GhL1L1 regulates the contents of unsaturated fatty acids by activating the expression of GhFAD2 genes in cotton.

Edible oils with high unsaturated fatty acids, particularly oleic acid, are beneficial to human health. Cotton is one of the top five oil crops in the world, but the mechanism of high-quality oil synthesis and regulatory networks in cotton are largely unclear. Here, we identified Leafy cotyledon1-like 1 (GhL1L1), a NF-YB subfamily gene that is specifically expressed during somatic embryogenesis and seed maturation in cotton. Overexpression of GhL1L1 regulates the contents of unsaturated fatty acids in cotton, especially in the seeds, which is associated with altered expression of the cotton fatty acid biosynthesis-related genes. GhL1L1 synergistically enhanced the expression of GhFAD2-1A by binding to the G-box in its promoter, leading to an increase in the content of linoleic acid. Furthermore, this activation could be enhanced by GhNF-YC2 and GhNF-YA1 by form a transcriptional complex. Collectively, these results contribute to provide new insights into the molecular mechanism of oil biosynthesis in cotton and can facilitate genetic manipulation of cotton varieties with enhanced oil content.

Comprehensive investigation of milk oligosaccharides in different mammalian species and the effect of breed and lactation period on sheep milk oligosaccharides.

Milk oligosaccharides (MOs) have unique health benefits for newborns, and MOs are important components in mammalian milk. The present study was conducted to provide a comprehensive analysis of MOs in important domestic animals, including goats, cows, camels and sheep. The comparison with human MOs was conducted simultaneously. Furthermore, analysis of the relative abundance of sheep MOs among different breeds (Hu sheep, East Friesen sheep, East Friesen-Hu crossbred sheep) and lactation periods (colostrum, mature milk) was performed. In general, 35, 24 19, 26, and 16 MOs were identified in human, goat, bovine, camel and sheep milk, respectively. The type of sheep MOs was not greatly influenced by the breeds and lactation period. Hu sheep colostrum had the highest abundance of MOs among six sheep milks, followed by East Friesen sheep colostrum, while East Friesen-Hu crossbred sheep mature milk had the lowest abundance of MOs. These findings provide evidence for the potential value of MOs from domestic animal milk for the commercial applications.

Characterization and anti-inflammatory effect of selenium-enriched probiotic Bacillus amyloliquefaciens C-1, a potential postbiotics.

A patented strain of Bacillus amyloliquefaciens C-1 in our laboratory could produce functional sodium selenite (Na2SeO3) under optimized fermentation conditions. With the strong stress resistance and abundant secondary metabolites, C-1 showed potential to be developed as selenium-enriched postbiotics. C-1 has the ability to synthesize SeNPs when incubated with 100 µg/ml Na2SeO3 for 30 h at 30 °C aerobically with 10% seeds-culture. The transformation rate from Na2SeO3 into SeNPs reached to 55.51%. After selenium enrichment, there were no significant morphology changes in C-1 cells but obvious SeNPs accumulated inside of cells, observed by scanning electron microscope and transmission electron microscope, verified by energy dispersive X-ray spectroscopy and X-ray photoelectron spectroscopy. SeNPs had antioxidant activity in radical scavenge of superoxide (O2-), Hydroxyl radical (OH-) and 1,1-diphenyl-2-picryl-hydrazine (DPPH), where scavenging ability of OH- is the highest. Selenium-enriched C-1 had obvious anti-inflammatory effect in protecting integrity of Caco-2 cell membrane destroyed by S. typhimurium; it could preventing inflammatory damage in Caco-2 stressed by 200 µM H2O2 for 4 h, with significantly reduced expression of IL-8 (1.687 vs. 3.487, P = 0.01), IL-1ß (1.031 vs. 5.000, P < 0.001), TNF-α (2.677 vs. 9.331, P < 0.001), increased Claudin-1 (0.971 vs. 0.611, P < 0.001) and Occludin (0.750 vs. 0.307, P < 0.001). Transcriptome data analysis showed that there were 381 differential genes in the vegetative growth stage and 1674 differential genes in the sporulation stage of C-1 with and without selenium-enrichment. A total of 22 ABC transporter protein-related genes at vegetative stage and 70 ABC transporter protein-related genes at sporulation stage were founded. Genes encoding MsrA, thiol, glutathione and thioredoxin reduction were significantly up-regulated; genes related to ATP synthase such as atpA and atpD genes showed down-regulated during vegetative stage; the flagellar-related genes (flgG, fliM, fliL, and fliJ) showed down-regulated during sporulation stage. The motility, chemotaxis and colonization ability were weakened along with synthesized SeNPs accumulated intracellular at sporulation stage. B. amyloliquefaciens C-1 could convert extracellular selenite into intracellular SeNPs through the oxidation-reduction pathway, with strong selenium-enriched metabolism. The SeNPs and selenium-enriched cells had potential to be developed as nano-selenium biomaterials and selenium-enriched postbiotics.

Effects of dietary tryptophan on the antioxidant capacity and immune response associated with TOR and TLRs/MyD88/NF-κB signaling pathways in northern snakehead, Channa argus (Cantor, 1842).

Introduction: Dietary tryptophan (Trp) has been shown to influence fish feed intake, growth, immunity and inflammatory responses. The purpose of this study was to investigate the effect and mechanism of Trp on immune system of juvenile northern snakehead (Channa argus Cantor, 1842). Methods: A total of 540 fish (10.21 ± 0.11 g) were fed six experimental diets containing graded levels of Trp at 1.9, 3.0, 3.9, 4.8, 5.9 and 6.8 g/kg diet for 70 days, respectively. Results and Discussion: The results showed that supplementation of 1.9-4.8 g/kg Trp in diets had no effect on the hepatosomatic index (HSI) and renal index (RI), while dietary 3.9 and 4.8 g/kg Trp significantly increased spleen index (SI) of fish. Dietary 3.9, 4.8, 5.9 and 6.8 g/kg Trp enhanced the total hemocyte count (THC), the activities of total antioxidant capacity (T-AOC) and superoxide dismutase (SOD). Malondinaldehyde (MDA) levels in the blood were significantly decreased by consuming 3.9 and 4.8 g/kg Trp. Fish fed with 3.0 and 3.9 g/kg Trp diets up-regulated interleukin 6 (il-6) and interleukin 8 (il-8) mRNA levels. The expression of tumor necrosis factor α (tnf-α) was highest in fish fed with 3.0 g/kg Trp diet, and the expression of interleukin 1ß (il-1ß) was highest in fish fed with 3.9 g/kg Trp diet. Dietary 4.8, 5.9 and 6.8 g/kg Trp significantly decreased il-6 and tnf-α mRNA levels in the intestine. Moreover, Trp supplementation was also beneficial to the mRNA expression of interleukin 22 (il-22). Additionally, the mRNA expression levels of target of rapamycin (tor), toll-like receptor-2 (tlr2), toll-like receptor-4 (tlr4), toll-like receptor-5 (tlr5) and myeloid differentiation primary response 88 (myd88) of intestine were significantly up-regulated in fish fed 1.9, 3.0 and 3.9 g/kg Trp diets, and down-regulated in fish fed 4.8, 5.9 and 6.8 g/kg Trp diets. Dietary 4.8 and 5.9 g/kg Trp significantly increased the expression of inhibitor of nuclear factor kappa B kinase beta subunit (ikkß) and decreased the expression of inhibitor of kappa B (iκbα), but inhibited nuclear transcription factor kappa B (nf-κb) mRNA level. Collectively, these results indicated that dietary 4.8 g/kg Trp could improve antioxidant capacity and alleviate intestinal inflammation associated with TOR and TLRs/MyD88/NF-κB signaling pathways.

Variations in Oligosaccharides and N/O-Glycans in Human Milk through the Eight-Month Lactation Period.

Oligosaccharides and N/O-glycans are abundant in human milk and have numerous biological functions (for instance sialylated glycans provide sialic acid for the growth of infant brains), but their variation trends during lactation need further exploration. Qualitative and quantitative analyses of oligosaccharides and N/O-glycans in human milk at different lactation stages (from 7 days to 8 months) were performed using UHPLC-ESI-MS/MS. Thirty-four oligosaccharides, twenty-three N-glycans, and six O-glycans were identified. Oligosaccharides showed the highest abundance in human colostrum and decreased with the progression of lactation, and the abundance of N/O-glycans fluctuated as lactation progressed, while a high abundance of sialylated oligosaccharides and sialylated N/O-glycans was observed in human colostrum. These findings provide evidence for breastfeeding support and contribute to the development of infant formula supplemented with human milk glycans.

Preparation, characteristic and anti-inflammatory effect of selenium nanoparticle-enriched probiotic strain Enterococcus durans A8-1.

BACKGROUND: Elemental selenium, a new type of selenium supplement, can be biosynthesized via microorganisms. This study is to characterize a patent probiotic bacteria Enterococcus durans A8-1, capable of reducing selenite (Se6+ or Se4+) to elemental selenium (Se0) with the formation of Se nanoparticles (SeNPs). METHODS: The selenium nanoparticles synthesized from A8-1 were characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy dispersive spectroscopy (EDS), and X-ray photoelectron energy (XPS). The Caco2 cells were used to investigate the effects of Se-enriched A8-1 on the viability, membrane integrity, and the regulation of cellular inflammation through MTT and ELISA assays. The selenium-enriched metabolic function of A8-1 was analyzed by transcriptome sequencing. RESULTS: E. durans A8-1 has the ability to synthesize intracellular SeNPs that are incubated with 60 mg/L sodium selenite for 18 h at 37 °C with 7 % inoculum under aerobic conditions. The selenium-enriched transformation rate increased to 43.46 %. After selenium enrichment, there were no significant morphological changes in E. durans A8-1 cells. The cells also exhibited no cytotoxicity when incubated with Caco-2 cells, and increased cellular proliferation. Furthermore, Se-enriched A8-1 cells antagonize the adhesion of S. typhimurium ATCC14028 onto the surface of Caco-2 cells protecting cell membrane integrity and was assessed by measuring LDH and AKP activities (P <0.001, P <0.001). Moreover, Se-enriched A8-1 could protect Caco-2 cells from inflammation induced by lipopolysaccharide and help the cells alleviate the inflammation through the reduced expression of cytokine IL-8 (P = 0.0012, P <0.001) and TNF-α (P <0.001, P <0.001). Based on transcriptome sequencing in Se-enriched E. durans A8-1 cells, there were 485 up-regulated genes and 322 down-regulated genes (Padj < 0.05). There were 19 predicted up-regulated genes that are highly related to the potential selenium metabolism pathway, which focuses on the transportation of Na2SeO3 by membrane proteins, and gradually reduces Na2SeO3 to elemental selenium aggregates that are deposited onto the membrane surface via the intracellular redox response. CONCLUSION: E. durans A8-1 could convert extracellular selenite into intracellular biological SeNPs via redox pathway with strong selenium-rich metabolism, and its biological SeNPs have anti-inflammatory properties, which have the potential for the development of composite selenium nanomaterials and can be further studied for the function of SeNPs with potential applications.

Effects of dietary Pediococcus acidilactici GY2 single or combined with Saccharomyces cerevisiae or/and ß-glucan on the growth, innate immunity response and disease resistance of Macrobrachium rosenbergii.

One Pediococcus acidilactici strain, named PA-GY2 was isolated from the gut of cultured Macrobrachium rosenbergii. In order to better examine the potential scope and applicability of this strain in M. rosenbergii culture, based on the control diet, four experimental diets containing single or combined immunostimulants were produced by supplementing with yeast (Saccharomyces cerevisiae, SC) or/and ß-glucan (G), then fed to the prawns (6.70 g ± 0.74) in five groups, which were named as group C (control group), P (PA-GY2), PS (PA-GY2 + SC, 1:1), PG (PA-GY2 + G) and PGS (PA-GY2 + SC + G), respectively. After a 60-day feeding trial, growth performance, feed utilization, immune response and disease resistance of prawns were evaluated in the present study. Results indicated that (1) The growth performance of the prawns in group PS and PGS were significantly improved. The prawns in group PGS presented the lowest feed coefficiency (FC), while prawns in group C presented the highest FC. (2) The protease activity was significantly improved by dietary immunostimulants supplementation, meanwhile, prawns in the group PS presented the highest lipase activity. (3) The highest total hemocyte count and respiratory burst activity were found in the group P and PG, respectively. The phagocytic index of the prawns in the group C was significantly lower than those in group P and PGS. (4) Dietary PA-GY2 single or combined with SC or/and ß-glucan increased the immune related genes expression, including some antibacterial and antioxidant enzymes, while decreased the tumor necrosis factor-α gene expression, which led to the decreased cumulative mortality rate of prawns during the Aeromonas hydrophila challenge test. Based on the results of growth performance, digestive enzymes activity and immune response of M. rosenbergii, PA-GY2 supplementation, single or combined with SC or/and ß-glucan could be suggested as promising immunostimulants in prawns farming.

Ginseng polysaccharides enhanced ginsenoside Rb1 and microbial metabolites exposure through enhancing intestinal absorption and affecting gut microbial metabolism.

ETHNOPHARMACOLOGICAL RELEVANCE: Polysaccharides and small molecules commonly co-exist in decoctions of traditional Chinese medicines (TCMs). Our previous study outlined that ginseng polysaccharides (GP) could interact with co-existing ginsenosides to produce synergistic effect in an over-fatigue and acute cold stress model via gut microbiota involved mechanisms. AIM OF THE STUDY: This study aimed to verify the interactions by examining the impact of GP on oral pharmacokinetics of ginsenoside Rb1 (Rb1), the dominant protopanoxadiol (PPD)-type ginsenoside in Ginseng, on a dextran sulphate sodium (DSS) induced experimental colitis model which was characterized by gut dysbiosis, and to delineate the underlying mechanisms in vitro. MATERIALS AND METHODS: Rats received drinking water (normal group), 5% DSS (UC group), or 5% DSS plus daily oral administration of GP (GP group) for 7 days and fecal samples were collected on day -3, 0 and 6. On day 7 all animals received an oral dosage of Rb1 and blood samples were withdrawn for pharmacokinetic study. The in vitro metabolism study of Rb1 in gut microbiota from normal and UC rats and the transport study of Rb1 across Caco-2 cell monolayer were carried out in presence/absence of GP. Rb1 and its bacterial metabolites ginsenoside Rd (Rd), ginsenoside F2 (F2), Compound K (CK) and PPD were determined using LC-MS/MS. Total and target bacteria in fecal samples were determined by using 16S rRNA-based RT-PCR. ß-Glucosidase activity was determined by measuring 4-nitrophenol formed from 4-nitrophenyl-ß-D-glucopyranoside hydrolysis. RESULTS: DSS induction did not alter AUC0-t and Cmax of Rb1, which, however, were doubled together with elevated AUC0-t of the metabolites, in particular Rd and CK, in GP group. GP influenced the microbial composition and showed a prebiotic-like effect. Accordingly, GP treatment could partially restore the ß-glucosidase activity which was reduced by DSS induction. The presence of GP resulted in quicker microbial metabolism of Rb1 and higher Rd formation in first 8 h of incubation, while the impact on F2 and CK formation/conversion became obvious after 8 h. More interestingly, GP slightly stimulated Caco-2 cell growth and facilitated Rb1 transport across the Caco-2 monolayer in both directions, increasing the Papp of Rb1 from 10-7 cm/s to 10-6 cm/s. CONCLUSIONS: GP alleviated DSS-induced colitis-like symptoms and enhanced the systemic exposure of Rb1 through enhancing microbial deglycosylation and intestinal epithelial absorption of Rb1. These findings further demonstrated the important role of gut microbiota in the multifaceted action of polysaccharides in the holistic actions of traditional decoction of TCMs.

Transcriptional analysis of degenerate strain Clostridium beijerinckii DG-8052 reveals a pleiotropic response to CaCO3-associated recovery of solvent production.

Degenerate Clostridium beijerinckii strain (DG-8052) can be partially recovered by supplementing CaCO3 to fermentation media. Genome resequencing of DG-8052 showed no general regulator mutated. This study focused on transcriptional analysis of DG-8052 and its response to CaCO3 treatment via microarray. The expressions of 5168 genes capturing 98.6% of C. beijerinckii NCIMB 8052 genome were examed. The results revealed that with addition of CaCO3 565 and 916 genes were significantly up-regulated, and 704 and 1044 genes significantly down-regulated at acidogenic and solventogenic phase of DG-8052, respectively. These genes are primarily responsible for glycolysis to solvent/acid production (poR, pfo), solventogensis (buk, ctf, aldh, adh, bcd) and sporulation (spo0A, sigE, sigma-70, bofA), cell motility and division (ftsA, ftsK, ftsY, ftsH, ftsE, mreB, mreC, mreD, rodA), and molecular chaperones (grpE, dnaK, dnaJ, hsp20, hsp90), etc. The functions of some altered genes in DG-8052, totalling 5.7% at acidogenisis and 8.0% at sovlentogenisis, remain unknown. The response of the degenerate strain to CaCO3 was suggested significantly pleiotropic. This study reveals the multitude of regulatory function that CaCO3 has in clostridia and provides detailed insights into degeneration mechanisms at gene regulation level. It also enables us to develop effective strategies to prevent strain degeneration in future.

Sesamin ameliorates hepatic steatosis and inflammation in rats on a high-fat diet via LXRα and PPARα.

Nonalcoholic fatty liver disease (NAFLD) is defined by a nonalcohol relevant pathological accumulation of fat in the liver. Previous studies have shown that sesamin exerts antioxidant effects and improves lipid metabolism of the fatty liver. In this study, we hypothesized that sesamin improves lipid homeostasis of Sprague-Dawley rats fed a high-fat diet (HFD) by regulating the expression of genes related to de novo lipogenesis and ß-oxidation. We induced NAFLD in rats with HFD and examined the effect of sesamin in vivo. The results showed that HFD rats accumulated total cholesterol and triacylglycerols in the liver and developed inflammation, as evidenced by the elevation of interleukin-6 and tumor necrosis factor-α in the liver and serum. Sesamin attenuated the disease progression by improving the blood lipid profile in a dose-dependent manner. Sesamin reduced the serum levels of total cholesterol, triacylglycerols, low-density lipoprotein cholesterol, and free fatty acid, whereas it increased the level of high-density lipoprotein cholesterol. Meanwhile, sesamin increased the activities of hepatic glutathione peroxidase and superoxide dismutase while reducing the level of malonaldehyde and cytochrome P450 2E1. Furthermore, higher doses of sesamin reduced the expression of liver X receptor α and its downstream target genes, whereas it upregulated the peroxisome proliferator-activated receptor α-mediated signaling. These findings suggest that sesamin attenuates diet-induced dyslipidemia and inflammation of NAFLD in rats via mechanisms regulated by liver X receptor α and peroxisome proliferator-activated receptor α.

Proteomic analysis to elucidate degeneration of Clostridium beijerinckii NCIMB 8052 and role of Ca(2+) in strain recovery from degeneration.

Degeneration of solventogenic Clostridium strains is one of the major barriers in bio-butanol production. A degenerated Clostridium beijerinckii NCIMB 8052 strain (DG-8052) was obtained without any genetic manipulation. Supplementation of CaCO3 to fermentation medium could partially recover metabolism of DG-8052 by more than 50 % increase of cell growth and solvent production. This study investigated the protein expression profile of DG-8052 and its response to CaCO3 treatment. Compared with WT-8052, the lower expressed proteins were responsible for disruption of RNA secondary structures and DNA repair, sporulation, signal transduction, transcription regulation, and membrane transport in DG-8052. Interestingly, accompanied with the decreased glucose utilization and lower solvent production, there was a decreased level of sigma-54 modulation protein which may indicate that the level of sigma-54 activity may be associated with the observed strain degeneration. For the addition of CaCO3, proteomic and biochemical study results revealed that besides buffer capacity, Ca(2+) could stabilize heat shock proteins, increase DNA synthesis and replication, and enhance expression of solventogenic enzymes in DG-8052, which has a similar contribution in WT-8052.

Anticancer activity of binary toxins from Lysinibacillus sphaericus IAB872 against human lung cancer cell line A549.

The inhibitory effect of binary toxic (Bin) protein produced by Lysinibacillus sphaericus IAB872 on cell proliferation of human lung, liver, stomach and cervical tumor cell lines was assessed using MTT assay. The effect of Bin protein on A549 cell proliferation, apoptosis, cell cycle, migration and invasion were examined by MTT assay, Western blotting, Immunocytochemical staining, flow cytometry assay and wound-healing assay. Results showed that Bin protein inhibits proliferation of a range of human cancer cells in vitro. The anti-proliferative effect of Bin is associated with cell apoptosis as a result of an increased ratio of cellular Bax/bcl-2, up-regulated CyclinB1 and down-regulated Cdc25c expression, and its anti-proliferative action was associated with cell cycle arrest in the G2/M-phase. Bin protein could promote apoptosis and inhibit motility and invasion of A549 cancer cells. The anti-proliferative effect of Bin protein was associated with the induction of apoptotic cell death and cell cycle disruption. These results show that Bin protein has the potential to be developed as a chemotherapeutic agent by induction of human tumor cell apoptosis.

Traceability of Italian garlic (Allium sativum L.) by means of HRMAS-NMR spectroscopy and multivariate data analysis.

(1)H High Resolution Magic Angle Spinning-Nuclear Magnetic Resonance (HRMAS-NMR) spectroscopy was used to analyse garlic (Allium sativum L.) belonging to red and white varieties and collected in different Italian regions, in order to address the traceability issue. 1D and 2D NMR spectra, performed directly on untreated small pieces of garlic, so without any sample manipulation, allowed the assignment of several compounds: organic acids, sugars, fatty acids, amino acids and the nutritionally important fructo-oligosaccharides and allyl-organosulphur compounds. Application of Partial Least Squares projections to latent structures-Discrimination Analysis provided an excellent model for the discrimination of both the variety and, most important, the place origin, allowing the identification of the metabolites contributing to such classifications. The presence of organosulphurs, allicin and some allyl-organosulphurs found by HRMAS-NMR, was confirmed also by SPME-GC-MS; 11 molecules were identified, containing from one up to three sulphur atoms and with and without allyl moieties.