on non stimulated cells. The exposure of synoviocytes to MSU crystals leads to a decrease of intracellular levels of the protein and an increase of extracellular chemokine concentration. The treatment of synoviocytes with HDL causes a dose-dependent inhibition of the release of MCP-1 which maintains its storage in granules. The same effect was observed pre-incubating cells with HDL 1 h before crystal activation. MP generated by stimulated T cells induce a production of IL-1ß, TNF? and IL-1Ra in monocyte cultures higher than those obtained by MP from unstimulated T lymphocytes. It has also been observed that monocytes stimulated with MP generated by activated HUT-78 release IL-1ß, TNF? and IL-1Ra in a dose-dependent manner. The treatment with HDL inhibits IL-1ß and TNF? levels, whereas the production of IL-1Ra remains unchanged. Conclusion: The inhibitory activity of HDL highlighted by the pre-treatment of cells is probably due to a direct action of lipoproteins on synoviocytes rather than to their adsorption on the surface of the crystals. By inhibiting MCP-1 release, HDL may limit the inflammatory process. The production of cytokines depends on the activation level of cells from which MP take origin. The almost complete inhibition of IL-1ß and TNF? levels by HDL lead us to hypothesize that HDL control cellular contact between MP and monocytes, as already observed in the lymphocytes T - monocytes interaction.