Brain xanthophyll content and exploratory gene expression analysis: subspecies differences in rhesus macaque

BACKGROUND: The dietary xanthophylls, lutein and zeaxanthin, accumulate in primate retina and brain, and emerging evidence indicates neural lutein content may be beneficial for cognition. Neural xanthophyll content in primates varies greatly among individuals, and genetic factors are likely to be significant contributors. Subspecies of rhesus macaques originating from different geographic locations are known to differ genetically, but the effect of origin on gene expression and carotenoid status has not been determined. The study objective was to determine whether xanthophyll status and expression of carotenoid-related genes, as well as genes with known variants between subspecies, differ between the brains of adult rhesus monkeys of Indian and Chinese origin. METHODS: Samples of prefrontal cortex, cerebellum, and striatum were collected from adult monkeys (n = 9) fed a standard stock diet containing carotenoids. Serum and brain carotenoids were determined using reverse-phase high-performance liquid chromatography. For each brain region, RNA sequencing and real-time quantitative polymerase chain reaction were used to determine differentially expressed genes between the subspecies. RESULTS: Indian-origin monkeys had higher xanthophyll levels in brain tissue compared to Chinese-origin monkeys despite consuming similar amounts of dietary carotenoids. In a region-specific manner, four genes related to carotenoid and fatty acid metabolism (BCO2, RPE65, ELOVL4, FADS2) and four genes involved in the immune response (CD4, CD74, CXCL12 LTBR) were differentially expressed between Indian- and Chinese-origin monkeys. Expression of all four genes involved in carotenoid and fatty acid metabolism were correlated with brain xanthophyll concentration in a region-specific manner. CONCLUSIONS: These results indicate that origin is related to differences in both gene expression and xanthophyll content in the brain. Findings from this study may have important implications regarding genetic diversity, lutein status, and cognition in primates

Effect of Carotenoid Supplemented Formula on Carotenoid Bioaccumulation in Tissues of Infant Rhesus Macaques: A Pilot Study Focused on Lutein

Lutein is the predominant carotenoid in the developing primate brain and retina, and may have important functional roles. However, its bioaccumulation pattern during early development is not understood. In this pilot study, we investigated whether carotenoid supplementation of infant formula enhanced lutein tissue deposition in infant rhesus macaques. Monkeys were initially breastfed; from 1 to 3 months of age they were fed either a formula supplemented with lutein, zeaxanthin, β-carotene and lycopene, or a control formula with low levels of these carotenoids, for 4 months (n = 2/group). All samples were analyzed by high pressure liquid chromatography (HPLC). Final serum lutein in the supplemented group was 5 times higher than in the unsupplemented group. All brain regions examined showed a selective increase in lutein deposition in the supplemented infants. Lutein differentially accumulated across brain regions, with highest amounts in occipital cortex in both groups. β-carotene accumulated, but zeaxanthin and lycopene were undetectable in any brain region. Supplemented infants had higher lutein concentrations in peripheral retina but not in macular retina. Among adipose sites, abdominal subcutaneous adipose tissue exhibited the highest lutein level and was 3-fold higher in the supplemented infants. The supplemented formula enhanced carotenoid deposition in several other tissues. In rhesus infants, increased intake of carotenoids from formula enhanced their deposition in serum and numerous tissues and selectively increased lutein in multiple brain regions

Lutein accumulates in subcellular membranes of brain regions in adult rhesus macaques: Relationship to DHA oxidation products

<div><p>Objectives</p><p>Lutein, a carotenoid with anti-oxidant functions, preferentially accumulates in primate brain and is positively related to cognition in humans. Docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid (PUFA), is also beneficial for cognition, but is susceptible to oxidation. The present study characterized the membrane distribution of lutein in brain regions important for different domains of cognitive function and determined whether membrane lutein was associated with brain PUFA oxidation.</p><p>Methods</p><p>Adult rhesus monkeys were fed a stock diet (~2 mg/day lutein or ~0.5 μmol/kg body weight/day) (n = 9) or the stock diet plus a daily supplement of lutein (~4.5 mg/day or~1 μmol/kg body weight/day) and zeaxanthin (~0.5 mg/day or 0.1 μmol/kg body weight/day) for 6–12 months (n = 4). Nuclear, myelin, mitochondrial, and neuronal plasma membranes were isolated using a Ficoll density gradient from prefrontal cortex (PFC), cerebellum (CER), striatum (ST), and hippocampus (HC). Carotenoids, PUFAs, and PUFA oxidation products were measured using HPLC, GC, and LC-GC/MS, respectively.</p><p>Results</p><p>All-<i>trans</i>-lutein (ng/mg protein) was detected in all regions and membranes and was highly variable among monkeys. Lutein/zeaxanthin supplementation significantly increased total concentrations of lutein in serum, PFC and CER, as well as lutein in mitochondrial membranes and total DHA concentrations in PFC only (P<0.05). In PFC and ST, mitochondrial lutein was inversely related to DHA oxidation products, but not those from arachidonic acid (P <0.05).</p><p>Discussion</p><p>This study provides novel data on subcellular lutein accumulation and its relationship to DHA oxidation in primate brain. These findings support the hypothesis that lutein may be associated with antioxidant functions in the brain.</p></div

The Subcellular Distribution of Alpha-Tocopherol in the Adult Primate Brain and Its Relationship with Membrane Arachidonic Acid and Its Oxidation Products

The relationship between α-tocopherol, a known antioxidant, and polyunsaturated fatty acid (PUFA) oxidation, has not been directly investigated in the primate brain. This study characterized the membrane distribution of α-tocopherol in brain regions and investigated the association between membrane α-tocopherol and PUFA content, as well as brain PUFA oxidation products. Nuclear, myelin, mitochondrial, and neuronal membranes were isolated using a density gradient from the prefrontal cortex (PFC), cerebellum (CER), striatum (ST), and hippocampus (HC) of adult rhesus monkeys (n = 9), fed a stock diet containing vitamin E (α-, γ-tocopherol intake: ~0.7 µmol/kg body weight/day, ~5 µmol/kg body weight/day, respectively). α-tocopherol, PUFAs, and PUFA oxidation products were measured using high performance liquid chromatography (HPLC), gas chromatography (GC) and liquid chromatography-gas chromatography/mass spectrometry (LC-GC/MS) respectively. α-Tocopherol (ng/mg protein) was highest in nuclear membranes (p &lt; 0.05) for all regions except HC. In PFC and ST, arachidonic acid (AA, µg/mg protein) had a similar membrane distribution to α-tocopherol. Total α-tocopherol concentrations were inversely associated with AA oxidation products (isoprostanes) (p &lt; 0.05), but not docosahexaenoic acid oxidation products (neuroprostanes). This study reports novel data on α-tocopherol accumulation in primate brain regions and membranes and provides evidence that α-tocopherol and AA are similarly distributed in PFC and ST membranes, which may reflect a protective effect of α-tocopherol against AA oxidation

Exploratory Metabolomic Analyses Reveal Compounds Correlated with Lutein Concentration in Frontal Cortex, Hippocampus, and Occipital Cortex of Human Infant Brain

<div><p>Lutein is a dietary carotenoid well known for its role as an antioxidant in the macula, and recent reports implicate a role for lutein in cognitive function. Lutein is the dominant carotenoid in both pediatric and geriatric brain tissue. In addition, cognitive function in older adults correlated with macular and postmortem brain lutein concentrations. Furthermore, lutein was found to preferentially accumulate in the infant brain in comparison to other carotenoids that are predominant in diet. While lutein is consistently related to cognitive function, the mechanisms by which lutein may influence cognition are not clear. In an effort to identify potential mechanisms through which lutein might influence neurodevelopment, an exploratory study relating metabolite signatures and lutein was completed. Post-mortem metabolomic analyses were performed on human infant brain tissues in three regions important for learning and memory: the frontal cortex, hippocampus, and occipital cortex. Metabolomic profiles were compared to lutein concentration, and correlations were identified and reported here. A total of 1276 correlations were carried out across all brain regions. Of 427 metabolites analyzed, 257 were metabolites of known identity. Unidentified metabolite correlations (510) were excluded. In addition, moderate correlations with xenobiotic relationships (2) or those driven by single outliers (3) were excluded from further study. Lutein concentrations correlated with lipid pathway metabolites, energy pathway metabolites, brain osmolytes, amino acid neurotransmitters, and the antioxidant homocarnosine. These correlations were often brain region—specific. Revealing relationships between lutein and metabolic pathways may help identify potential candidates on which to complete further analyses and may shed light on important roles of lutein in the human brain during development.</p></div

Mean (±SEM) carotenoid concentrations (ng/mg protein) in different regions of the brain from (A) stock diet-fed (n = 9) and (B) lutein/zeaxanthin (L/Z) supplemented (n = 4) adult rhesus macaques.

<p>*Lutein was significantly greater than all other carotenoids within each brain region. Bars with different superscripts across brain regions for each individual carotenoid are significantly different according to Tukey’s HSD test (P<0.05).</p

Lysophospholipid correlations with lutein in human infant brain.^a

<p>^aSubject ages ranged from 1 to 488 days old, and the median age was 100.5 days. 30 total decedents were studied and included both male (n = 21) and female (n = 9) infants. A total of 81 tissues were analyzed from three brain regions, including the frontal cortex (FC, n = 29), hippocampus (HPC, n = 24), and the occipital cortex (OC, n = 28).</p><p>^bAll values are significant (P < 0.05). Lysophospholipid metabolites:lutein correlations with r values ≥ |0.45| and P < 0.05 are reported.</p><p>Lysophospholipid correlations with lutein in human infant brain.<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0136904#t003fn001" target="_blank">^a</a></p

Rhesus monkey characteristics (mean ± SEM) in stock diet and L/Z supplemented treatment groups.

<p>Rhesus monkey characteristics (mean ± SEM) in stock diet and L/Z supplemented treatment groups.</p