Exploration of mechanism of total triterpenoids from fruits of Chaenomeles speciosa against senescent GES-1 cells induced by D-galactose based on Gln/GLS1/α-KG metabolic axis and mitochondrial apoptosis signaling pathway
Total triterpenoids from the fruits of Chaenomeles speciosa(TCS)are active components in the prevention and treatment of gastric mucosal damage,which have potential anti-aging effects.However,it is still unclear whether TCS can improve gastric aging,especially its molecular mechanism against gastric aging.On this basis,this study explored the effect and mechanism of TCS on senes-cent GES-1 cells induced by D-galactose(D-gal)to provide scientific data for the clinical use of TCS to prevent gastric aging.GES-1 cells cultured in vitro and those transfected with overexpression GLS1(GLS1-OE)plasmid of glutaminase 1(GLS1)were induced to aging by D-gal,and then TCS and or GLS1 inhibitor bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide(BPTES)were given.Cell survival rate,positive rate ofβ-galactosidase(SA-β-gal)staining,mitochondrial membrane potential(MMP),and apopto-sis were investigated.GLS1 activity,levels of glutamine(Gln),glutamate(Glu),α-ketoglutarate(α-KG),urea,and ammonia in supernatant and cells were detected by enzyme-linked immunosorbent assay(ELISA)and colorimetric methods.The mRNA and pro-tein expressions of GLS1 and the related genes of the mitochondrial apoptosis signaling pathway were measured by real-time fluorescence quantitative PCR and Western blot.The results manifested that compared with the D-gal model group and GLS1-OE D-gal model group,TCS significantly decreased the SA-β-gal staining positive cell rate and MMP of D-gal-induced senescent GES-1 cells and GLS1-OE senescent GES-1 cells,inhibited the survival of senescent cells,and promoted their apoptosis(P＜0.01).It decreased the activity of GLS1 and the content of Gin,Glu,α-KG,urea,and ammonia in supernatant and cell(P＜0.01),reduced the concentration of cy-tochrome C(Cyto C)in mitochondria and the mRNA and protein expressions of GLS1 and proliferating nuclear antigen in cells(P＜0.01).The mRNA expression of Bcl-2 and Bcl-xl,the protein expression of pro-caspase-9 and pro-caspase-3,and the ratio of Bcl-2/Bax and Bcl-xl/Bad in cells were decreased(P＜0.01).Cyto C concentration in the cytoplasm,the mRNA expressions of Bax,Bad,apoptosis protease activating factor 1(Apaf-1),and protein expressions of cleaved-caspase-9,cleaved-caspase-3,cleaved-PARP-1 were increased(P＜0.01).The aforementioned results indicate that TCS can counteract the senescent GES-1 cells induced by D-gal,and its mechanism may be closely related to suppressing the Gln/GLS1/α-KG metabolic axis,activating the mitochondrial apoptosis pathway,and thereby accelerating the apoptosis of the senescent cells and eliminating senescent cells.
total triterpenoids from the fruits of Chaenomeles speciosaGES-1 cellsD-galactosesenescenceglutamine/glutami-nase 1/α-ketoglutarate metabolic axismitochondrial apoptosis signaling pathway
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