Effects of fluoroquinolones on cultured articular chondrocytes flow cytometric analysis of free radical production.

Using flow cytometry, we previously established in an ex vivo model that fluoroquinolones induce a stimulation of the oxidative metabolism in immature chondrocytes. To assess these findings in an in vitro model, primary cultures of immature articular chondrocytes were incubated with four quinolone solutions: ofloxacin, ciprofloxacin, nalidixic acid at 10 micrograms/ml for 24 hr and pefloxacin at 1, 10 and 100 micrograms/ml for various periods of time (2, 4, 6, 12, 24 and 48 hr). Three fluorochromes were used: DCFH-DA, reflecting cellular production of H2O2, rhodamine 123 (Rh123) and 10-N-nonyl-acridine orange (NAO), which are specific for mitochondrial activity and mass, respectively. In immature chondrocyte cultures treated with pefloxacin, ofloxacin and nalidixic acid at 10 micrograms/ml for 24 hr, levels of cellular fluorescent dichlorofluorescein DCF (oxidized form of DCFH-DA) were significantly higher than in control cells. No significant increase could be registered with ciprofloxacin. In the same experimental conditions, incorporation of Rh123 and NAO was not significantly modified. Pefloxacin (10 micrograms/ml, 24 hr) did not induce any significant increase of DCFH-DA processing either in mature chondrocytes or in alveolar macrophages removed from immature rabbits. Quinolones induce in vitro an early stimulation of the oxidative metabolism in immature but not in mature chondrocytes, a phenomenon that could explain juvenile onset of quinolone arthropathy. This in vitro model could be proposed as an easy and reproducible method for screening potential arthrotoxicity of antimicrobial agents, capable of stimulating the formation of H2O2.