The mechanism of low-concentration sodium nitroprusside-mediated protection of chondrocyte death

Hyun A Kim,Ki Byoung Lee,Sang-Cheol Bae

doi:10.1186/ar1705

Abstract

Sodium nitroprusside (SNP), a widely used nitric oxide donor, has recently been shown to mediate chondrocyte apoptosis by generating reactive oxygen species, whereas more potent nitric oxide donors do not induce chondrocyte apoptosis. The present study was performed to investigate the protective effect of a low concentration of SNP upon the cytotoxicity of chondrocytes to higher concentrations of SNP, and to elucidate the underlying mechanism. Human osteoarthritis chondrocytes were cultured as monolayers, and first-passage cells were used for the experiments. Chondrocyte death induced by 1 mM SNP was completely inhibited by pretreating with 0.1 mM SNP. This protective effect of SNP was replicated by the guanosine-3',5'κ-cyclic monophosphate analog, DBcGMP. Protection from chondrocyte death conferred by 0.1 mM SNP was mediated by heme oxygenase 1 (HO-1), as was revealed by the increased expression of HO-1 in 0.1 mM SNP pretreated chondrocytes and by the reversal of this protective effect by the HO-1 inhibitor, zinc protoporphyrin. SNP-mediated chondrocyte protection correlated with the downregulation of both extracellular signal-regulated protein kinase 1/2 and p38 kinase activation. SNP at 0.1 mM induced significant NF-κB activation as revealed by electrophoretic mobility shift assays, and the inhibition of NF-κB by MG132 or Bay 11-7082 nullified 0.1 mM SNP-mediated chondrocyte protection. The upregulation of p53 and the downregulation of Bcl-XL and Mcl-1 by 1 mM SNP were reversed by 0.1 mM SNP pretreatment at the protein level by western blotting. Our study shows that priming with 0.1 mM SNP confers complete protection against cell death induced by 1 mM SNP in human articular chondrocytes. This protective effect was found to be correlated with the upregulation of both HO-1 and NF-κB and with the concomitant downregulation of both extracellular signal-regulated protein kinase 1/2 and p38 activation.

Highlights

Articular cartilage consists of chondrocytes, the only cell type present, which are responsible for repairing tissue damage
Chondrocyte death does not correlate with the amount of nitric oxide (NO) released by NO donors A nitrate/nitrite assay kit was used to determine the amount of NO generated by the various NO donor compounds, sodium nitroprusside (SNP), NOC-5, SIN-1, and SNAP [15]
The amounts of NO produced by 2 mM SIN-1 or 2 mM SNAP were 10-fold and 8.9-fold higher than that produced by 2 mM SNP, respectively, but the levels of cell death induced were not as profound as that produced by 2 mM SNP

Summary

Introduction

Articular cartilage consists of chondrocytes, the only cell type present, which are responsible for repairing tissue damage. Chondrocyte death and the pertinent signaling pathway involved have been the focus of interest recently as pathogenetic factors leading to joint cartilage degradation in various forms of arthritides [1,2]. Several stimuli involved in the pathophysiology of arthritis, including nitric oxide (NO), Fas receptor ligation, and ceramide, have been reported to induce chondrocyte death in vitro [3,4,5]. The pathogenetic involvement of NO in arthritis was first demonstrated when levels of nitrite, a stable end product of NO metabolism, were shown to be elevated in serum and synovial fluid samples of rheumatoid arthritis patients and osteoarthritis patients [6]. Because osteoarthritic cartilage produces large amounts of NO, it could serve as a powerful initiator of chondrocyte death. In addition to the negative effects of NO on cartilage matrix synthesis (i.e. the inhibition of cartilage matrix macromolecule neosynthesis), the enhancement of matrix metalloproteinase activity, and the reduction of IL-1

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