Fermentation Broth Of Serpula Similis Protect Cells From Senescence Via AMPK-dependent Autophagy Restoration

Objective:To investigate the effect of Fermentation broth of Serpula similis（FBS）on autophagy of oxidative stress-induced senescent cells and explored whether FBS could inhibit cell senescence and its mechanism.Methods:Senescence induced by hydrogen peroxide（H₂O₂）in NIH3T3 fibroblasts to investigate the anti-senescent effect of the Fermentation broth of Serpula similis.Real-time PCR and Western blot were used to detect the m RNA and protein expression,respectively.Cell Senescence was examined by SA-gal staining,cell viability was detected by CCK-8,SAHF phenomenon in nucleus was detected by DAPI staining,and autophagy was detected by immunofluorescence.Results:1.H₂O₂ induced senescence in fibroblastsCompared with the Ctrl,the percentage of SA-β-Gal positive cells and SAHF positive cells were significantly increased;the expression levels of two senescence associated genes（IL6 and p21）and proteins（p53 and p16）were increased.2.Fermentation broth of Serpula similis（FBS）inhibits H₂O₂-induced NIH3T3 cells senescenceCompared with the Vehicle,the cytotoxicity did not change significantly with the increase of FBS,the percentage of SA-β-Gal positive cells were significantly decreased with the concentration went up;the expressions of three senescence associated genes,IL6,p53 and p21 also obviously decreased in senescent cells.3.Fermentation broth of Serpula similis（FBS）prevented senescence by enhancing the H₂O₂-impaired autophagy in senescent cellsCompared with the Vehicle,the expression levels of two autophagy-related proteins（p62and LC3）were decreased with FBS treatment.meanwhile,increased GFP luorescence.This showed that FBS enhanced autophagic degradation.4.Fermentation broth of Serpula similis（FBS）ameliorate H₂O₂-induced autophagic dysfunction in NIH3T3 cells by activating AMPKCompared with the Vehicle,the protein levels of p62 and LC3 were decreased with FBS treatment,but protein the level p-AMPK significantly elevated and weakened GFP fluorescence,indicating an increase in cell activity.The expression of autophagy-associated Protein P62 and LC3 was consistent with that of Vehicle when AMPK inhibitor was added,that is,autophagy was inhibited,indicating that the anti-aging effect of FBS was AMPK dependent.Conclusions:The Cell Senescence induced by oxidative stress can be inhibited by fermentation of dry rot fungi,In this study,we found that H₂O₂ induced senescence and decreased autophagic flux in NIH3T3s.Fermentation broth of Serpula similis（FBS）inhibited in H₂O₂-induced senescent cells and this anti-aging effect is related to activate AMPK and restore AMPK-dependent autophagic activity.The anti-aging effect of FBS expands the scope of the research on the mechanism of aging,and provides a new basis and perspective for the research and development of new anti-aging drugs.