Valvason C, Musacchio E, Pozzuoli A, Ramonda R, Aldegheri R, Punzi L
Rheumatology Unit, Department of Clinical and Experimental Medicine, University of Padova, Via Giustiniani, 2-35128 Padova, Italy.
Rheumatology (Oxford). 2008 Jan;47(1):31-5. doi: 10.1093/rheumatology/kem289.
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.
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)-1beta and simultaneously treated with GS. Moreover, HO-1 protein and total nitrite production were evaluated.
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-1beta stimulation induced expression of iNOS reverted by 1 and 10 mmol/l GS. Moreover, HO-1 gene expression was down-regulated by IL-1beta and 10 mmol/l GS restored baseline values. These data were confirmed by evaluating HO-1 protein level and nitrite production.
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.
活性氧（ROS）是关节软骨改变的主要决定因素。在细胞保护机制中，血红素加氧酶（HO-1）的诱导型同工型发挥着特别重要的作用。另一方面，烟酰胺腺嘌呤二核苷酸磷酸（NADPH）系统的酶活性可能有助于ROS的产生。硫酸氨基葡萄糖（GS）是用于治疗骨关节炎的药物之一；然而，其作用机制仍 largely 未知。本研究的目的是研究 GS 对原代人软骨细胞的体外影响，特别是关于 HO-1、p22(Phox)（NADPH 复合物的一个亚基）和诱导型一氧化氮合酶（iNOS）表达的影响。
用不同浓度的 GS 处理原代人软骨细胞；通过逆转录-聚合酶链反应法评估 HO-1、p22(Phox)和 iNOS 的基因表达。在另一组实验中，用人重组白细胞介素（IL）-1β刺激细胞并同时用 GS 处理。此外，评估了 HO-1 蛋白和总亚硝酸盐的产生。
24 小时时，10 mmol/l 的 GS 上调了 HO-1 基因表达（相对于对照组增加 40%，P < 0.001），而 10 mmol/l 的 GS 下调了 p22(Phox)基因表达，在处理 48 小时后观察到最大抑制作用。IL-1β刺激诱导的 iNOS 表达被 1 和 10 mmol/l 的 GS 逆转。此外，IL-1β下调了 HO-1 基因表达，10 mmol/l 的 GS 恢复了基线值。通过评估 HO-1 蛋白水平和亚硝酸盐产生证实了这些数据。
本研究中观察到的 GS 对氧化应激的影响揭示了一种可能的新作用机制，似乎与对软骨细胞群体的潜在保护作用一致。
