Pentosan polysulfate inhibits IL-1β-induced iNOS, c-Jun and HIF-1α upregulation in canine articular chondrocytes

Osteoarthritic (OA) chondrocytes are shown to express inducible nitric oxide synthase (iNOS) which produces high concentrations of nitric oxide (NO), particularly when stimulated with proinflammatory cytokines. NO is involved in OA cartilage degradation. On the other hand, c-Jun N-terminal Kinase (JNK) pathway mediates the activation and transcription of c-Jun, which is required for interleukin-1 (IL-1)-induction of matrix metalloproteinases-13 (MMP-13) in OA pathogenesis. Therefore, the selective inhibition of iNOS and c-Jun is a promising target for treatment and prevention of OA. The purpose of the study was to investigate the inhibitory effects of pentosan polysulfate (PPS) on IL-1 beta-induced iNOS, c-Jun and HIF-alpha isoforms upregulation in canine articular chondrocytes (CACs). Primary (P0) chondrocytes were isolated and cultured from femoral head cartilages of three (3) dogs. First passage (P1) chondrocytes were preincubated with 0, 1, 5, 15 and 40 mu g/mL of PPS for 4 hr before treatment with 10 ng/mL rhIL-1 beta for a further 8 hr. In addition, we evaluated the effects of single and multiple cytokine with or without LPS on iNOS protein induction. PPS significantly inhibited (P < 0.05) IL-1 beta-induced iNOS, c-Jun and HIF-1 alpha mRNA upregulation in a dose-dependent pattern. iNOS mRNA was significantly inhibited at 15 and 40 mu g/mL whereas c-Jun and HIF-1 alpha were significantly downregulated at 5, 15 and 40 mu g/mL of PPS compared to chondrocytes treated with only rhIL-1 beta. Intriguingly, CACs were recalcitrant to single IL-1 beta, TNF-alpha or LPS-induction of iNOS protein including to a combination of IL-1 beta+TNF-alpha, IL-1 beta+LPS except to TNF-alpha+LPS and IL-1 beta+TNF-alpha+LPS suggestive of a protective mechanism from iNOS detrimental effects on perpetuating OA. IL-1 beta+TNF-alpha+LPS-induced iNOS protein expression was significantly abrogated by PPS. We demonstrate for the first time that PPS is a novel inhibitor of IL-1 beta-induced iNOS, c-Jun, and HIF-1 alpha mRNA upregulation and iNOS protein induction which may be beneficial for prevention and treatment OA.