In vitro stimulation of human intervertebral disc (IVD) cells.Objective.
To investigate the oxidative/nitrosative effects of peroxynitrite on human nucleus pulposus (NP) cells.Summary of Background Data.
Peroxynitrite is an important tissue-damaging species generated at sites of inflammation and degeneration. The aim of this study was to examine the effects of oxidative/nitrosative stress caused by peroxynitrite and the peroxynitrite donor SIN-1 in human NP cells.Methods.
Degenerated human IVD tissue was analyzed for nitrosylation by immunofluorescence. In addition, human NP cells were isolated from IVDs, expanded and stimulated either with peroxynitrite itself or a stable peroxynitrite donor (SIN-1). Nitrosylation, accumulation of intracellular reactive oxygen species, NF-κB nuclear translocation, and cell viability were analyzed by fluorescence. Gene expression of TNF-α, IL-1β, IL-6, IL-8, and IL-10 was quantified by real-time (RT)-PCR.Results.
Degenerated IVD tissue showed strong nitrosylation, especially in the NP. Isolated human NP cells showed a strong signal for nitrosylation and intracellular reactive oxygen species on stimulation with peroxynitrite or SIN-1. NF-κB/p65 sustained nuclear translocation of NF-κB/p65 and stimulation of IL-1β, IL-6, and IL-8 expression was noted on treatment of cells with SIN-1.Conclusion.
This study provides evidence that peroxynitrite may play a role in disc degeneration and discogenic back pain development by an increased synthesis of proinflammatory cytokines. Nuclear translocation of NF-κB was identified as the potential underlying pathway. Therefore, neutralizing peroxynitrite and its derivatives (e.g., via the use of antioxidants) may be a novel treatment option for discogenic back pain.