A sulfate-arsenical-ferruginous water affects apoptosis, oxidative stress and the gene expression of inflammatory mediators and of a panel of MicroRNA in IL-1β stimulated human osteoarthritic chondrocytes

Background: The beneficial effects of balneotherapy in osteoarthritis (OA) using the sulfate-arsenicalferruginous Levico water (LW) have been recognized by several clinical studies. However, the specific biological properties of LW are partially understood. Objective: The present study aimed at evaluating the ability of LW in the regulation of apoptosis, oxidative stress and of the gene expression of a panel of articular cartilage markers and MicroRNA (miRNA) in human osteoarthritic chondrocytes exposed to interleukin (IL)-1β. Methods: Chondrocytes were obtained from the femoral heads of patients with OA who underwent surgery for total hip prostheses. The cells were incubated for 24 h and 48 h with different concentrations of LW, alone or in combination with IL-1β (10 ng/ml). Apoptosis and mitochondrial superoxide anion production were detected by cytometry. Gene levels of antioxidant enzymes, B-cell lymphoma (BCL)2, cytokines [IL-1β, IL-6, tumor necrosis factor (TNF)-α], metalloproteinases (MMPs)1 and 13, type II collagen (Col2a1), aggrecan, and of a pattern of miRNA were evaluated by Quantitative Real-Time PCR analysis (PCR). The involvement of nuclear factor (NF)-κB was investigated by PCR and immunofluorescence and by pre-incubation with a specific inhibitor (BAY 117082, IKKα/β). Results: LW at 50% or 25% concentration counteracted the negative effects of IL-1β on cell viability, apoptosis and mitochondrial superoxide anion production and on the gene expression of antioxidant enzymes, in a dose-dependent manner. The treatment of chondrocytes with 50% of LW significantly inhibited the gene expression of pro-inflammatory cytokines, MMPs and of miR-34a and miR-181a, while it upregulated miR-140 in IL-1β stimulated cells. The pre-incubation with BAY 11-7082, IKKα/β reduced the damage induced by IL-1β, and, interestingly, potentiated the properties of LW. Conclusions: Our data demonstrated the ability of LW to regulate apoptosis, oxidative stress and gene expression of main factors implicated in OA pathogenesis through a possible modulation of NF-κB signaling pathway. This study supports the use of balneotherapy with a sulfate-arsenical-ferruginous mineral water in the treatment of OA.