p-Coumaric acid suppresses reactive oxygen species-induced senescence in nucleus pulposus cells

p-Coumaric acid (PCA) is a phenolic acid that is widely present in numerous plants and human diets. Studies have demonstrated the antioxidant and anti-senescence effects of PCA in different cell types. However, the anti-senescence effects of PCA in nucleus pulposus (NP) cells have remained to be determined. In the present study, reverse transcription-quantitative PCR was used to measure the gene expression of Cyclooxygenase-2 (Cox-2), inducible nitric oxide synthase (iNOS), p53, p16, aggrecan and collagen-2 in NP cells. Immunofluorescence staining was used to evaluate the protein expression of p53, p16 and collagen-2 in NP cells. In addition, cell cycle of NP cells was measured by flow cytometry. β-galactosidase staining were used to investigate the senescence of NP cells. Preliminary results indicated that PCA suppressed ROS-induced senescence in NP cells via both the p16 and p53 pathways. NP cells were pretreated with PCA at a concentration of 10 or 50 µg/ml prior to stimulation with 200 µM hydrogen peroxide (H₂O₂). Pretreatment with PCA significantly inhibited H₂O₂-induced cell cycle arrest in a dose-dependent manner. PCA also reduced the gene expression of Cox-2, iNOS, p53 and p16 induced by H₂O₂. By contrast, aggrecan and collagen-2 expression in NP cells was upregulated after PCA treatment. Furthermore, PCA suppressed H₂O₂-induced changes in the protein expression of p16, p53 and collagen-2. H₂O₂ stimulation of NP cells increased senescence-associated β-galactosidase (SA-β-gal) activities, while PCA treatment markedly reversed these SA-β-gal activities. Collectively, the present results indicated that PCA attenuated H₂O₂-induced oxidative stress and cellular senescence, suggesting a potential therapeutic utility of PCA in intervertebral disc degeneration.