Study On The Mechanism Of Epigallocatechin-3-gallate On Alleviating Acrylamide-induced Liver And Nerve Damage In Rats

Acrylamide(ACR)is a raw material for the production of polyacrylamide.ACR and its polymers are widely used in industry,and they are utilized in paper production,soil conditioning and water treatment.In addition to residual in environmental,ACR is one of the products formed via the Maillard reaction during food processing.ACR induces liver and nervous system damage as a result of its hepatotoxicity and neurotoxicity.ACR exposure is unavoidable,therefore,it is great significant to explore how to attenuate ACR-induced damage.Green tea is a traditional beverage,and epigallocatechin-3-gallate(EGCG)is the most abundant catechin in green tea with many bioactive properties.As a natural antioxidant,EGCG has protective effects on liver and nervous system.However,the effect and mechanism of EGCG on ACR-induced liver and nerve damage are not clear.In this paper,PC 12 cells and Sprague-Dawley rats were used to establish vitro and vivo experimental models respectively to research the inhibitory effect of EGCG on ACR-induced hepatotoxicity and neurotoxicity,and to delve into the mechanism from several aspects,such as oxidative stress,apoptosis and inflammation.The main results of this study are as follows:(1)EGCG inhibits ACR-induced decrease in rat cell viability and acetylcholinesterase(AChE)activity.EGCG attenuates cell morphology damage against ACR by observing and comparing under microscope.Using flow cytometry to analysis cell apoptotic state,it is found that EGCG significantly reduces percentage of apoptotic and necrotic cells,increases mitochondrial membrane potential and reduces intracellular calcium level,cytochrome c release,caspase 3 activity and expression of Bax mRNA,which means that EGCG inhibits ACR-induced apoptosis through protecting mitochondrial function.EGCG reduces reactive oxygen species(ROS)level while enhancing superoxide dismutase(SOD)activity and glutathione(GSH)content,thereby attenuates oxidative stress induced by ACR in cells,including decreases lipid peroxidation.The inhibitory effect of EGCG on ACR-induced cytotoxicity in rat cells may be related to the inhibition of ACR-induced oxidative stress in cells.(2)EGCG inhibits ACR-induced increase in aspartate transaminase and alanine transaminase activities in both serum and liver of rate.Using terminal uridine nick-end labeling(TUNEL)assay,it is found that EGCG decreases the rate of TUNEL-positive cells in liver,which means that EGCG inhibits ACR-induced apoptosis.From the perspective of inflammatory injury,EGCG reduces the expression of inducible nitric oxide synthase(iNOS)and cyclooxygenase-2(COX-2),which indicates that EGCG inhibits ACR-induced inflammatory respond in liver.EGCG decreases lactate dehydrogenase activity and the expression of cytochrome P450 1A2,which means that EGCG has inhibitory effect on ACR-induced metabolic dysfunction in liver.EGCG attenuates oxidative stress induced by ACR in liver through reducing ROS level,increasing activities of antioxidant enzymes,such as SOD,catalase(CAT),glutathione peroxidase(GSH-Px),and lipid peroxidation and DNA oxidative damage are thus decreased.The inhibitory effect of EGCG on ACR-induced liver damage may be related to the inhibition of ACR-induced oxidative stress in liver.(3)ACR induces characteristic neurotoxic symptoms in rats,such as loss weight,exhibits external rotation of hind limbs and has dull corneas,while EGCG maintains weight and clinical state of rats.EGCG increases the rate of Nissl-positive cells and AChE activity,which means that EGCG has inhibitory effect on ACR-induced cerebral cortex damage.EGCG is demonstrated to decrease the rate of glial fibrillary acidic protein-positive cells using immunohistochemistry,which means that EGCG inhibits ACR-induced astrogliosis.Results from TUNEL and western blotting indicate that EGCG attenuates ACR-induced apoptosis in cerebral cortex by regulating the expression of Bax,Bcl-2,cytochrome c and caspase 3.From the perspective of inflammatory injury,EGCG inhibits ACR-induced inflammatory injury by reducing the expression of tumor necrosis factor-a,iNOS and COX-2.EGCG decreases the rate of senescence-associated ?-galactosidase-positive cells,denonstrating that EGCG has inhibitory effect on senescence induced by ACR in cerebral cortex.EGCG significantly increases the rate of brain-derived neurotrophic factor-positive cells and decreases the rate of nestin-positive cells,which indicates that EGCG promotes regeneration in cerebral cortex.EGCG also inhibits ACR-induced oxidative damage of lipid,protein and DNA through reducing ROS level and increasing activities of SOD,CAT and GSH-Px and GSH content in cerebral cortex.The inhibitory effect of EGCG on ACR-induced nerve damage may be related to the inhibition of ACR-induced oxidative stress in cerebral cortex.