Study On The Mechanism Of Resveratrol In Promoting Peripheral Nerve Repair

BackgroundPeripheral nerve injury is a common clinical disease.Peripheral nerve injury leads to patients'mobility inconvenience or even disability,which brings patients not only with life health problems,but also causes huge social pressure and economic burden.Different from the central nervous system,the peripheral nerve has the ability of regeneration after injury.Wallerian degeneration occurs at the distal end of the injured nerve,which is considered to be a key step in the regeneration of the injured nerve.Autophagy widely exists in many kinds of cells.It can maintain homeostasis by degrading the accumulation of proteins and damaged organelles.During autophagy,these organelles or proteins are encapsulated in the bilayer membrane and degraded in lysosomes.In the nervous system,autophagy occurs widely in the normal physiological process,and the development of myelin sheath of peripheral nerve also needs the participation of autophagy process.Under pathological conditions,the reduction of autophagy level can lead to a variety of neurodegenerative diseases;after nerve trauma,the inhibition of autophagy lysosomal system can also lead to neuronal necrosis and degeneration.In the peripheral nervous system,autophagy has been studied more and more widely.Schwann cells play an important role in the process of nerve injury and repair.It can not only secrete a variety of neurotrophic factors,but also has evidence that it can occur autophagy.After peripheral nerve injury,the myelin sheath at the distal end of injured nerve degraded with the rupture of tissue fragments.The degradation of myelin depends not only on the phagocytosis of macrophages,but also on the autophagy of Schwann cells.Resveratrol is a natural product extracted from some foods,which has antioxidant properties,reduces inflammation and slows down cell aging.In recent years,more and more attention has been paid to the effect of resveratrol on autophagy.Resveratrol has a protective effect on the nervous system,which has been proved in the animal models of spinal cord injury and cerebral ischemia-reperfusion injury.In the peripheral nervous system,resveratrol can accelerate the process of nerve regeneration through the vascular endothelial growth factor signal pathway.Therefore,whether resveratrol can promote peripheral nerve regeneration through other ways remains to be further studied.ObjectsIn this study,we established a rat model of sciatic nerve crush injury.The effects of autophagy on nerve injury and repair were studied by using autophagy inhibitors and agonists,and the effects of resveratrol on nerve injury and repair were studied by using resveratrol.Finally,RSC96 cells were stimulated by H₂O₂ to study the protective effect of resveratrol on cells under oxidative stress.Methods1.The animal model of sciatic nerve crush injury in SD rats was established.2.The recovery of motor nerve function was analyzed by Sciatic Function Indexes?SFI?.3.The regeneration nerve fiber and the thickness of myelin sheath after regeneration were analyzed by electron microscopy and toluidine blue staining.4.Western blot and immunofluorescence staining were used to identify the expression of autophagy related protein and myelin protein.Toluidine blue staining was used to observe the early degradation process of myelin and to determine the effect of early Wallerian degeneration on the regeneration rate.5.The effect of autophagy and resveratrol on proliferation after peripheral nerve injury of Schwann cells was evaluated by immunohistochemistry.6.The drug toxicity of resveratrol was studied in cells and animal experiments7.The protective effect of resveratrol on cells was detected by CCK-8 method.8.To investigate the effect of resveratrol on the production of ROS by flow cytometry.9.To study the protective effect of resveratrol and its mechanism by Western blot and immunofluorescence staining.Results1.Autophagy agonists and resveratrol can promote the recovery of motor nerve function after injury;autophagy inhibitors affect nerve regeneration in the early stage,but in the second week,the recovery speed is not statistically different from the control group.2.Morphology study confirmed that autophagy agonists and resveratrol increased the number of nerve regeneration and the thickness of myelin sheath on the14th day after injury.3.Compared with the control group,autophagy activator and resveratrol accelerated the early degradation of myelin sheath,and the level of MPZ decreased.Toluidine blue staining suggested that the degradation of myelin sheath structure was more complete.At the same time,toluidine blue staining showed that the number of intact myelin sheath was more than that of the control group.Resveratrol promoted autophagy of Schwann cells in the early stage of injury.Immunofluorescence staining showed that the expression of LC3 was significantly increased,Western blot showed that the expression of autophagy related protein LC3B-II/LC3B-I was significantly increased;electron microscopy showed that the double membrane structure of autophagy was widespread in Schwann cells.4.Autophagy agonists and resveratrol promoted the proliferation of Schwann cells in the early stage of injury.5.At 25?M concentration,rsc96 cells'activity had no significant difference with the control group;after continuous intraperitoneal administration at the highest concentration of 200mg/kg for one week,HE staining of heart,liver,spleen,kidney and lung tissue of the rats did not show significant damage,indicating that resveratrol has low toxicity,suggesting that resveratrol may be used in future clinical research.6.Resveratrol increased the cell survival rate stimulated by H₂O₂,which could be blocked by autophagy inhibitors.7.Resveratrol reduced the production of ROS in cells stimulated by H₂O₂.8.Under the action of resveratrol,the level of autophagy related protein increased,p62 decreased significantly,mTOR phosphorylation decreased;resveratrol reduced the expression level of scorch related protein,NLRP3,caspase-1 and IL-1?;with the increase of concentration,resveratrol inhibited the level of TXNIP.Conclusion1.Resveratrol can promote the recovery of motor neuron function after crush injury.2.Resveratrol can promote the autophagy of Schwann cells after injury,promote the degradation process of myelin sheath in the early stage of injury,thus accelerate the speed of nerve regeneration;at the same time,resveratrol can also promote the proliferation of Schwann cells.3.Resveratrol has low cytotoxicity and good clinical application prospect.4.Resveratrol can promote autophagy,reduce oxidative stress level,and reduce pyroptosis by inhibiting mTOR phosphorylation;resveratrol can enhance autophagy.At the same time,resveratrol can directly reduce the expression of TXNIP in the cells.