| Aging and related chronic diseases have always attracted people's attention.Although aging is an irreversible process,people can delay aging and prevent agerelated chronic diseases through dietary intervention.Increased consumption of fruits and vegetables has been associated with reduced risk of age-related functional decline and chronic diseases,primarily attributed to their phytochemicals.Blueberries and apples are rich in phytochemicals with a wide range of biological activities and health benefits.However,the anti-aging effects of the combination of blueberries and apples have not been reported.The objectives of this study were to explore the anti-aging and anti-oxidative stress activities of the combination of blueberry and apple peel extracts,and to investigate the underlying mechanisms of action,based on the Caenorhabditis elegans model.The main findings are as follows:(1)Blueberry and apple peel extracts exhibited strong antioxidant activities detected by Oxygen Radical Absorbance Capacity(ORAC),Peroxyl radical Scavenging Capacity(PSC)and Cellular Antioxidant Activity(CAA)methods.The antioxidant capacity was positively correlated with the phenolic content,and was closely related to the phytochemical composition.At the same time,the combination of blueberry and apple peel extracts could significantly inhibit the proliferation of Hep G2 cells,and regulate the expression levels of proliferation-related genes p53 and c-myc.When the inhibition rate was 95%,the combination index(CI)value was 0.96 ± 0.11,and the dose-reduction index(DRI)values for bluberry and apple peel extracts were 2.24 ± 0.17 and 1.92 ± 0.20-fold,indicating that the combination of blueberry and apple peel extracts had a synergistic effect in inhibiting the proliferation of Hep G2 cells.(2)The anti-aging activities of the combination of blueberry and apple peel extracts were conducted using the Caenorhabditis elegans aging model.The results showed that the combination of blueberry and apple peel extracts could synergistically extend the mean lifespan of C.elegans,reducing the accumulation of lipofuscin,and increase health span of animals including improving motility and enhancing resistance to heat stress and UV-B radiation in C.elegans;Meanwhile,treatment with the combination of blueberry and apple peel extracts could regulate the expression of antiaging related genes,including daf-2,age-1,akt-2,dod17,daf-16,sir-2.1,jnk-1 and hsp-16.2,and promote the migration of DAF-16 into the nucleus;In addition,administration with the combination of blueberry and apple peel extracts abolished the extension of the lifespan of daf-2(e1370),hsf-1(sy441),jnk-1(gk7),daf-16(mu86);daf-2(e1370)mutants,and inhibited the expression of daf-16 downstream gene,including sod-3,ctl-2 and dod17.In conclusion,the combination of blueberry and apple peel extracts could synergistically increase the lifespan,improve health span and enhance stress resistance via insulin/IGF signaling pathway and DAF-16 in C.elegans.(3)Based on the skn-1(zu135)mutant model,the combination of blueberry and apple peel extracts to improve the body's ability to resist oxidative stress was studied.The results showed that the combination of blueberry and apple peel extracts could synergistically ameliorate oxidative stress by improving antioxidant enzyme activities and enhancing resistance to paraquat,and regulate the expression levels of reactive oxygen species(ROS)and antioxidant related genes(sod-3,cat-1,ctl-1,skn-1,mev-1 and isp-1).At the same time,administration with the combination of blueberry and apple peel extracts abolished the extension of the lifespan of skn-1(zu135)mutants,and eliminated its improvement on ROS and its resistance to paraquat;In addition,supplementation with the combination of blueberry and apple peel extracts could promote the migration of SKN-1 into the nucleus,thereby regulating the expression of skn-1 downstream genes,including gcs-1,gst-4 and gst-7.In short,the combination of blueberry and apple peel extracts could synergistically improve oxidative stress in C.elegans via the SKN-1/Nrf2 pathway.(4)Used RNAi technology to establish a model of nematode mitochondrial dysfunction,and explored whether the treatment with the combination of blueberry and apple peel extracts improved the lifespan and oxidative stress of C.elegans through the regulation by mitochondrial function.Our results demonstrated that if the clk-1 genes were silenced,the combination of blueberry and apple peel extracts to extend the lifespan of nematodes was abolished.Meanwhile,treatment with the combination of blueberry and apple peel extracts could reduce the ROS content in clk-1(e2519)mutants,and regulate the expression level of clk-1 downstream genes(nuo-2,cco-1,cyc-1 and atp-3),thereby affecting the normal function of the mitochondrial respiratory chain,improving its oxidative stress ability,and regulating the aging process.In addition,the combination of blueberry and apple peel extracts could prolong the lifespan of RNAi(mev-1)C.elegans,and improve mitochondrial function,while there is no significant change for RNAi(isp-1)C.elegans.In short,the treatment with the combination of blueberry and apple peel extracts improved the oxidative stress and extend the lifespan of nematodes,which were regulated by mitochondrial function.In summary,the treatment with the combination of blueberry and apple peel extracts could synergistically extend the lifespan via insulin signaling pathway and DAF-16 transcription factor in C.elegans,and synergistically improve oxidative stress in C.elegans via the SKN-1/Nrf2 pathway.The extension of nematode lifespan and its resistance to oxidative stress were regulated through mitochondrial function.This study provided the scientific basis to explore the health benefits,and deep processing utilization of blueberries and apples,and offered theoretical support for the prevention and treatment of aging related diseases. |
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