Study On The Improvement Effect Of Sodium Butyrate Combined With Berberine On Non-alcoholic Fatty Liver Disease

Sodium butyrate(NaB)is a soluble sodium salt of butyric acid,which can be metabolized as butyric acid in vivo after oral administration.It has a variety of biological activities such as improving glucose and lipid metabolism,weight loss,anti-inflammatory,antioxidant and tumor prevention.Berberine(BBR)is an isoquinoline alkaloid derived from natural products.This study aims to explore whether or not the combination of BBR and NaB can improve nonalcoholic fatty liver disease(NAFLD)better than the either agent alone,and to explore the preliminarily mechanisms of the combination.The study is divided into two parts:in vitro experiments and in vivo experiments.For in vitro experiments,the effects of NaB,BBR and NaB+BBR on the AMP-activated protein kinase(AMPK)signaling pathway and the expression levels of lipogenic and lipolytic genes were determined in human liver cell line HL-7702.The steatosis model of HL-7702 cells was established by induction of oleic acid(OA),and the effect of NaB,BBR and NaB+BBR on lipid accumulation was evaluated by Oil Red O(ORO)staining and intracellular triglyceride(TG)content determination.Meanwhile,the indexes of oxidative stress induced by OA were detected by kits.For in vivo experiments,C57BL/6J mice were given a high fat diet(HFD)for 7 weeks to establish a NAFLD model,while control mice were given a standard mouse maintenance diet(normal diet,ND).At the same time,mice were given normal saline(normal saline,NS),NaB(200 mg·kg-1),BBR(100 mg·kg-1),or NaB(200 mg.kg-1)+BBR(100 mg.kg-1)by gavage.Fatty acid components of the liver were detected by 1H-proton magnetic resonance spectroscopy(1H-MRS).Hematoxylin-eosin staining(H&E)and ORO staining were performed on liver tissue sections.Western blot and real-time fluorescent reverse transcription polymerase chain reaction(RT-PCR)were used to determine the levels of the target proteins or mRNAs,and the corresponding kits were used to detect the indexes related to oxidative stress level in the liver of mice.The in vitro experiments showed that BBR combined with NaB significantly activated the AMPK signaling pathway and reduced steatosis in HL-7702 cells,and they also significantly improved the degree of oxidative stress of HL-7702 cells after OA induction.The efficacies of combination were stronger as compared with BBR or NaB alone.For in vivo experiments,the results showed that as compared with ND group,HFD+NS group of mice had obvious fat accumulation in liver,accompanied with fatty acid composition abnormality and liver function damage.In liver of the model group of mice,the AMPK pathway was inhibited,the expression levels of lipogenic genes(FAS,SCD1)increased significantly,while those of lipolytic genes(CPT1A,ACOX1)decreased significantly.Meanwhile,the expression levels of liver proinflammatory factors(IL-1β,IL-6,TNFα)and oxidative stress indexes increased greatly in the model group.Combination of NaB and BBR significantly reduced lipid accumulation in the liver;meanwhile,they greatly activated the liver AMPK pathway,and alleviated liver inflammation and oxidative stress.The effcacies of the combination regimen were significantly superior to those of NaB or BBR alone.In conclusion,combination of NaB and BBR effectively improves hepatocellular steatosis or NAFLD,both in vitro and in vivo,and the efficacies are stronger than BBR or NaB alone.This combination regimen may have certain prospects for development and application in the future.