Protein Ingredient Quality within Infant Formulas Impacts Plasma Amino Acid Concentrations in Neonatal Minipiglets.

BACKGROUND: Infant formulas (IFs), the only adequate substitute to human milk, are complex matrices that require numerous ingredients and processing steps that may impact protein digestion and subsequent amino acid (AA) absorption. OBJECTIVES: The objective was to understand the impact of the protein ingredient quality within IFs on postprandial plasma AA profiles. METHODS: Four isonitrogenous and isocaloric IFs were produced at a semi-industrial scale using whey proteins from different origins (cheese compared with ideal whey) and denaturation levels (IF-A, -B, -C), and caseins with different supramolecular organizations (IF-C, -D). Ten Yucatan minipiglets (12- to 27-d-old) were used as a human infant model and received each IF for 3 d according to a Williams Latin square followed by a 2-d wash-out period. Jugular plasma was regularly sampled from 10 min preprandial to 4 h postprandial on the third day to measure free AAs, urea, insulin, and glucose concentrations. Data were statistically analyzed using a mixed linear model with diet (IFs), time, and sex as fixed factors and piglet as random factor. RESULTS: IFs made with cheese whey (IF-A and -B) elicited significantly higher plasma total and essential AA concentrations than IFs made with ideal whey (IF-C and -D), regardless of the pre- and postprandial times. Most of the differences observed postprandially were explained by AA homeostasis modifications. IFs based on cheese whey induced an increased plasma concentration of Thr due to both a higher Thr content in these IFs and a Thr-limiting degrading capability in piglets. The use of a nonmicellar casein ingredient led to reduced plasma content of AA catabolism markers (IF-D compared with IF-C). CONCLUSIONS: Overall, our results highlight the importance of the protein ingredient quality (composition and structure) within IFs on neonatal plasma AA profiles, which may further impact infant protein metabolism.

Obesogenic diet leads to luminal overproduction of the complex IV inhibitor H2 S and mitochondrial dysfunction in mouse colonocytes.

Obesity is characterized by systemic low-grade inflammation associated with disturbances of intestinal homeostasis and microbiota dysbiosis. Mitochondrial metabolism sustains epithelial homeostasis by providing energy to colonic epithelial cells (CEC) but can be altered by dietary modulations of the luminal environment. Our study aimed at evaluating whether the consumption of an obesogenic diet alters the mitochondrial function of CEC in mice. Mice were fed for 22 weeks with a 58% kcal fat diet (diet-induced obesity [DIO] group) or a 10% kcal fat diet (control diet, CTRL). Colonic crypts were isolated to assess mitochondrial function while colonic content was collected to characterize microbiota and metabolites. DIO mice developed obesity, intestinal hyperpermeability, and increased endotoxemia. Analysis of isolated colonic crypt bioenergetics revealed a mitochondrial dysfunction marked by decreased basal and maximal respirations and lower respiration linked to ATP production in DIO mice. Yet, CEC gene expression of mitochondrial respiration chain complexes and mitochondrial dynamics were not altered in DIO mice. In parallel, DIO mice displayed increased colonic bile acid concentrations, associated with higher abundance of Desulfovibrionaceae. Sulfide concentration was markedly increased in the colon content of DIO mice. Hence, chronic treatment of CTRL mouse colon organoids with sodium sulfide provoked mitochondrial dysfunction similar to that observed in vivo in DIO mice while acute exposure of isolated mitochondria from CEC of CTRL mice to sodium sulfide diminished complex IV activity. Our study provides new insights into colon mitochondrial dysfunction in obesity by revealing that increased sulfide production by DIO-induced dysbiosis impairs complex IV activity in mouse CEC.

Exposure to a sensory functional ingredient in the pig model modulates the blood-oxygen-level dependent brain responses to food odor and acute stress during pharmacological MRI in the frontostriatal and limbic circuits.

Introduction: In the present study, we examined the effects of a supplementation with a sensory functional ingredient (FI, D16729, Phodé, France) containing vanillin, furaneol, diacetyl and a mixture of aromatic fatty acids on the behavioural and brain responses of juvenile pigs to acute stress. Methods: Twenty-four pigs were fed from weaning with a standard granulated feed supplemented with the functional ingredient D16729 (FS animals, N = 12) or a control formulation (CT animals, N = 12). After a feed transition (10 days after weaning), the effects of FI were investigated on eating behaviour during two-choice feed preference tests. Emotional reactivity to acute stress was then investigated during openfield (OF), novel suddenly moving object (NSO), and contention tests. Brain responses to the FI and the two different feeds' odour, as well as to an acute pharmacological stressor (injection of Synacthen®) were finally investigated with functional magnetic resonance imaging (fMRI). Results: FS animals tended to spend more time above the functional feed (p = 0.06) and spent significantly more time at the periphery of the arena during NSO (p < 0.05). Their latency to contact the novel object was longer and they spent less time exploring the object compared to CT animals (p < 0.05 for both). Frontostriatal and limbic responses to the FI were influenced by previous exposure to FI, with higher activation in FS animals exposed to the FI feed odor compared to CT animals exposed to a similarly familiar feed odor without FI. The pharmacological acute stress provoked significant brain activations in the prefrontal and thalamic areas, which were alleviated in FS animals that also showed more activity in the nucleus accumbens. Finally, the acute exposure to FI in naive animals modulated their brain responses to acute pharmacological stress. Discussion: Overall, these results showed how previous habituation to the FI can modulate the brain areas involved in food pleasure and motivation while alleviating the brain responses to acute stress.

The ghrelin system follows a precise post-natal development in mini-pigs that is not impacted by dietary medium chain fatty-acids.

The ghrelin-ghrelin receptor (GHSR1) system is one of the most important mechanisms regulating food intake and energy balance. To be fully active, ghrelin is acylated with medium-chain fatty acids (MCFA) through the ghrelin-O-acetyl transferase (GOAT). Several studies reported an impact of dietary MCFA on ghrelin acylation in adults. Our study aimed at describing early post-natal development of the ghrelin system in mini-pigs as a model of human neonates and evaluating the impact of dietary MCFA. Suckled mini-pigs were sacrificed at post-natal day (PND) 0, 2, 5, and 10 or at adult stage. In parallel, other mini-pigs were fed from birth to PND10 a standard or a dairy lipid-enriched formula with increased MCFA concentration (DL-IF). Plasma ghrelin transiently peaked at PND2, with no variation of the acylated fraction except in adults where it was greater than during the neonatal period. Levels of mRNA coding pre-proghrelin (GHRL) and GOAT in the antrum did not vary during the post-natal period but dropped in adults. Levels of antral pcsk1/3 (cleaving GHRL into ghrelin) mRNA decreased significantly with age and was negatively correlated with plasma acylated, but not total, ghrelin. Hypothalamic ghsr1 mRNA did not vary in neonates but increased in adults. The DL-IF formula enriched antral tissue with MCFA but did not impact the ghrelin system. In conclusion, the ghrelin maturation enzyme PCSK1/3 gene expression exhibited post-natal modifications parallel to transient variations in circulating plasma ghrelin level in suckling piglets but dietary MCFA did not impact this post-natal development.

Regular exposure to a Citrus-based sensory functional food ingredient alleviates the BOLD brain responses to acute pharmacological stress in a pig model of psychosocial chronic stress.

Psychosocial chronic stress is a critical risk factor for the development of mood disorders. However, little is known about the consequences of acute stress in the context of chronic stress, and about the related brain responses. In the present study we examined the physio-behavioural effects of a supplementation with a sensory functional food ingredient (FI) containing Citrus sinensis extract (D11399, Phodé, France) in a pig psychosocial chronic stress model. Female pigs underwent a 5- to 6-week stress protocol while receiving daily the FI (FI, n = 10) or a placebo (Sham, n = 10). We performed pharmacological magnetic resonance imaging (phMRI) to study the brain responses to an acute stress (injection of Synacthen®, a synthetic ACTH-related agonist) and to the FI odour with or without previous chronic supplementation. The olfactory stimulation with the ingredient elicited higher brain responses in FI animals, demonstrating memory retrieval and habituation to the odour. Pharmacological stress with Synacthen injection resulted in an increased activity in several brain regions associated with arousal, associative learning (hippocampus) and cognition (cingulate cortex) in chronically stressed animals. This highlighted the specific impact of acute stress on the brain. These responses were alleviated in animals previously supplemented by the FI during the entire chronic stress exposure. As chronic stress establishes upon the accumulation of acute stress events, any attenuation of the brain responses to acute stress can be interpreted as a beneficial effect, suggesting that FI could be a viable treatment to help individuals coping with repeated stressful events and eventually to reduce chronic stress. This study provides additional evidence on the potential benefits of this FI, of which the long-term consequences in terms of behaviour and physiology need to be further investigated.

fMRI-Based Brain Responses to Olfactory Stimulation with Two Putatively Orexigenic Functional Food Ingredients at Two Different Concentrations in the Pig Model.

Natural plant extracts are increasingly used as functional feed ingredients in animal husbandry and food ingredients in human alternative medicine to improve welfare and health. We investigated in 20 growing pigs via functional magnetic resonance imaging (fMRI) the brain blood oxygen level-dependent (BOLD) responses to olfactory stimulation with two sensory functional feed ingredients, A and B, at two different concentrations. Functional ingredient A contained extracts from Citrus sinensis (60% to 80%), and ingredient B contained a mixture of extracts Oreganum vulgarae (40% to 55%) and Cymbopogon flexuosus (20% to 25%). Increased concentration of ingredients induced a higher activation in reward and cognitive areas compared to lower concentrations. Moreover, considering both ingredients at the highest concentration, the ingredient A elicited higher brain responses in brain areas involved in hedonism/pleasantness compared to ingredient B, and more specifically in the caudate nucleus and orbitofrontal cortex. Our findings shed new light in the scope of emotion regulation through olfactory modulation via sensory functional ingredients, which opens the way to further preclinical studies in animal models and translational research in the context of nutrition, welfare, and health. PRACTICAL APPLICATION: Functional food/feed ingredients are gaining interest for improving health and welfare in humans and animals. Besides representing an alternative to antibiotics for example, food ingredients and their sensory characteristics might have a positive impact on emotions and consequently on well-being. Functional brain imaging in large animals such as in the pig model is a promising approach to investigate the central and behavioural effects of food ingredients, and determine the most effective blends and concentrations to modulate internal and emotional states.

Locomotion and eating behavior changes in Yucatan minipigs after unilateral radio-induced ablation of the caudate nucleus.

The functional roles of the Caudate nucleus (Cd) are well known. Selective Cd lesions can be found in neurological disorders. However, little is known about the dynamics of the behavioral changes during progressive Cd ablation. Current stereotactic radiosurgery technologies allow the progressive ablation of a brain region with limited adverse effects in surrounding normal tissues. This could be of high interest for the study of the modified behavioral functions in relation with the degree of impairment of the brain structures. Using hypofractionated stereotactic radiotherapy combined with synchrotron microbeam radiation, we investigated, during one year after irradiation, the effects of unilateral radio-ablation of the right Cd on the behavior of Yucatan minipigs. The right Cd was irradiated to a minimal dose of 35.5 Gy delivered in three fractions. MRI-based morphological brain integrity and behavioral functions, i.e. locomotion, motivation/hedonism were assessed. We detected a progressive radio-necrosis leading to a quasi-total ablation one year after irradiation, with an additional alteration of surrounding areas. Transitory changes in the motivation/hedonism were firstly detected, then on locomotion, suggesting the influence of different compensatory mechanisms depending on the functions related to Cd and possibly some surrounding areas. We concluded that early behavioral changes related to eating functions are relevant markers for the early detection of ongoing lesions occurring in Cd-related neurological disorders.

fMRI-Based Brain Responses to Quinine and Sucrose Gustatory Stimulation for Nutrition Research in the Minipig Model: A Proof-of-Concept Study.

The minipig model is of high interest for brain research in nutrition and associated pathologies considering the similarities to human nutritional physiology, brain structures, and functions. In the context of a gustatory stimulation paradigm, fMRI can provide crucial information about the sensory, cognitive, and hedonic integration of exteroceptive stimuli in healthy and pathological nutritional conditions. Our aims were (i) to validate the experimental setup, i.e., fMRI acquisition and SPM-based statistical analysis, with a visual stimulation; (ii) to implement the fMRI procedure in order to map the brain responses to different gustatory stimulations, i.e., sucrose (5%) and quinine (10 mM), and (ii) to investigate the differential effects of potentially aversive (quinine) and appetitive/pleasant (sucrose) oral stimulation on brain responses, especially in the limbic and reward circuits. Six Yucatan minipigs were imaged on an Avanto 1.5-T MRI under isoflurane anesthesia and mechanical ventilation. BOLD signal was recorded during visual or gustatory (artificial saliva, sucrose, or quinine) stimulation with a block paradigm. With the visual stimulation, brain responses were detected in the visual cortex, thus validating our experimental and statistical setup. Quinine and sucrose stimulation promoted different cerebral activation patterns that were concordant, to some extent, to results from human studies. The insular cortex (i.e., gustatory cortex) was activated with both sucrose and quinine, but other regions were specifically activated by one or the other stimulation. Gustatory stimulation combined with fMRI analysis in large animals such as minipigs is a promising approach to investigate the integration of gustatory stimulation in healthy or pathological conditions such as obesity, eating disorders, or dysgeusia. To date, this is the first intent to describe gustatory stimulation in minipigs using fMRI.