Dietary Oligosaccharides Attenuate Stress-Induced Disruptions in Immune Reactivity and Microbial B-Vitamin Metabolism.

Background: Exposure to stressful stimuli dysregulates inflammatory processes and alters the gut microbiota. Prebiotics, including long-chain fermentable fibers and milk oligosaccharides, have the potential to limit inflammation through modulation of the gut microbiota. To determine whether prebiotics attenuate stress-induced inflammation and microbiota perturbations, mice were fed either a control diet or a diet supplemented with galactooligosaccharides, polydextrose and sialyllactose (GOS+PDX+SL) or sialyllactose (SL) for 2 weeks prior to and during a 6-day exposure to a social disruption stressor. Spleens were collected for immunoreactivity assays. Colon contents were examined for stressor- and diet- induced changes in the gut microbiome and metabolome through 16S rRNA gene sequencing, shotgun metagenomic sequencing and UPLC-MS/MS. Results: Stress increased circulating IL-6 and enhanced splenocyte immunoreactivity to an ex vivo LPS challenge. Diets containing GOS+PDX+SL or SL alone attenuated these responses. Stress exposure resulted in large changes to the gut metabolome, including robust shifts in amino acids, peptides, nucleotides/nucleosides, tryptophan metabolites, and B vitamins. Multiple B vitamins were inversely associated with IL-6 and were augmented in mice fed either GOS+PDX+SL or SL diets. Stressed mice exhibited distinct microbial communities with lower abundances of Lactobacillus spp. and higher abundances of Bacteroides spp. Diet supplementation with GOS+PDX+SL, but not SL alone, orthogonally altered the microbiome and enhanced the growth of Bifidobacterium spp. Metagenome-assembled genomes (MAGs) from mice fed the GOS+PDX+SL diet unveiled genes in a Bifidobacterium MAG for de novo B vitamin synthesis. B vitamers directly attenuated the stressor-induced exacerbation of cytokine production in LPS-stimulated splenocytes. Conclusions: Overall, these data indicate that colonic metabolites, including B vitamins, are responsive to psychosocial stress. Dietary prebiotics reestablish colonic B vitamins and limit stress-induced inflammation.

Early life diet containing prebiotics and bioactive whey protein fractions increased dendritic spine density of rat hippocampal neurons.

Early life nutrition plays an important role in brain development. Emerging research in rodents, piglets and humans suggest that prebiotics, milk fat globule membrane and lactoferrin may each play unique roles in brain development and cognitive functions. However, knowledge of their combined impact is lacking. We show here that providing weanling rats with a diet containing milk fat globule membrane, lactoferrin and a polydextrose/galactooligosaccharide prebiotic blend led to a significant increase in total dendritic spine density in hippocampal dentate gyrus neurons. Region-specific alterations in dendritic spine density and morphology could provide a mechanistic basis underlying broader cognitive benefits, but further research is required to demonstrate functional consequences of these observations.

Dietary Alpha-Lipoic Acid Alters Piglet Neurodevelopment.

INTRODUCTION: Alpha-lipoic acid (a-LA) is an antioxidant shown to ameliorate age-associated impairments of brain and cardiovascular function. Human milk is known to have high antioxidant capacity; however, the role of antioxidants in the developing brain is largely uncharacterized. This exploratory study aimed to examine the dose-response effects of a-LA on piglet growth and neurodevelopment. METHODS: Beginning at 2 days of age, 31 male pigs received 1 of 3 diets: control (CONT) (0 mg a-LA/100 g), low a-LA (LOW) (120 mg a-LA/100 g), or high a-LA (HIGH) (240 mg a-LA/100 g). From 14 to 28 days of age, pigs were subjected to spatial T-maze assessment, and macrostructural and microstructural neuroimaging procedures were performed at 31 days of age. RESULTS: No differences due to diet were observed for bodyweight gain or intestinal weight and length. Spatial T-maze assessment did not reveal learning differences due to diet in proportion of correct choices or latency to choice measures. Diffusion tensor imaging revealed decreased (P = 0.01) fractional anisotropy (FA) in the internal capsule of HIGH-fed pigs compared with both the CONT (P < 0.01)- and LOW (P = 0.03)-fed pigs, which were not different from one another. Analysis of axial diffusivity (AD) within the internal capsule revealed a main effect of diet (P < 0.01) in which HIGH-fed piglets exhibited smaller (P < 0.01) rates of diffusion compared with CONT piglets, but HIGH-fed piglets were not different (P = 0.12) than LOW-fed piglets. Tract-based spatial statistics, a comparison of FA values along white matter tracts, revealed 1,650 voxels where CONT piglets exhibited higher (P < 0.05) values compared with HIGH-fed piglets. CONCLUSION: The lack of differences in intestinal and bodyweight measures among piglets indicate a-LA supplementation does not impact overall growth, regardless of concentration. Additionally, no observed differences between CONT- and LOW-fed piglets in behavior and neuroimaging measures indicate a low concentration of a-LA does not affect normal brain development. Supplementation of a-LA at a high concentration appeared to alter white matter maturation in the internal capsule, which may indicate delayed neurodevelopment in these piglets.

Dietary Isomers of Sialyllactose Increase Ganglioside Sialic Acid Concentrations in the Corpus Callosum and Cerebellum and Modulate the Colonic Microbiota of Formula-Fed Piglets.

BACKGROUND: Sialyllactose is a key human milk oligosaccharide and consists of sialic acid (SA) bound to a lactose molecule. Breastfed infants have increased accumulation of ganglioside-bound SA compared with formula-fed infants. OBJECTIVE: This study aimed to determine whether different isomers of sialyllactose enrich brain SA and modulate the microbiome of developing neonatal piglets. METHODS: Day-old pigs were randomly allocated to 6 diets (control, 2 or 4 g 3'-sialyllactose/L, 2 or 4 g 6'-sialyllactose/L, or 2 g polydextrose/L + 2 g galacto-oligosaccharides/L; n = 9) and fed 3 times/d for 21 d. Pigs were killed, and the left hemisphere of the brain was dissected into cerebrum, cerebellum, corpus callosum, and hippocampus regions. SA was determined by using a modified periodic acid-resorcinol reaction. Microbial composition of the intestinal digesta was analyzed with the use of 16S ribosomal DNA Illumina sequencing. RESULTS: Dietary sialyllactose did not affect feed intake, growth, or fecal consistency. Ganglioside-bound SA in the corpus callosum of pigs fed 2 g 3'-sialyllactose or 6'-sialyllactose/L increased by 15% in comparison with control pigs. Similarly, ganglioside-bound SA in the cerebellum of pigs fed 4 g 3'-sialyllactose/L increased by 10% in comparison with control pigs. Significant (P < 0.05, Adonis Test) microbiome differences were observed in the proximal and distal colons of piglets fed control compared with 4-g 6'-sialyllactose/L formulas. Differences were attributed to an increase in bacterial taxa belonging to species Collinsella aerofaciens (phylum Actinobacteria), genera Ruminococcus and Faecalibacterium (phylum Firmicutes), and genus Prevotella (phylum Bacteroidetes) (Wald test, P < 0.05, DeSeq2) compared with piglets fed the control diet. Taxa belonging to families Enterobacteriaceae and Enterococcaceae (phylum Proteobacteria), as well as taxa belonging to family Lachnospiraceae and order Lactobacillales (phylum Firmicutes), were 2.3- and 4-fold lower, respectively, in 6'-sialyllactose-fed piglets than in controls. CONCLUSIONS: Supplementation of formula with 3'- or 6'-sialyllactose can enrich ganglioside SA in the brain and modulate gut-associated microbiota in neonatal pigs. We propose 2 potential routes by which sialyllactose may positively affect the neonate: serving as a source of SA for neurologic development and promoting beneficial microbiota.

Effect of perinatally supplemented flavonoids on brain structure, circulation, cognition, and metabolism in C57BL/6J mice.

Evidence suggests that flavanol consumption can beneficially affect cognition in adults, but little is known about the effect of flavanol intake early in life. The present study aims to assess the effect of dietary flavanol intake during the gestational and postnatal period on brain structure, cerebral blood flow (CBF), cognition, and brain metabolism in C57BL/6J mice. Female wild-type C57BL/6J mice were randomly assigned to either a flavanol supplemented diet or a control diet at gestational day 0. Male offspring remained on the corresponding diets throughout life and performed cognitive and behavioral tests during puberty and adulthood assessing locomotion and exploration (Phenotyper and open field), sensorimotor integration (Rotarod and prepulse inhibition), and spatial learning and memory (Morris water maze, MWM). Magnetic resonance spectroscopy and imaging at 11.7T measured brain metabolism, CBF, and white and gray matter integrity in adult mice. Biochemical and immunohistochemical analyses evaluated inflammation, synaptic plasticity, neurogenesis, and vascular density. Cognitive and behavioral tests demonstrated increased locomotion in Phenotypers during puberty after flavanol supplementation (p = 0.041) but not in adulthood. Rotarod and prepulse inhibition demonstrated no differences in sensorimotor integration. Flavanols altered spatial learning in the MWM in adulthood (p = 0.039), while spatial memory remained unaffected. Additionally, flavanols increased diffusion coherence in the visual cortex (p = 0.014) and possibly the corpus callosum (p = 0.066) in adulthood. Mean diffusion remained unaffected, a finding that corresponds with our immunohistochemical data showing no effect on neurogenesis, synaptic plasticity, and vascular density. However, flavanols decreased CBF in the cortex (p = 0.001) and thalamus (p = 0.009) in adulthood. Brain metabolite levels and neuroinflammation remained unaffected by flavanols. These data suggest that dietary flavanols results in subtle alterations in brain structure, locomotor activity and spatial learning. Comparison of these data to published findings in aging or neurodegeneration suggests that benefits of dietary flavanols may increase with advancing age and in disease.

Impact of dietary n-3 polyunsaturated fatty acids on cognition, motor skills and hippocampal neurogenesis in developing C57BL/6J mice.

Maternal intake of omega-3 polyunsaturated fatty acids (n-3 PUFA) is critical during perinatal development of the brain. Docosahexaenoic acid (DHA) is the most abundant n-3 PUFA in the brain and influences neuronal membrane function and neuroprotection. The present study aims to assess the effect of dietary n-3 PUFA availability during the gestational and postnatal period on cognition, brain metabolism and neurohistology in C57BL/6J mice. Female wild-type C57BL/6J mice at day 0 of gestation were randomly assigned to either an n-3 PUFA deficient diet (0.05% of total fatty acids) or an n-3 PUFA adequate diet (3.83% of total fatty acids) containing preformed DHA and its precursor α-linolenic acid. Male offspring remained on diet and performed cognitive tests during puberty and adulthood. In adulthood, animals underwent (31)P magnetic resonance spectroscopy to assess brain energy metabolites. Thereafter, biochemical and immunohistochemical analyses were performed assessing inflammation, neurogenesis and synaptic plasticity. Compared to the n-3 PUFA deficient group, pubertal n-3 PUFA adequate fed mice demonstrated increased motor coordination. Adult n-3 PUFA adequate fed mice exhibited increased exploratory behavior, sensorimotor integration and spatial memory, while neurogenesis in the hippocampus was decreased. Selected brain regions of n-3 PUFA adequate fed mice contained significantly lower levels of arachidonic acid and higher levels of DHA and dihomo-γ-linolenic acid. Our data suggest that dietary n-3 PUFA can modify neural maturation and enhance brain functioning in healthy C57BL/6J mice. This indicates that availability of n-3 PUFA in infant diet during early development may have a significant impact on brain development.

alpha-Tocopherol and selenium facilitate recovery from lipopolysaccharide-induced sickness in aged mice.

The elderly suffer a decline in immune function that increases their vulnerability to infections. Because antioxidants improve some age-related deficits in immune and cognitive function, our goal was to determine whether dietary alpha-tocopherol (alpha-T) and selenium inhibit LPS-induced sickness behavior in aged mice. Male BALB/c mice were fed modified AIN93-M diets that were low, adequate, or high in both alpha-T (10, 75, or 500 mg/kg) and selenium (0.05, 0.15, or 2 mg/kg) from 18 to 21 mo of age. Sickness was quantified by measuring time in social exploration of a novel juvenile conspecific. The lipopolysaccharide treatment reduced social exploration by 74% at 2 h, regardless of diet. By 4 h, aged mice fed the low diet were 88% less social, whereas mice fed the adequate and high diets displayed only approximately 40% reductions due to LPS treatment. Mice fed the low diet had greater LPS-induced weight loss than mice fed the high diet. Plasma alpha-T concentration and glutathione peroxidase (GPX) activity increased with each increment in alpha-T and selenium 24 h post-LPS treatment. Brain alpha-T concentration and GPX activity were lower in mice fed the low diet than in those fed the adequate or high diet. Regardless of diet, interleukin (IL)-6, IL-1beta, and tumor necrosis factor (TNF)alpha mRNA levels were elevated by LPS approximately 3-fold in cortex, cerebellum, striatum, and hippocampus. Thus, antioxidants inhibit sickness behavior independently of IL-6, IL-1beta, and TNFalpha mRNA levels 2 h post-LPS in the brain regions analyzed. Taken together, these findings suggest that adequate intake of dietary alpha-T and selenium may help promote recovery from gram-negative bacterial infection in the aged.

Alpha-tocopherol attenuates lipopolysaccharide-induced sickness behavior in mice.

Antioxidants protect cells from oxidative damage and reduce lipopolysaccharide (LPS)-induced expression of inflammatory cytokines. Because inflammatory cytokines induce sickness behavior, we hypothesized that antioxidants, namely alpha-tocopherol (alpha-T) and selenium would inhibit sickness behavior caused by LPS. In an initial study, mice were injected intraperitoneal (i.p.) with vehicle, 2, or 20mg alpha-T for 3 consecutive days and then challenged with vehicle, 1, 10, or 100 microg of LPS. Sickness behavior was quantified by measuring social exploratory behavior. Vehicle pretreated mice injected with LPS showed a marked reduction in social behavior at 4h (p < .01). However, sickness behavior induced by the lowest dose of LPS was partially or completely blocked by 2 or 20mg alpha-T, respectively. alpha-T did not prevent sickness from higher doses of LPS. In a second study, mice were fed AIN93-M modified diets containing 10, 75, and 500 mg alpha-T/kg and 0.05, 0.15, and 2mg selenium/kg for 8 weeks and then challenged with saline or LPS (1 microg). The highest concentration of dietary alpha-T and selenium tended (p = .1) to reduce LPS-induced sickness behavior. Mice fed diets low in antioxidants had reduced plasma alpha-T levels and glutathione peroxidase activity (p = .08 and p < .01, respectively) and elevated liver thiobarbituric acid reactive substances (p < .001) 24h post LPS. Collectively, these data indicate that alpha-T improved the oxidative status after exposure to LPS, which may explain its ability to ameliorate symptoms of sickness.