Scutellarin Protects Against Mitochondrial ROS-Dependent NLRP3 Inflammasome Activation To Attenuate Intervertebral Disc Degeneration

Objective:Intervertebral disc degeneration(IVDD)is a common degenerative disease that is the main cause of intervertebral disc herniation and carries a heavy burden on society and families.Although there are many surgical methods to treat patients with intervertebral disc herniation,it still has a high recurrence rate after surgery because of the unique structure of the intervertebral disc,which increases the economic burden on patients.IVDD is a complex process,with the inflammatory reaction of nucleus pulposus cells,mitochondrial dysfunction,and extracellular matrix(ECM)degradation.Maintaining the metabolic balance of nucleus pulposus cells and reducing the degradation of ECM have become potential therapeutic strategies for intervertebral disc degeneration.Scutellarin,the active ingredient extracted from a kind of traditional Chinese medicine Scutellaria baicalensis,has been reported to produce therapeutic potential in some degenerative diseases by inhibiting the inflammatory reaction and regulating cell metabolism.However,whether scutellarin can improve IVDD is still unknown.Therefore,we aim to explore the therapeutic potential and possible mechanism of scutellarin in intervertebral disc degeneration through in vitro and in vivo experiments.Methods:We obtained degenerated nucleus pulposus tissue of human Pfirmann grade Ⅰ toⅡ and extracted human primary nucleus pulposus cells(HNPCs)to culture in vitro.TNF-αstimulation was used to simulate the inflammatory reaction in intervertebral disc degeneration,and scutellarin was added or not to explore whether scutellarin has a protective effect.Additionally,we established a rat acupuncture model to simulate the environment of nucleus pulposus prolapse and injected Scutellarin into the rat’s caudal intervertebral disc.After a certain period,MRI and X-ray examinations were performed on the rat’s caudal vertebra,and a histological examination was performed to verify the protective function of scutellarin on IVDD.Results:In vitro experiments,RT-PCR,Western Blot,and immunofluorescence showed that TNF-α enhances the inflammatory reaction and catabolism of primary nucleus pulposus cells,while scutellarin can weaken the inflammatory reaction and retain the main components of IVD.In vivo,the injection of scutellarin into the intervertebral disc maintained the high signal and the height of the intervertebral disc on the T2-weighted MRI image.In terms of mechanism,Scutellarin reduces the production of reactive oxygen species(ROS),reduces the damage of mitochondria,and decreases the expression level of apoptosis-related biomarkers under the stimulation of TNF-α.Additionally,Scutellarin can also antagonize the activation of the NF-κB signal pathway and MAPK signal pathway and inhibit the TNF-α-mediated activity of NLRP3 inflammatory corpuscles.Conclusion:During intervertebral disc degeneration,the activity of pro-inflammatory factors such as TNF-α is enhanced,which in turn promotes the production of downstream inflammatory factors,leading to increased decomposition of IVD and accelerated degradation of ECM.After treatment with scutellarin,we found that scutellarin can inhibit the inflammatory response of HNPCs,reduce the loss of ECM components,and protect mitochondrial function by reducing the production of ROS,thus reducing the activation of NLRP3 inflammatory body and the expression of apoptosis-related markers,which has a protective effect on IVDD.This provides a new treatment strategy for future intervertebral disc degeneration.