Dietary L-Methionine modulates the gut microbiota and improves the expression of tight junctions in an in vitro model of the chicken gastrointestinal tract.

BACKGROUND: The poultry industry encounters a number of factors that affect growth performance and productivity; nutrition is essential for sustaining physiological status and protecting against stressors such as heat, density, and disease. The addition of vitamins, minerals, and amino acids to the diet can help restore productivity and support the body's defense mechanisms against stress. Methionine (Met) is indispensable for poultry's energy metabolism, physiology, performance, and feed utilization capacity. Through this study, we aimed to examine the physiological effects of methionine supplementation on poultry as well as alterations of intestinal microbiome. METHODS: We utilized the DL- and L- form of methionine on Caenorhabditis elegans and the FIMM (Fermentor for intestine microbiota model) in-vitro digesting system. A genomic-analysis of the transcriptome confirmed that methionine supplementation can modulate growth-related physiological metabolic pathways and immune responses in the host poultry. The C. elegans model was used to assess the general health benefits of a methionine supplement for the host. RESULTS: Regardless of the type or concentration of methionine, supplementation with methionine significantly increased the lifespan of C. elegans. Feed grade L-Methionine 95%, exhibited the highest lifespan performance in C. elegans. Methionine supplementation increased the expression of tight junction genes in the primary intestinal cells of both broiler and laying hens, which is directly related to immunity. Feed grade L-Methionine 95% performed similarly or even better than DL-Methionine or L-Methionine treatments with upper doses in terms of enhancing intestinal integrity. In vitro microbial cultures of healthy broilers and laying hens fed methionine revealed changes in intestinal microflora, including increased Clostridium, Bacteroides, and Oscillospira compositions. When laying hens were given feed grade L-Methionine 95% and 100%, pathogenic Campylobacter at the genus level was decreased, while commensal bacteria were increased. CONCLUSIONS: Supplementation of feed grade L-Methionine, particularly L-Methionine 95%, was more beneficial to the host poultry than supplementing other source of methionine for maintaining intestinal integrity and healthy microbiome.

Solanum americanum genome-assisted discovery of immune receptors that detect potato late blight pathogen effectors.

Potato (Solanum tuberosum) and tomato (Solanum lycopersicon) crops suffer severe losses to late blight caused by the oomycete pathogen Phytophthora infestans. Solanum americanum, a relative of potato and tomato, is globally distributed and most accessions are highly blight resistant. We generated high-quality reference genomes of four S. americanum accessions, resequenced 52 accessions, and defined a pan-NLRome of S. americanum immune receptor genes. We further screened for variation in recognition of 315P. infestans RXLR effectors in 52 S. americanum accessions. Using these genomic and phenotypic data, we cloned three NLR-encoding genes, Rpi-amr4, R02860 and R04373, that recognize cognate P. infestans RXLR effectors PITG_22825 (AVRamr4), PITG_02860 and PITG_04373. These genomic resources and methodologies will support efforts to engineer potatoes with durable late blight resistance and can be applied to diseases of other crops.

Physiology of lipid digestion and absorption in poultry: An updated review on the supplementation of exogenous emulsifiers in broiler diets.

Lipids are a concentrated source of energy with at least twice as much energy as the same amount of carbohydrates and protein. Dietary lipids provide a practical alternative toward increasing the dietary energy density of feeds for high-performing modern broilers. However, the digestion and absorption of dietary lipids are much more complex than that of the other macronutrients. In addition, young birds are physiologically limited in their capacity to utilise dietary fats and oils effectively. The use of dietary emulsifiers as one of the strategies aimed at improving fat utilisation has been reported to elicit several physiological responses including improved fat digestibility and growth performance. In practical terms, this allows for the incorporation of lipids into lower-energy diets without compromising broiler performance. Such an approach may potentially lower feed costs and raise revenue gains. The current review revisits lipids and the different roles that they perform in diets and whole-body metabolism. Additional information on the process of dietary lipid digestion and absorption in poultry; and the physiological limitation brought about by age on lipid utilisation in the avian gastrointestinal tract have been discussed. Subsequently, the physiological responses resulting from the dietary supplementation of exogenous emulsifiers as a strategy for improved lipid utilisation in broiler nutrition are appraised. Suggestions of nascent areas for a better understanding of exogenous emulsifiers have been highlighted.

Efficacy and Equivalency of Phytase for Available Phosphorus in Broilers Fed an Available Phosphorus-Deficient Diet.

This study was conducted to assess the effectiveness of phytase on the performance, carcass traits, nutrient digestibility, tibia characteristics, and inositol phosphorus (IP) degradation in broiler chickens. Additionally, the available phosphorus (AP) equivalency of phytase in AP-deficient diets was estimated for 35 days after hatching. A total of 336 one-day-old Ross 308 broiler chicks were allocated to one of seven dietary treatments with six replications with eight birds per cage. The dietary treatments were as follows: (1) positive control containing 0.45% AP of the starter and 0.42% AP of the grower diet (PC), (2) 0.10% AP deficiency from the PC (NC-1), (3) 0.15% AP deficiency from the PC (NC-2), (4) 0.20% AP deficiency from the PC (NC-3), (5) NC-3 +phytase (500 FTU/kg; NC-3-500), (6) NC-3 + phytase (1000 FTU/kg; NC-3-1000), and (7) NC-3 + phytase (1500 FTU/kg; NC-3-1500). On d 35, the NC-3 diet exhibited lower tibia weight compared to the other treatments (p < 0.001). The NC-3-1500 group had higher calcium and phosphorus contents in the tibia than the other treatments on d 35 (p < 0.01). Phytase supplementation led to a reduced IP6 concentration and increased IP3 concentrations in different sections of the gastrointestinal tract on d 21 and 35 compared to the control diet (p < 0.05). In conclusion, based on the tibia phosphorus content, this study determined that 500 FTU/kg phytase was equivalent to 0.377% and 0.383% AP in the diet on d 21, and 0.317% and 0.307% AP in the diet on d 35, respectively. Likewise, 1000 FTU/kg was determined to be equivalent to 0.476% and 0.448% AP on d 21, and 0.437% and 0.403% AP on d 35, respectively. Furthermore, 1500 FTU/kg was determined to be equivalent to 0.574% and 0.504% AP on d 21, and 0.557% and 0.500 AP on d 35, respectively.

Exogenous emulsifiers and multi-enzyme combination improves growth performance of the young broiler chickens fed low energy diets containing vegetable oil.

OBJECTIVE: The present study examined the effects of exogenous emulsifiers and multi-enzyme supplementation into a low energy density diet on growth performance, visceral organ parameters, blood metabolites, ileal morphology, and nutrient digestibility in broiler chickens from hatch to 21 days. METHODS: One hundred and sixty-eight one-day-old Ross 308 broiler chickens were allocated in a completely randomized design to 24 pens and each pen was assigned to one of four dietary treatments to give six replications with seven chickens in a cage. Dietary treatments were: i) positive control with standard energy level (PC); ii) negative control with 100 kcal/kg lower energy of the standard level (NC); iii) NC diet supplemented 0.05% calcium stearoyl-2 lactylate as an emulsifier (NC+E); and iv) NC diet supplemented with both 0.05% calcium stearoyl-2 lactylate and 0.05% multi-enzyme (NC+E+M). Corn and soybean meal-based control diets containing vegetable oil were formulated to meet the Ross 308 nutrition specification. Chickens were fed ad-libitum with the treatment diets and sampling was conducted on day 21. RESULTS: Our results revealed that emulsifier and multi-enzyme supplementation into NC diets improved (p<0.05) feed efficiency of the broiler chickens compared to the broiler chickens fed NC diets from hatch to 21 days. Supplementation of emulsifier and multi-enzyme into NC diet improved (p<0.05) nutrient digestibility of the broiler chickens. However, emulsifier and multi-enzymesupplementation into diet did not influence (p>0.05) visceral organ weight, blood metabolites, and intestinal morphology in broiler chickens fed NC diets. CONCLUSION: Supplementation of emulsifier and multi-enzyme in the NC diet would support improving growth performance in young broiler chickens with improved feed efficiency and increased nutrient digestibility thereby curtailing the negative impact of energy reduction in the diets.

Supplementation of live yeast culture modulates intestinal health, immune responses, and microbiota diversity in broiler chickens.

The present study investigated the effects of live yeast cultures (LYC) on growth performance, gut health indicators, and immune responses in broiler chickens. A total of 720 mixed-sex broilers (40 birds/pen; 9 replicates/treatment) were randomly allocated to two dietary treatments: (1) a basal diet based on corn-soybean meal (CON) and (2) CON with 1 g/kg LYC. At 35 d of age, one bird per replicate pen was chosen for biopsy. LYC group tended (P < 0.10) to increase average daily gain during the grower phase compared with CON group. Broilers fed LYC diet had increased (P = 0.046) duodenal villus height and area but reduced (P = 0.003) duodenal crypt depth compared with those fed CON diet. Birds fed LYC diet presented alleviated (P < 0.05) serum TNF-α, IL-1ß, and IL-6 levels compared with those fed CON diet. Further, birds fed LYC diet exhibited upregulated (P < 0.05) ileal tight junction-related proteins and pro-inflammatory cytokines in the ileal tissue compared with those fed CON diet. Inverse Simpson's diversity (P = 0.038) revealed that birds fed CON diet had a more diverse microbiota community in the ileal digesta, compared with those fed LYC diet, while no significant difference between the treatments on Chao1 and Shannon's indices was observed. Based on the weighted UniFrac distance, the PCoA showed that microbiota in the ileal digesta of the LYC group was different from that of the CON group. LYC group increased the abundance of the phyla Firmicutes and genera Lactobacillus, Prevotella, and Enterococcus compared with CON group. The present study demonstrated that supplemental LYC as a feed additive provide supportive effects on enhancing gut functionality by improving the upper intestinal morphology and gut integrity, and modulating the immune system and microbiota communities of birds.

Live yeast culture (LYC) is composed of Saccharomyces cerevisiae and its metabolites such as mannan-oligosaccharides, peptides, nucleotides, vitamins and unknown growth factors. The supplementation of LYC is expected to exert health benefits in animals; however, the responses of broiler chickens to supplemental LYC is not fully explored. Thus, the present study evaluated the effects of LYC supplementation on growth performance, immune responses and intestinal health in broiler chickens. Based on the results from the present study, supplementation of LYC to a corn-based diet did not affect growth performance. Nonetheless, supplemental LYC improved intestinal morphology, upregulated tight junction-related protein genes and altered ileal microbiota diversity, suggesting its health benefits in improving gut health. In addition, supplemental LYC modulated serum immune responses and ileal cytokine genes expression, presenting its immunomodulatory potential.

The comparisons of expression pattern reveal molecular regulation of fruit metabolites in S. nigrum and S. lycopersicum.

Solanum nigrum, known as black nightshade, is a medicinal plant that contains many beneficial metabolites in its fruit. The molecular mechanisms underlying the synthesis of these metabolites remain uninvestigated due to limited genetic information. Here, we identified 47,470 unigenes of S. nigrum from three different tissues by de novo transcriptome assembly, and 78.4% of these genes were functionally annotated. Moreover, gene ontology (GO) analysis using 18,860 differentially expressed genes (DEGs) revealed tissue-specific gene expression regulation. We compared gene expression patterns between S. nigrum and tomato (S. lycopersicum) in three tissue types. The expression patterns of carotenoid biosynthetic genes were different between the two species. Comparison of the expression patterns of flavonoid biosynthetic genes showed that 9 out of 14 enzyme-coding genes were highly upregulated in the fruit of S. nigrum. Using CRISPR-Cas9-mediated gene editing, we knocked out the R2R3-MYB transcription factor SnAN2 gene, an ortholog of S. lycopersicum ANTHOCYANIN 2. The mutants showed yellow/green fruits, suggesting that SnAN2 plays a major role in anthocyanin synthesis in S. nigrum. This study revealed the connection between gene expression regulation and corresponding phenotypic differences through comparative analysis between two closely related species and provided genetic resources for S. nigrum.

Feeding a calcium-enriched fatty acid could ameliorate the growth performance of broilers under the chronic heat stress.

The current study was conducted to evaluate the effect of calcium-enriched fatty acid supplementation on the growth performance, blood metabolites, intestinal morphology, carcass traits, and nutrient digestibility of broilers subjected to chronic heat stress. A total of 210 one-day-old broiler chicks (40.12 ± 0.25 g) were randomly allocated to one of five dietary treatments, to obtain six replicates per treatment. Broilers were subjected to chronic heat stress from day 21 to day 35, post-hatching, at 34°C for 9 h per day. The body weight (BW) and feed intake of the experimental broilers were recorded weekly, and the average daily gain (ADG) and feed conversion ratio (FCR) were calculated accordingly. Rectal temperature was measured to compare the basal body temperatures between individuals, and blood samples were collected on days 21 and 35 to evaluate basal body temperature, serum total cholesterol, and the triglyceride content of the broilers. On days 21 and 35, one broiler from each cage (n=6) was euthanized to measure carcass trait parameters, nutrient digestibility in digesta, and intestinal morphology. On days 14, 28, and 35, the broilers fed 2.0% calcium-enriched fatty acids had higher BW (p < 0.05) than those fed the other diets. However, no differences (p > 0.05) were found in the average daily feed intake (ADFI) between dietary treatments over the 35 experimental days. On the other hand, on day 21, post-hatching, the broilers fed the 2.0% calcium-enriched fatty acid diet had improved (p < 0.05) dietary feed efficiencies compared to the other treatments. On day 28, the broilers fed the 5.0% of calcium-enriched fatty acid diet also had higher (p < 0.05) dietary feed efficiencies than those fed with the other dietary treatments. No effects (p > 0.05) on carcass weight, nutrient digestibility, intestinal morphology, or blood parameters were found between broilers fed with dietary treatments. This study demonstrated that the inclusion of an additive, containing 2.0% calcium-enriched fatty acid, to broiler diet could ameliorate the negative growth performance of broilers; and no interaction (p > 0.05) was observed between the calcium-enriched fatty acid and nutrient digestibility, digestive anatomy, blood metabolism, and carcass traits of broilers subjected to chronic heat stress conditions for 35 days post-hatching.

Osteoinductive function of fucoidan on periodontal ligament stem cells: Role of PI3K/Akt and Wnt/ß-catenin signaling pathways.

BACKGROUND/OBJECTIVES: Fucoidan has been focused as a multifunctional therapeutic uses including bone health supplements. However, the critical molecular mechanisms of fucoidan for bone therapeutic agents have not been fully understood. We investigated the osteoinductive effect of fucoidan on periodontal ligament stem cells (PDLSCs) and how this polymer encouraged PDLSC osteogenesis. MATERIALS AND METHODS: Osteogenic induction of PDLSCs was processed by culturing cells with fucoidan treatment. Osteogenic differentiation of PDLSCs was verified by alkaline phosphatase (ALP) activity, matrix mineralization assay, intracellular calcium levels, and mRNA expression and protein levels of osteogenic markers. RESULTS: Fucoidan treatment showed higher osteogenic activity in the PDLSCs than the control groups. PDLSCs with fucoidan also presented increased levels of the phosphatidylinositol-3-kinase (PI3K) isoforms, p110α and p110γ compared to control cells. The phosphorylation of Akt, a PI3K downstream effector, was significantly increased at 90 min of fucoidan induction. Expression of ß-catenin, a coactivator of canonical Wnt pathways, was increased in PDLSCs with fucoidan. ß-catenin was found to link with PI3K activation during the fucoidan stimulation. When cells were blocked by PI3K inhibitor or ß-catenin-specific siRNA, fucoidan-induced osteogenic activity of PDLSCs was significantly attenuated. CONCLUSION: These findings suggest that the fucoidan stimulates osteogenic differentiation of PDLSCs via the PI3K/Akt and Wnt/ß-catenin pathways.

Moderate dietary boron supplementation improved growth performance, crude protein digestibility and diarrhea index in weaner pigs regardless of the sanitary condition.

OBJECTIVE: The study was conducted to investigate the impact of boron supplementation on nutrient digestibility, inflammatory responses, blood metabolites and diarrhea index, and their relevance to growth performance in weaned pigs housed in good and poor sanitary environments for 14 days after weaning. METHODS: A total of 108 male pigs (Duroc×[Yorkshire×Landrace]) weaned at 21 days of age were used in a randomized complete block design with 2×3 factorial arrangement. Pigs were assigned to three boron treatments (0, 5, and 10 mg/kg) under two environments (good and poor sanitary) to give six replicates per treatment (3 pigs per replicate). On 0, 7, and 14 days, one pig per replicate was euthanized to collect, ileum tissue samples, and rectal fecal samples. RESULTS: Boron supplementation quadratically influenced (p<0.001) feed intake and weight gain in pigs housed in good sanitary conditions from 1 to 14 days post-weaning where pigs offered 5 mg/kg boron optimized weight gain and feed intake. There is a quadratic interaction (p = 0.019) on feed intake for 1 to 14 days post-weaning where 5 mg/kg boron increased feed intake in good sanitary conditions. Pigs housed in the poor sanitary environment decreased (p<0.001) villus height and crypt depth in ileum at days 7 and 14. On day 7 and 14, crude protein digestibility was quadratically influenced (p<0.05) by boron supplementation. Boron supplementation linearly increased (p<0.05) plasma calcium and cholesterol levels whilst linearly (p = 0.005) reducing plasma triglyceride concentrations. Diarrhea index was quadratically influenced (p<0.05) by boron supplementations regardless of sanitary conditions where 5 mg/kg boron inclusion achieved the lowest diarrhea index. CONCLUSION: Pigs offered 5 mg/kg of boron increased weight gain which may be deduced by improved dry matter, crude protein, and energy digestibility regardless of the sanitary conditions.

Aristolactam BIII, a naturally derived DYRK1A inhibitor, rescues Down syndrome-related phenotypes.

BACKGROUND: Dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) is a significant pathogenic factor in Down syndrome (DS), wherein DYRK1A is overexpressed by 1.5-fold because of trisomy of human chromosome 21. Thus, DYRK1A inhibition is considered a therapeutic strategy to modify the disease. PURPOSE: This study aims to identify a novel DYRK1A inhibitor and validate its therapeutic potential in DS-related pathological conditions. STUDY DESIGN: In order to identify a novel DYRK1A inhibitor, we carried out two-step screening: a structure-based virtual screening of > 300,000 chemical library (first step) and cell-based nuclear factor of activated T-cells (NFAT)-response element (RE) promoter assay (second step). Primary hits were evaluated for their DYRK1A inhibitory activity using in vitro kinase assay and Tau phosphorylation in mammalian cells. Confirmed hit was further evaluated in pathological conditions including DYRK1A-overexpressing fibroblasts, flies, and mice. RESULTS: We identified aristolactam BIII, a natural product derived from herbal plants, as a novel DYRK1A inhibitor. It potently inhibited the kinase activity of DYRK1A in vitro (IC50 = 9.67 nM) and effectively suppressed DYRK1A-mediated hyperphosphorylation of Tau in mammalian cells. Aristolactam BIII rescued the proliferative defects of DYRK1A transgenic (TG) mouse-derived fibroblasts and neurological and phenotypic defects of DS-like Drosophila models. Oral administration of aristolactam BIII acutely suppressed Tau hyperphosphorylation in the brain of DYRK1A TG mice. In the open field test, aristolactam BIII significantly ameliorated the exploratory behavioral deficit of DYRK1A TG mice. CONCLUSION: Our work revealed that aristolactam BIII as a novel DYRK1A inhibitor rescues DS phenotypes in cells and in vivo and suggested its therapeutic potential for the treatment of DYRK1A-related diseases.

Multi-tissue transcriptomic analysis reveals that L-methionine supplementation maintains the physiological homeostasis of broiler chickens than D-methionine under acute heat stress.

The objective of this study was to compare the effects of supplementation with two methionine isoforms, L-methionine (L-Met) or D-methionine (D-Met), on transcriptome expression in broiler chickens under acute heat stress. A total of 240 one-day-old chicks were randomly assigned to one of four treatments in a 2 × 2 factorial arrangement: thermo-neutral vs. acute heat-stress and L-Met vs. D-Met supplementation. On day 14, the heat-stressed group was exposed to 32°C for 5 h, while the others remained at 25°C. Six chicks were randomly selected per treatment and total RNA was isolated from whole blood, ileum, and liver tissues. Two RNA samples from each tissue of each treatment group were randomly selected and pooled in equal amounts. A total of 1.87 billion raw reads obtained from 36 samples (four treatments × three tissues × three composited replicates) were mapped to the reference genome build (Gallus_gallus-5.0) and used to identify differentially expressed genes (DEGs) using DESeq2. Functional enrichment of DEGs was tested using DAVID. Comparing the two isoforms of supplemented methionine, two, three, and ten genes were differentially expressed (> 1 or < -1 log2 fold change) in whole blood, ileum, and liver, respectively. A total of 38, 71, and 16 genes were differentially expressed in response to the interaction between heat stress and Met isoforms in the blood, ileum, and liver, respectively. Three-tissue-specific DEGs were functionally enriched for regulation of cholesterol homeostasis and metabolism, glucose metabolism, and vascular patterning. Chicks fed with L-Met had lower immune (e.g., IL4I1 and SERPINI1) and intestinal angiogenic responses (e.g., FLT1 and FGD5), and stable glucose and lipid metabolism (e.g., PCK1 and LDLR) under heat stress conditions. In conclusion, unlike D-Met, L-Met supplementation seems to help maintain physiological homeostasis and enhances cellular defense systems against external stresses like high environmental temperature.

Stimbiotic supplementation improved performance and reduced inflammatory response via stimulating fiber fermenting microbiome in weaner pigs housed in a poor sanitary environment and fed an antibiotic-free low zinc oxide diet.

This study investigated whether the inclusion of a stimbiotic (STB) can improve performance, influence intestinal microbiota and fermentation activity, and reduce pro-inflammatory cytokines in piglets fed a low zinc oxide diet without antimicrobial growth promotors compared to fructo-oligosaccharide (FOS) and mannan-oligosaccharide (MOS) when housed either in good sanitary (GS) or poor sanitary (PS) environments. One hundred forty-four male pigs (28-day-old) were sorted by initial body weight (BW) and allocated to one of six experimental treatments: 1) GS environment without any additive (GS-CTR); 2) GS environment with 0.01% stimbiotic (GS-STB); 3) PS environment (without cleaning and disinfection of a previously populated room) without any additive (PS-CTR); 4) PS environment with 0.01% STB (PS-STB); 5) PS environment with 0.1% MOS (PS-MOS); and 6) PS environment with 0.2% FOS (PS-FOS). Each treatment had six replicates, with four animals each. Three feeding phases, based on corn, wheat, and soybean meal were available ad libitum for the 42-days of the study. Housing piglets under PS conditions negatively influenced performance, increased plasma tumor necrosis factor alpha (TNF-α), affected the fecal microbial populations and increased concentrations of branched-chain fatty acids (BCFA) compared to GS. Stimbiotic improved 42-d-BW under PS conditions (P < 0.05) whereas MOS or FOS had no effect. On d35, plasma TNF-α was reduced with STB in PS (P < 0.05). The ratio between VFA:BCFA increased (P < 0.05) with STB, MOS or FOS in PS, and under GS condition, STB also increased the ratio. Stimbiotic increased the proportion of Clostridiales Family XIII Incertae Sedis and Clostridiaceae, while MOS and FOS increased Selenomonadaceae, Catabacteriaceae and Fibrobacteraceae. These results indicate that STB shifted the intestinal microbiome to favor fiber fermentation which likely contributed to reduced inflammatory response and improved performance, particularly in piglets reared in PS conditions.

Supplement of nitric oxide through calcium carbonate-based nanoparticles contributes osteogenic differentiation of mouse embryonic stem cells.

This study investigated the delivery of S-nitrosothiol (GSNO) as a nitric oxide (NO) donor loaded into calcium carbonate-based mineralized nanoparticles (GSNO-MNPs) to regulate cell signaling pathways for the osteogenic differentiation of mouse embryonic stem cells (ESCs). GSNO-MNPs were prepared by an anionic block copolymer template-mediated calcium carbonate (CaCO3) mineralization process in the presence of GSNO. GSNO-MNPs were spherical and had a narrow size distribution. GSNO was stably loaded within the MNPs without denaturation. TEM analysis also demonstrated the localization of GSNO-MNPs within membrane-bound structures in the cell, indicating the successful introduction of GSNO-MNPs into the cytosol of ESCs. Intracellular levels of NO and cGMP were significantly increased upon treatment with GSNO-MNPs, compared with the control group. When cells were exposed to GSNO-MNPs, the effects of nanoparticles on cell viability were not statistically significant. GSNO-MNPs treatment increased ALP activity assay and intracellular calcium levels. Real-time RT-PCR also revealed highly increased expression levels of the osteogenic target genes ALP, osteocalcin (OCN), and osterix (OSX) in GSNO-MNP-treated ESCs. The protein levels of OSX and Runt-related transcription factor 2 (RUNX2) showed similar patterns of expression based on real-time RT-PCR. These results indicate that GSNO-MNPs influenced the osteogenic differentiation of ESCs. Transcriptome profiling identified several significantly enriched and involved biological networks, such as RAP1, RAS, PI3K-AKT, and MAPK signaling pathways. These findings suggest that GSNO-MNPs can modulate osteogenic differentiation in ESCs via complex molecular pathways.

Effects of Adding Phytase from Aspergillus niger to a Low Phosphorus Diet on Growth Performance, Tibia Characteristics, Phosphorus Excretion, and Meat Quality of Broilers 35 days after hatching.

This study was designed to determine the effect of phytase extracted from Aspergillus niger (Natuphos® E) on growth performance, bone mineralization, phosphorous excretion, and meat quality parameters in broilers fed available phosphorous (aP)-deficient diet. In total, 810 one-day-old Indian River broilers were randomly allotted into one of three dietary treatments, with six replicates per treatment. The three dietary treatments were 1) control group (CON: basal diet with sufficient aP), 2) low phytase (LPY: available phosphorus-deficient diet supplemented with 0.01% phytase), and 3) high phytase (HPY: available phosphorus-deficient diet supplemented with 0.02% phytase). Average daily gain and, feed intake, and feed conversion ratio were measured for 35 days. Excreta were collected from each pen on day 35. One broiler from each cage was euthanized to collect visceral organs and tibia samples. Broiler chickens fed LPY and HPY showed improved (P<0.05) growth performance compared to broilers fed CON on day 35. The tibia length of HPY-fed broilers was more than those of broilers fed other diets on day 35 (P<0.05). However, tibia calcium and phosphorous contents in LPY-fed broilers was higher (P<0.05) than in CON and HPY-fed broilers. Tibia length and calcium and phosphorous content showed a positive correlation (P<0.05) with the weight gain of broilers on day 35. Phosphorous level in the excreta of LPY- and HPY-fed broilers was lesser than those of CON broilers on day 35 (P<0.05). Furthermore, HPY-fed broilers showed lower (P<0.05) phosphorous content in the excreta than LPY-fed broilers. LPY- and HPY-fed broilers showed higher (P<0.05) liver weight than the CON broilers. In conclusion, broilers fed aP-deficient diet supplemented with phytase from Aspergillus excreted less phosphorus, which enhanced growth performance and tibia development from time of hatching to day 35 post-hatching.

Effect of calcium stearoyl-2 lactylate and lipase supplementation on growth performance, gut health, and nutrient digestibility of broiler chickens.

OBJECTIVE: To evaluate calcium stearoyl-2 lactylate (CSL) performance as an exogenous emulsifier together with lipase for broiler diets. METHODS: In total, 252 one-day-old Ross 308 broiler chickens were allocated in a completely randomized design to give 6 replications per treatment with 7 birds in each cage. There were six dietary treatments representing a 2×3 factorial arrangement consisted of two energy levels (standard energy [positive control, PC] and -100 kcal/kg of the requirement level [negative control, NC]) and three dietary treatments (without additives [CON], CON+CSL [CSL], and CON+CSL+lipase [CSL-Lipase]). Corn and soybean meal-based experimental diets containing vegetable oil were formulated. Growth performance, blood parameters, visceral organ weights, ileal morphology, nutrient digestibility, and cytokine gene expression were measured. RESULTS: Birds fed a diet including CSL increased (p<0.05) lipase level in blood compared to birds fed a diet including CSL-Lipase on day 21. Similarly, higher (p<0.05) liver weight was observed in birds fed a diet including either CSL or CSL-Lipase on day 21. Birds fed NC diet with CSL improved (p<0.05) nutrient digestibility compared to the NC diet on day 21. However, birds fed a diet supplemented with CSL or CSL-Lipase did not affect (p>0.05) the weight gain, feed efficiency, ileal morphology, and cytokine concentrations during the experiment period, regardless of dietary energy levels. CONCLUSION: Our results indicated that CSL has a role in improving nutrient digestibility in young birds when supplemented to a corn-soybean meal based broiler diet.

Broilers fed a low protein diet supplemented with synthetic amino acids maintained growth performance and retained intestinal integrity while reducing nitrogen excretion when raised under poor sanitary conditions.

The present study investigated the effects of supplementing a low protein (LP) diet supplemented with key essential amino acids (AA) to broilers on growth performance, intestinal tract function, blood metabolites, and nitrogen excretion when the animals were maintained under various sanitary conditions for 35 D after hatching. Three hundred eighty-four one-day-old male broilers (Ross 308) were randomly allotted to groups that received one of 6 dietary treatments in a 2 × 3 factorial arrangement (i.e., 2 environmental conditions and 3 dietary treatments) to give 8 replicates per treatment. Broilers were challenged with 2 environmental conditions (sanitary vs. poor sanitary). The dietary treatments were (1) high protein (HP) diet, (2) LP diet, and (3) LP diet with synthetic key essential AA (LPA): the LP diet was supplemented with synthetic AA up to the required levels for broilers. On day 14, birds consumed the LP diet impaired growth performance compared with those fed the HP diet, while the average daily weight gain-to-feed conversion ratio of birds fed the LPA diet improved to the level of birds fed the HP diet under poor sanitary conditions (P < 0.05). Broilers raised under poor sanitary conditions and fed the LP diet displayed higher (P < 0.05) zonula occludens (ZO-1) expression on day 14 than broilers fed either the HP or LPA diet. Under sanitary conditions, birds fed HP and LPA diets showed higher villus height and crypt depth compared with those of broilers fed the LP diet on day 35. Moreover, broilers raised in the poor sanitary environment had higher (P < 0.05) serum endotoxins than those raised in the sanitary environment. Broilers fed the LPA diet showed reduced (P < 0.05) nitrogen excretion on days 14 and 35 compared with those fed the LP and HP diets independent of the environment. In conclusion, the LPA diet did not impair growth performance under poor sanitary conditions for 14 D after hatch while resulting in lower nitrogen excretion in any environment conditions throughout the experiment.

Identification of small-molecule compounds targeting the dishevelled PDZ domain by virtual screening and binding studies.

The Dishevelled (Dvl) protein, which conveys signals from receptors to the downstream effectors, is a critical constituent of the Wnt/ß-catenin signaling pathway. Because the PDZ domain of Dvl protein functions through associations with a wide range of protein partners, Dvl protein involved in the Wnt signaling pathway has been considered to be therapeutic targets in cancers. In this study, we performed structure-based pharmacophore model of the Dvl PDZ domain to discover novel small-molecule binders and identified eight compounds with micromolar affinity. The most potent compound identified, BMD4702, efficiently bound to the Dvl PDZ domain with 11.2µM affinity and had a 0.186µM KD value according to surface plasmon resonance and fluorescence spectroscopy, respectively. Combining both structural-kinetic relationship analyses and docking studies, we fourmulated that the ligand-binding site is composed of three H-bonds and three hydrophobic features. Thus, our approach led to the identification of potent binders of the Dvl PDZ domain and the findings provide novel insights into structure-based approaches to design high-affinity binders for the Dvl PDZ domain.

Small molecule inhibitors of the Dishevelled-CXXC5 interaction are new drug candidates for bone anabolic osteoporosis therapy.

Bone anabolic agents promoting bone formation and rebuilding damaged bones would ideally overcome the limitations of anti-resorptive therapy, the current standard prescription for osteoporosis. However, the currently prescribed parathyroid hormone (PTH)-based anabolic drugs present limitations and adverse effects including osteosarcoma during long-term use. Also, the antibody-based anabolic drugs that are currently being developed present the potential limits in clinical application typical of macromolecule drugs. We previously identified that CXXC5 is a negative feedback regulator of the Wnt/ß-catenin pathway via its interaction with Dishevelled (Dvl) and suggested the Dvl-CXXC5 interaction as a potential target for anabolic therapy of osteoporosis. Here, we screened small-molecule inhibitors of the Dvl-CXXC5 interaction via a newly established in vitro assay system. The screened compounds were found to activate the Wnt/ß-catenin pathway and enhance osteoblast differentiation in primary osteoblasts. The bone anabolic effects of the compounds were shown using ex vivo-cultured calvaria. Nuclear magnetic resonance (NMR) titration analysis confirmed interaction between Dvl PDZ domain and KY-02061, a representative of the screened compounds. Oral administration of KY-02327, one of 55 newly synthesized KY-02061 analogs, successfully rescued bone loss in the ovariectomized (OVX) mouse model. In conclusion, small-molecule inhibitors of the Dvl-CXXC5 interaction that block negative feedback regulation of Wnt/ß-catenin signaling are potential candidates for the development of bone anabolic anti-osteoporosis drugs.

Effects of dietary Forsythia suspensa on feed utilization, rumen fermentation, and immune response of Korean native goats (Capra hircus) / Efectos de la adición dietaria de Forsythia suspensa sobre utilización del alimento, fermentación ruminal y respuesta inmune en cabras nativas coreanas (Capra hircus) / Efeitos na dieta pela utilização de Forsythia suspensa nos alimentos, fermentação ruminal e resposta imune em caprinos nativos coreanos (Capra hircus)

Background: plant extracts of Forsythia suspensa, which contain pinoresinol, have been proposed as an alternative to antibiotics due to their antioxidant, anti-inflammatory, and microflora modulating effects. Objective: to evaluate the effects of dietary F. suspensa on feed utilization, rumen fermentation, blood parameters and immune response of Korean native goats (Capra hircus). Methods: a total of nine Korean native goats were assigned to one of three dietary treatments: 1) a basal diet without F. suspensa, 2) a basal diet added with 0.25% F. suspensa, or 3) a basal diet added with 0.50% F. suspensa. A change-over design was used in three consecutive periods to give 9 replicates per treatment. Each period lasted 18 days, including 15 days of adaptation to feed and environmental conditions and three days of total collection of feces and urine. Rumen fluid and blood samples were also collected. Nutrient digestibility, nitrogen retention, ruminal content, and blood metabolites—including immunoglobulins—were measured. Results: F. suspensa supplementation had no effect on nutrient digestibility, whole body nitrogen retention rate, ruminal pH, acetate, propionate, isobutyrate, butyrate, isovalerate, or valerate content (p>0.05). However, F. suspensa supplementation decreased total volatile fatty acids (VFA) concentration compared with the control diet, regardless of F. suspensa concentrations (p<0.05). Goats fed a diet containing 0.25% F. suspensa had greater alkaline phosphatase (ALP) concentration than those fed a control or a diet with 0.50% F. suspensa (p<0.05). Feeding different concentrations of dietary F. suspensa did not influence plasma IgA and IgM levels (p>0.05), although goats receiving both 0.25% and 0.50% F. suspensa diets had greater plasma IgG than those fed the control diet (p<0.05). Conclusion: supplementation of 0.25% F. suspensa significantly decreased total VFA concentrations and increased plasma IgG in Korean native goats, compared with those fed the control diet.

Antecedentes: los extractos de la planta Forsythia suspensa contienen pinoresinol. Dicha planta se ha propuesto como una alternativa a los antibióticos debido a su contenido de antioxidantes, antiinflamatorios y sus efectos moduladores de la microflora. Objetivo: evaluar los efectos dietarios de F. suspensa sobre la utilización del alimento, fermentación ruminal, parámetros sanguíneos y respuesta inmune en cabras nativas coreanas (Capra hircus). Métodos: un total de nueve cabras nativas coreanas fueron asignadas a 1 de 3 tratamientos: (1) una dieta basal sin F. suspensa, (2) una dieta basal con 0,25%, o (3) 0,50% de F. suspensa dietaría en un diseño change-over por tres periodos consecutivos con nueve réplicas por tratamiento. Cada periodo se prolongó por 18 días, incluyendo 15 días de adaptación al alimento y a las condiciones ambientales, y tres días de colección total de heces y orina. Tambien se colectó fluido ruminal y muestras sanguineas. Se midió la digestibilidad de nutrientes, nitrógeno retenido, contenido ruminal y metabolitos sanguíneos —incluyendo inmunoglobulinas—. Resultados: la suplementación con F. suspensa no tuvo efecto sobre la digestibilidad de los nutrientes, la tasa de retención de N, pH ruminal, ni sobre los contenidos de acetato, propionato, isobutirato, butirato, isovalerato o valerato (p>0,05). Sin embargo, la suplementación con F. suspensa disminuyo la concentración total de ácidos grasos volátiles (VFA) en comparacion con la dieta control, sin tener en cuenta las concentraciones de F. suspensa (p<0,05). Las cabras alimentadas con la dieta de 0,25% de F. suspensa tuvieron mayor concentración de fosfatasa alcalina (ALP) que cuando fueron alimentadas con la dieta control o con la de 0,50% de F. suspensa (p<0,05). Aunque la alimentación con diferentes concentraciones de F. suspensa no influyo en los niveles de IgA y IgM en plasma (p>0,05), las cabras que recibieron 0,25 y 0,50% de F. suspensa tuvieron mayor concentración de IgG en plasma que aquellas alimentadas con la dieta control (p<0,05). Conclusiones: la suplementación de 0,25% de F. suspensa disminuyo significativamente la concentración total de VFA e incrementó la IgG en el plasma de las cabras nativas coreanas, en comparación con aquellas que consumieron la dieta control.

Antecedentes: a Forsythiae suspensa é uma planta e seu extrato contém pinoresinol. Tem sido proposta como uma alternativa aos antibióticos existentes, devido ao seu teor de antioxidantes, anti-inflamatórias e efeitos moduladores da microflora, com base em estudos feitos in vivo e in vitro. Objetivo: avaliar os efeitos na dieta da inclusão de F. suspensa no aproveitamento alimentar, fermentação ruminal, parâmetros sanguíneos e resposta imune em caprinos coreanos nativos (Capra hircus). Métodos: um total de nove cabras coreanas nativas foram assignadas a um de três tratamentos na dieta: (1) uma dieta basal sem F. suspensa, (2) uma dieta basal com 0,25% de F. suspensa, ou (3) com 0,50% de F. suspensa na dieta. Se fez um desenho change-over por três períodos consecutivos com nove repetições por tratamento. Cada período de pesquisa consistiu de 18 dias, incluindo 15 dias de adaptação às condições alimentares e ambientais, e três dias de coleta total de fezes e urina. Também foram coletadas amostras de líquido ruminal e sangue. Mediou-se a digestibilidade dos nutrientes, o nitrogênio (N) retido, o conteúdo ruminal, e os parâmetros sanguíneos —incluindo imunoglobulinas—. Resultados: a suplementação com F. suspensa não teve nenhum efeito sobre a digestibilidade dos nutrientes, a taxa de retenção de N, o pH do rúmen, nem sobre os conteúdos de: acetato, propionato, isobutirato, butirato, isovalerato e valerato (p>0,05). No entanto, a suplementação de F. suspensa diminuiu a concentração total de ácidos gordurosos voláteis (AGV) em comparação com a dieta controle, independentemente das concentrações de F. suspensa (p<0,05). As cabras alimentadas com 0,25% F. suspensa na dieta tiveram maiores quantidades da enzima fosfatasse alcalina (ALP) do que quando elas foram alimentadas com a dieta controle ou com 0,50% de F. suspensa (p<0,05). Ainda que a alimentação com diferentes níveis de F. suspensa não influenciou os níveis de IgA e IgM no plasma (p>0,05), as cabras que receberam na dieta 0,25 e 0,50% de F. suspensa tiveram uma maior concentração de IgG no plasma comparadas as cabras alimentadas com a dieta controle (p<0,05). Conclusões: a suplementação com 0,25% e 50% diminuiu significativamente a concentração de AGV e incrementou a concentração em plasma de IgG em cabras nativas coreanas comparadas com aquelas que consumiram a dieta controle.