Blueberries Improve Neuroinflammation and Cognition differentially Depending on Individual Cognitive baseline Status.

Daily supplementation of blueberries (BBs) reverses age-related deficits in behavior in aged rats. However, it is unknown whether BB is more beneficial to one subset of the population dependent on baseline cognitive performance and inflammatory status. To examine the effect of individual differences on the efficacy of BB, aged rats (17 months old) were assessed for cognition in the radial arm water maze (RAWM) and divided into good, average, and poor performers based on navigation errors. Half of the rats in each cognitive group were then fed a control or a 2% BB diet for 8 weeks before retesting. Serum samples were collected, pre-diet and post-diet, to assess inflammation. Latency in the radial arm water maze was significantly reduced in the BB-fed poor performers (p < .05) and preserved in the BB-fed good performers. The control-fed good performers committed more working and reference memory errors in the post-test than pretest (p < .05), whereas the BB-fed good performers showed no change. An in vitro study using the serum showed that BB supplementation attenuated lipopolysaccharide (LPS)-induced nitrite and tumor necrosis factor-alpha, and cognitive performance was associated with innate anti-inflammatory capability. Therefore, consumption of BB may reverse some age-related deficits in cognition, as well as preserve function among those with intact cognitive ability.

Nutritional Factors Affecting Adult Neurogenesis and Cognitive Function.

Adult neurogenesis, a complex process by which stem cells in the hippocampal brain region differentiate and proliferate into new neurons and other resident brain cells, is known to be affected by many intrinsic and extrinsic factors, including diet. Neurogenesis plays a critical role in neural plasticity, brain homeostasis, and maintenance in the central nervous system and is a crucial factor in preserving the cognitive function and repair of damaged brain cells affected by aging and brain disorders. Intrinsic factors such as aging, neuroinflammation, oxidative stress, and brain injury, as well as lifestyle factors such as high-fat and high-sugar diets and alcohol and opioid addiction, negatively affect adult neurogenesis. Conversely, many dietary components such as curcumin, resveratrol, blueberry polyphenols, sulforaphane, salvionic acid, polyunsaturated fatty acids (PUFAs), and diets enriched with polyphenols and PUFAs, as well as caloric restriction, physical exercise, and learning, have been shown to induce neurogenesis in adult brains. Although many of the underlying mechanisms by which nutrients and dietary factors affect adult neurogenesis have yet to be determined, nutritional approaches provide promising prospects to stimulate adult neurogenesis and combat neurodegenerative diseases and cognitive decline. In this review, we summarize the evidence supporting the role of nutritional factors in modifying adult neurogenesis and their potential to preserve cognitive function during aging.

Modulation of oxidative stress, inflammation, autophagy and expression of Nrf2 in hippocampus and frontal cortex of rats fed with açaí-enriched diets.

OBJECTIVE: Açaí (Euterpe spp.), an exotic palm fruit, has recently emerged as a promising source of natural antioxidants with wide pharmacological and nutritional value. In this study, two different species of açaí pulp extracts, naturally grown in two distinct regions of the Amazon, namely, Euterpe oleracea Mart. (habitat: Brazilian floodplains of the Amazon) and Euterpe precatoria Mart. (habitat: Bolivian Amazon), were studied for their effects on brain health and cognition. METHODS: Neurochemical analyses were performed in critical brain regions associated with memory and cognition of 19-month-old açaí-fed rats, in whom the cognitive benefits of açaí had been established. RESULTS: Results indicated significant reductions (P< 0.05) in prooxidant NADPH-oxidoreductase-2 (NOX2) and proinflammatory transcription factor NF-κB in açaí-fed rats. Measurement of Nrf2 expression, a transcription factor for antioxidant enzymes, and a possible link between oxidative stress, neuroinflammation and autophagy mechanisms, indicated significant overexpression (P<0.005) in the hippocampus and frontal cortex of the açaí-fed rats. Furthermore, significant activation of endogenous antioxidant enzymes GST and SOD were also observed in the açaí-fed animals when compared to control. Analysis of autophagy markers such as p62, phospho-mTOR, beclin1 and MAP1B-LC3 revealed differential expression in frontal cortex and hippocampus, mostly indicating an upregulation in the açaí-fed rats. DISCUSSION: In general, results were more profound for EP than EO in hippocampus as well as frontal cortex. Therefore, an açaí-enriched diet could possibly modulate Nrf2, which is known to modulate the intracellular redox status, thereby regulating the ubiquitin-proteosomal pathway, ultimately affecting cognitive function in the aging brain.

Serum metabolites from walnut-fed aged rats attenuate stress-induced neurotoxicity in BV-2 microglial cells.

The shift in equilibrium towards excess reactive oxygen or nitrogen species production from innate antioxidant defenses in brain is a critical factor in the declining neural function and cognitive deficit accompanying age. Previous studies from our laboratory have reported that walnuts, rich in polyphenols, antioxidants, and omega fatty acids such as alpha-linolenic acid and linoleic acid, improve the age-associated declines in cognition and neural function in rats. Possible mechanisms of action of these effects include enhancing protective signaling, altering membrane microstructures, decreasing inflammation, and preventing accumulation of polyubiquitinated protein aggregates in critical regions of the brain. In the current study, we investigated whether the serum collected from aged animals fed with walnut diets (0, 6, and 9%, w/w) would enhance protection on stressed BV-2 microglia in vitro. In the growth medium, fetal bovine serum was substituted with the serum collected from 22-month-old rats fed per protocol for 12 weeks. Walnut diet serum (6 and 9%) significantly attenuated lipopolysaccharide-induced nitrite release compared to untreated control cells and those treated with serum from rats fed 0% walnut diets. The results also indicated a significant reduction in pro-inflammatory tumor necrosis factor-alpha, cyclooxygenase-2, and inducible nitric oxide synthase. These results suggest antioxidant and anti-inflammatory protection or enhancement of membrane-associated functions in brain cells by walnut serum metabolites.

Dietary supplementation with the polyphenol-rich açaí pulps (Euterpe oleracea Mart. and Euterpe precatoria Mart.) improves cognition in aged rats and attenuates inflammatory signaling in BV-2 microglial cells.

OBJECTIVES: The present study was carried out to determine if lyophilized açaí fruit pulp (genus, Euterpe), rich in polyphenols and other bioactive antioxidant and anti-inflammatory phytochemicals, is efficacious in reversing age-related cognitive deficits in aged rats. METHODS: The diets of 19-month-old Fischer 344 rats were supplemented for 8 weeks with 2% Euterpe oleracea (EO), Euterpe precatoria (EP), or a control diet. Rats were tested in the Morris water maze and then blood serum from the rats was used to assess inflammatory responses of BV-2 microglial cells. RESULTS: After 8 weeks of dietary supplementation with 2% EO or EP, rats demonstrated improved working memory in the Morris water maze, relative to controls; however, only the EO diet improved reference memory. BV-2 microglial cells treated with blood serum collected from EO-fed rats produced less nitric oxide (NO) than control-fed rats. Serum from both EO- and EP-fed rats reduced tumor necrosis factor-alpha (TNF-α). There is a relationship between performance in the water maze and the production of NO and TNF-α by serum-treated BV-2 cells, such that serum from rats with better performance was more protective against inflammatory signaling. DISCUSSION: Protection of memory during aging by supplementation of lyophilized açaí fruit pulp added to the diet may result from its ability to influence antioxidant and anti-inflammatory signaling.

Effects of pterostilbene and resveratrol on brain and behavior.

Age is the greatest universal risk factor for neurodegenerative diseases. During aging, these conditions progress from minor loss of function to major disruptions in daily life, loss of independence and ultimately death. Because approximately 25% of the world population is expected to be older than age 65 by 2050, and no treatments exist to halt or reverse ongoing neurodegeneration, the need for effective prevention strategies is more pressing that ever before. A growing body of research supports the role of diet in healthy aging, particularly diets rich in bioactive phytochemical compounds. Recently, stilbenes such as resveratrol (3, 5, 4'-trans-trihydroxystilbene) and its analogue, pterostilbene, have gained a significant amount of attention for their potent antioxidant, anti-inflammatory, and anticarcinogenic properties. However, evidence for the beneficial effects of stilbenes on cerebral function is just beginning to emerge. In this review, we summarize the current knowledge on the role of resveratrol and pterostilbene in improving brain health during aging, with specific focus on antioxidant and anti-inflammatory signaling and behavioral outcomes.

Protective effects of blueberry- and strawberry diets on neuronal stress following exposure to (56)Fe particles.

Particles of high energy and charge (HZE particles), which are abundant outside the magnetic field of the Earth, have been shown to disrupt the functioning of neuronal communication in critical regions of the brain. Previous studies with HZE particles, have shown that irradiation produces enhanced indices of oxidative stress and inflammation as well as altered neuronal function that are similar to those seen in aging. Feeding animals antioxidant-rich berry diets, specifically blueberries and strawberries, countered the deleterious effects of irradiation by reducing oxidative stress and inflammation, thereby improving neuronal signaling. In the current study, we examined the effects of exposure to (56)Fe particles in critical regions of brain involved in cognitive function, both 36h and 30 days post irradiation. We also studied the effects of antioxidant-rich berry diets, specifically a 2% blueberry or strawberry diet, fed for 8 weeks prior to radiation as well as 30 days post irradiation. (56)Fe exposure caused significant differential, neurochemical changes in critical regions of the brain, such as hippocampus, striatum, frontal cortex, and cerebellum, through increased inflammation, and increased oxidative stress protein markers. (56)Fe exposure altered the autophagy markers, and antioxidant-rich berry diets significantly reduced the accumulation of p62 in hippocampus, a scaffold protein that co-localizes with ubiquitinated protein at the 30 days post irradiation time-point. Exposure to (56)Fe particles increased the accumulation of disease-related proteins such as PHF-tau in the hippocampus of animals fed the control diet, but not in the irradiated animals fed the blueberry diet. These results indicate the potential protective effects of antioxidant-rich berry diets on neuronal functioning following exposure to HZE particles.

Restoration of stressor-induced calcium dysregulation and autophagy inhibition by polyphenol-rich açaí (Euterpe spp.) fruit pulp extracts in rodent brain cells in vitro.

OBJECTIVES: Oxidative damage to lipids, proteins, and nucleic acids in the brain often causes progressive neuronal degeneration and death that are the focal traits of chronic and acute pathologies, including those involving cognitive decline. The aim of this study was to investigate the specific effects of both Euterpe oleracea and Euterpe precatoria açaí fruit pulp on restoring stressor-induced calcium dysregulation, stunted growth of basal dendrites, and autophagy inhibition using embryonic hippocampal and HT22 hippocampal neurons. METHODS: Water-soluble whole fruit pulp extracts from two açaí species were applied to rat primary neurons and HT22 hippocampal neurons with varied time and concentrations. Recovery of neurons from dopamine-induced Ca(2+) dysregulation was measured by live cell imaging using fluorescent microscopy. The effect of açaí fruit pulp extracts on neurons following chemically-induced autophagy inhibition was measured using both immunofluorescence and immunohistochemical techniques. RESULTS: It has been postulated that at least part of the loss of cognitive function in aging may depend on a dysregulation in calcium ion (Ca(2+)) homeostasis and a loss of autophagy function in the brain, which affects numerous signaling pathways and alters protein homeostasis. In the present study, polyphenol-rich fruit pulp extracts from two species of açaí, Euterpe precatoria and Euterpe oleracea, when applied to rat hippocampal primary neuronal cells (E18), caused a significant (P < 0.05) recovery of depolarized brain cells from dopamine-induced Ca(2+) influx. Autophagy, a protein homeostasis mechanism in brain, when blocked by known inhibitors such as bafilomycin A1 or wortmannin, caused a significant reduction in the growth of primary basal dendrites in rodent primary hippocampal neurons and significant accumulation of polyubiquitinated proteins in mouse HT22 hippocampal neurons in culture. However, pretreatment with açaí extracts up to 1 mg/mL significantly increased the length of basal dendrites and attenuated the inhibitor-induced autophagy dysfunction. Açaí extracts activated the phosphorylation of mammalian target of rapamycin, increased the turnover of autophagosomes and MAP1 B LC3-II, and decreased accumulation of LC3-ubiquitin binding P62/SQSTM1. CONCLUSION: Although the polyphenol profile of Euterpe precatoria showed substantially higher concentrations of major flavonoids han Euterpe oleracea, the relative effects were essentially similar for both species. The study adds to growing evidence that supports the putative health effects of açaí fruit species on brain cells.

Role of walnuts in maintaining brain health with age.

Because of the combination of population growth and population aging, increases in the incidence of chronic neurodegenerative disorders have become a societal concern, both in terms of decreased quality of life and increased financial burden. Clinical manifestation of many of these disorders takes years, with the initiation of mild cognitive symptoms leading to behavioral problems, dementia and loss of motor functions, the need for assisted living, and eventual death. Lifestyle factors greatly affect the progression of cognitive decline, with high-risk behaviors including unhealthy diet, lack of exercise, smoking, and exposure to environmental toxins leading to enhanced oxidative stress and inflammation. Although there exists an urgent need to develop effective treatments for age-related cognitive decline and neurodegenerative disease, prevention strategies have been underdeveloped. Primary prevention in many of these neurodegenerative diseases could be achieved earlier in life by consuming a healthy diet, rich in antioxidant and anti-inflammatory phytochemicals, which offers one of the most effective and least expensive ways to address the crisis. English walnuts (Juglans regia L.) are rich in numerous phytochemicals, including high amounts of polyunsaturated fatty acids, and offer potential benefits to brain health. Polyphenolic compounds found in walnuts not only reduce the oxidant and inflammatory load on brain cells but also improve interneuronal signaling, increase neurogenesis, and enhance sequestration of insoluble toxic protein aggregates. Evidence for the beneficial effects of consuming a walnut-rich diet is reviewed in this article.

Anthocyanin-rich açai (Euterpe oleracea Mart.) fruit pulp fractions attenuate inflammatory stress signaling in mouse brain BV-2 microglial cells.

Age-related diseases of the brain compromise memory, learning, and movement and are directly linked with increases in oxidative stress and inflammation. Previous research has shown that supplementation with berries can modulate signaling in primary hippocampal neurons or BV-2 mouse microglial cells. Because of their high polyphenolic content, fruit pulp fractions of açai ( Euterpe oleracea Mart.) were explored for their protective effect on BV-2 mouse microglial cells. Freeze-dried açai pulp was fractionated using solvents with different polarities and analyzed using HPLC for major anthocyanins and other phenolics. Fractions extracted using methanol (MEOH) and ethanol (ETOH) were particularly rich in anthocyanins such as cyanidin, delphinidin, malvidin, pelargonidin, and peonidin, whereas the fraction extracted using acetone (ACE) was rich in other phenolics such as catechin, ferulic acid, quercetin, resveratrol, and synergic and vanillic acids. Studies were conducted to investigate the mitigating effects of açai pulp extracts on lipopolysaccharide (LPS, 100 ng/mL) induced oxidative stress and inflammation; treatment of BV-2 cells with acai fractions resulted in significant (p < 0.05) decreases in nitrite production, accompanied by a reduction in inducible nitric oxide synthase (iNOS) expression. The inhibition pattern was emulated with the ferulic acid content among the fractions. The protection of microglial cells by açai pulp extracts, particularly that of MEOH, ETOH, and ACE fractions, was also accompanied by a significant concentration-dependent reduction in cyclooxygenase-2 (COX-2), p38 mitogen-activated protein kinase (p38-MAPK), tumor necrosis factor-α (TNFα), and nuclear factor κB (NF-κB). The current study offers valuable insights into the protective effects of açai pulp fractions on brain cells, which could have implications for improved cognitive and motor functions.

Exposure to 16O-particle radiation causes aging-like decrements in rats through increased oxidative stress, inflammation and loss of autophagy.

Exposing young rats to particles of high energy and charge (HZE particles), a ground-based model for exposure to cosmic rays, enhances indices of oxidative stress and inflammation, disrupts the functioning of neuronal communication, and alters cognitive behaviors. Even though exposure to HZE particles occurs at low fluence rates, the cumulative effects of long-term exposure result in molecular changes similar to those seen in aged animals. In the present study, we assessed markers of autophagy, a dynamic process for intracellular degradation and recycling of toxic proteins and organelles, as well as stress and inflammatory responses, in the brains of Sprague-Dawley rats irradiated at 2 months of age with 5 and 50 cGy and 1 Gy of ionizing oxygen particles ((16)O) (1000 MeV/n). Compared to nonirradiated controls, exposure to (16)O particles significantly inhibited autophagy function in the hippocampus as measured by accumulation of ubiquitin inclusion bodies such as P62/SQSTM1, autophagosome marker microtubule-associated protein 1 beta light chain 3 (MAP1B-LC3), beclin1 and proteins such as mammalian target of rapamycin (mTOR). The molecular changes measured at short (36 h) and long (75 days) intervals after (16)O-particle exposure indicate that the loss of autophagy function occurred shortly after exposure but was recovered via inhibition of mTOR. However, HZE-particle radiation caused significant sustained loss of protein kinase C alpha (PKC-α), a key G protein modulator involved in neuronal survival and functions of neuronal trophic factors. Exposure to (16)O particles also caused substantial increases in the levels of nuclear factor kappa B (NF-κB) and glial fibrillary acidic protein (GFAP), indicating glial cell activation 75 days after exposure. This is the first report to show the molecular effects of (16)O-particle radiation on oxidative stress, inflammation and loss of autophagy in the brain of young rats.

Inhibition of colon cancer cell growth and antioxidant activity of bioactive compounds from Poncirus trifoliata (L.) Raf.

Recently several plant derived natural compounds have been screened for their anticancer activity in order to identify putative compounds with novel structures or mechanism of action. In the present study, fruits of Poncirus trifoliata were extracted with acetone and loaded onto silica gel column chromatography. The column was eluted with different solvents to obtain two bioactive compounds. The purity of compounds was analyzed by HPLC and their structures were identified by 1H and 13C NMR experiments as beta-sitosterol and 2-hydroxy-1,2,3-propanetricarboxylic acid 2-methyl ester (HPCME). beta-Sitosterol, HPCME, and trolox were tested for their antioxidant capacity by oxygen radical absorbance capacity (ORAC) measurement. Further, these compounds were tested for their inhibition of cancer cell proliferation and apoptosis using human colon cancer cell line (HT-29). These results were compared with the corresponding activity on non-cancerous (COS-1 fibroblast) cells. Cell proliferation and induction of apoptosis were determined by MTT assay and nuclear staining. The MTT assay indicated that both the compounds exhibited differential inhibition at various concentrations. Significant arrest of cell growth was observed with beta-sitosterol even at low concentration such as 0.63 microM in 48 h and none of the compounds exerted any apparent cytostatic effects on the non-cancerous COS-1 fibroblast cells. Growth inhibition assay suggested the potential use of bioactive compounds as cancer chemopreventive and therapeutic agents. This is the first report on HPCME isolation and identification from Rutaceae family and beta-sitosterol from P. trifoliata.

Antiproliferative effects of citrus limonoids against human neuroblastoma and colonic adenocarcinoma cells.

Limonoids, a family of triterpenoids with putative anticancer properties, occur in fruits as glucosides and aglycones. Both highly purified forms were isolated from seeds and molasses of citrus fruits and tested for toxic effects against two human cancer cell lines, SH-SY5Y neuroblastoma and Caco-2 colonic adenocarcinoma, and a noncancerous mammalian epithelial Chinese hamster ovary (CHO) cells. Viability, as quantified by 3-[4,5-dimethylthiazol- 2-yl]-2,5-diphenyltetrazolium reduction and light microscopy, was shortened significantly (P < 0.001) in cancer cells exposed to aglycones, viz., limonin, nomilin, obacunone, and deacetylnomilin. SH-SY5Y cells were more sensitive than Caco-2 cells to the limonoids, whereas noncancerous CHO cells showed hardly any change in cell numbers or cell morphology. Aglycone toxicity was dose dependent, but below the killing potential of glucosides. This observation correlated with a slower rate of induction of caspase 3/7 activity by aglycones. A flow cytometric analysis of SH-SY5Y cells treated with glucosides and aglycones showed an increased ploidy, which is consistent with enhancing chromosomal abnormalities. The results confirm that limonoids exert a strong multifaceted lethal action against cancer cells, but are relatively ineffective against CHO cells. Of the two, metabolites derived from glucosides are the more likely progenitors of an apoptosis response in situ.

Furocoumarins from grapefruit juice and their effect on human CYP 3A4 and CYP 1B1 isoenzymes.

Bioactive compounds present in grapefruit juice are known to increase the bioavailability of certain medications by acting as potent CYP 3A4 inhibitors. An efficient technique has been developed for isolation and purification of three furocoumarins. The isolated compounds have been tested for the inhibition of human CYP 1B1 isoform using specific substrates. Grapefruit juice was extracted with ethyl acetate (EtOAc) and the dried extract was loaded onto silica gel column chromatography. Further, column fractions were subjected to preparative HPLC to obtain three compounds. The purity of these compounds was analyzed by HPLC and structures were determined by NMR studies. The identified compounds, bergamottin, 6',7'-dihydroxybergamottin (DHB), and paradisin-A, were tested for their inhibitory effects on hydroxylase and O-dealkylase activities of human cytochrome P450 isoenzymes CYP 3A4 and CYP 1B1. Paradisin-A was found to be a potent CYP 3A4 inhibitor with an IC50 of 1.2 microM followed by DHB and bergamottin. All three compounds showed a substantial inhibitory effect on CYP 3A4 below 10 microM. Inhibitory effects on CYP 1B1 exhibited a greater variation due to the specificity of substrates. Paradisin A showed an IC50 of 3.56+/-0.12 microM for the ethoxy resorufin O-dealkylase (EROD) activity and 33.56+/-0.72 microM for the benzyloxy resorufin (BROD). DHB and bergamottin showed considerable variations for EROD and BROD activities with an IC50 of 7.17 microM and 13.86 microM, respectively.

Citrus limonoids induce apoptosis in human neuroblastoma cells and have radical scavenging activity.

Citrus limonoid glucosides, a family of fruit bioactive compounds, were postulated to have free radical-scavenging and apoptosis-inducing properties against certain types of cancers. Four highly purified limonoid glucosides, limoin 17beta D-glucopypranoside (LG), obacunone 17beta D-glucopyranoside (OG), nomilinic acid 17beta D-glucopyranoside (NAG), and deacetylnomilinic acid 17beta D-glucopyranoside (DNAG) were tested for superoxide radical (O(2)(-))-quenching activity and cytotoxic action against undifferentiated human SH-SY5Y neuroblastoma cells in culture. All 4 scavenged O(2)(-) as measured by inhibition of pyrogallol decomposition in a spectrophotometric assay. Quenching by NAG in particular emulated an equivalent concentration of vitamin C. When added to the medium of SH-SY5Y cells in culture, micromolar amounts of LG and OG, compared with untreated controls, caused a cessation of cell growth and rapid cell death (P < 0.001); NAG and DNAG were better tolerated, but nonetheless toxic as well. Cytotoxicity was related to a concentration- and time-dependent increase in caspase 3/7 activity, suggesting that limonoid glucosides were capable of inducing apoptosis. Arrested cell growth and the induction of apoptosis were confirmed by flow cytometry and DNA fragmentation analysis. Importantly, caspase induction at 12 h correlated with cell survival at 24 h (P = 0.046), suggesting that apoptosis was the primary cause of cell death. We conclude that citrus limonoid glucosides are toxic to SH-SY5Y cancer cells. Cytotoxicity is exerted through apoptosis by an as yet unknown mechanism of induction. Individual limonoid glucosides differ in efficacy as anticancer agents, and this difference may reside in structural variations in the A ring of the limonoid molecule.