| Summary: | 博士 === 國立中興大學 === 食品暨應用生物科技學系 === 97 === Anthocyanins are natural occurring pigments in lots of vegetables and fruits. It has been suggested that anthocyanins possess multifunctional and beneficial activities such as antioxidaion, anti-inflammation, anti-cancer, and cardio-protection. The modern high fat and carbonydrate diet habits result in the perturbation of lipid metabolism nomeostasis. The glucotoxicity- and lipotoxicity-induced oxidative stress is found to be involved in the development of metabolic syndrome. Accumulation of oxidative damage may cause dysfunction of genes and/or cells relevant to lipid metabolism. Besides, oxidative stress influences health and disease. This might be of special relevance for ageing. Alzheimer''s disease (AD) is one of popular age-related dementia affecting hundred million people worldwide. The neurophathological hallmark of AD is an extracellular aggregation of amyloid β-protein (Aβ). Thus, one of the strategies to prevent AD is to inhibit Aβ-induced neurotoxicity. Mulberry (Morus atropurpurea) is a local fruit with the highest antioxidant capacity in Taiwan. It has pharmacological functions including antioxidation, anti-diabetes, and anti-cancer. Anthocyanins are the active compounds abundantly found in mulberry. Based upon the above research, we suggest that the decrease in oxidative stress and increase in antioxdiant capacity are beneficial for protecting against radical-related diseases. However, the regulatory effects on antioxidant enzymes expressions and the chemopreventive mechanisms of anthocyanins have not yet been verified. The effects of anthocyanins on the regulation of genes relevant to lipid metabolism are also unclear. Thus, the biological benefits of mulberry are worth further investigation and evaluation. The aim of the present study was: (1) to investigate the promotion effects of anthocyanins on antioxidant and detoxifying enzymes and the involved mechanism, (2) to evaluate the combination of anthocyanin and the lipid-lowering drug, trolitazone, on the expression of genes relevant to lipid metabolism in hepatocytes upon H2O2 stimulaiton, (3) the neuroprotective effects of anthocyanins on Aβ-induced neurotoxicity (4) to investigate the relation between ageing and antioxidant defense systems and the oxidant status in senescence-accelarated mice (SAMP8) with anthocyanin-rich mulberry extracts.
We found that (1) rat liver Clone 9 cells treated with anthocyanins had positive effects on elevating their antioxidant capacity, including activated expression of glutathione-related enzymes (glutathione reductase, glutathione peroxidase, and glutathione S-transferase) and recruited GSH content. The activity of NAD(P)H:quinone oxidoreductase was also promoted upon anthocyanins stimulation. Among the ten anthocyanins, cyanidin, cyanidin-3-O-glucoside (kuromanin), delphinidin, and malvidin showed positive representation. With the technologies of gene cloning, silencing, and protein inhibition, we demonstrated that extracellular signal-regulated kinase (ERK1/2) and c-Jun N-terminal kinase (JNK) played critical roles in the regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) activation, which resulted in the translocation of Nrf2 from cytosol into nuclei and combined with antioxidant response element (ARE). The expression of antioxidant and detoxifying enzymes were then promoted.
(2) Hyperlipidemia-lowering drug, i.e. troglitazone, is the ligand of peroxisome proliferated-activated receptor (PPAR), which regulates genes relevant to lipid metabolism. We found that treatment with H2O2 caused increasing in ROS production and apoptotic cell death in Clone 9 cells. Even the dose of troglitazone and anthocyanin were reduced to half, co-treated cells with trolitazone and anthocyanin still showed potent PPARγ and Nrf2 activation ability. H2O2-mediated down-regulation in genes PPARγ, RXR, acyl-CoA oxidase, and PMP70 were ameliorated by the combined treatment of troglitazone and anthocyanin. The data suggested that the synergistic effects of anthocyanin aglycone, its ability to activate the PPAR-regulated genes expression by PPAR agonist, and its potent characteristics to prevent H2O2-induced cytoxicity. The results implied that anthocyanins had a potential role in the prevention of oxidative stress-involved metabolic syndrome.
(3) Neuro-2A neuronal cells were treated with Aβ to investigate the neuroprotective effects of anthocyanins. Aβcombination (Aβ 25-35 and Aβ 1-40) induced a decrease in cell viability through elevating intracellar ROS formation and disrupting Ca2+ homeostasis, and then resulted in cell cycle arrested at S and G2/M phases. The mRNA expressions of LXRα, ApoE, ApoE transporter ABCA1, and seladin-1, a key regulator in cellular response to oxidative stress, were all significantly down-regulated by Aβ stimulation. Furthermore, Bcl-2, which maintains the integrity of mitochondria, was also decreased upon Aβ treatment. Besides, β-secretase, a critical enzyme involved in Aβ formaiotn, was promoted in the Aβ-treated cells. For the duration of neurotoxicity induced by Aβ, we added anthocyanins (malvidin or malvidin 3-O-glucoside) to disrupt the disservice-induced by Aβ. The data suggested that anthocyanins had beneficial effects against Aβ-induced neurotoxicity through the blocking of ROS formation, preserving Ca2+ homeostasis, and preventing the Aβ-mediated perturbation in genes involved in Aβ metabolism and cellular defence. Therefore, anthocyanins might show a potential role in prevention of AD.
(4) The data revealed that oxidant status in the livers of aged SD rats were worse than those of young and middle-aged rats. The lipid and protein oxidation levels were more severe and the antioxidant enzymes activities were relatively lower in aged-rats. As compared with young rats, the expressions of hepatic ERK1/2 and Nrf2 were lowered while those of p38 were increased. The results suggested that accumulation of oxidative stress was accompaning with age. To investigate the effects of anthocyanins rich-mulberry extracts on the cognitive promotion and antioxidant activity in senescence-accelerated prone mice (SAMP8), we fed SAMP8 mice with mulberry extracts (500 mg/kg bw) for 12 weeks. The data implied that dietary supplementation with mulberry extracts increased antioxidant enzyme expression and reduced oxidative damage. The Aβ deposition in hippocampus was also inhibited upon mulberry treatment. Furthermore, the JNK/p38 levels were decreased and ERK1/2 and Nrf2 levels were elevated upon mulberry consumption. Overall, treatment with mulberry might be advantageous to the induction of antioxidant defense system and to the improvement of memory deterioration in ageing animals.
In this study, we demonstrated that chemopreventive effects of anthocyanins against oxidative stress-induced cytotoxicity were mediated through the Nrf2-ARE pathway to promote antioxidant enzymes expression. Besides, the combined treatment of anthocyanin and hyperlipidemia-lowering drug ameliorated the oxidative stress-related perturbation of genes involved in lipid metabolism and decreased the dosage of drug. While, oxidative damage accumulated with age, dietary supplementation with mulberry extracts promoted antioxidant capacity and improved the cognitive ability by inhibiting the aggregation of Aβ. Finally, we found that anthocyanins had neuroprotective effects aginst Aβ-induced radical burst and disintegration of Ca2+ homeostasis in neural cells. The neuroprotective effects of anthocaynins from the Aβ-mediated perturbation in genes involved in Aβ metabolism and cellular defence were remarkable. Therefore, anthocyanins might show a potential role in prevention of AD. We suggested that anthocyanins had protective effects on oxidative stress-induced damage, and mulberry possessed potential benefits against ageing-related dysfunction in antioxidant defence system and dementia. This study would contribute to the development as well as the application of functional phytochemicals in mulberry.
|
|---|
