BTB and CNC homology 1 deficiency disrupts intestinal IgA secretion through regulation of polymeric immunoglobulin receptor expression.

Immunoglobulin A (IgA)-mediated mucosal immunity is important for the host because it contributes to reducing infection risk and to establishing host-microbe symbiosis. BTB and CNC homology 1 (Bach1) is a transcriptional repressor with physiological and pathophysiological functions that are of particular interest for their relation to gastrointestinal diseases. However, Bach1 effects on IgA-mediated mucosal immunity remain unknown. For this study using Bach1-deficient (Bach1-/-) mice, we investigated the function of Bach1 in IgA-mediated mucosal immunity. Intestinal mucosa, feces, and plasma IgA were examined using immunosorbent assay. After cell suspensions were prepared from Peyer's patches and colonic lamina propria, they were examined using flow cytometry. The expression level of polymeric immunoglobulin receptor (pIgR), which plays an important role in the transepithelial transport of IgA, was evaluated using Western blotting, quantitative real-time PCR, and immunohistochemistry. Although no changes in the proportions of IgA-producing cells were observed, the amounts of IgA in the intestinal mucosa were increased in Bach1-/- mice. Furthermore, plasma IgA was increased in Bach1-/- mice, but fecal IgA was decreased, indicating that Bach1-/- mice have abnormal secretion of IgA into the intestinal lumen. In fact, Bach1 deficiency reduced pIgR expression in colonic mucosa at both the protein and mRNA levels. In the human intestinal epithelial cell line LS174T, suppression of Bach1 reduced pIgR mRNA stability. In contrast, overexpression of Bach1 increased pIgR mRNA stability. These results demonstrate that Bach1 deficiency causes abnormal secretion of IgA into the intestinal lumen via suppression of pIgR expression.

Prebiotic effect of galacto-N-biose on the intestinal lactic acid bacteria as enhancer of acetate production and hypothetical colonization.

Galacto-N-biose (GNB) is an important core structure of glycan of mucin glycoproteins in the gastrointestinal (GI) mucosa. Because certain beneficial bacteria inhabiting the GI tract, such as bifidobacteria and lactic acid bacteria, harbor highly specialized GNB metabolic capabilities, GNB is considered a promising prebiotic for nourishing and manipulating beneficial bacteria in the GI tract. However, the precise interactions between GNB and beneficial bacteria and their accompanying health-promoting effects remain elusive. First, we evaluated the proliferative tendency of beneficial bacteria and their production of beneficial metabolites using gut bacterial strains. By comparing the use of GNB, glucose, and inulin as carbon sources, we found that GNB enhanced acetate production in Lacticaseibacillus casei, Lacticaseibacillus rhamnosus, Lactobacillus gasseri, and Lactobacillus johnsonii. The ability of GNB to promote acetate production was also confirmed by RNA-seq analysis, which indicated the upregulation of gene clusters that catalyze the deacetylation of N-acetylgalactosamine-6P and biosynthesize acetyl-CoA from pyruvate, both of which result in acetate production. To explore the in vivo effect of GNB in promoting acetate production, antibiotic-treated BALB/cA mice were administered with GNB with L. rhamnosus, resulting in a fecal acetate content that was 2.7-fold higher than that in mice administered with only L. rhamnosus. Moreover, 2 days after the last administration, a 3.7-fold higher amount of L. rhamnosus was detected in feces administered with GNB with L. rhamnosus than in feces administered with only L. rhamnosus. These findings strongly suggest the prebiotic potential of GNB in enhancing L. rhamnosus colonization and converting L. rhamnosus into higher acetate producers in the GI tract. IMPORTANCE: Specific members of lactic acid bacteria, which are commonly used as probiotics, possess therapeutic properties that are vital for human health enhancement by producing immunomodulatory metabolites such as exopolysaccharides, short-chain fatty acids, and bacteriocins. The long residence time of probiotic lactic acid bacteria in the GI tract prolongs their beneficial health effects. Moreover, the colonization property is also desirable for the application of probiotics in mucosal vaccination to provoke a local immune response. In this study, we found that GNB could enhance the beneficial properties of intestinal lactic acid bacteria that inhabit the human GI tract, stimulating acetate production and promoting intestinal colonization. Our findings provide a rationale for the addition of GNB to lactic acid bacteria-based functional foods. This has also led to the development of therapeutics supported by more rational prebiotic and probiotic selection, leading to an improved healthy lifestyle for humans.

Activity difference of three labdane diterpenoids on human constitutive androstane receptor.

The constitutive androstane receptor (CAR) regulates enzyme transcription related to drug metabolism; therefore, natural compound clarification in food that interacts with CAR is significant for drug development. We revealed that 13-epimanool, which is a compound found in the common sage, is bound to hCAR based on differential scanning fluorometry (DSF) measurements using recombinant hCAR protein. Similar labdane diterpenoids were examined, which revealed that manool and sclareol, which were both natural compounds contained in herbs, are bound to hCAR. They exhibited different effects for CAR activity in the luciferase assay despite the structural similarity. Manool was a partial agonist, 13-epimanool was a weak partial agonist, and sclareol was an antagonist. The activity of hCAR may be regulated by slight differences in the bound compound.

Water-soluble dietary fiber alleviates cancer-induced muscle wasting through changes in gut microenvironment in mice.

Cancer cachexia and the associated skeletal muscle wasting are considered poor prognostic factors, although effective treatment has not yet been established. Recent studies have indicated that the pathogenesis of skeletal muscle loss may involve dysbiosis of the gut microbiota and the accompanying chronic inflammation or altered metabolism. In this study, we evaluated the possible effects of modifying the gut microenvironment with partially hydrolyzed guar gum (PHGG), a soluble dietary fiber, on cancer-related muscle wasting and its mechanism using a colon-26 murine cachexia model. Compared with a fiber-free (FF) diet, PHGG contained fiber-rich (FR) diet-attenuated skeletal muscle loss in cachectic mice by suppressing the elevation of the major muscle-specific ubiquitin ligases Atrogin-1 and MuRF1, as well as the autophagy markers LC3 and Bnip3. Although tight-junction markers were partially reduced in both FR and FF diet-fed cachectic mice, the abundance of Bifidobacterium, Akkermansia, and unclassified S24-7 family increased by FR diet, contributing to the retention of the colonic mucus layer. The reinforcement of the gut barrier function resulted in the controlled entry of pathogens into the host system and reduced circulating levels of lipopolysaccharide-binding protein (LBP) and IL-6, which in turn led to the suppression of proteolysis by downregulating the ubiquitin-proteasome system and autophagy pathway. These results suggest that dietary fiber may have the potential to alleviate skeletal muscle loss in cancer cachexia, providing new insights for developing effective strategies in the future.

Role of Lipoteichoic Acid from the Genus Apilactobacillus in Inducing a Strong IgA Response.

Lactic acid bacterium-containing fermentates provide beneficial health effects by regulating the immune response. A naturally fermented vegetable beverage, a traditional Japanese food, reportedly provides health benefits; however, the beneficial function of its bacteria has not been clarified. Apilactobacillus kosoi is the predominant lactic acid bacterium in the beverage. Using murine Peyer's patch cells, we compared the immunoglobulin A (IgA)-inducing activity of A. kosoi 10HT to those of 29 other species of lactic acid bacteria and found that species belonging to the genus Apilactobacillus (A. kosoi 10HT, A. apinorum JCM30765T, and A. kunkeei JCM16173T) possessed significantly higher activity than the others. Thereafter, lipoteichoic acids (LTAs), important immunostimulatory molecules of Gram-positive bacteria, were purified from the three Apilactobacillus species, and their IgA-inducing activity was compared to those of LTAs from Lactiplantibacillus plantarum JCM1149T and a probiotic strain, Lacticaseibacillus rhamnosus GG. The results revealed that LTAs from Apilactobacillus species had significantly higher activity than others. We also compared the LTA structure of A. kosoi 10HT with that of L. plantarum JCM1149T and L. rhamnosus GG. Although d-alanine or both d-alanine and carbohydrate residues were substituents of free hydroxyl groups in the polyglycerol phosphate structure in LTAs from strains JCM1149T and GG, d-alanine residues were not found in LTA from strain 10HT by 1H nuclear magnetic resonance (NMR) analysis. Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) analysis of the glycolipid structure of LTA revealed that LTA from strain 10HT contained dihexosyl glycerol, whereas trihexosyl glycerol was detected in LTAs from other strains. These structural differences may be related to differences in IgA-inducing activity. IMPORTANCE The components of lactic acid bacteria that exert immunostimulatory effects are of increasing interest for therapeutic and prophylactic options, such as alternatives to antibiotics, cognitive enhancements, and vaccine adjuvants. LTAs act as immunostimulatory molecules in the host innate immune system by interacting with pattern recognition receptors. However, as LTA structures differ among species, detailed knowledge of the structure-function relationship for immunostimulatory effects is required. Comparisons of the IgA-inducing activity of LTAs have demonstrated that LTAs from the genus Apilactobacillus possess distinctive activities to stimulate mucosal immunity. The first analysis of the LTA structure from the genus Apilactobacillus suggests that it differs from structures of LTAs of related species of lactic acid bacteria. This knowledge is expected to aid in the development of functional foods containing lactic acid bacteria and pharmaceutical applications of immunostimulatory molecules from lactic acid bacteria.

Lycopene intake induces colonic regulatory T cells in mice and suppresses food allergy symptoms.

BACKGROUND: Food allergy (FA) is a common disease in children; thus, a high level of safety is required for its prevention and treatment. Colonic regulatory T cells (Tregs) have been suggested to attenuate FA. We investigated the Treg-inducing ability and anti-FA effects of carotenoids, a pigment contained in vegetables and fruits. METHODS: C57BL/6N mice were fed a diet containing 0.01% (w/w) of lycopene, ß-carotene, astaxanthin or lutein for 4 weeks, and the population of colonic Tregs was assessed. Subsequently, to evaluate the Treg-inducing ability of lycopene, splenic naïve CD4+ T cells from BALB/c mice were cultured with anti-CD3/CD28 antibody, TGF-ß and lycopene, and the frequencies of Tregs were examined. The effect of 0.1% (w/w) lycopene containing diet on FA was investigated in OVA-induced FA model BALB/c mice. RESULTS: In screening, only lycopene significantly increased the frequency and number of colonic Tregs. Lycopene also increased Treg differentiation in splenic naïve CD4+ T cells. In FA mice, lycopene feeding significantly increased the number of colonic Tregs and attenuated allergic symptoms. The expression levels of IL-4, IL-9 and IL-13 mRNA in colonic mucosa were also significantly reduced by lycopene. IL-9 is known to induce proliferation of mast cells, and we found that lycopene feeding significantly reduced the number of mast cells in the colonic mucosa of FA mice. CONCLUSION: Our results suggest that lycopene, a carotenoid present in many common foods on the market, may have the potential to induce colonic Tregs and suppress FA symptoms.

Oligosaccharides from agar extends lifespan through activation of unfolded protein response via SIR-2.1 in Caenorhabditis elegans.

PURPOSE: Agaro-oligosaccharides (AGO), hydrolysis products of agarose, is known to have antioxidant and anti-inflammatory properties. Speculating that AGO is effective for preventing aging, we investigated the longevity-supporting effects of AGO and their mechanisms using Caenorhabditis elegans. METHODS: Caenorhabditis elegans were fed AGO from young adulthood. The lifespan, locomotory activity, lipofuscin accumulation, and heat stress resistance of the worms were examined. To elucidate mechanisms of AGO-mediated longevity, we conducted comprehensive expression analysis using microarrays. Moreover, we used quantitative real-time PCR (qRT-PCR) to verify the genes showing differential expression levels. Furthermore, we measured the lifespan of loss-of-function mutants to determine the genes related to AGO-mediated longevity. RESULTS: AGO extended the lifespan of C. elegans, reduced lipofuscin accumulation, and maintained vigorous locomotion. The microarray analysis revealed that the endoplasmic reticulum-unfolded protein response (ER-UPR) and insulin/insulin-like growth factor-1-mediated signaling (IIS) pathway were activated in AGO-fed worms. The qRT-PCR analysis showed that AGO treatment suppressed sir-2.1 expression, which is a negative regulator of ER-UPR. In loss-of-function mutant of sir-2.1, AGO-induced longevity and heat stress resistance were decreased or cancelled completely. Furthermore, the pro-longevity effect of AGO was decreased in loss-of-function mutants of abnormal Dauer formation (daf) -2 and daf-16, which are IIS pathway-related genes. CONCLUSION: AGO delays the C. elegans aging process and extends their lifespan through the activations of ER-UPR and the IIS pathway.

Streptococcus thermophilus extends lifespan through activation of DAF-16-mediated antioxidant pathway in Caenorhabditis elegans.

Streptococcus thermophilus bacteria, which are widely used as fermented starter for dairy production, exert various beneficial health effects. Nevertheless, even though pro-longevity effects of various probiotics have been reported, no report has described Streptococcus thermophilus effects on longevity. This study was conducted to evaluate Streptococcus thermophilus effects on lifespan extension and to elucidate the Streptococcus thermophilus-mediated longevity mechanism using Caenorhabditis elegans worms as a model animal. They were fed standard food (Escherichia coli OP50) or Streptococcus thermophilus from the young adult stage. Feeding with Streptococcus thermophilus, compared to Escherichia coli OP50, to Caenorhabditis elegans extend the lifespan, reduced lipofuscin accumulation, and maintain vigorous locomotion. Feeding with Streptococcus thermophilus did not alter the worm growth curve or the offspring number, indicating that the Streptococcus thermophilus-mediated lifespan extension is not attributable to caloric restriction. The qRT-PCR data showed that Streptococcus thermophilus increased the expression of daf-16 and some of its downstream antioxidant genes. Furthermore, the pro-longevity effects of Streptococcus thermophilus were decreased in loss-of-function mutant of daf-16. Results show that Streptococcus thermophilus extends the lifespan of Caenorhabditis elegans through DAF-16-mediated antioxidant pathway activation.

Red-Beet Betalain Pigments Inhibit Amyloid-ß Aggregation and Toxicity in Amyloid-ß Expressing Caenorhabditis elegans.

Betalain pigments are mainly produced by plants belonging to the order of Caryophyllales. Betalains exhibit strong antioxidant activity and responds to environmental stimuli and stress in plants. Recent reports of antioxidant, anti-inflammatory and anti-cancer properties of betalain pigments have piqued interest in understanding their biological functions. We investigated the effects of betalain pigments (betanin and isobetanin) derived from red-beet on amyloid-ß (Aß) aggregation, which causes Alzheimer's disease. Non-specific inhibition of Aß aggregation against Aß40 and Aß42 by red-beet betalain pigments, in vitro was demonstrated using the thioflavin t fluorescence assay, circular dichroism spectroscopy analysis, transmission electron microscopy and nuclear magnetic resonance (NMR) analysis. Furthermore, we examined the ability of red-beet betalain pigments to interfere with Aß toxicity by using the transgenic Caenorhabditis elegans model, which expresses the human Aß42 protein intracellularly within the body wall muscle. It responds to Aß-toxicity with paralysis and treatment with 50 µM red-beet betalain pigments significantly delayed the paralysis of C. elegans. These results suggest that betalain pigments reduce Aß-induced toxicity.

Similarities and differences in the conformational stability and reversibility of ORF8, an accessory protein of SARS-CoV-2, and its L84S variant.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has the characteristic accessory protein ORF8. Although clinical reports indicate that ORF8 variant strains (Δ382 and L84S variants) are less likely to cause severe illness, functional differences between wild-type and variant ORF8 are unknown. Furthermore, the physicochemical properties of the ORF8 protein have not been analyzed. In this study, the physicochemical properties of the wild-type ORF8 and its L84S variant were analyzed and compared. Using the tobacco BY-2 cell production system, which has been successfully used to produce the wild-type ORF8 protein with a single conformation, was used to successfully produce the ORF8 L84S variant protein at the same level as wild-type ORF8. The produced proteins were purified, and their temperature and pH dependencies were examined using nuclear magnetic resonance spectra. Our data suggested that the wild-type and L84S variant ORF8 structures are highly stable over a wide temperature range. Both proteins displayed an aggregated conformation at higher temperature that reverted when the temperature was decreased to room temperature. Moreover, ORF8 precipitated at acidic pH and this precipitation was reversed when the solution pH was shifted to neutral. Interestingly, the L84S variant exhibited greater solubility than wild-type ORF8 under acidic conditions. Thus, the finding indicated that conformational stability and reversibility of ORF8 are key properties related to function in oppressive environments.

Rectal administration of carbon monoxide inhibits the development of intestinal inflammation and promotes intestinal wound healing via the activation of the Rho-kinase pathway in rats.

The inhalation of carbon monoxide (CO) gas and the administration of CO-releasing molecules were shown to inhibit the development of intestinal inflammation in a murine colitis model. However, it remains unclear whether CO promotes intestinal wound healing. Herein, we aimed to evaluate the therapeutic effects of the topical application of CO-saturated saline enemas on intestinal inflammation and elucidate the underlying mechanism. Acute colitis was induced with trinitrobenzene sulfonic acid (TNBS) in male Wistar rats. A CO-saturated solution was prepared via bubbling 50% CO gas into saline and was rectally administrated twice a day after colitis induction; rats were sacrificed 3 or 7 days after induction for the study of the acute or healing phases, respectively. The distal colon was isolated, and ulcerated lesions were measured. In vitro wound healing assays were also employed to determine the mechanism underlying rat intestinal epithelial cell restitution after CO treatment. CO solution rectal administration ameliorated acute TNBS-induced colonic ulceration and accelerated ulcer healing without elevating serum CO levels. The increase in thiobarbituric acid-reactive substances and myeloperoxidase activity after induction of acute TNBS colitis was also significantly inhibited after CO treatment. Moreover, the wound healing assays revealed that the CO-saturated medium enhanced rat intestinal epithelial cell migration via the activation of Rho-kinase. In addition, the activation of Rho-kinase in response to CO treatment was confirmed in the inflamed colonic tissue. Therefore, the rectal administration of a CO-saturated solution protects the intestinal mucosa from inflammation and accelerates colonic ulcer healing through enhanced epithelial cell restitution. CO may thus represent a novel therapeutic agent for the treatment of inflammatory bowel disease.

Identification of colorectal neoplasia by using serum bile acid profile.

BACKGROUND: Colonoscopy is the gold standard for detecting earlier stages of CRC, although screening of patients is difficult because of invasiveness, low compliance and procedural health risks. Therefore, the need for new screening methods for CRC is rising. Previous studies have demonstrated the diagnostic ability of serum BAs; however, the results have been inconsistent. In this study, we conducted a comprehensive analysis of serum BAs from patients with CRC and verified their diagnostic ability to detect CRC. METHODS: A total of 56 CRC patients (n = 14 each of stages I-IV), 59 patients with colonic adenoma and 60 healthy controls were included. Age and sex were matched for each group. Serum BA compositions were measured by LC-MS/MS and serum concentration of 30 types of BAs were analysed by discriminant analysis with multidimensional scaling method. RESULTS: Free CA, 3epi-DCA&CDCA, 3-dehydro CA, GCA and TCA were extracted as principal component (PC) 1 and free 3-dehydroDCA as PC 2 by canonical discriminant function coefficients. The verification of discriminability using cross-validation method revealed that the correct classification rate was 66.3% for original data and 52.6% for cross-validation data. CONCLUSIONS: A combined analysis using comprehensive serum BA concentration can be an efficient method for screening CRC.

Production of ORF8 protein from SARS-CoV-2 using an inducible virus-mediated expression system in suspension-cultured tobacco BY-2 cells.

KEY MESSAGE: Production of ORF8 protein from SARS-CoV-2 in tobacco BY-2 cells.

Dietary intake of yacon roots (Smallanthus sonchifolius) affects gut microbiota and fecal mucin and prevents intestinal inflammation in mice.

Consumption of yacon (Smallanthus sonchifolius) is associated with beneficial effects such as prevention of metabolic diseases. Yacon root is known to contain various bioactive components including indigestible carbohydrates, but the alteration of intestinal environment after treatment with yacon has not been fully investigated. This study investigated yacon-containing diet effects on the intestinal environment in mice, including microbial composition, short-chain fatty acid levels, and mucus content. After mice were administered yacon-containing diet for 4 weeks, 16S rRNA gene sequencing analyses revealed their fecal microbiota profiles. Organic acid concentrations in cecal contents were measured using an HPLC system. Compared to the control group, yacon-containing diet-received mice had significantly higher the concentrations of succinic acid, lactic acid, acetic acid, and propionic acid. The fecal mucin content was also higher in yacon-containing diet-received mice. Results of 16S rRNA gene sequencing analyses showed that the relative abundances of 27 taxa differed significantly in yacon-containing diet-received mice. Furthermore, results show effects of yacon administration on intestinal inflammation using 2,4,6-trinitrobenzene sulfonic acid induced colitis model in mice. Increased colonic damage and myeloperoxidase activity after 2,4,6-trinitrobenzene sulfonic acid treatment were suppressed in yacon-containing diet-received mice. Results suggest that oral intake of yacon root modulates the intestinal environment, thereby inhibiting intestinal inflammation.

Trans-unsaturated fatty acid activates NLRP3 inflammasome in macrophages and exacerbates intestinal inflammation in mice.

Higher consumption of trans fatty acid (TFA) is a risk factor for several inflammatory diseases including inflammatory bowel disease (IBD). However, the detailed mechanisms by which TFA intake affects IBD pathology remain unclear. We demonstrate here that elaidate, a trans-isomer of oleate, enhances interleukin (IL)-1ß production through the activation of NLRP3 inflammasome in mouse bone marrow-derived macrophages (BMDMs). Oleate has no effect on IL-1ß production. Elaidate also induces oxidative stress and activates endoplasmic reticulum stress in BMDMs. The elaidate-induced IL-1ß production is suppressed by co-treatments with antioxidants and a chemical chaperone. Furthermore, we analyze the effects of elaidate administration on intestinal inflammation using 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model in mice. Increased colonic damage and myeloperoxidase activity after TNBS treatment are elevated by elaidate administration. Also, TNBS treatment induces IL-1ß production in colonic mucosa; elaidate administration enhances the induction. We believe that these data reveal some mechanisms by which the TFA intake is associated with increased risk for IBD.

Cholestyramine, a Bile Acid Sequestrant, Increases Cecal Short Chain Fatty Acids and Intestinal Immunoglobulin A in Mice.

Bile acid sequestrants are used as medicinal drugs to treat dyslipidemia and type 2 diabetes. We found that cholestyramine, a bile acid sequestrant, increases cecal short-chain fatty acid (SCFA) production and intestinal immunoglobulin A (IgA) in C57BL/6J mice. In a 12-week high-fat diet study, feeding cholestyramine (2% (w/w)) significantly promoted C2-C4 SCFAs in the cecum by approximately 1.6-fold and fecal IgA by 1.8-fold. In an 8-week normal-fat diet study, feeding cholestyramine (1 and 2%) increased the cecal propionic acid content by approx. 2.0-fold. Fecal IgA was also significantly increased at 4 weeks (1%: 1.7-fold; 2%: 2.1-fold) and 8 weeks (1%: 1.8-fold; 2%: 2.0-fold) in the normal-fat diet study. These results indicate that bile acid sequestrants may exert their physiological functions, such as intestinal IgA production, through SCFA-dependent signaling pathways.

Administration of Enterococcus faecium HS-08 increases intestinal acetate and induces immunoglobulin A secretion in mice.

A probiotic is considered a live microbial feed supplement that has beneficial effects on the host. In this study, the probiotic property by which Enterococcus faecium HS-08 strengthens the immune system was investigated. Using a murine model, we evaluated the abilities of this strain to increase intestinal short-chain fatty acid contents and to induce the production of mucosal immunoglobulin A (IgA), which are crucial for mucosal immune systems. Various amounts (0%, 0.0038%, 0.038%, or 0.38%) of strain HS-08 cells were administered to BALB/cAJcl mice, which resulted in a dose-dependent increase of fecal IgA levels. A qRT-PCR analysis of Peyer's patch cells revealed that the gene expression of retinal-dehydrogenase, interleukin 6, B-cell-activating factor, and a proliferation-inducing ligand were increased, which leads to IgA secretion via a T-cell-independent mechanism. The administration of 0.038% and 0.38% of strain HS-08 cells also increased fecal acetate levels, which plays an important role for maintaining immune functions. This cecal floral analysis and the stability of strain HS-08 against gastrointestinal digestion suggest that this strain can inhabit the host intestine. In conclusion, the administration of E. faecium HS-08 increased intestinal acetate levels and enhanced IgA secretion, which may result in strengthening of the mucosal immune system.

Isolation of amaranthin synthetase from Chenopodium quinoa and construction of an amaranthin production system using suspension-cultured tobacco BY-2 cells.

Betalains are plant pigments primarily produced by plants of the order Caryophyllales. Because betalain possesses anti-inflammatory and anticancer activities, it may be useful as a pharmaceutical agent and dietary supplement. Recent studies have identified the genes involved in the betalain biosynthesis of betanin. Amaranthin and celosianin II are abundant in the quinoa (Chenopodium quinoa Willd.) hypocotyl, and amaranthin comprises glucuronic acid bound to betanin; therefore, this suggests the existence of a glucuronyltransferase involved in the synthesis of amaranthin in the quinoa hypocotyl. To identify the gene involved in amaranthin biosynthesis, we performed a BLAST analysis and phylogenetic tree analysis based on sequences homologous to flavonoid glycosyltransferase, followed by expression analysis on the quinoa hypocotyl to obtain three candidate proteins. Production of amaranthin in a transient Nicotiana benthamiana expression system was evaluated for these candidates and one was identified as having the ability to produce amaranthin. The gene encoding this protein was quinoa amaranthin synthetase 1 (CqAmaSy1). We also created a transgenic tobacco bright yellow-2 (BY-2) cell line wherein four betalain biosynthesis genes were introduced to facilitate amaranthin production. This transgenic cell line produced 13.67 ± 4.13 µm (mean ± SEM) amaranthin and 26.60 ± 1.53 µm betanin, whereas the production of isoamaranthin and isobetanin could not be detected. Tests confirmed the ability of amaranthin and betanin to slightly suppress cancer cell viability. Furthermore, amaranthin was shown to significantly inhibit HIV-1 protease activity, whereas betanin did not.

Combined treatment of dipeptidyl peptidase-4 inhibitor and exercise training improves lipid profile in KK/Ta mice.

NEW FINDINGS: What is the central question of this study? Exercise for type 2 diabetes patients treated with insulin therapy involves the risk of hypoglycaemia. Dipeptidyl peptidase-4 (DPP-4) inhibitors can be effective in combination with exercise because they reduce the incidence of hypoglycaemia. We evaluated the effect of this combination of treatments on hepatic lipid metabolism in diabetic KK/Ta mice. What is the main finding and its importance? The combination of a DPP-4 inhibitor and exercise, which lowers the risk of hypoglycaemia, is useful for improving insulin resistance by inhibiting excess insulin secretion and decreasing hepatic lipid accumulation, validated by downregulated CD36. ABSTRACT: The role of exercise training in prevention of diabetes and/or dyslipidaemia has been firmly established. Dipeptidyl peptidase-4 (DPP-4) inhibitors improve insulin sensitivity and have attracted attention as therapeutics for hepatic lipid accumulation. The effect of a combination of DPP-4 inhibitor and exercise training on the prevention and treatment of hepatic lipid accumulation is unclear. Here, we investigated whether alogliptin, a DPP-4 inhibitor, enhances the preventive effect of exercise-induced hepatic lipid accumulation in diabetic mice. Balb/c and KK/Ta mice were fed a high-fat diet. Mice were divided into the following five groups: B, Balb/c mice; K, KK/Ta mice; K-A, KK/Ta mice with alogliptin (0.01%); K-Ex, KK/Ta mice with exercise training (3 days week-1 , 15-20 m min-1 for 30 min); and K-Ex+A, KK/Ta mice with alogliptin and exercise training (n = 8 or 9 mice per group). After 8 weeks, glucose, insulin and triglyceride concentrations in the blood and triglyceride levels in the liver were significantly lower in the K-Ex+A group than in the K group. The liver expression level of PPAR-γ in the K group was significantly higher than that in the other groups. Additionally, the liver CD36 expression level was significantly lower in the K-Ex+A and B groups than in the K group. Thus, combined therapy of a DPP-4 inhibitor with exercise training was effective against high-fat diet-induced hepatic lipid accumulation in KK/Ta mice. The results of this study provide useful support for the practice of safe exercise therapy even in diabetic patients who require treatment with a DPP-4 inhibitor.

Green tea polyphenol (epigallocatechin-3-gallate) improves gut dysbiosis and serum bile acids dysregulation in high-fat diet-fed mice.

Gut microbiota have profound effects on bile acid metabolism by promoting deconjugation, dehydrogenation, and dehydroxylation of primary bile acids in the distal small intestine and colon. High-fat diet-induced dysbiosis of gut microbiota and bile acid dysregulation may be involved in the pathology of steatosis in patients with non-alcoholic fatty liver disease. Epigallocatechin-3-gallate (EGCG), the most abundant polyphenolic catechin in green tea, has been widely investigated for its inhibitory or preventive effects against fatty liver. The aim of the present study was to investigate the effects of EGCG on the abundance of gut microbiota and the composition of serum bile acids in high-fat diet-fed mice and determine the specific bacterial genera that can improve the serum bile acid dysregulation associated with EGCG anti-hepatic steatosis action. Male C57BL/6N mice were fed with the control diet, high-fat diet, or high-fat diet + EGCG at a concentration of 0.32% for 8 weeks. EGCG significantly inhibited the increases in weight, the area of fatty lesions, and the triglyceride content in the liver induced by the high-fat diet. Principal coordinate analysis revealed significant differences in microbial structure among the groups. At the genus level, EGCG induced changes in the microbiota composition in high-fat diet-fed mice, showing a significantly higher abundance of Adlercreutzia, Akkermansia, Allobaculum and a significantly lower abundance of Desulfovibrionaceae. EGCG significantly reversed the decreased population of serum primary cholic acid and ß-muricholic acid as well as the increased population of taurine-conjugated cholic acid, ß-muricholic acid and deoxycholic acid in high-fat diet-fed mice. Finally, the correlation analysis between bile acid profiles and gut microbiota demonstrated the contribution of Akkermansia and Desulfovibrionaceae in the improvement of bile acid dysregulation in high-fat diet-fed mice by treatment with EGCG. In conclusion, the present study suggests that EGCG could alter bile acid metabolism, especially taurine deconjugation, and suppress fatty liver disease by improving the intestinal luminal environment.