Synovial tissues were obtained from 10 OA patients (4 females and 6 males; median age was 73.3 [77.5–69.5] years old) and 10 OA-DB patients (2 females and 8 males; median age was 74.6 [83.4–65.8] years old) who underwent joint replacement surgery and gave informed consent. This study was reviewed and approved by the Local Ethics Committee. Samples were subsequently embedded in paraffin for obtaining 4 µm-thick histological section. For in vitro experiments, we used the immortalized cell line of monocytes TPH-1 that was maintained in Roswell Park Memorial Institute Medium (RPMI)-1640 (ThermoFisher, Madrid, Spain) containing 10% fetal bovine serum (FBS), 2 mM l-glutamine, 100 mg/mL streptomycin, and 100 U/mL penicillin (Lonza, Basel, Switzerland). TPH-1 were differentiated into macrophages after treatment with phorbol-12-myristate-13-acetate (PMA; 500 nM) (Sigma-Aldrich, St Lois, USA) for 3 h [8 (link)]. Thereafter, macrophages were incubated in RPMI-1640 2% FBS with 1 g/l glucose (normal glucose, NG) or 4.5 g/l (high glucose, HG) and stimulated with bacterial lipopolysaccharides (LPS; 1 ug/ml) (Sigma-Aldrich, San Luis, MO, USA), a classical inductor of M1-like macrophages [14 (link), 29 ], in the presence or absence of a slow-releasing H2S donor, GYY-4137 (500 µM) (SantaCruz Biotechnology, Heidelberg, Germany), based on previous research [46 , 66 (link)].
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