The Neuroprotective Effects Of Sinomenine Against Secondary Brain Injury In The Mice Model Of Traumatic Brain Injury

BACKGROUD:Traumatic brain injury(TBI)remains a major public health problem in modern society,with high morbidity and mortality,imposing ecomic burden to the family and society.The pathological process of TBI involves primary and secondary injury.After primary brain insult,a complex series of endogenous events referred to as secendary brain injury are triggered,including oxidative stress,glutamate excitotoxicity,activation of the inflammatory response,loss of ionic homeostasis,and increased vascular permeability,leading to further neuronal degeneration and apoptosis.Despite great efforts were made to seek effective ways to alleviate the secondary injury,to date,most approaches to the treatment of TBI targeting a single injury mechanism have failed in clinical trials.Oxidative stress plays an important role in secondary injury,which occurs when the production of reactive oxygen species(ROS)exceeds the detoxification of cell.The excessive production of ROS damages cellular components including lipids,proteins,and DNA,leading to a decline in physiological function and cell death.The transcription factor nuclear factor erythroid 2-related factor 2(Nrf2)is a key protein in the reduction of oxidative stress.Under normal conditions,kelch ECH associating protein 1(KEAP1),a cytosolic repressor protein that binds to Nrf2,retains Nrf2 in the cytosol and facilitates its proteasomal degradation.Once the cell encounters stimulations such as oxidative stress,Nrf2 dissociates from KEAP1 and translocates into the nucleus.By binding to the antioxidant response element(ARE),Nrf2 induces the production of a battery of endogenous enzymes,such as superoxide dismutase(SOD),glutathione peroxidase(GPx),nicotinamide adenine dinucleotide phosphate,quinine oxidoreductase-1(NQO1),and heme oxygenase-1(HO-1).Together,these free radical scavenging enzymes represent a powerful antioxidant defense mechanism.The Nrf2-ARE signaling pathway is activated in several central nervous system(CNS)diseases,including TBI,and is considered a protective molecular mechanism against TBI.Sinomenine(SIN)is an active alkaloid isolated from the Chinese medicinal herb Sinomeniumacutum,and used for the clinical treatment of rheumatoid arthritis and mesangial proliferative nephritis in China.SIN has a variety of pharmacological properties including immunosuppression,anti-inflammation and cytoprotection.SIN exerts neuroprotection in several CNS disease models,including cerebral ischemia,intracerebral hemorrhage,and neurodegenerative diseases.However,few studies have addressed the neuroprotective effect of SIN in TBI.Therefore,the purpose of the present study was to determine whether SIN administration after TBI could attenuate brain injury in a TBI model.Objective:In the present study,we investigated the effects of sinomenine in the mice model of traumatic brain injury,and then clarified the underlying molecular mechanisms.Methods:(1)Adult ICR mice were randomly assigned to the following groups:1)Sham(n = 42);2)TBI(n = 42);3)TBI + vehicle(n = 42);4)TBI + SIN(30 mg/kg i.p.)(n = 42).The model of TBI used in the present study was based on Feeney's weight-drop model with some modifications.(2)Observing the effects of SIN on neurological function and brain edema following TBI.(3)Western blotting was conducted to measure levels of the apoptosis-related proteins Bcl-2 and cleaved caspase-3.NeuN/TUNEL double immunofluorescence staining was performed to evaluate the neuronal apoptosis.(4)The levels of MDA,GPx and SOD,indicators of lipid peroxidation and antioxidant enzyme activity in the brain tissues,were measured.(5)Expression of the NQO-1,HO-1,and Nrf2 were also investigated by Western blotting.Then the localization of Nrf2 were detected by NeuN and Nrf2 double immunofluorescence.(6)The mRNA level of NQO-1 and HO-1 were measure by qRT-PCR.Results:(1)Compared with the TBI group,administration of SIN significantly improved neurological function(P<0.01),alleviated cerebral edema(P<0.001),and inhibited neuronal apoptosis(P<0.001).(2)SIN markedly mitigated TBI-induced oxidative stress,which is represented by the reduced level of MDA and the enhanced activity of GPx and SOD.(3)The translocation of Nrf2 from the cytoplasm to the nucleus after TBI,and SIN further promoted this translocation,subsequently increased the expression of downstream factors at both mRNA and protein levels.Conclusion:Our data demonstrate that SIN treatment 30 min after TBI ameliorates secondary brain injury via improving neurologic function,reducing brain edema,combating oxidative stress and attenuating neuronal apoptosis.These effects might correlate with translocation of Nrf2 from the cytoplasm to the nucleus and subsequent activation of downstream proteins.