Effect of High Glucose-induced Oxidative Stress and Inflammatory Response on Apoptosis and Extracellular Matrix Metabolic Imbalance in Nucleus Pulposus Cells
Objective To investigate the effect of high glucose treatment on apoptosis and extra-cellular matrix(ECM)metabolism in rat nucleus pulposus cells(NPC)and to preliminarily explore possible underlying mechanisms.Methods NPC were isolated from lumbar spine tissues of male Wistar rats.NPC were divided into 6 groups:Control group,Glucose-treated groups(treated with 5 mmol/L,15 mmol/L,and 25 mmol/L glucose,respectively),high glucose+NAC group(treated with 25 mmol/L glucose and 3 mmol/L antioxidant NAC),high glucose+PDTC group(treated with 25 mmol/L glucose and 10 μmol/L NF-κB pathway inhibitor PDTC).Cell viability and apoptosis were detected by cell counting kit-8 and flow cytometry,respectively.Fluorescent probes were used to detect intracellular reactive oxygen species(ROS)levels.Enzyme-linked immunosorbent assay was used to detect IL-1β,IL-6 and TNF-α levels.Western blotting was used to detect relevant protein ex-pression levels.Results Compared to the cell viability and apoptosis in the control group,no signifi-cant change was observed in the 5 mmol/L and 15 mmol/L glucose groups,however,a significant decrease in cell viability and a significant increase in apoptosis were seen in the 25 mmol/L glucose group(P＜0.05).The expression levels of Bcl-2-associated X protein(Bax),Caspase-3,MMP-3,MMP-9,MMP-13,A disintegrin and metalloproteinase with thrombospondin 4(ADAMTS-4),and ADAMTS-5 were significantly decreased in the glucose+NAC and glucose+PDTC groups when compared with those in the 25 mmol/L glucose group,whereas the expression levels of Bcl-2,colla-gen Ⅱ(COL Ⅱ),and aggrecan were significantly increased(P＜0.05).The intracellular ROS levels and supernatant levels of the IL-1 β,IL-6,and TNF-α in the glucose+NAC and glucose+PDTC groups were significantly lower when compared with those in the 25 mmol/L glucose group(P＜0.05).In addition,the cellular levels of p-P65/P65 and p-IκBα/IκBα were significantly reduced in the glucose+NAC and glucose+PDTC groups when compared with those in the 25 mmol/L glucose group(P＜0.05).Conclusion High glucose treatment promotes NPC apoptosis and ECM degrada-tion by inducing oxidative stress and inflammatory responses,which involves the activation of ROS/NF-κB signaling pathway.
high glucoseintervertebral disc degenerationnucleus pulposus cellsoxidative stressinflammationextracellular matrix
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