Comparative study the alleviated effects of various oligosaccharides on colitis in mice.

Oligosaccharides, namely, chitosan oligosaccharides (COS), fructooligosaccharides (FOS), and 2'-fucosyllactose (2-FL) were used to prevent the dextran sulfate sodium (DSS)-induced colitis in vivo based on antioxidant properties and anti-inflammatory activities, further comparing their alleviating effects to investigate the optimal anti-inflammatory agent. The results showed COS demonstrated the highest antioxidant properties, with a DPPH scavenging rate of 37.4% and an ABTS scavenging rate of 46.4% in these oligosaccharides. Consequently, COS exhibited the best anti-inflammatory activities on inflamed RAW 264.7 cells. Furthermore, the COS intervention demonstrated the best attenuated effects on decrease in the body weight and increase in DAI score, as well as on the overexpressed inflammatory factors and underexpressed short-chain fatty acids (SCFAs) compare to FOS and 2-FL. Therefore, these beneficial changes help prevent the damage to the inflammatory lesions in colonic histopathology. Additionally, COS significantly increased the diversity of gut microbiota and the ratio of Firmicutes/Bacteroidetes at phylum level. It also up-regulated the abundance of Lactobacillaceae and down-regulated Helicobacteraceae and Desulfovibrionaceae more effectively at family level to maintain oral tolerance against DSS. In short, COS intervention could be a promising nutritional strategy for alleviating colitis.

[Components and lipid-lowering effect of total saponins from underground part of Gynostemma pentaphyllum].

The saponins in different parts of Gynostemma pentaphyllum were analyzed via UPLC-Q-TOF-MS~E. A total of 46 saponins were identified, and the underground part had 26 saponins more than the aboveground part, most of which were trisaccharide saponins. The rat model of hyperlipidemia was established with high-fat diet. This study explored the lipid-lowering activity of total saponins in the underground part of G. pentaphyllum, so as to provide a theoretical basis for the comprehensive utilization of the underground part of G. pentaphyllum. A total of 99 healthy SD rats were randomly assigned into a blank group, a model group, a positive drug group, an aboveground total saponins group, and low-, medium-, and high-dose underground total saponins groups. Except the blank group, the other groups were fed with high-fat diet for 6 weeks. Then, the blood was collected from the orbital cavity to determine whether the modeling was successful according to the serum levels of total cholesterol(TC) and triglyceride(TG). After intragastric administration of the corresponding agents for 30 continuous days, the physical state of the rats were observed, and the body weight and liver specific gravity were measured. Furthermore, the levels of TC, TG, low-density lipoprotein cholesterol(LDL-C), high-density lipoprotein cholesterol(HDL-C), alanine transaminase(ALT), aspartate transaminase(AST), bilirubin, and total bile acids in serum, as well as the levels of superoxide dismutase(SOD), malondialdehyde(MDA), peroxidase proliferator-activated receptor(PPAR-γ) in the liver tissue, were determined. The pathological changes of liver was observed via HE staining. The results showed that the aboveground total saponins and medium-and high-dose underground total saponins can treat hepatocyte steatosis, lower TC, TG, LDL-C, ALT, AST, total bilirubin, MDA, and PPAR-γ levels, and increase HDL-C and SOD levels in the model rats. The effect tended to be more obvious with the increase in dosage. Therefore, the total saponins in the underground part of G. pentaphyllum have good pharmacological effect of reducing blood lipid, which provides a theoretical basis for the comprehensive utilization of the underground part of G. pentaphyllum.

Pharmacological mechanisms of chitotriose as a redox regulator in the treatment of rat inflammatory bowel disease.

Although extensive development has been made in the treatment of inflammatory bowel disease (IBD), adverse effects and incomplete efficacy of currently used medications are continuous challenge. Accumulated reports on the benefits of chitosan oligosaccharides in intestinal disorders make chitotriose (COS) a breakthrough in the development of new IBD drugs. This study aimed to investigate the biosafety, efficacy and pharmacological mechanisms of COS in the treatment of experimental IBD in compare with the commercial 5-Aminosalicylic acid (5-ASA). In this study, COS effectively relieved active inflammation, restored epithelial function, and reduced intestinal fibrosis. Further investigation demonstrated that COS treatment regulated redox state of the colon tissue by stimulating the transcription factor nuclear factor E2-related factor 2 (Nrf2), increasing production of endogenous antioxidants, and alleviating oxidative stress. The offset of oxidative stress shut down the nuclear factor kappa-B (NF-ĸB) inflammatory pathway, mitophagy of epithelial cells, M2 macrophage polarization in pre-fibrotic inflammation, and myofibroblast activation in intestinal fibrogenesis. In conclusion, COS is a safe and effective therapeutic agent for experimental IBD as a redox regulator. Our results expand the current understanding of the pharmacology of chitosan oligosaccharides for IBD treatment and provides experimental basis for the medicinal development of small molecule carbohydrates.

Main active components of Si-Miao-Yong-An decoction (SMYAD) attenuate autophagy and apoptosis via the PDE5A-AKT and TLR4-NOX4 pathways in isoproterenol (ISO)-induced heart failure models.

Heart failure (HF), the main cause of death in patients with many cardiovascular diseases, has been reported to be closely related to the complicated pathogenesis of autophagy, apoptosis, and inflammation. Notably, Si-Miao-Yong-An decoction (SMYAD) is a traditional Chinese medicine (TCM) used to treat cardiovascular disease; however, the main active components and their relevant mechanisms remain to be discovered. Based on our previous ultra-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS) results, we identified angoriside C (AC) and 3,5-dicaffeoylquinic acid (3,5-DiCQA) as the main active components of SMYAD. In vivo results showed that AC and 3,5-DiCQA effectively improved cardiac function, reduced the fibrotic area, and alleviated isoproterenol (ISO)-induced myocarditis in rats. Moreover, AC and 3,5-DiCQA inhibited ISO-induced autophagic cell death by inhibiting the PDE5A/AKT/mTOR/ULK1 pathway and inhibited ISO-induced apoptosis by inhibiting the TLR4/NOX4/BAX pathway. In addition, the autophagy inhibitor 3-MA was shown to reduce ISO-induced apoptosis, indicating that ISO-induced autophagic cell death leads to excess apoptosis. Taken together, the main active components AC and 3,5-DiCQA of SMYAD inhibit the excessive autophagic cell death and apoptosis induced by ISO by inhibiting the PDE5A-AKT and TLR4-NOX4 pathways, thereby reducing myocardial inflammation and improving heart function to alleviate and treat a rat ISO-induced heart failure model and cell heart failure models. More importantly, the main active components of SMYAD will provide new insights into a promising strategy that will promote the discovery of more main active components of SMYAD for therapeutic purposes in the future.

Interventions for preventing diarrhoea-associated haemolytic uraemic syndrome.

BACKGROUND: Haemolytic uraemic syndrome (HUS) is a common cause of acquired kidney failure in children and rarely in adults. The most important risk factor for development of HUS is a gastrointestinal infection by Shiga toxin-producing Escherichia coli (STEC). This review addressed the interventions aimed at secondary prevention of HUS in patients with diarrhoea who were infected with a bacteria that increase the risk of HUS. OBJECTIVES: Our objective was to evaluate evidence regarding secondary preventative strategies for HUS associated with STEC infections. In doing so, we sought to assess the effectiveness and safety of interventions as well as their potential to impact the morbidity and death associated with this condition. SEARCH METHODS: We searched the Cochrane Kidney and Transplant Register of Studies up to 12 November 2020 through contact with the Information Specialist using search terms relevant to this review. Studies in the Register are identified through searches of CENTRAL, MEDLINE, and EMBASE, conference proceedings, the International Clinical Trials Register (ICTRP) Search Portal and ClinicalTrials.gov. SELECTION CRITERIA: Studies were considered based on the methods, participants, and research goals. Only randomised controlled trials were considered eligible for inclusion. The participants of the studies were paediatric and adult patients with diarrhoeal illnesses due to STEC. The primary outcome of interest was incidence of HUS. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures as recommended by Cochrane. Summary estimates of effect were obtained using a random-effects model, and results were expressed as risk ratios (RR) and their 95% confidence intervals (CI) for dichotomous outcomes. Confidence in the evidence was assessed using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach. MAIN RESULTS: We identified four studies (536 participants) for inclusion that investigated four different interventions including antibiotics (trimethoprim-sulfamethoxazole), anti-Shiga toxin antibody-containing bovine colostrum, Shiga toxin binding agent (Synsorb Pk: a silicon dioxide-based agent), and a monoclonal antibody against Shiga toxin (urtoxazumab). The overall risk of bias was unclear for selection, performance and detection bias and low for attrition, reporting and other sources of bias. It was uncertain if trimethoprim-sulfamethoxazole reduced the incidence of HUS compared to no treatment (47 participants: RR 0.57, 95% CI 0.11-2.81, very low certainty evidence). Adverse events relative to this review, need for acute dialysis, neurological complication and death were not reported. There were no incidences of HUS in either the bovine colostrum group or the placebo group. It was uncertain if bovine colostrum caused more adverse events (27 participants: RR 0.92, 95% CI 0.42 to 2.03; very low certainty evidence). The need for acute dialysis, neurological complications or death were not reported. It is uncertain whether Synsorb Pk reduces the incidence of HUS compared to placebo (353 participants: RR 0.93, 95% CI 0.39 to 2.22; very low certainty evidence). Adverse events relevant to this review, need for acute dialysis, neurological complications or death were not reported. One study compared two doses of urtoxazumab (3.0 mg/kg and 1.0 mg/kg) to placebo. It is uncertain if either 3.0 mg/kg urtoxazumab (71 participants: RR 0.34, 95% CI 0.01 to 8.14) or 1.0 mg/kg urtoxazumab (74 participants: RR 0.95, 95% CI 0.79 to 1.13) reduced the incidence of HUS compared to placebo (very low certainty evidence). Low certainty evidence showed there may be little or no difference in the number of treatment-emergent adverse events with either 3.0 mg/kg urtoxazumab (71 participants: RR 1.00, 95% CI 0.84 to 1.18) or 1.0 mg/kg urtoxazumab (74 participants: RR 0.95, 95% CI 0.79 to 1.13) compared to placebo. There were 25 serious adverse events reported in 18 patients: 10 in the placebo group, and 9 and 6 serious adverse events in the 1.0 mg/kg and 3.0 mg/kg urtoxazumab groups, respectively. It is unclear how many patients experienced these adverse events in each group, and how many patients experienced more than one event. It is uncertain if either dose of urtoxazumab increased the risk of neurological complications or death (very low certainty evidence). Need for acute dialysis was not reported. AUTHORS' CONCLUSIONS: The included studies assessed antibiotics, bovine milk, and Shiga toxin inhibitor (Synsorb Pk) and monoclonal antibodies (Urtoxazumab) against Shiga toxin for secondary prevention of HUS in patients with diarrhoea due to STEC. However, no firm conclusions about the efficacy of these interventions can be drawn given the small number of included studies and the small sample sizes of those included studies. Additional studies, including larger multicentre studies, are needed to assess the efficacy of interventions to prevent development of HUS in patients with diarrhoea due to STEC infection.

2'-fucosyllactose inhibits imiquimod-induced psoriasis in mice by regulating Th17 cell response via the STAT3 signaling pathway.

Psoriasis is a chronic immune-mediated inflammatory cutaneous disorder with Th17 cells and Th17-related cytokines playing an important role in its development. 2'-FL (2'-fucosyllactose), which makes up about 30% of all HMOs (human milk oligosaccharides) in blood type secretor positive maternal milk, plays an essential role in supporting aspects of immune development and regulation. To explore the immunomodulatory effect of 2'-FL in psoriasis, we employed the imiquimod (IMQ)-induced psoriasis-like mouse model. Our data showed that mice administered with 2'-FL exhibited attenuated skin damage and inflammation, characterized by significantly decreased erythema and thickness and reduced recruitment of pro-inflammatory cytokines, when compared to control mice. The alleviated skin inflammation in 2'-FL treated mice was associated with a reduced proportion of Th17 cells and decreased production of Th17-related cytokines. Furthermore, we have demonstrated that 2'-FL reduced the phosphorylation of STAT3 in the skin tissue from mice with IMQ stimulation, which could account for the decreasing recruitment of Th17 cells. In vitro studies showed that 2'-FL inhibited differentiation of Th17 cells, phosphorylation of STAT3, and RORγt mRNA levels in T cells under Th17 polarization. Our results indicate that 2'-FL ameliorates IMQ-induced psoriasis by inhibiting Th17 cell immune response and Th17-related cytokine secretion via modulation of the STAT3 signaling pathway.

The Protective Effects of 2'-Fucosyllactose against E. Coli O157 Infection Are Mediated by the Regulation of Gut Microbiota and the Inhibition of Pathogen Adhesion.

As the richest component in human milk oligosaccharides (HMOs), 2'-fucosyllactose (2'-FL) can reduce the colonization of harmful microbiota in vivo, thus lowering the risk of infection; however, the mechanism for this is still unclear. In this study, a model of Escherichia coli O157 infection in healthy adult mice was established to explore the effect of 2'-FL intervention on E. coli O157 colonization and its protective effects on mice. The results showed that 2'-FL intake reduced E. coli O157 colonization in mice intestine by more than 90% (p < 0.001), and it also reduced intestinal inflammation, increased the content of fecal short-chain fatty acids, and enhanced intestinal barrier function. These beneficial effects were attributed to the increased expression of mucins such as MUC2 (increased by more than 20%, p < 0.001), and inhibition of E. coli O157 cell adhesion (about 30% reduction, p < 0.001), and were associated with the modulation of gut microbiota composition. 2'-FL significantly increased the abundance of Akkermansia, a potential probiotic, which may represent the fundamental means by which 2'-FL enhances the expression of mucin and reduces the colonization of harmful bacteria. The current study may support the use of 2'-FL in the prevention of foodborne pathogen infections in human.

A Novel Role for Necroptosis in the Pathogenesis of Necrotizing Enterocolitis.

BACKGROUND & AIMS: Necrotizing enterocolitis (NEC) is a devastating disease of premature infants characterized by Toll-like receptor 4 (TLR4)-dependent intestinal inflammation and enterocyte death. Given that necroptosis is a proinflammatory cell death process that is linked to bacterial signaling, we investigated its potential role in NEC, and the mechanisms involved. METHODS: Human and mouse NEC intestine were analyzed for necroptosis gene expression (ie, RIPK1, RIPK3, and MLKL), and protein activation (phosphorylated RIPK3). To evaluate a potential role for necroptosis in NEC, the effects of genetic (ie, Ripk3 knockout or Mlkl knockout) or pharmacologic (ie, Nec1s) inhibition of intestinal inflammation were assessed in a mouse NEC model, and a possible upstream role of TLR4 was assessed in Tlr4-deficient mice. The NEC-protective effects of human breast milk and its constituent milk oligosaccharides on necroptosis were assessed in a NEC-in-a-dish model, in which mouse intestinal organoids were cultured as either undifferentiated or differentiated epithelium in the presence of NEC bacteria and hypoxia. RESULTS: Necroptosis was activated in the intestines of human and mouse NEC in a TLR4-dependent manner, and was up-regulated specifically in differentiated epithelium of the immature ileum. Inhibition of necroptosis genetically and pharmacologically reduced intestinal-epithelial cell death and mucosal inflammation in experimental NEC, and ex vivo in the NEC-in-a-dish system. Strikingly, the addition of human breast milk, or the human milk oligosaccharide 2 fucosyllactose in the ex vivo system, reduced necroptosis and inflammation. CONCLUSIONS: Necroptosis is activated in the intestinal epithelium upon TLR4 signaling and is required for NEC development, and explains in part the protective effects of breast milk.

MytiLec-1 Shows Glycan-Dependent Toxicity against Brine Shrimp Artemia and Induces Apoptotic Death of Ehrlich Ascites Carcinoma Cells In Vivo.

MytiLec-1, a 17 kDa lectin with ß-trefoil folding that was isolated from the Mediterranean mussel (Mytilus galloprovincialis) bound to the disaccharide melibiose, Galα(1,6) Glc, and the trisaccharide globotriose, Galα(1,4) Galß(1,4) Glc. Toxicity of the lectin was found to be low with an LC50 value of 384.53 µg/mL, determined using the Artemia nauplii lethality assay. A fluorescence assay was carried out to evaluate the glycan-dependent binding of MytiLec-1 to Artemia nauplii. The lectin strongly agglutinated Ehrlich ascites carcinoma (EAC) cells cultured in vivo in Swiss albino mice. When injected intraperitoneally to the mice at doses of 1.0 mg/kg/day and 2.0 mg/kg/day for five consecutive days, MytiLec-1 inhibited 27.62% and 48.57% of cancer cell growth, respectively. Antiproliferative activity of the lectin against U937 and HeLa cells was studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in vitro in RPMI-1640 medium. MytiLec-1 internalized into U937 cells and 50 µg/mL of the lectin inhibited their growth of to 62.70% whereas 53.59% cell growth inhibition was observed against EAC cells when incubated for 24 h. Cell morphological study and expression of apoptosis-related genes (p53, Bax, Bcl-X, and NF-κB) showed that the lectin possibly triggered apoptosis in these cells.

Biosynthesis of the anti-diabetic metabolite montbretin A: glucosylation of the central intermediate mini-MbA.

Type 2 diabetes (T2D) affects over 320 million people worldwide. Healthy lifestyles, improved drugs and effective nutraceuticals are different components of a response against the growing T2D epidemic. The specialized metabolite montbretin A (MbA) is being developed for treatment of T2D and obesity due to its unique pharmacological activity as a highly effective and selective inhibitor of the human pancreatic α-amylase. MbA is an acylated flavonol glycoside found in small amounts in montbretia (Crocosmia × crocosmiiflora) corms. MbA cannot be obtained in sufficient quantities for drug development from its natural source or by chemical synthesis. To overcome these limitations through metabolic engineering, we are investigating the genes and enzymes of MbA biosynthesis. We previously reported the first three steps of MbA biosynthesis from myricetin to myricetin 3-O-(6'-O-caffeoyl)-glucosyl rhamnoside (mini-MbA). Here, we describe the sequence of reactions from mini-MbA to MbA, and the discovery and characterization of the gene and enzyme responsible for the glucosylation of mini-MbA. The UDP-dependent glucosyltransferase CcUGT3 (UGT703E1) catalyzes the 1,2-glucosylation of mini-MbA to produce myricetin 3-O-(glucosyl-6'-O-caffeoyl)-glucosyl rhamnoside. Co-expression of CcUGT3 with genes for myricetin and mini-MbA biosynthesis in Nicotiana benthamiana validated its biological function and expanded the set of genes available for metabolic engineering of MbA.

The Combination of 2'-Fucosyllactose with Short-Chain Galacto-Oligosaccharides and Long-Chain Fructo-Oligosaccharides that Enhance Influenza Vaccine Responses Is Associated with Mucosal Immune Regulation in Mice.

BACKGROUND: A critical role for host-microbe interactions and establishment of vaccine responses has been postulated. Human milk oligosaccharides, of which 2'-fucosyllactose (2'FL) is the most prevalent, are known to alter host-associated microbial communities and play a critical role in the immunologic development of breastfed infants. OBJECTIVES: Dietary supplementation with a combination of 2'FL and prebiotic short-chain (sc) galacto-oligosaccharides (GOS) and long-chain (lc) fructo-oligosaccharides (FOS) was employed to examine human milk oligosaccharide effects on immune responsiveness, within a murine influenza vaccination model. METHODS: Female mice (6 wk old, C57Bl/6JOlaHsd) were fed either control diet (CON) or scGOS/lcFOS/2'FL-containing diet (GF2F) for 45 d. After starting dietary intervention (day 14), mice received a primary influenza vaccination (day 0) followed by a booster vaccination (day 21), after which ear challenges were conducted to measure vaccine-specific delayed type hypersensitivity (DTH). Serum immunoglobulin (Ig) levels, fecal and cecal microbial community structure, short-chain fatty acids, host intestinal gene expression and cellular responses in the mesenteric lymph nodes (MLNs) were also measured. RESULTS: Relative to CON, mice fed the GF2F diet had increased influenza vaccine-specific DTH responses (79.3%; P < 0.01), higher levels of both IgG1 (3.2-fold; P < 0.05) and IgG2a (1.2-fold; P < 0.05) in serum, and greater percentages of activated B cells (0.3%; P < 0.05), regulatory T cells (1.64%; P < 0.05), and T-helper 1 cells (2.2%; P < 0.05) in their MLNs. GF2F-fed mice had elevated cecal butyric (P < 0.05) and propionic (P < 0.05) acid levels relative to CON, which correlated to DTH responses (R2 = 0.22; P = 0.05 and R2 = 0.39; P < 0.01, respectively). Specific fecal microbial taxa altered in GF2F diet fed mice relative to CON were significantly correlated with the DTH response and IgG2a level increases. CONCLUSIONS: Dietary GF2F improved influenza vaccine-specific T-helper 1 responses and B cell activation in MLNs and enhanced systemic IgG1 and IgG2a concentrations in mice. These immunologic changes are correlated with microbial community structure and metabolites.

Biological Activities of Lactose-Derived Prebiotics and Symbiotic with Probiotics on Gastrointestinal System.

Lactose-derived prebiotics provide wide ranges of gastrointestinal comforts. In this review article, the probable biochemical mechanisms through which lactose-derived prebiotics offer positive gastrointestinal health are reported along with the up-to-date results of clinical investigations; this might be the first review article of its kind, to the best of our knowledge. Lactose-derived prebiotics have unique biological and functional values, and they are confirmed as 'safe' by the Food and Drug Administration federal agency. Medical practitioners frequently recommend them as therapeutics as a pure form or combined with dairy-based products (yoghurt, milk and infant formulas) or fruit juices. The biological activities of lactose-derived prebiotics are expressed in the presence of gut microflora, mainly probiotics (Lactobacillus spp. in the small intestine and Bifidobacterium spp. in the large intestine). Clinical investigations reveal that galacto-oligosaccharide reduces the risks of several types of diarrhea (traveler's diarrhea, osmotic diarrhea and Clostridium difficile associated relapsing diarrhea). Lactulose and lactosucrose prevent inflammatory bowel diseases (Crohn's disease and ulcerative colitis). Lactulose and lactitol reduce the risk of hepatic encephalopathy. Furthermore, lactulose, galacto-oligosaccharide and lactitol prevent constipation in individuals of all ages. It is expected that the present review article will receive great attention from medical practitioners and food technologists.

α1,2-Fucosyllactose Does Not Improve Intestinal Function or Prevent Escherichia coli F18 Diarrhea in Newborn Pigs.

OBJECTIVES: Infectious diarrhea, a leading cause of morbidity and deaths, is less prevalent in breastfed infants compared with infants fed infant formula. The dominant human milk oligosaccharide (HMO), α-1,2-fucosyllactose (2'-FL), has structural homology to bacterial adhesion sites in the intestine and may in part explain the protective effects of human milk. We hypothesized that 2'-FL prevents diarrhea via competitive inhibition of pathogen adhesion in a pig model for sensitive newborn infants. METHODS: Intestinal cell studies were coupled with studies on cesarean-delivered newborn pigs (n = 24) without (control) or with inoculation of enterotoxigenic Escherichia coli F18 (7.5 × 10/day for 8 days) fed either no (F18) or 10 g/L 2'-FL (2FL-F18). RESULTS: In vitro studies revealed decreased pathogen adhesion to intestinal epithelial cells with 2'-FL (5 g/L; P < 0.001). F18 pigs showed more diarrhea than control pigs (P < 0.01). Administration of 2'-FL to F18 pigs failed to prevent diarrhea, although the relative weight loss tended to be reduced (-19 vs -124 g/kg, P = 0.12), higher villi were observed in the distal small intestine (P < 0.05), and a trend toward increased proportion of mucosa and activities of some brush border enzymes in the proximal small intestine. In situ abundance of α-1,2-fucose and E coli was similar between groups, whereas sequencing showed higher abundance of Enterobacteriaceae in F18, Enterococcus in control and Lachnospiraceae in 2FL-F18 pigs. CONCLUSIONS: 2'-FL inhibited in vitro adhesion of E coli F18 to epithelial cells, but had limited effects on diarrhea and mucosal health in newborn pigs challenged with E coli F18.

Host-microbiome interactions in human type 2 diabetes following prebiotic fibre (galacto-oligosaccharide) intake.

Aberrant microbiota composition and function have been linked to several pathologies, including type 2 diabetes. In animal models, prebiotics induce favourable changes in the intestinal microbiota, intestinal permeability (IP) and endotoxaemia, which are linked to concurrent improvement in glucose tolerance. This is the first study to investigate the link between IP, glucose tolerance and intestinal bacteria in human type 2 diabetes. In all, twenty-nine men with well-controlled type 2 diabetes were randomised to a prebiotic (galacto-oligosaccharide mixture) or placebo (maltodextrin) supplement (5·5 g/d for 12 weeks). Intestinal microbial community structure, IP, endotoxaemia, inflammatory markers and glucose tolerance were assessed at baseline and post intervention. IP was estimated by the urinary recovery of oral 51Cr-EDTA and glucose tolerance by insulin-modified intravenous glucose tolerance test. Intestinal microbial community analysis was performed by high-throughput next-generation sequencing of 16S rRNA amplicons and quantitative PCR. Prebiotic fibre supplementation had no significant effects on clinical outcomes or bacterial abundances compared with placebo; however, changes in the bacterial family Veillonellaceae correlated inversely with changes in glucose response and IL-6 levels (r -0·90, P=0·042 for both) following prebiotic intake. The absence of significant changes to the microbial community structure at a prebiotic dosage/length of supplementation shown to be effective in healthy individuals is an important finding. We propose that concurrent metformin treatment and the high heterogeneity of human type 2 diabetes may have played a significant role. The current study does not provide evidence for the role of prebiotics in the treatment of type 2 diabetes.

Oral supplementation of healthy adults with 2'-O-fucosyllactose and lacto-N-neotetraose is well tolerated and shifts the intestinal microbiota.

The gut microbiota has been established as an important player influencing many aspects of human physiology. Breast milk, the first diet for an infant, contains human milk oligosaccharides (HMO) that shape the infant's gut microbiota by selectively stimulating the growth of specific bacteria, especially bifidobacteria. In addition to their bifidogenic activity, the ability of HMO to modulate immune function and the gut barrier makes them prime candidates to restore a beneficial microbiota in dysbiotic adults and provide health benefits. We conducted a parallel, double-blind, randomised, placebo-controlled, HMO-supplementation study in 100 healthy, adult volunteers, consuming chemically produced 2'-O-fucosyllactose (2'FL) and/or lacto-N-neotetraose (LNnT) at various daily doses and mixes or placebo for 2 weeks. All participants completed the study without premature discontinuation. Supplementation of 2'FL and LNnT at daily doses up to 20 g was shown to be safe and well tolerated, as assessed using the gastrointestinal symptoms rating scale. 16S rRNA sequencing analysis showed that HMO supplementation specifically modified the adult gut microbiota with the primary impact being substantial increases in relative abundance of Actinobacteria and Bifidobacterium in particular and a reduction in relative abundance of Firmicutes and Proteobacteria. This study provides the first set of data on safety, tolerance and impact of HMO on the adult gut microbiota. Collectively, the results from this study show that supplementing the diet with HMO is a valuable strategy to shape the human gut microbiota and specifically promote the growth of beneficial bifidobacteria.

Evaluation of the Antihyperglycemic Effect of Minor Steviol Glycosides in Normoglycemic and Induced-Diabetic Wistar Rats.

Steviol glycosides are a family of compounds found in Stevia rebaudiana Bertoni that are responsible for sweetness capacity. The antihyperglycemic effect of the two major steviol glycosides, Rebaudioside A and Stevioside, has been studied and it has been found that despite having the same common structure, only Stevioside exerts an antihyperglycemic effect. Although other steviol derivatives are found in smaller amounts (minor steviol glycosides) in S. rebaudiana, whether or not they possess antihyperglycemic activity has not been evaluated. The aim of this study was to evaluate the antihyperglycemic effect of minor steviol glycosides in normoglycemic and diabetic (streptozotocin/nicotinamide) Wistar rats. Rats were subjected to an intraperitoneal glucose tolerance test (IPGTT) both before and after chronic treatment (28 days). After 6 h of fasting, IPGTT was conducted in pentobarbital-anesthetized rats using 1 g/kg of glucose plus 20 mg/kg of the minor glycoside (Dulcoside A, Rebaudioside B, C, D, or Steviolbioside) or control treatment (distilled water, glibenclamide, or metformin); the blood of the tip of the tail was collected at time 0, 15, 30, 60, and 120 min.; and blood glucose was measured, and its net area under the curve (AUCnet) was calculated. After 28-day chronic oral administration, IPGTT was again performed. Differences were considered significant at P < .05 by one-way ANOVA. Acute intraperitoneal or chronic oral administration of 20 mg/kg of minor steviol glycosides had no antihyperglycemic effect in normoglycemic or induced-diabetic Wistar rats. Considering the dose tested, it is unlikely that these glycosides have an effect on glucose in diabetic or normoglycemic humans.

Novel Combination of Prebiotics Galacto-Oligosaccharides and Inulin-Inhibited Aberrant Crypt Foci Formation and Biomarkers of Colon Cancer in Wistar Rats.

The selectivity and beneficial effects of prebiotics are mainly dependent on composition and glycosidic linkage among monosaccharide units. This is the first study to use prebiotic galacto-oligosaccharides (GOS) that contains ß-1,6 and ß-1,3 glycosidic linkages and the novel combination of GOS and inulin in cancer prevention. The objective of the present study is to explore the role of novel GOS and inulin against various biomarkers of colorectal cancer (CRC) and the incidence of aberrant crypt foci (ACF) in a 1,2-dimethyl hydrazine dihydrochloride (DMH)-induced rodent model. Prebiotic treatments of combined GOS and inulin (57 mg each), as well as individual doses (GOS: 76-151 mg; inulin 114 mg), were given to DMH-treated animals for 16 weeks. Our data reveal the significant preventive effect of the GOS and inulin combination against the development of CRC. It was observed that inhibition of ACF formation (55.8%) was significantly (p ≤ 0.05) higher using the GOS and inulin combination than GOS (41.4%) and inulin (51.2%) treatments alone. This combination also rendered better results on short-chain fatty acids (SCFA) and bacterial enzymatic activities. Dose-dependent effects of prebiotic treatments were also observed on cecum and fecal bacterial enzymes and on SCFA. Thus, this study demonstrated that novel combination of GOS and inulin exhibited stronger preventive activity than their individual treatments alone, and can be a promising strategy for CRC chemoprevention.

Sialylated galacto-oligosaccharides and 2'-fucosyllactose reduce necrotising enterocolitis in neonatal rats.

Necrotising enterocolitis (NEC) is one of the most frequent and fatal intestinal disorders in preterm infants and has very limited treatment options. Breast-fed infants are at a 6-10-fold lower NEC risk than formula-fed infants, and we have previously shown that human milk oligosaccharides (HMO) improved survival and reduced pathology in a rat NEC model. The HMO disialyllacto-N-tetraose (DSLNT) was most effective, and sialylation was shown to be essential for its protective effect. Galacto-oligosaccharides (GOS), currently added to some infant formula, but not containing sialic acid, had no effect. In addition to DSLNT, our previous work also showed that the neutral HMO fraction, which contains high concentrations of 2'-fucosyllactose (2'FL), slightly improved pathology scores. Here, we assessed the in vivo efficacy of 2'FL, as well as of GOS that we enzymatically sialylated (Sia-GOS). Neonatal rats were randomised into the following study groups - dam-fed (DF), formula-fed (FF), FF containing pooled HMO (10 mg/ml), GOS (8 mg/ml), Sia-GOS (500 µm) or 2'FL (2 mg/ml) - and subjected to the established NEC protocol. The DF and HMO groups had the lowest pathology scores with mean values of 0·67 (sd 0·34) and 0·90 (sd 0·47), respectively. The FF group had significantly elevated pathology scores of 2·02 (sd 0·63). Although the addition of GOS to the formula had no protective effect and generated scores of 2·00 (sd 0·63), the addition of Sia-GOS or 2'FL significantly lowered pathology scores to 1·32 (sd 0·56) (P<0·0034) and 1·43 (sd 0·51) (P<0·0040), respectively. The results warrant further studies to investigate the underlying mechanisms and to assess safety and efficacy in human neonates.

Human Milk Oligosaccharides and Synthetic Galactosyloligosaccharides Contain 3'-, 4-, and 6'-Galactosyllactose and Attenuate Inflammation in Human T84, NCM-460, and H4 Cells and Intestinal Tissue Ex Vivo.

BACKGROUND: The immature intestinal mucosa responds excessively to inflammatory insult, but human milk protects infants from intestinal inflammation. The ability of galactosyllactoses [galactosyloligosaccharides (GOS)], newly found in human milk oligosaccharides (HMOS), to suppress inflammation was not known. OBJECTIVE: The objective was to test whether GOS can directly attenuate inflammation and to explore the components of immune signaling modulated by GOS. METHODS: Galactosyllactose composition was measured in sequential human milk samples from days 1 through 21 of lactation and in random colostrum samples from 38 mothers. Immature [human normal fetal intestinal epithelial cell (H4)] and mature [human metastatic colonic epithelial cell (T84) and human normal colon mucosal epithelial cell (NCM-460)] enterocyte cell lines were treated with the pro-inflammatory molecules tumor necrosis factor-α (TNF-α) or interleukin-1ß (IL-1ß) or infected with Salmonella or Listeria. The inflammatory response was measured as induction of IL-8, monocyte chemoattractant protein 1 (MCP-1), or macrophage inflammatory protein-3α (MIP-3α) protein by ELISA and mRNA by quantitative reverse transcriptase-polymerase chain reaction. The ability of HMOS or synthetic GOS to attenuate this inflammation was tested in vitro and in immature human intestinal tissue ex vivo. RESULTS: The 3 galactosyllactoses (3'-GL, 4-GL, and 6'-GL) expressed in colostrum rapidly declined over early lactation (P < 0.05). In H4 cells, HMOS attenuated TNF-α- and IL-1ß-induced expression of IL-8, MIP-3α, and MCP-1 to 48-51% and pathogen-induced IL-8 and MCP-1 to 26-30% of positive controls (P < 0.001). GOS reduced TNF-α- and IL-1ß-induced inflammatory responses to 25-26% and pathogen-induced IL-8 and MCP-1 to 36-39% of positive controls (P < 0.001). GOS and HMOS mitigated nuclear translocation of nuclear transcription factor κB (NF-κB) p65. HMOS quenched the inflammatory response to Salmonella infection by immature human intestinal tissue ex vivo to 26% and by GOS to 50% of infected controls (P < 0.01). CONCLUSION: Galactosyllactose attenuated NF-κB inflammatory signaling in human intestinal epithelial cells and in human immature intestine. Thus, galactosyllactoses are strong physiologic anti-inflammatory agents in human colostrum and early milk, contributing to innate immune modulation. The potential clinical utility of galactosyllactose warrants investigation.

Efeito do galactooligossacarídeo sobre os sintomas de constipação / Effect of 4′galactooligosaccharide on constipation symptoms

Resumo Objetivo A adição de frutooligossacarídeos e galactooligossacarídeos a fórmulas infantis pode diminuir a consistência fecal e aumentar a frequência das evacuações. O objetivo do presente estudo foi determinar o efeito do galactooligossacarídeo em crianças com constipação crônica. Métodos Entre 2010 e 2012, 20 pacientes constipados (4-16 anos), atendidos numa unidade básica de saúde, completaram ensaio clínico duplo cego, placebo-controlado e de delineamento crossover. Onze pacientes receberam galactooligossacarídeo (1,7 g) por 30 dias, seguidos por 15 dias de washout, e, após, placebo (maltodextrina) por 30 dias; nove pacientes receberam placebo 30 dias, seguidos de 15 dias de washout e 30 dias de galactooligossacarídeo (1,7 g). Os desfechos primários foram frequência semanal de evacuações, esforço evacuatório e consistência fecal, classificada por escala numérica elaborada para este estudo e compilada no primeiro, 15̊ e 30̊ dias de cada período de crossover. Análise estatística foi feita por método de análise de variância (Anova) para medidas repetidas. Resultados Intensidade dos sintomas nos grupos foi semelhante no início do estudo (p = 0,45). Durante a ingestão de galactooligossacarídeo constatou-se maior frequência de evacuações, p < 0,0001, menor dificuldade evacuatória, p < 0,0001 e diminuição da consistência fecal, p = 0,0014. Efeitos colaterais não foram referidos durante a ingestão do prebiótico. Conclusão Durante a ingestão de galactooligossacarídeo os sintomas clínicos da constipação em crianças e adolescentes foram significantemente aliviados.

Abstract Objective Fructooligosacharides and galactooligosacharides soften fecal bolus and increase frequency of depositions when added to infant formula. This study aimed to determine the effects of galactooligosaccharide in pediatric patients with chronic constipation. Methods From 2010 to 2012, 20 constipated patients (4-16 years of age) attended to at a primary healthcare unit were enrolled in a double-blinded, placebo-controlled crossover trial. Eleven children ingested galactooligosaccharide (1.7 g) for 30 days, followed by a 15-day washout period, and a 30-day period of placebo (maltodextrin). Nine patients ingested maltodextrin for 30 days, followed by 15-day washout period, and galactooligosaccharide (1.7 g) for 30 days. Constipation symptoms were considered as primary outcomes: bowel movements/week, straining during defecation, and stool consistency. Outcome symptoms were ranked according to a numerical scale elaborated for this study. Data were recorded at baseline, and on days 15 and 30 of each 30-day crossover period. Repeated-measures analysis of variance (ANOVA) was used to analyze symptoms along time. Results At baseline, there was no significant difference in symptoms severity between groups (p = 0.45). Galactooligosaccharide ingestion was related to increase of the bowel movement frequency, p < 0.0001; relief of defecation straining, p < 0.0001; and decrease in stool consistency, p = 0.0014, compared to placebo ingestion. Patients reported no side effects from galactooligosaccharide. Conclusion Galactooligosaccharide was effective at improving clinical symptoms in this group of constipated children.