Influence of glucosamine sulphate on oxidative stress in human osteoarthritic chondrocytes: effects on HO-1, p22Phox and iNOS expression

Abstract

Objective. Reactive oxygen species (ROS) are major determinants in the alteration of articular cartilage. Among protective cellular mechanisms, the inducible isoform of haem oxygenase (HO-1) plays a particularly relevant role. On the other hand, the enzymatic activity of the Nicotinamide adenine dinucleotide phosphate (NADPH) system could contribute to the generation of ROS. Glucosamine sulphate (GS) is one of the drugs used in the treatment of osteoarthritis; however, its mechanism of action is still largely unknown. The aim of the present study was to investigate the effects of GS on primary human chondrocytes in vitro, in particular with regard to HO-1, p22^Phox (a subunit of NADPH complex) and inducible nitric oxide synthase (iNOS) expression. Methods. Primary human chondrocytes were treated with different concentrations of GS; gene expression of HO-1, p22^Phox and iNOS was assessed by the reverse transcriptase–polymerase chain reaction method. In a separate set of experiments, the cells were stimulated with human recombinant interleukin (IL)-1β and simultaneously treated with GS. Moreover, HO-1 protein and total nitrite production were evaluated. Results. HO-1 gene expression was up-regulated (+40% with respect to the controls, P < 0.001) by 10 mmol/l GS at 24 h, while p22^Phox gene expression was down-regulated by 10 mmol/l GS with a maximum inhibitory effect observed after 48 h treatment. IL-1β stimulation induced expression of iNOS reverted by 1 and 10 mmol/l GS. Moreover, HO-1 gene expression was down-regulated by IL-1β and 10 mmol/l GS restored baseline values. These data were confirmed by evaluating HO-1 protein level and nitrite production. Conclusions. The influence of GS on oxidative stress observed in this study discloses a possible new mechanism of action and seems to be in keeping with a potential protective effect on chondrocyte population.