Nobiletin Inhibits Hypoxia-Induced Placental Damage via Modulating P53 Signaling Pathway.

In this study, we aimed to evaluate the effect of Nobiletin (NOB) on the placenta of Sprague-Dawley (SD) rats that had undergone reduced uterine perfusion pressure (RUPP) surgery and to evaluate the safety of NOB intervention during pregnancy. The results showed that NOB alleviated placental hypoxia, attenuated placental cell apoptosis, and inhibited placental damage in RUPP rats. No side effect of NOB intervention during pregnancy was observed. BeWo cell lines with P53 knockdown were then constructed using lentiviral transfection, and the P53 signaling pathway was found to be essential for NOB to reduce hypoxia-induced apoptosis of the BeWo cell lines. In summary, NOB attenuated hypoxia-induced placental damage by regulating the P53 signaling pathway, and those findings may contribute some insights into the role of NOB in placental development and the prevention of placental-related diseases.

The effects of high-density lipoprotein and oxidized high-density lipoprotein on forskolin-induced syncytialization of BeWo cells.

INTRODUCTION: The negative relationship between maternal high-density lipoprotein-cholesterol (HDL-c) level during pregnancy and infant birth weight has been found. Syncytialization (differentiation and fusion) of trophoblast cells is important to fetal development. HDL has an antioxidant effect, and has been proved to protect trophoblast functions including hormone secretion and invasion. However, HDL is susceptible to oxidation, and high concentrations of HDL impair cell growth and oxidized HDL (oxHDL) inhibits cell proliferation and migration. Moreover, the effects of HDL and oxHDL on trophoblast syncytialization have not been characterized. The aim of this study was to investigate the effects of HDL and oxHDL on trophoblast syncytialization. METHODS: Human choriocarcinoma trophoblasts (BeWo cells) were treated with human HDL or oxHDL and then induced to differentiate by forskolin in syncytialization assays. Expression levels of mRNAs and proteins regulating syncytialization were detected by real-time PCR and western blotting, respectively. RESULTS: Treatments of HDL at high concentrations reduced human chorionic gonadotropin (hCG) secretion, placental alkaline phosphatase activity and fusion rates, and decreased the expressions of GCM1 and ERVW-1 mRNA as well as phospho-MAPK1/3 (p-MAPK1/3) and total MAPK1/3 protein in the forskolin-induced syncytialization of BeWo cells. Furthermore, treatment of oxHDL (20 µg/ml) decreased hCG secretion, but increased the expression of p-MAPK1/3 protein. DISCUSSION: These data suggested that both HDL at high concentrations and oxHDL inhibited BeWo cells syncytialization, and might be harmful to placental and fetal development.

Elaidic acid induces cell apoptosis through induction of ROS accumulation and endoplasmic reticulum stress in SH­SY5Y cells.

Elaidic acid, which is a major trans fatty acid, has been reported to be involved in neurotoxicity; however, the underlying molecular mechanisms underlying its neurotoxic effects remain largely unknown. Therefore, the present study aimed to investigate the potential mechanisms underlying elaidic acid­induced neuronal damage in vitro. The SH­SY5Y neuroblastoma cell line was used as a model in the present study. Following treatment of cells with various concentrations of elaidic acid or with vehicle for 24 h, cell viability was measured using the MTT assay. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) release were measured using flow cytometry. Cell apoptosis was measured by Annexin V­fluorescein isothiocyanate/propidium iodide double staining, and cellular redox status was determined using ELISA analysis. Furthermore, western blotting was used to detect the protein expression levels of factors associated with oxidative damage and components of the endoplasmic reticulum (ER) stress/unfolded protein response (UPR) signaling pathways. The results demonstrated that elaidic acid treatment inhibited cell viability, elevated cell apoptosis and resulted in a loss of MMP. In addition, elaidic acid induced marked alterations in cellular redox status. Treatment with high doses of elaidic acid treatment also enhanced the release of ROS, and upregulated lipid peroxide and malondialdehyde levels; however, it reduced superoxide dismutase and glutathione peroxidase activities. Furthermore, elaidic acid resulted in upregulation of nuclear factor erythroid 2­related factor 2 and downregulation of heme oxygenase 1, which are two key antioxidative factors. Elaidic acid treatment also induced or inhibited the expression of numerous ER stress/UPR­associated molecules. It induced glucose­regulated protein 78 (GRP78) expression, whereas the expression levels of activating transcription factor 4 (ATF4) and CCAAT/enhancer­binding protein homologous protein (CHOP) were upregulated and then downregulated following treatment with various doses of elaidic acid. These results indicated that elaidic acid inhibited SH­SY5Y cell growth and induced apoptosis by enhancing oxidative stress and activating the ER stress/UPR signaling pathway and the GRP78/ATF4/CHOP pathway.

Modulation of cholesterol transport by maternal hypercholesterolemia in human full-term placenta.

The significance of maternal cholesterol transporting to the fetus under normal as well as pathological circumstances is less understood. The objective of this study was to observe the effects of maternal hypercholesterolemia on placental cholesterol transportation. Human full-time placenta, maternal and venous cord blood were sampled at delivery from the pregnant women with serum total cholesterol (TC) concentrations at third trimester higher than 7.25 mM (n = 19) and the pregnant women with normal TC concentrations (n = 19). Serum lipids and expression of genes related to cholesterol transportation were measured by western blot or real-time PCR. The results indicated that serum TC, high density lipoprotein cholesterol (HDL-C), and low density lipoprotein cholesterol (LDL-C) levels were significantly increased, in pregnancies, but decreased in cord blood in hypercholesterolemic group compared to the matched control group. All the subjects were no-drinking, non-smoker, and gestational disease free. The mRNA expression of lipoprotein receptors, including LDLR and VLDLR were significantly increased, while the protein expression of PCSK9 was significantly increased in hypercholesterolemic placenta. In conclusion, maternal hypercholesterolemia might decrease the transportation of cholesterol from mother to fetus because of the high levels of PCSK9 protein expression.

27-Hydroxycholesterol regulates cholesterol synthesis and transport in C6 glioma cells.

The oxysterol 27-Hydroxycholesterol (27-OHC) is a major cholesterol metabolite that can cross the blood brain barrier (BBB) from peripheral circulation to the brain. Currently, the role of 27-OHC on cholesterol homeostasis in astrocytes and the underlying mechanisms are not defined. Since all brain cholesterol is essentially synthesized in brain itself and astrocytes as net producers of cholesterol are essential for normal brain function, here we investigated the effects of 27-OHC on cholesterol synthesis and transport in C6 glioma cells. C6 cells were treated with 5, 10 and 20µM 27-OHC for 24h and the cell viability and apoptosis, the cholesterol levels and metabolism-related mediators, genes and proteins were subsequently assessed using cell-counting kit (CCK)-8, Amplex red, ELISA, real-time PCR and Western blot, respectively. We found that 27-OHC decreased cholesterol levels by down-regulating the expression of sterol-regulated element binding protein-1 (SREBP-1a), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CR) and low density lipoprotein receptor (LDLR) and promoted cholesterol transport by up-regulating the expression of peroxisome proliferator-activated receptors-γ (PPAR-γ), liver X receptor-α (LXR-α), ATP-binding cassette transporter protein family member A1 (ABCA1) and apolipoprotein E (ApoE)genes. Our results suggested that 27-OHC may represent a sensitive modulator of cholesterol metabolism disorder by suppressing cholesterol synthesis and stimulating cholesterol transport in astrocytes.

Neurocalcin-delta: a potential memory-related factor in hippocampus of obese rats induced by high-fat diet.

INTRODUCTION: Aberrant protein expression within the hippocampus has recently been implicated in the pathogenesis of obesity-induced memory impairment. OBJECTIVES: The objective of the current study was to search for specific memory-related factors in the hippocampus in obese rats. METHODS: Sprague-Dawley (SD) rats were fed either a high-fat (HF) diet or normal-fat (NF) diet for 10 weeks to obtain the control (CON), diet-induced obese rats (DIO) and diet-resistant (DR) rats. D-galactose was injected subcutaneously for 10 weeks to establish model (MOD) rats with learning and memory impairment. After the hippocampus of the rats sampling, the proteome analysis was conducted using two-dimensional get electrophoresis (2-DE) combined with peptide mass fingerprinting (PMF). RESULTS: We found 15 differential proteins that expressed in the hippocampus in rats induced by HF diet from the 2-DE map. In addition, Neurocalcin-delta (NCALD) was nearly down-regulated in the DR rats compared with CON rats and MOD rats, which was further confirmed by Western blot, real-time PCR and ELISA results. CONCLUSION: Our data demonstrates that the differential memory-related proteins were a reflection of the HF diet, but not potential factors in obesity proneness or obesity resistance. Furthermore, NCALD is proved to be a potential hippocampus-memory related factor related to obesity.

Soy milk powder supplemented with phytosterol esters reduced serum cholesterol level in hypercholesterolemia independently of lipoprotein E genotype: a random clinical placebo-controlled trial.

Phytosterols (PSs) are reported to lower the serum total cholesterol and low-density lipoprotein cholesterol concentrations enriched in some fatty foods, such as margarine. However, these high-fat foods are not very suitable for older people. Soy milk is the favorite food for elderly people in China; therefore, we hypothesized that the consumption of soy milk powder supplemented with PSs would decrease the serum cholesterol levels in older Chinese people independent of the genotypes of apolipoprotein E (ApoE). Mild to moderate hyperlipidemic patients (n = 170) were recruited from different communities and treated with placebo soy milk powder or 3.4 g PS esters-enriched soy milk powder (2.0 g/d free PS in 30 g soy milk powder). The fasting serum lipid profiles at the baseline and after 3 and 6 months of intervention were measured. The ApoE genotype was also determined. After 3 months of PS intervention, the serum lipid profile was not changed significantly in either group. The serum total cholesterol, low-density lipoprotein cholesterol, and non- high-density lipoprotein cholesterol levels decreased by 9.3%, 11.4%, and 12.6%, respectively, in the PS group at the end of the intervention (6 months) compared with the control group, whereas the serum high-density lipoprotein cholesterol and triglyceride levels were not affected significantly. In the PS group, both the ApoE3 and ApoE4 carriers had a similar response to PS consumption. These findings suggested that PS-fortified soy milk powder was effective in lowering the serum cholesterol levels in older Chinese volunteers with mild to moderate hypercholesterolemia in both the ApoE3 and ApoE4 carriers.

Genistein Inhibits Aß25-35-Induced Synaptic Toxicity and Regulates CaMKII/CREB Pathway in SH-SY5Y Cells.

Genistein (Gen), as a functional food in human diet, has shown many beneficial effects on neurodegenerative diseases such as Alzheimer's disease (AD). But the neuroprotective mechanism of Gen is not clear. Because synaptic failure is considered as the earliest phase in the pathogenesis of AD, we try to validate our hypothesis that synapse may be one target of Gen on protecting neurons. In this study, SH-SY5Y cells were pre-incubated with or without Gen for 2 h followed by the incubation with Aß25-35 (25 µmol/L) for another 24 h. Flow cytometry, Western Blots, and RT-PCR analysis were used to test the synaptic factors. The data showed that Gen pre-treatment could reverse the Aß25-35-induced down-regulation of synaptophysin and postsynaptic marker postsynaptic density-95. In addition, the down-regulation of NR1 and NR2B induced by Aß25-35 which are subunits of N-methyl-D-aspartate receptor also could be antagonized by pre-treatment of Gen. Moreover, the factors of CaMKII/CREB signaling pathway were detected. The results showed that mRNA and protein expressions of (Ca(2+))/calmodulin(CaM), CaMKII/pCaMKII, and CREB/pCREB were significantly down-regulated by Aß25-35, but they were all restored by the pre-treatment of Gen. Furthermore, Gen also maintained the intracellular Ca(2+) concentration which was disturbed by Aß25-35. In conclusion, these results suggested that Gen could protect synaptic dysfunction induced by Aß, and the mechanism might be associated with the regulation of synaptic markers and Ca(2+) level through activating CaM/CaMK/CREB signaling pathway.

Global DNA methylation was changed by a maternal high-lipid, high-energy diet during gestation and lactation in male adult mice liver.

An epigenetic mechanism has been suggested to explain the effects of the maternal diet on the development of disease in offspring. The present study aimed to observe the effects of a maternal high-lipid, high-energy (HLE) diet on the DNA methylation pattern of male offspring in mice. Female C57BL/6J mice were fed an HLE diet during gestation and lactation. The genomic DNA methylations at promoter sites of genes in the liver, mRNA and protein levels of selected genes related to lipid and glucose metabolism were measured by microarray, real-time PCR and Western blot. The results indicated that the percentage of methylated DNA in offspring from dams that were fed an HLE diet was significantly higher than that from dams that were fed a chow diet, and most of these genes were hypermethylated in promoter regions. The nuclear protein content and mRNA levels of hypermethylated genes, such as PPARγ and liver X receptor α (LXRα), were decreased significantly in offspring in the HLE group. The results suggested that the DNA methylation profile in adult offspring livers was changed by the maternal HLE diet during gestation and lactation.

The oxysterol 27-hydroxycholesterol increases oxidative stress and regulate Nrf2 signaling pathway in astrocyte cells.

The disturbance in cholesterol metabolism has been considered as a cause of alzheimer's disease (AD), which dues to the oxidative damage and cell apoptosis in the brain. We aimed to investigate the toxicity and mechanism of AD-like pathology caused by cholesterol oxidation metabolite 27-hydroxycholesterol (27-OHC) in astrocyte cells. C6 cells were treated with 0, 5, 10, 20 µM 27-OHC for 24 h (h). The cell viability was monitored by using methyl thiazolyl tetrazolium test, generation of reactive oxygen species (ROS) was measured by using 2', 7'-dichlorodihydrofluorescein diacetate fluorescent probe under flow cytometry. The concentrations of 8-hydroxyl deoxyguanosine, the anti-oxidative enzymes such as total superoxide dismutase (tSOD), reduced glutathione (rGSH) and glutathione peroxidase (GSH-Px) were tested by using enzyme-linked immunosorbent assay and enzymic method, respectively. The gene and protein expression of nuclear factor E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NAD(P)H dehydrogenase quinone 1 (NQO1) and γ-glutamylcysteine synthetase (γ-GCS) in C6 cells were detected by quantitative western blot analysis and real-time PCR analysis. Moreover, the Nrf2 expressions in both of the cytoplasm and nucleus were detected with western blot analysis, and the localization of Nrf2 was performed by immunocytochemistry and confocal microscopy. 27-OHC increased the levels of ROS and decreased the levels of tSOD, rGSH, GSH-Px in C6 cells dose-dependently. In addition, 27-OHC down regulated the expressions of Nrf2, HO-1, NQO1 and γ-GCS at both of gene and protein levels, while Nrf2 expression in the cytoplasm showed decreased trend after incubated for 24 h with 27-OHC. The cholesterol metabolite 27-OHC is toxic to C6 cells and contributed to oxidative damage via regulating the Nrf2 signaling pathway. Our results suggest that 27-OHC may represent a common pathogenic factor in AD.

Soy isoflavone antagonizes the oxidative cerebrovascular injury induced by ß-amyloid peptides 1-42 in rats.

Numerous evidences have shown that the antioxidative properties of soy isoflavone (SIF) have beneficial effects on prophylaxis of neurodegeneration, however, the mechanism is still not fully illustrated. As cerebrovascular dysfunction could initiate a cascade of events leading to pathogenesis of Alzheimer's disease, we tried to investigate whether SIF could protect the cerebrovascular system due to antagonizing oxidative damage induced by Aß1-42 in present study. In addition, NF-E2-related factor 2 (Nrf2) signaling pathways in the cerebrovascular tissue of Wistar rats were investigated to identify the potential cerebrovascular protective targets of SIF. Research results showed that SIF reduced the excessive production of nitrotyrosine in cerebrovascular tissue induced by Aß1-42, and maintained redox homeostasis by increasing the level of GSH and GSH/GSSG. Moreover, SIF could alleviate the down-regulation of Nrf2, γ-glutamylcysteine synthetase, Heme oxygenase-1 expressions in cerebrovascular tissue induced by Aß1-42 and suppress the increase of Kelch like ECH protein-1 (Keap1). These data suggested that SIF might reduce the cerebrovascular oxidative damage induced by Aß1-42 through regulating the Nrf2 signaling pathway. The mechanisms of SIF modulating the potential target Nrf2 might be associated with Keap1 expression.

Beta amyloid peptide (25-35) leading to inflammation through Toll-like receptors and the anti-inflammatory effect of genistein in BV-2 cells.

Genistein, the main soy isoflavone component, has received much attention for its potential multifunction. Here, we reported that in BV-2 cells, genistein significantly inhibited beta amyloid peptides 25-35 (Aß25-35)-induced inflammatory response. The results indicated that Aß25-35-stimulated BV-2 cells upregulated Toll-like receptors 2 and 4, Myd88, and IKK gene expression with the increasing expression of IL-6 and decreasing expression of TGF-ß and IL-10. Further, inhibiting TLR4 expression with small interfering RNA prevented the inflammatory response induced by Aß25-35, indicating the key role of TLRs in Aß-mediated inflammation. Genistein pre-treated BV-2 cells showed less inflammatory response when exposed to Aß25-35. These results suggested that Aß induced BV-2 cells inflammation though TLRs and genistein has an anti-inflammatory effect in vitro.

Adaptive responses by mouse fetus to a maternal HLE diet by downregulating SREBP1: a microarray- and bio-analytic-based study.

Maternal diet has long been recognized as a significant factor affecting offspring development and health, but the target genes affected by a maternal high-lipid diet are currently unknown. In this study, the gene expression profile of neonatal mouse liver was analyzed using gene chips to identify genes with significant up- or downregulated expression levels due to maternal high-fat diet during gestation. Real-time PCR and Western blotting were used to measure key genes selected using microarray. Serum lipid, glucose, and insulin levels in adult offspring from dams fed with chow or a high-lipid diet were measured using commercial kits. Results indicate that the expression of genes involved in cholesterol and fatty acid synthesis were significantly inhibited, while the expression of genes involved in glycolysis were significantly decreased by maternal high-lipid diet during gestation. SREBP1 might be the key gene regulating genes involved in fatty acid, glucose, and cholesterol metabolism in response to a maternal high-fat diet.

The effects of phytosterol supplementation on serum LDL-C levels and learning ability in mice fed a high-fat, high-energy diet from gestation onward.

A high-fat, high-energy (HFE) diet may be deleterious to the cardiovascular system and mental health. We previously reported that serum cholesterol levels and escape latency were significantly increased in mice by feeding them an HFE diet from gestation onward. In this study, we examined whether an HFE diet supplemented with phytosterols fed to pregnant C57BL/6j dams and their offspring would protect the HFE-diet-induced compromise of the offspring's learning capability. We measured serum cholesterol levels, brain N-methyl-D-aspartate receptor (NMDAR1) mRNA and protein expression and liver sterol 27-hydroxylase (Cyp27a1) mRNA expression, as well as a Morris water maze performance. The results showed that, compared to mice consuming the HFE diet alone, those also consuming phytosterols (the HFE + PS diet) significantly decreased mean serum low-density lipoprotein cholesterol levels and altered brain NMDAR1 mRNA and protein expression and liver Cyp27a1 mRNA expression. The Morris water maze experiments indicated that dietary phytosterol supplementation slightly decreased the escape latency (p = 0.07). Collectively, these observations suggest that consumption of phytosterols from early in life may help alleviate the detrimental effects of HFE diets in mice.

Soy isoflavone alleviates Aß1-42-induced impairment of learning and memory ability through the regulation of RAGE/LRP-1 in neuronal and vascular tissue.

The neuroprotective properties of soy isoflavone (SIF) have been demonstrated by our previous studies and others, but its potential mechanism is not clear. Because of the key role of neurovascular dysfunction in the pathogenesis of Alzheimer's disease (AD), we hypothesized neurovascular tissue might be one neuroprotective target of SIF. In the present study, learning and memory ability, ß-amyloid (Aß) expressions both in neurovascular tissue and plasma, the receptor for advanced glycation end products (RAGE), low-density lipoprotein receptor-related protein (LRP)-1, nuclear factor-κB p65 (NF-κB p65), tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) expressions in neurovascular tissue were measured in Wistar rats following lateral cerebral ventricle administration of Aß1-42 by miniosmotic pump with or without intragastric administration of SIF from 14 days before surgery to the end of experiment. The results showed that SIF could improve the impairment of learning and memory of rats induced by Aß1-42, maintain Aß homeostasis in brain, regulate the disordered expressions of RAGE/LRP-1 and restrain RAGE related NF-κB and inflammatory cytokines activation in neurovascular structure. These results suggested that SIF could protect Aß-impaired learning and memory in rats, and its mechanism might be associated with the regulation of vascular Aß transportation and vascular inflammatory reaction.

Genistein alleviates the mitochondria-targeted DNA damage induced by ß-amyloid peptides 25-35 in C6 glioma cells.

Reactive oxygen species (ROS) are mainly produced by mitochondria which can cause oxidative stress. It has been considered that mitochondrial damage induced by oxidative stress is related to Alzheimer's disease (AD). Besides, mitochondrial DNA (mtDNA) is more vulnerable to oxidative damage than other biomacromolecules, causing serious dysfunction to mitochondria. ß-amyloid peptides (Aß) is a main factor responsible for the occurrence and development of AD. Astrocytes is an important target cell for Aß' toxicity and can be activated to neglect their normal fountain in the central nervous system. Genistein (Gen), a main active ingredient of soybean isoflavone, has been shown to have neuroprotective effects by antagonizing oxidative damage induced by Aß. Thus, in the present study, we evaluated Aß25-35 induced mitochondrial DNA (mtDNA) damage and the protective effect of Gen in C6 glioma cells (C6 cells). The study design was consisted of four groups: control group (vehicle), Aß group treated with Aß25-35, Gen + Aß group treated with Gen + Aß25-35 and Gen group treated with Gen only. C6 cells were pre-incubated with or without Gen (50 µM) for 2 h followed by the incubation with Aß25-35 (25 µM) for another 24 h. Then the cells were harvested and processed to perform the analysis according to protocols. The mitochondrial ROS in C6 cells were measured by fluorescence spectrometer. Enzyme-linked immunosorbent assay (ELISA) was used to detect the mitochondrial reduced glutathione (GSH) and oxidized glutathione (GSSG) in C6 cells, then the ratio of GSH and GSSG was calculated. The levels of 8-hydroxydeoxyguanosine (8-OHdG) in C6 cells was also detected by ELISA. In addition, mtDNA deletion was detected by polymerase chain reaction (PCR). The mRNA and protein expression of 8-oxoguanine DNA glycosylase (OGG1) in both C6 cells and its mitochondria, and manganese superoxide dismutase (MnSOD) in mitochondria were detected by using reverse transcription-PCR and Western blot. The results showed that the increased mitochondrial ROS accumulation in C6 cells induced by Aß was profoundly reversed by pre-treaded with Gen (p < 0.05). The ratio of GSH and GSSG in mitochondria was significantly increased in both Gen + Aß group and Gen group compared with Aß group (p < 0.05). The levels of 8-OHdG in C6 cells and mtDNA deletion were decreased after pre-treated with Gen (p < 0.05). Gen could also up-regulate the mRNA and protein expression of OGG1 in both C6 cells and its mitochondria and mitochondrial MnSOD compared with the Aß group (p < 0.05). These results confirmed that Gen could alleviate the mitochondria-targeted oxidative damage induced by ß-amyloid 25-35 in C6 cells which might be useful for the treatment of neurodegenerative diseases.

Soy isoflavone attenuates brain mitochondrial oxidative stress induced by ß-amyloid peptides 1-42 injection in lateral cerebral ventricle.

The aim of this study is to investigate whether soy isoflavone (SIF) reduces oxidative stress and improves the antioxidant ability in mitochondria of rat brain damaged by injection of beta-amyloid peptides 1-42 (Aß1-42). Forty Wistar rats were randomly divided into control, Aß1-42, SIF + Aß1-42, and SIF groups according to body weight. The rats in the SIF + Aß1-42 group and SIF group were intragastrically administered SIF suspension in 0.5% CMC-Na for 28 days, whereas the rats in control group and Aß1-42 group were administered the same volume of 0.5% CMC-Na. On day 14, the rats in the Aß1-42 group and SIF + Aß1-42 group were injected with Aß1-42 into the lateral cerebral ventricle with physiological saline. The rat brains were then sampled, and brain mitochondria were isolated. After this, the mitochondrial membrane potential (MMP) and mitochondrial redox state were measured. The contents of brain nuclear factor E2-related factor (Nrf2) and heme oxygenase-1 (HO-1) protein in brain tissue were quantitated by Western blot. The results showed that SIF maintained the MMP, elevated the reduced glutathione/oxidized glutathione (GSH/GSSG) ratio, and increased glutathione peroxidase (GPx) and manganese superoxide dismutase (MnSOD) protein expression in brain mitochondria. Additionally, SIF reversed the Aß1-42-induced downregulation of the protein expression of Nrf2 and HO-1 in brain tissue. These results indicated that SIF could alleviate the oxidative damage and maintain the redox imbalance in brain mitochondria damaged by Aß1-42. This might result from regulation of the Nrf2/HO-1 pathway.

Antagonizing effects of soybean isoflavones on ß-amyloid peptide-induced oxidative damage in neuron mitochondria of rats.

Soybean isoflavone (SIF) has been demonstrated to have neuroprotective effects induced by ß-amyloid peptides (Aß) through suppressing oxidative stress; however, the explicit mechanisms still remain uncovered. In the present study, 32 Wistar rats were randomly divided into four groups: an Aß1-42-treated group, a SIF + Aß1-42 group, a SIF-treated group and a control group. We measured the protein content of 8-hydroxydeoxyguanosine (8-OhdG) and mRNA expression of 8-oxoguanine DNA glycosylase (OGG1). The protein expression of OGG1, Bcl-xl, Bad, beta subunit of ATP synthase (ATPB) and pyruvate dehydrogenase (PDH) in brain was also measured. The results showed that the level of 8-OHdG in both SIF groups was significantly decreased compared to the Aß1-42-treated group (p < 0.05), while the mRNA and protein expression of OGG1 in the SIF + Aß1-42 groups were up-regulated compared with the Aß1-42-treated groups (p < 0.05). The expression of Bcl-xl was up-regulated in the SIF-treated group compared with the Aß1-42-treated groups (p < 0.05), while the expression of Bad was down-regulated in the two SIF-treated groups (p < 0.05). Aß1-42 significantly down-regulated the expression of ATPase and PDH proteins compared with the control group (p < 0.05). SIF reversed the down-regulation effects on the mitochondrial energy metabolic enzymes induced by Aß1-42 (p < 0.05) in the rats. These results suggest that SIF alleviate the oxidative stress in neurons and mitochondria of rat brains mediated by Aß1-42, and these protective effects might be associated with the regulation of OGG1, Bad, Bcl-xl, ATPB and PDH.

Flavonoids protect cerebrovascular endothelial cells through Nrf2 and PI3K from ß-amyloid peptide-induced oxidative damage.

ß-amyloid peptides (Aß) induced cerebrovascular dysfunction has been recognized as a vital factor involved in the pathogenesis of neurodegeneration. Genistein, a flavonoid, has antioxidative properties to prevent neurodegeneration induced by ß-amyloid peptides. In this study, we were investigating whether genistein could antagonize oxidative damage induced by ß-amyloid peptide 25-35 (Aß25-35) in bEND.3 cells, and also identifying the potential neuroprotective targets of genistein. Vitamin E was used as the positive control. The bEND.3 cells were pre-incubated with/out genistein or vitamin E for 2 h followed by the incubation with 25 µM A 25-35 for another 24 h. The reactive oxygen species (ROS), nitrotyrosine, cell redox state, mRNA or protein expressions of the factors on Nrf2 signaling pathway were measured after Aß25-35 treatment. The results showed that genistein alleviated the increase of ROS and nitrotyrosine production induced by Aß25-35, and maintained bEND.3 cell redox state by increasing GSH level and GSH/GSSG. Genistein could reverse the down-regulation of total protein and mRNA expression of NF-E2-related factor 2 (Nrf2), nuclear Nrf2, γ-glutamylcysteine synthetase (γ-GCS), phosphatidylinositol 3-kinase (PI3K) induced by Aß25-35; while PI3K inhibitor LY294002 could attenuate the activation effects of genistein on Nrf2, especially for the promotion of nuclear translocation. These results suggested that genistein could protect cerebrovascular endothelial cells from Aß25-35-induced oxidative damage. The potential mechanisms might be associated with the activation of Nrf2 signaling pathway by modulating PI3K activity.

Transcription factor GATA-6 recruits PPARα to cooperatively activate Glut4 gene expression.

Peroxisome proliferator-activated receptor α (PPARα) is a nuclear hormone receptor that regulates energy metabolism, but its precise mechanisms remain unknown. Here, we demonstrate that the PPARα agonist fenofibrate activated expression of the glucose transporter Glut4. Moreover, PPARα was associated with the Glut4 promoter through GATA sites upon fenofibrate stimulation in cardiomyocytes. This occupancy is achieved through an interaction between amino acids 1-136 of PPARα with amino acids 276-443 of the cardiac transcription factor GATA-6. In addition, the interaction of PPARα with GATA-6 activated Glut4 gene expression, improved glucose consumption, and enhanced activity of mitochondrial citrate synthase in C2C12 myoblasts; both mutants of PPARα (1-101 aa) and GATA-6 (227-331 aa) were unable to cooperate in Glut4 activation. Thus, GATA-6 is an important component of the transcription network required for energy metabolism mediated by PPARα, and these findings provide a molecular basis for understanding the role of GATA-6 proteins in muscle development and disease.