Effects Of Moutan Seed Coat Extracts And Suffruticosol B On Cognitive Impairment In Mice And The Mechanism Based On Omics Technology

Objective:Alzheimer’s disease(AD)is a neurodegenerative disease characterized by learning and memory dysfunction.Our laboratory has previously found that stilbenoids,abundant in the seed coat extract of Paeonia suffruticosa(PSCE),could improve cognitive impairment in vivo and in vitro.Therefore,in this study,PSCE and suffruticosol B(SB),one of its main components,as the research objects,were taken as the research object to explore its improving effect on cognitive impairment and its mechanism,in order to provide new ideas for the research and development of AD prevention and treatment drugs.Method:C57BL/6 mice were injected intraperitoneally with scopolamine(1.5 mg/kg/d)to induce cognitive impairment in mice and orally administered with PSCE(30 and 120 mg/kg/d)or SB(15 and 60 mg/kg/d)for 22 days.The learning and memory ability of mice were evaluated by behavioral tests and biochemical indexes.Then KM mice were injected subcutaneously with D-gal(120 mg/kg/d)for 110 days to induce cognitive impairment in mice and orally administered with PSCE(30,60 and 120 mg/kg/d)or SB(10,30 and 90 mg/kg/d)for 70 days.The learning and memory ability of mice were evaluated by behavioral tests,brain tissue staining and biochemical indexes.Transcriptome sequencing was used to analyze the effects of drugs on differentially expressed genes and signaling pathways in D-gal-induced hippocampal tissue of mice.Finally,we collected mice feces and analyzed the regulation of drugs on intestinal flora by microbiome technology based on 16S rRNA amplicon sequencing,and analyzed the changes of drugs on metabolites in mice feces by UPLC-Q-TOF/MS metabolomics technology.In addition,UPLC technique was used to determine the change of PSCE composition after incubation with intestinal flora in vitro.Results:1 PSCE and SB could improve scopolamine and D-gal-induced cognitive dysfunction in mice,respectively.In the scopolamine induced cognitive impairment model,high dose group PSCE and SB could significantly increase the time of exploring new objects in NOT test,and significantly shorten the escape latency in MWM test,while only part of the low dose group had significant difference.The results of biochemical indexes showed that PSCE and SB could improve scopolamine-induced cognitive impairment by improving cholinergic system(ACh,ChAT and AChE),anti-inflammatory(IL-6,TNF-α and IL-1β)and anti-oxidation(CAT,SOD and GSH)levels.In D-gal-induced cognitive impairment model,medium and high doses of PSCE and SB could significantly increase the exploration time of new objects in the NOR test,prolong the incubation period of the mice and reduce the number of errors in the PAT,increase the spontaneous alternation rate of mice in Y maze test,and shorten the search latency of mice in MWM test,but the low dose only partially showed significant difference.The results of HE staining and Nissl staining showed that PSCE and SB could improve the pathological damage of the hippocampus and cerebral cortex of mice to varying degrees,such as the reduction of cell number,loose arrangement of cells,pyknosis of cells,and deep color.The results of biochemical indexes showed that PSCE and SB could significantly increase the anti-inflammatory(IL-6,TNF-α and IL-1β)and antioxidant(CAT,SOD and GSH)levels in serum and cerebral cortex of mice.2 PSCE and SB could effectively improve D-gal-induced oxidative stress and neuroinflammation in mouse brain tissue,regulate the corresponding signaling pathways,and thus improve cognitive impairment by reshaping mouse intestinal flora and regulating the level of microbial metabolites.Based on transcriptome sequencing technology,PSCE and SB could improve D-gal induced brain tissue damage in mice by regulating oxidative stress and neuroinflammatory pathways.KEGG pathway enrichment analysis showed that differentially expressed genes were mainly involved in neurodegenerative disease,oxidative stress,neuroinflammation,cell proliferation and apoptosis signaling pathways.Pathways related to oxidative stress include FoxO signaling pathway,and pathways related to neuroinflammation include Fc epsilon RI signaling pathway,MAPK signaling pathway and Hippo signaling pathway.Studies based on microbiome techniques of 16S rRNA amplification sequencing showed that PSCE and SB could regulate D-gal-induced intestinal flora disturbances in mice,and regulate the flora abundance associated with short-chain fatty acids and lipopolysaccharides(e.g.Firmicutes,Bacteroidetes,Muribaculaceae and Lactobacillus).Reduce the symbiosis of inflammation-related flora(such as Lachnoclostridium)and regulate the metabolic pathway of intestinal flora in mice.The results of RDA analysis showed that the bacteria in PSCE group and SB group were negatively correlated with inflammatory factors(IL-1β,IL-6,TNF-α),and positively correlated with antioxidants(SOD,CAT,GSH).Based on UPLC-Q-TOF/MS metabonomics,PSCE and SB could improve the disturbance of endogenous metabolites induced by D-gal in mice,down-regulated metabolites such as sphingomyelin and prostaglandin,and up-regulated hyaluronic acid,lysophosphatidic acid and 6-hydroxymelatonin metabolites.Histidine metabolism,fatty acid metabolism and bile acid biosynthesis were regulated to improve intestinal metabolic disorders.The stilbenoids in PSCE could undergo biotransformation under in vitro incubation conditions,and the conversion rate of the main active ingredient SB within 48 h was 17.38%.Conclusion:PSCE and one of its main active components,SB,could improve scopolamine and D-gal-induced cognitive dysfunction,and regulate the metabolism of short-chain fatty acids,lipopolysaccharides and lipids by regulating the abundance and symbiosis of intestinal flora and effectively improve D-gal-induced oxidative stress and neuroinflammation in mouse brain tissue,thus improving cognitive impairment.The involved oxidative and inflammatory signaling pathways might be FoxO signaling pathway,Fc epsilon RI signaling pathway,MAPK signaling pathway and Hippo signaling pathway.This study laid a solid foundation for the improvement of cognitive impairment by PSCE and one of its main active components,suffruticosol B,and preliminarily revealed the complex pharmacodynamic mechanism,which indicated that the stilbenoids in moutan seed coat had the important potential to be developed into AD drugs.