Protective Effect And Mechanism Of Rosmarinik Acid On Acute Liver Injury Induced By APAP In Mice

There are many predisposing factors for acute liver injury,including drugs,chemistry,microorganisms,etc.,which can lead to acute liver failure or even life-threatening in severe cases.Acetaminophen(APAP)is one of the most common forms of drug-induced acute liver damage,which can occur in large doses or over a long period of time,and excessive inflammation and oxidative stress levels are considered to be important factors.Rosmarinic acid(RA)is a natural polyphenolic drug compound with various pharmacological activities such as antiinflammatory,anti-oxidant,anti-microbial,anti-depressant,etc.Several studies have suggested that Rosmarinic acid plays a protective role in different types of liver injury models.Therefore,this study took RA as the main research object,established a mouse model of acute liver injury with high dose APAP to explore the protective effect and mechanism of RA on acute liver injury.Methods:(1)Establishment of APAP-induced acute liver injury model:Two doses of 300 mg/kg and 400 mg/kg were selected and APAP was given by intraperitoneal injection.Samples were taken at 3 h,6 h,and 8 h respectively,and serum liver injury markers ALT,AST level and the degree of liver tissue pathological section damage were used as the basis to determine the appropriate APAP concentration and duration of action,and established an acute liver injury model.(2)The protective effect and mechanism of RA on APAP-induced acute liver injury in mice: After successfully established on the basis of acute liver injury,the mice were administed RA pretreatment at three does of 10,20,40mg/kg,the levels of serum cytokines IL-1β,IL-6,IL-10,and TNF-α in mice were detected by ELISA kits.The antioxidant capacity of mouse liver GSH-Px,GSH,CAT,SOD,T-AOC activity and protein expression of lipid peroxidation product MDA and inflammation-related enzyme MPO were detected by ELISA or biochemical kits,q RT-PCR detected cytokines IL-1β,IL-6,IL-10,i NOS and enzymes MPO,GSH-Px,CAT,SOD and APAP in liver tissue metabolic enzyme CYP2E1 m RNA transcription level,while quantifying the m RNA transcription levels of apoptosis-related genes Bax,Bcl-2 and antioxidant genes GCLC,GCLM,and immunohistochemical technique to detect the antioxidant protection genes Nrf2 and HO-1 in liver tissue expression and Western blot to detect the protein levels of GCLC and GCLM.The results showed:(1)APAP concentration at 400 mg/kg and the action time of 6 h significantly increased the levels of ALT and AST in the mouse serum.Histopathological observation revealed that the degree of liver damage increased with the increase of APAP action time.(2)Antiinflammatory ability:RA significantly reduced the levels of inflammatory factors IL-1β,IL-6,IL-10,TNF-α and inflammasy-related enzyme MPO,as well as the m RNA transcription levels of IL-1β,IL-6,IL-10,MPO and i NOS in mice with liver injury.(3)Anti-oxidative stress ability: RA improved the vitality of antioxidants GSH-Px,GSH,CAT,SOD and total antioxidant capacity T-AOC and the m RNA expression of GSH-Px,CAT and SOD in liver injury of mice,and reduced lipids peroxidation indicator MDA level.(4)Protection mechanism: RA can significantly increase the protein expression levels of Nrf2,HO-1,GCLC and GCLM,and increase the m RNA expression of GCLC and GCLM at the transcription level in liver injury of mice.RA can up-regulate the transcription level of anti-apoptotic protein Bcl-2 and down-regulate the transcription level of pro-apoptotic protein Bax.RA can down-regulated the transcription level of CYP2E1,which is the main metabolic enzyme of APAP in liver injury.To sum up,RA plays an anti-inflammatory,antioxidant and anti-apoptotic role in the body.Conclusion:(1)In this study,the dose of APAP 400 mg/kg were used for 6hours to successfully establish an APAP-induced acute liver injury model in mice.(2)RA had a good anti-inflammatory and antioxidant effect on acute liver injury induced by APAP in mice.(3)RA played an antioxidant role by regulating the gene expression of antioxidant Nrf2 and its downstream HO-1,GCLC and GCLM,and had a protective effect on liver injury by regulating the transcriptional level of CYP2E1 and apoptose-related proteins Bax and Bcl-2.