IL RUOLO DEL PATHWAY SOCS-1/TRAF-6 NELL'INCREMENTO DELL'OSTEOCLASTOGENESI IN PAZIENTI HIV+ IN CART

Up to 30% of cART treated HIV-infected patients show an early onset osteopenia/osteoporosis, leading to a significant increase in the risk of fractures. The pathogenic molecular mechanisms remain poorly understood. However, it has been shown that, in HIV transgenic rats model, osteoclasts (OCs) express high levels of suppressor of cytokine signaling-1 (SOCS-1) and tumor necrosis factor receptor-associated factor-6 (TRAF-6), inducing an increased osteoclastogenesis through IFN-γ pathway inhibition. Given these premises, our study aims to investigate osteoclastogenesis and SOCS-1/TRAF-6 pathways in human peripheral blood-derived-OCs of HIV positive cART-treated patients with and without reduced BMD (Bone Mineral Density). Material and Methods. We consecutively enrolled 40 HIV+ patients on virally-effective cART (HIV-RNA <40cp/ml) matched for viro-immunologic/demographic features: 16 resulted with normal BMD (nBMD) by dual x-ray absorptiometry (osteopenia, z-score -1/-2.5; osteoporosis, z-score <-2.5), 24 with reduced BMD (rBMD, including osteopenic and osteoporotic patients). We enrolled 15 HIV-negative as healthy controls (HIV-). Circulating OC precursors (OCPs; CD14+CD11b+CD51/61+ and C14+/CD16++) were studied by flow-cytometry. OCs were differentiated from peripheral blood-purified CD14+ supplemented with M-CFS and RANKL (8 days). OCs differentiation/maturation was evaluated by: TRAP staining and dentin resorption. RANK, SOCS-1, TRAF6, MMP-9 and CTSK expression of OCs ex vivo differentiated were analyzed by Real-Time PCR and Western-Blot with and without IFN-γ stimulation (100 U, 2 h). Kruskal-Wallis, Mann–Whitney and ANCOVA were used. Results. Higher levels of OCPs (CD14+CD11b+CD51/61+ and C14+/CD16++) were observed in HIV+ rBMD group compared to HIV+ nBMD (respectively, p=.0001; p=.0068). CD14+ cultures from HIV+ rBMD were enriched in large multinucleated TRAP+ OCs versus nBMD and HIV- (p=.012; p=.023), as confirmed by TRAP quantification (p=.038; p=.04). Interestingly, OCs from HIV+ rBMD expressed significantly higher SOCS-1 and TRAF-6: > 2.14, >1.65 fold vs HIV-, respectively, significantly higher than nBMD (p=.027; p=.014). IFN-γ challenge resulted in an up-regulation of SOCS-1 in HIV+ rBMD/nBMD OCs and HIV- OCs, respectively (p=.0022; p=.0057; p=.0013). Despite this, only in HIV+ rBMD we detected an increasing in TRAF-6 (p=.024). No difference was found in MMP-9, on the contrary we observed an increasing trend of CTSK mRNA expression in HIV+ rBMD OCs. Interestingly, ANCOVA test demonstrated an interaction between SOCS-1/ TRAF-6 upregulation, HIV infection and bone impairment (p=.003, p=.025). Conclusion. HIV+ rBMD patients show increased circulating OCPs and OCs ex-vivo differentiation, indicating heightened osteoclastogenesis in treated HIV+ osteopenic/osteoporotic patients. Relevantly, we show that OCs from HIV+ rBMD express high SOCS-1/TRAF-6 levels, that are further upregulated upon IFN-γ challenge. Together, these findings identify deregulated SOCS-1/TRAF-6 as molecular pro-osteoclastogenesis pathway that might be sustained by abnormal interferon-mediated inflammation in successfully-treated HIV.