Manuel Oliveira

(+)-Catechin is more bioavailable than (−)-catechin: Relevance to the bioavailability of catechin from cocoa

Catechin is a flavonoid present in fruits, wine and cocoa products. Most foods contain the (+)-enantiomer of catechin but chocolate mainly contains ( − )-catechin, in addition to its major flavanol, ( − )-epicatechin. Previous studies have shown poor bioavailability of catechin when consumed in chocolate. We compared the absorption of ( − ) and (+)-catechin after in situ perfusion of 10, 30 or 50 μmol/l of each catechin enantiomer in the jejunum and ileum in the rat. We also assayed 23 samples of chocolate for (+) and ( − )-catechin. Samples were analyzed using HPLC with a Cyclobond I-2000 RSP chiral column. At all concentrations studied, the intestinal absorption of ( − )-catechin was lower than the intestinal absorption of (+)-catechin (p < 0.01). Plasma concentrations of ( − )-catechin were significantly reduced compared to (+)-catechin (p < 0.05). The mean concentration of ( − )-catechin in chocolate was 218 ± 126 mg/kg compared to 25 ± 15 mg/kg (+)-catechin. Our findings provide an explanation for the poor bioavailability of catechin when consumed in chocolate or other cocoa containing products.

Lactoferrin reduces in vitro osteoclast differentiation and resorbing activity.

Lactoferrin (LF) is a key modulator of inflammatory response. Since bone and immune systems are genetically and functionally linked, we were interested to know if LF could influence bone remodeling. Bovine LF (bLF) inhibited in vitro bone resorbing activity (IC50, 200 microg/ml) in a rabbit mixed bone cell culture, consisting of authentic osteoclasts in an environment of osteoblast and stromal cells. Using human CD14 selected cells committed toward osteoclasts, bLF (10 microg/ml) stimulated cell proliferation, however, led to an inhibition of calcitonin receptor mRNA expression, a main marker of osteoclast phenotype, and decreased the global resorbing activity. No modulation of RANK mRNA expression was observed and mRNA for RANKL and OPG were not detected in this culture system, suggesting that bLF inhibits osteoclastogenesis and reduces bone resorption through a mechanism independent of OPG/RANKL/RANK. In conclusion, bLF appears to modulate bone remodeling. Its mechanism of action remains to be elucidated.

Influence of gut microbiota on mouse B2 B cell ontogeny and function

A complex interplay between the microbiota and the host immune system is evidenced to shape the immune system throughout life, but little is known about the microbial effect on key players of the adaptive immune system, the B2 B cells. In the presented study, we have evaluated the effect of commensal bacteria on B cell ontogeny and function, with the focus on B2 B cells of spleen and Peyers patches. We have compared germ-free mice to mice that are exposed to a normal complex bacterial community from the day of birth and combined classical immunological assessment with advanced genome-wide expression profiling. Despite a preservation of all B cell subsets and phenotype, our results show that microbiota strongly impact mucosal B cell physiology and lead to higher serum Ig concentrations. We show that this microbial influence comprises downregulation of transcription factors involved in early B cell activation steps and upregulation of genes and proteins involved in later stages of B cell response. In summary, we show an influence of the gut microbiota on function of mucosal B2 B cells, involving mechanisms downstream of B cell activation and proliferation.

Benefits of Structured and Free Monoacylglycerols to Deliver Eicosapentaenoic (EPA) in a Model of Lipid Malabsorption

In the present study, we used a preclinical model of induced lipolytic enzyme insufficiency, and hypothesized that the use of monoacylglycerols (MAG) will enhance their bioavailability and delivery to the tissues. Experimental diets containing 20% lipids were fed to rats for 21 days with or without Orlistat. The control diet of fish oil (FO), a source of EPA and DHA, was tested against: structured (A) vanillin acetal of sn-2 MAG (Vanil + O) and (B) diacetyl derivative of sn-2 MAG (Acetyl + O) and (C) free MAG (MAG + O). FA profiles with an emphasis on EPA and DHA levels were determined in plasma, red blood cells (RBC), liver, spleen, brain and retina. We observed significant reduction of lipid absorption when rats co-consumed Orlistat. As expected, the FO groups with and without Orlistat showed the biggest difference. The Vanil + O, Acetyl + O and MAG + O groups, demonstrated higher levels of EPA (5.5 ± 1.9, 4.6 ± 1.6 and 5.6 ± 0.6, respectively) in RBC compared with FO + O diets (3.3 ± 0.2, 2.6 ± 0.2). Levels of EPA incorporation, in plasma, were similar to those obtained for RBC, and similar trends were observed for the collected tissues and even with DHA levels. These observations with two MAG derivatives providing the fatty acid esterified in the sn-2 position, show that these molecules are efficient vehicles of EPA in malabsorption conditions which is in line with our hypothesis. Free MAG, characterized as having exclusively sn-1(3) isomers of EPA, demonstrated better absorption efficiencies and accretion to tissues when compared to structured MAG. The study demonstrated that structured and free MAG can be used efficiently as an enteral vehicle to supply bioactive fatty acids such as EPA and DHA in lipid malabsorption where diminished lipolytic activity is the underlying cause.

Glucuronidation of the Green Tea Catechins, (-)-epigallocatechin-3-gallate and (-)-epicatechin-3-gallate, by Rat Hepatic and Intestinal Microsomes

The flavonoids (-)-epigallocatechin-3-gallate (EGCg) and (-)-epicatechin-3-gallate (ECg) are major components of green tea and show numerous biological effects. We investigated the glucuronidation of these compounds and of quercetin by microsomes. Quercetin was almost fully glucuronidated by liver microsomes after 3 h, whereas ECg and ECGg were conjugated to a lesser extent ([Formula: See Text] and [Formula: See Text] respectively). The intestinal microsomes also glucuronidated quercetin much more efficiently than ECg and EGCg. Although the rates were lower than quercetin, intestinal microsomes exhibited higher activity on the galloyl group of ECg and EGCg compared to the flavonoid ring, whereas hepatic glucuronidation was higher on the flavonoid ring of EGCg and ECg compared to the galloyl groups. The low glucuronidation rates could partially explain why these flavanols are present in plasma as unconjugated forms.

Protective effects of dietary EPA and DHA on ischemia–reperfusion-induced intestinal stress

The immunoregulatory effects of dietary omega-3 fatty acids are still not fully characterized. The aim of this study was to determine whether dietary eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) intake limits intestinal ischemia-reperfusion (IR) injury. To test this, rats were fed either control or EPA/DHA supplemented diet for 3 weeks following which they underwent either a sham or an IR surgical protocol. A significant reduction in mucosal damage was observed after EPA/DHA supplemented diet as reflected by maintenance of total protein content. To address the underlying mechanisms of protection, we measured parameters of oxidative stress, intestinal and serological cytokines and intestinal eicosanoids. Interestingly, EPA/DHA fed animals displayed a higher activity of oxidative stress enzyme machinery, i.e., superoxide dismutase and catalase in addition to a reduction in total nitrate/nitrite content. While no changes in cytokines were observed, eicosanoid analyses of intestinal tissue revealed an increase in metabolites of the 12-lipoxygenase pathway following IR. Further, IR in EPA/DHA fed animals was accompanied by a significant increase of 17,18-epoxyeicosatetraenoic acid, 8-Iso prostaglandin F(3α) and thromboxane B(3), by more than 12-, 6-, 3-fold, respectively. Thus, the data indicate that EPA/DHA supplementation may be able to reduce early intestinal IR injury by anti-oxidative and anti-inflammatory mechanisms.

Lactobacillus paracasei Reduces Intestinal Inflammation in Adoptive Transfer Mouse Model of Experimental Colitis

Studies showed that specific probiotics provide therapeutic benefits in inflammatory bowel disease. In vitro evidence suggested that Lactobacillus paracasei also called ST11 (CNCM I-2116) is a potent strain with immune modulation properties. However, little is known about its capacity to alleviate inflammatory symptoms in vivo In this context, the main objective of this study was to investigate the role of ST11 on intestinal inflammation using the adoptive transfer mouse model of experimental colitis. Rag2−/− recipient mice were fed with ST11 (109 CFU/day)a month prior toinduce colitis by adoptive transfer of naive T cells. One month later, in clear contrast to nonfed mice, weight loss was significantly reduced by 50% in ST11-fed mice. Further analysis of colon specimens revealed a significant reduction neutrophil infiltration and mucosal expression of IL1β, IL-6, and IL12 proinflammatory cytokines, whereas no consistent differences in expression of antibacterial peptides or tight junction proteins were observed between PBS and ST11-fed mice. All together, our results demonstrate that oral administration of ST11 was safe and had a significant preventive effect on colitis. We conclude that probiotics such as Lactobacillus paracasei harbor worthwhile in vivo immunomodulatory properties to prevent intestinal inflammation by nutritional approaches.

Lipase inhibitor orlistat decreases incorporation of eicosapentaenoic and docosahexaenoic acids in rat tissues.

Orlistat is a gastric and pancreatic lipases inhibitor that is often prescribed to obese subjects. Orlistat has been shown to decrease the absorption of biologically important lipophilic micronutrients such as liposoluble vitamins. We hypothesized that long-term administration of orlistat may lower the incorporation of n-3 long-chain polyunsaturated fatty acids (LC-PUFA) in blood lipids and tissues. This hypothesis was tested in rats fed a diet supplemented with fish oil as a source of n-3 LC-PUFA. Male Wistar rats (n = 18) were divided into 3 groups and fed experimental high-fat diets containing fish oil (control diet) or fish oil plus orlistat (200 and 400 mg/kg of diet) over the course of 3 weeks. Fat absorption and the level of eicosapentaenoic acid (EPA) and docosahexaenoic acid, among other fatty acids, in red blood cells, plasma, liver, and spleen, were measured at the end of the experimental period. The results show that at 200 mg and 400 mg/kg of diet orlistat lowers fat absorption by 9% (P = .008) and 54% (P = .008). Orlistat given at the higher level induced a reduction of the incorporation of EPA in red blood cell (-45%; P = .006) and in plasma (-34%; P = .026) compared to the control group. Our results confirmed that administration of orlistat reduces incorporation of n-3 LC-PUFA in blood lipids and tissues in a rat model.

Metabolic Phenotyping of an Adoptive Transfer Mouse Model of Experimental Colitis and Impact of Dietary Fish Oil Intake

Inflammatory bowel diseases are acute and chronic disabling inflammatory disorders with multiple complex etiologies that are not well-defined. Chronic intestinal inflammation has been linked to an energy-deficient state of gut epithelium with alterations in oxidative metabolism. Plasma-, urine-, stool-, and liver-specific metabonomic analyses are reported in a naïve T cell adoptive transfer (AT) experimental model of colitis, which evaluated the impact of long-chain n-3 polyunsaturated fatty acid (PUFA)-enriched diet. Metabolic profiles of AT animals and their controls under chow diet or fish oil supplementation were compared to describe the (i) consequences of inflammatory processes and (ii) the differential impact of n-3 fatty acids. Inflammation was associated with higher glycoprotein levels (related to acute-phase response) and remodeling of PUFAs. Low triglyceride levels and enhanced PUFA levels in the liver suggest activation of lipolytic pathways that could lead to the observed increase of phospholipids in the liver (including plasmalogens and sphingomyelins). In parallel, the increase in stool excretion of most amino acids may indicate a protein-losing enteropathy. Fecal content of glutamine was lower in AT mice, a feature exacerbated under fish oil intervention that may reflect a functional relationship between intestinal inflammatory status and glutamine metabolism. The decrease in Krebs cycle intermediates in urine (succinate, α-ketoglutarate) also suggests a reduction in the glutaminolytic pathway at a systemic level. Our data indicate that inflammatory status is related to this overall loss of energy homeostasis.

Characterization of Selected Metabolic and Immunologic Markers FollowingExclusive Enteral Nutrition of Pediatric CrohnâÂÂs Disease Patients

Objectives: Exclusive enteral nutrition (EEN) is one of the first-line therapeutic options for pediatric patients with active Crohn’s disease (CD). However, only sparse data exist on plasma metabolic changes from patients on EEN therapy. Thus to gain mechanistic insights, we have characterized selected markers in pediatric CD patients treated with EEN for induction of disease remission. Methods: Plasma levels of 18 cytokines and chemokines and 163 metabolites in 10 pediatric CD patients receiving 8 weeks of EEN therapy were measured. Measurements were performed at three time points: a) at baseline (V0); b) during EEN intervention at week 4 (V4) and; c) at 4 weeks after completion of EEN therapy at week 12 (V12). Results: Comparisons between V0 and V4 levels identified changes in 20 different molecules, including increases in 7 metabolites of the diacyl phosphatidylcholine (PC) class and 6 metabolites of the alkylacyl PC class. However, most of these changes were not sustained after returning to a normal diet, i.e. only 7 out of the 20 molecules with significant changes between V0 and V4 still retained significance upon comparing V0 and V12. Conclusions: The changes in the plasma levels of various phospholipids at different time-points reflect the nutritional intervention and improved health status of pediatric CD patients.