Research On Anti-fatigue Effect And Mechanism Of Sea Cucumber Peptide

Fatigue affects the quality life and mental state of modern people.It is a hot research spot to develop nutritional supplements with effective anti-fatigue function.Sea cucumber peptides have strong anti-fatigue effect,but the anti-fatigue mechanism is still unclear.In this study,the anti-fatigue mechanism of sea cucumber peptides was explored from the aspects of anti-oxidative stress,promoting lipocatabolism,promoting gluconeogenesis,and enhancing mitochondrial plasticity.In addition,bioinformatics method was used to analyze the biological process and core targets of anti-fatigue effect of sea cucumber peptides,and experimental verification was carried out.The main research results are as follows:1.Research on the mechanism of sea cucumber peptides against animal fatigue.The molecular weight of the enzymatic hydrolysis product of sea cucumber body wall is less than1998 Da(92.59%),and the main component of the hydrolysis product is sea cucumber peptides(71.32%).64 healthy male mice aged 7 weeks were randomly divided into control group,low sea cucumber peptides dose group(0.7 mg/g·d),medium sea cucumber peptides dose group(1.4 mg/g·d)and high sea cucumber peptides dose group(2.8 mg/g·d).On the 21st day of feeding,the forelimb grip strength test was conducted.The fatigue rotating rod test was conducted on the 25th day.The isochronous swimming test was conducted on the 30th day to construct a sports fatigue model.After swimming,the blood and tissue samples were taken for analysis.The main results were as follows:the sea cucumber peptides increased the feed intake of mice,but there was no significant difference in body weight between groups(p>0.05).Compared with the control group,the forelimb grip strength in the H-Scp group and the time of fatigue rotating rod in each sea cucumber peptides group were significantly increased(p<0.05),and both showed a dose-effect relationship.Compared with the control group,the sea cucumber peptides groups improved the levels of oxidative stress and inflammatory factors,the levels of blood glucose and tissue glycogen,and the contents of fatigue metabolic markers after isochronous swimming.It was further found that the sea cucumber peptides enhanced the catabolism of fatty acids in liver,heart and skeletal muscle.The expressions of PPAR?,PGC1?,CPT1,LPL and PDK4 in the liver were up-regulated,the expressions of PPAR?and CPT1 in the heart were up-regulated,and the expressions of PPAR?and CPT1 in the skeletal muscle were up-regulated.The sea cucumber peptides increased the gluconeogenesis in liver,and the expressions of G6Pase and PEPCK were up-regulated.In addition,the sea cucumber peptides increased mitochondrial copy number(mt DNA)levels in the heart and skeletal muscle.After isochronic swimming,the activity of ATPase and the expressions of COXI,PGC1?,NRF2,NRF1,TFAM,AMPK and SIRT1 in heart and skeletal muscle of the sea cucumber peptides groups were higher than those in the control group.The results showed that the sea cucumber peptides maintained energy homeostasis during exercise and improved mitochondrial ATP production capacity.2.Bioinformatics study of the anti-fatigue sea cucumber peptides.Firstly,the sea cucumber peptides with the strongest DPPH·scavenging ability were identified by UPLC-Q-TOF-MS/MS,and the peptides were predicted their biological target.Secondly,the antifatigue target set was constructed by transcriptome.Finally,the structure-activity targets of the sea cucumber peptides and anti-fatigue targets were analyzed by bioinformatics method.(1)The134 sea cucumber peptides sequences identified by chromatographic analysis were predicted by SEA and Target Net servers,and the 488 effective targets were obtained.(2)Through the differential gene analysis of the transcriptomes GSE59088,GSE9013,and GSE155271 related to endurance training,a target data set containing 1762 genes was constructed.(3)After the combination,the protein interaction network of the potential anti-fatigue targets of sea cucumber peptides was obtained.The main biological processes include"regulation of inflammatory response","regulation of small molecule metabolism","response to hypoxia and different oxygen levels","regulation of calcium ion homeostasis",etc.;"Regulate fat metabolism","response to reactive oxygen species","inflammation regulation"and"calcium ion homeostasis"are consistent with the results of animal experiments.The analysis also showed that NF-?B/STAT3/RELA/SRC,together act on the production of IL-6 and TNF-?.Further analysis of the protein interaction network of the sea cucumber peptides to interfere with fatigue,the key gene ESR1 of the core module may be related to the effect of the sea cucumber peptides in increasing feed intake.3.Synthesize sea cucumber peptides for cell verification test.According to the results of the 134 sea cucumber peptide functional analysis,we chose to synthesize sea cucumber peptides P1(G(Hyp)LQADY),P2(FD(Hyp)GA),P3(LPGSDDF)and P4(APGLTY)to intervene in the C2C12 oxidative damage model induced by H₂O₂,verifying the intervention effect on the redox system and inflammation-related targets.The results showed that compared with model injury group,the P1 and P2 peptides at 50?g/m L significantly increased cell viability,antioxidant capacity,significantly reduced the content and expression of IL-6 and TNF-?content,reduced the phosphorylation level of NF-?B and increased the phosphorylation level of STAT3.The results indicate that the P1 and P2 peptides have anti-inflammatory effects,which are in line with the function prediction results of biological information.In summary,based on the results of animal,cell and bioinformatics analysis experiments in this study,sea cucumber peptides play anti-fatigue effects by slowing down glycogen consumption,accelerating fat catabolism,increasing mitochondrial copy number and ATP production capacity,and reducing oxidative stress and inflammatory factor levels.