Proinflammatory Polarization of Macrophages Causes Intestinal Inflammation in Low-Birth-Weight Piglets and Mice.

BACKGROUND: Low-birth-weight (LBW) animals suffer from intestinal damage and inflammation in their early life. OBJECTIVES: The aim of this study was to investigate the role of macrophages in intestinal inflammation in LBW piglets and mice. METHODS: Major genes involved in intestinal barrier function such as claudin-1, zonula occludens-1 (ZO-1), occludin, and mucin 2 and inflammatory cytokines such as IL-1ß, TNF-α, IL-10, and IL-13 were evaluated in 21-day-old, normal-birth-weight (NBW) and LBW piglets and mice. Macrophage markers such as CD16/32, CD163, and CD206 were also assessed by immunofluorescence and flow cytometry. Polarized and unpolarized macrophages were further transferred into NBW and LBW mice, followed by an evaluation of intestinal permeability and inflammation. RESULTS: Claudin-1 mRNA in LBW piglets as well as claudin-1, occludin, ZO-1, and mucin 2 mRNAs in LBW mice, was significantly downregulated. IL-1ß and TNF-α were significantly upregulated in LBW piglets (P < 0.05). LBW mice showed a reduced expression of IL-10 and IL-13 (P < 0.05), with a heightened IL-6 level (P < 0.01) in the jejunum. CD16, a marker for M1 macrophages, was significantly elevated in the jejunum of LBW piglets, whereas CD163, a marker for M2 macrophages, was significantly decreased (P < 0.05). Similarly, LBW mice had more CD11b+CD16/32+ M1 macrophages (P < 0.05) and fewer CD206+ M2 macrophages (P < 0.01) than NBW mice. Moreover, the transfer of M1 macrophages exacerbated intestinal inflammation in LBW mice. Furthermore, 2 major glycolysis-associated genes, hexokinase 2 (HK2) and lactate dehydrogenase A (LDHA), were significantly upregulated in LBW piglets and mice (P < 0.05). CONCLUSIONS: This study revealed for the first time that the intestinal macrophages are polarized toward a proinflammatory phenotype in LBW piglets and mice, contributing to intestinal inflammation. The findings of this study provide new options for the management of intestinal inflammation in LBW animals.

The protective effect of Eleutheroside E against the mechanical barrier dysfunction triggered by lipopolysaccharide in IPEC-J2 cells.

Eleutheroside E (EE) exhibits immunocompetence, antioxidant, and anti-inflammatory activity. Lipopolysaccharide (LPS) can elicit a strong immune response. In vitro experiments were used to explore whether EE protects intestinal porcine jejunum epithelial cells (IPEC-J2) barriers from LPS stress. The experiment was divided into group C (control group: complete medium), group E (group C + 0.1 mg/mL EE), group L (group C + 10 µg/mL LPS), and group EL (adding 0.1 mg/mL EE for 6 h, and then adding 10 µg/mL LPS for culture). Finally, the cell proliferation, permeability, mRNA expression of cytokines, mRNA and protein expression of tight junctions (TJs) were analyzed. The result show that, when compared to the C group, EE significantly promoted the proliferation of IPEC-J2 at 58 h and showed low permeability (P < 0.05), the anti-inflammatory cytokines IL-10 and TGF-ß mRNA expression were increased extremely significantly, the inflammatory cytokines IL-6, TNF-α, and IFN-γ mRNA expression were extremely significantly decreased (P < 0.01), the mRNA and protein expression of TJ were significantly increased in group E (P < 0.05). However, LPS showed a damaging effect. EL group compared with L group, the cell index (CI) value was higher at 58 h (P < 0.05), the permeability was significantly lower (P < 0.05), the mRNA expressions of the inflammatory cytokines were down-regulated(P < 0.01), and the TJ mRNA and protein relative expression were increased (P < 0.05). In summary, the addition of EE protects the LPS-induced increase in permeability of IPEC-J2, potentially by expressing high levels of TJ proteins and inhibiting the increase of inflammatory cytokines.

Melatonin protects porcine oocyte from copper exposure potentially by reducing oxidative stress potentially through the Nrf2 pathway.

Copper is widely used as a feeding additive to promote livestock growth. However, excessive copper can be excreted with feces, causing heavy metal pollution and aggravating environmental problems. At the same time, studies have found that excess copper can cause damage to reproductive function and reduce gamete quality. Here, we explored the effects of adding different concentrations of copper to the culture medium on porcine oocytes. First polar body extrusion rate, embryo development, and intracellular levels of reactive oxygen species (ROS), mitochondrial membrane potential (MMP) ΔΨm, adenosine triphosphate(ATP) content, and acetylation of lysine 9 on histone H3 protein subunit (H3K9ac) were assessed. Results demonstrated that Cu exposure causes abnormalities in mitochondrial function and epigenetic modification, resulting in increased oxidative stress and levels of ROS, ultimately leading to a decreased porcine oocyte quality. In addition, we found melatonin can protect porcine oocytes from those damages. Notably, Nrf2 protein expression was significantly increased by copper exposure, meanwhile, Nrf2 signaling pathway inhibitor ML385 significantly attenuated the protective role of melatonin on oxidative stress induced by copper exposure. In summary, our study demonstrates that copper activates the Nrf2 pathway and impairs oocyte maturation by inducing oxidative stress, leading to poor quality of porcine oocytes, and the changes can be reversed by melatonin.

Effects of pyruvate on early embryonic development and zygotic genome activation in pigs.

Pyruvate is an important energy substance during early embryonic development of mammals. However, the underlying mechanisms of pyruvate during early embryonic development in pigs and its role in zygotic genome activation (ZGA) are not fully understood. Here, based on a previous RNA-seq dataset of porcine early embryos, we found that pyruvate metabolism-related genes started to be expressed at the 4-cell stage and that pyruvate metabolism-related genes were correlated with porcine ZGA marker genes. To determine the function of pyruvate in porcine embryos, in vitro fertilization (IVF) embryos were cultured in PZM-3 medium (control group); modified PZM-3 medium that only contains pyruvate and lactate plus salts (+P group); or modified PZM-3 medium lacking pyruvate (-P group). The 4-cell arrest rate at 72 h was significantly increased in the -P group compared to the +P group (P < 0.05). In addition, we observed that the reactive oxygen species (ROS) level was significantly increased and that the adenosine triphosphate (ATP) level was significantly (P < 0.05) decreased in the -P group compared to the +P group. Moreover, the expression of ZGA marker genes and SIRT1 protein in embryos was significantly decreased in the -P group compared to the +P group (P < 0.05). Furthermore, the acetylation level of H3K9 was significantly decreased (P < 0.05) and the methylation level of H3K9 was significantly increased (P < 0.05) in the -P group compared to the +P group. In summary, our findings demonstrate that pyruvate affects early embryonic development in pigs by promoting ZGA and reducing oxidative stress levels.

Mitigative effects of Eleutheroside E against the mechanical barrier dysfunction induced by soybean agglutinin in IPEC-J2 cell line.

Soybean agglutinin (SBA) is an anti-nutritional factor which decreases the mechanical barrier function in intestinal porcine jejunum epithelial cells (IPEC-J2). Eleutheroside E (EE) is a key part of Acanthopanax senticosus to exert pharmacological effects. This study aims to investigate the effects of EE on the barrier function in IPEC-J2 cells and to determine the ability of EE to enhance the protective effect of barrier function against SBA exposure. The IPEC-J2 cells were cultured in mediums with concentration of 0.1 mg/ml EE, 0.5 ml/ml SBA and 0.1 mg/ml EE pre-treated then treated with 0.5 mg/ml SBA. Then, the transepithelial electric resistance (TEER) value, inflammatory cytokines mRNA expression, tight junction mRNA and protein expression were tested by epithelial Voltohm meter, q-PCR and Western blot method respectively. The results showed that cells treated with 0.1 mg/ml EE had lower permeability (p < 0.05) while 0.5 mg/ml SBA treatment had higher permeability through tested TEER, and higher tight junction proteins (Claudin-3 and ZO-1) expressions and genes (Claudin-3, Occludin and ZO-1) expressions (p < 0.05) in 0.1 mg/ml EE group. IPEC-J2 cells pre-treated with 0.1 mg/ml EE could significantly improve the inflammatory response caused by 0.5 mg/ml SBA by up-regulation for IL-10, TGF-ß, and down-regulation gene expression of IL-6, TNF-α and IFN-γ (p < 0.05). In conclusion, 0.1 mg/ml EE can improve the mechanical barrier function and could protect the effects while 0.5 mg/ml of SBA-induced barrier dysfunction in IPEC-J2.

N-Acetyl-d-galactosamine prevents soya bean agglutinin-induced intestinal barrier dysfunction in intestinal porcine epithelial cells.

Soya bean agglutinin (SBA) is a glycoprotein and the main anti-nutritional component in most soya bean feedstuffs. It is mainly a non-fibre carbohydrate-based protein and represents about 10% of soya bean-based anti-nutritional effects. In this study, we sought to determine the effects of N-Acetyl-D-galactosamine (GalNAc or D-GalNAc) on the damage induced by SBA on the membrane permeability and tight junction proteins of piglet intestinal epithelium (IPEC-J2) cells. The IPEC-J2 cells were pre-cultured with 0, 0.125 × 10-4 , 0.25 × 10-4 , 0.5 × 10-4 , 1.0 × 10-4 and 2.0 × 10-4  mmol/L GalNAc at different time period (1, 2, 4 and 8 hr) before being exposed to 0.5 mg/ml SBA for 24 hr. The results indicate that pre-incubation with GalNAc mitigates the mechanical barrier injury as reflected by a significant increase in trans-epithelial electric resistance (TEER) value and a decrease in alkaline phosphatase (ALP) activity in cell culture medium pre-treated with GalNAc before incubation with SBA as both indicate a reduction in cellular membrane permeability. In addition, mRNA levels of the tight junction proteins occludin and claudin-3 were lower in the SBA-treated groups without pre-treatment with GalNAc. The mRNA expression of occludin was reduced by 17.3% and claudin-3 by 42% (p < 0.01). Moreover, the corresponding protein expression levels were lowered by 17.8% and 43.5% (p < 0.05) respectively. However, in the GalNAc pre-treated groups, occludin and claudin-3 mRNAs were reduced by 1.6% (p > 0.05) and 2.7% (p < 0.01), respectively, while the corresponding proteins were reduced by 4.3% and 7.2% (p < 0.05). In conclusion, GalNAc may prevent the effect of SBA on membrane permeability and tight junction proteins on IPEC-J2s.

Eleutheroside B increase tight junction proteins and anti-inflammatory cytokines expression in intestinal porcine jejunum epithelial cells (IPEC-J2).

Eleutheroside B (EB) is a phenylpropanoid glycoside with anti-inflammatory properties, neuroprotective abilities, immunomodulatory effects, antinociceptive effects, and regulation of blood glucose. The aim of this study was to investigate the effects of EB on the barrier function in the intestinal porcine epithelial cells J2 (IPEC-J2). The IPEC-J2 cells were inoculated into 96-well plates at a density of 5 × 103 cells per well for 100% confluence. The cells were cultured in the presence of EB at concentrations of 0, 0.05, 0.10, and 0.20 mg/ml for 48 hr. Then, 0.10 mg/ml was selected as the suitable concentration for the estimation of transepithelial electric resistance (TEER) value, alkaline phosphatase activity, proinflammatory cytokines mRNA expression, tight junction mRNA and protein expression. The results of this study indicated that the supplementation of EB in IPEC-J2 cells decreased cellular membrane permeability and mRNA expression of proinflammatory cytokines, including interleukin-6 (IL-6), interferon-γ (INF-γ), and tumour necrosis factor-α (TNF-α). The supplementation of EB in IPEC-J2 cells increased tight junction protein expression and anti-inflammatory cytokines, interleukin 10 (IL-10) and transforming growth factor beta (TGF-ß). In addition, the western blotting and real-time quantitative polymerase chain reaction (RT-qPCR) results indicated that EB significantly (p < 0.05) increased the mRNA and protein expression of intestinal tight junction proteins, Claudin-3, Occludin, and Zonula Occludins protein-1 (ZO-1). Therefore, dietary supplementation of EB may increase intestinal barrier function, tight junction protein expression, anti-inflammatory cytokines, and decrease proinflammatory cytokines synthesis in IPEC-J2 cells.

Effects of pulverized oyster mushroom (Pleurotus ostreatus) on diarrhea incidence, growth performance, immunity, and microbial composition in piglets.

BACKGROUND: Pleurotus ostreatus mushroom (POM) is an edible mushroom with rich nutritional components and vital pharmacological properties. The present study comprised 100 cross-bred piglets, weaned at 28 days old, who were randomly assigned to four POM diets with five replicates per diet and five piglets per pen. RESULTS: POM supplementation (P < 0.05) decreased the incidence of diarrhea, and also increased the average daily feed intake and average daily gain of pigs. Fecal acetate, butyrate and propionate increased with the addition of POM. Interleukin-2, immunoglobulin G, immunoglobulin M, tumor necrosis factor-α and immunoglobulin A increased (P < 0.05) with the addition of POM. The 16S rDNA sequencing results showed that the Bacteroidetes and Firmicutes were the dominant microbial strains in the fecal samples, irrespective of POM supplementation. Shannon diversity, whole tree phylogenetic diversity, observed species and Chao1 analysis exhibited significant variation in species richness across the treatments. Principal coordinates analysis showed a significant (P < 0.1) increase in the microbial communities amongst all of the treatment groups. CONCLUSION: The results of the present study suggest that the supplementation of POM in the diet of piglets might increase feed consumption, gut microbial composition and diversity, as well as short-chain fatty acids synthesis, consequently preventing the occurrence of diarrhea and increasing the growth of piglets. © 2019 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Interactions of Dietary Fibre with Nutritional Components on Gut Microbial Composition, Function and Health in Monogastrics.

The relation between dietary fibre and the well-being of human and other monogastrics has recently became a hot topic as shown by the increasing number of publications of the related research. The aim of this review is to describe - through a logical approach - the scientific suggestion linking possible benefits of dietary fibre on nutritional components and their effect on the gastrointestinal composition in relation to disease conditions in humans and animals. Dietary fibre plays a key role in: influencing blood glucose or insulin concentrations, stool bulkiness, reducing the pH within the digestive tract, synthesising volatile fatty acids (VFA), reducing intestinal transit time, stimulating growth of intestinal microbes, and constructively enhancing various blood parameters. The available literature suggests that fibre influences the bioavailability of nutrients and maintains the host's well-being by controlling disorders and disease prevalent with a Western way of living such as constipation and diarrhoea, diabetes, obesity, gastrointestinal inflammation, atherosclerosis, and colon cancer. Although there are some studies demonstrating that dietary fibre may be effective in the prevention and treatment of these disorders, the mechanisms involved are yet to be understood.

Effects of Dietary Iron Modulation on Gut Microbial Composition and Function in Monogastrics: A Review.

Dietary iron is a crucial nutrient element for biological processes of both hosts and gut microbiota. Deficiency in dietary iron is a highly common disorder in the developing locations of the world and can be healed by oral iron administration or complementary iron diet. While the redundant iron that enters the gut lumen leads to negative effects, and modulates the gut microbial composition and function. Such modulation led to a significant effect on vital biological pathways of the host, including metabolic disease (obesity and type 2 diabetes), metabolites (SCFA, blood glucose and cholesterol), bile acid metabolism, endocrine, neural, and other well-being patterns. This review covers the multifaceted aspects of different nutritional iron stress on the composition and function of microbial gut in monogastrics and consequential health conditions as well as it reveals unclear points that need further studies.

Effects of Dietary Non-Fiber Carbohydrates on Composition and Function of Gut Microbiome in Monogastrics: A Review.

Non-fiber carbohydrates (NFC) have a crucial function on the gut health of monogastrics. This paper aims to review the relevant published materials on the influence of NFCs on the gut's microbial population and composition in monogastrics, and points out the areas of the required research. Total bacteria count and Lactobacillus sp. were decreased with an increase in composition of dietary NFC intake, as well as accompanied by a decrease in the short-chain fatty acids (SCFAs) levels. Consequently, some metabolites were affected by the accumulation of the bile acids, including molecules which control different gene expression levels, as regulators involved in glucose (FXR and TGR5) and fat metabolism (cholesterol). Cell proliferation rate of both gastrointestinal epithelium and microbiome cells was negatively correlated with the dietary NFC levels in many species of monogastric animals. Low levels of NFC diet are negatively associated with digestibility, total gut weight, and gastrointestinal secretions. High levels of dietary NFC have negative effects on the digestion and absorption of macronutrients, with an increase of the contact time of the carcinogens in the intestinal lumen. The data obtained from different animals' studies did not give the same results. In conclusion, dietary NFC should be adjusted to the optimal consumption levels as the human and the monogastric animals are anatomically and physiologically different. Digestion, metabolism, host wellbeing, and host behavior were negatively affected by the administration of high NFC levels. The relations between sulphate-reducing bacteria and some metabolic diseases such as diabetes mellitus and obesity need further exploration.

Elevated expression of vascular endothelial growth factor (VEGF) 120 in parthenogenetic porcine placentas.

Most mammalian parthenogenetic embryos are unable to develop to term due to placental defects, potentially caused by decreased vasculogenesis and angiogenesis of the parthenogenetic placenta. Here we have compared the expression status of vascular endothelial growth factor (VEGF) and angiopoietin family members between normally developing and parthenogenetic porcine placentas. The result showed significantly reduced expression of these genes but elevated expression of VEGF 120 in the parthenogenetic porcine placenta (p < 0.05). We postulate that the abnormal expression levels of VEGF and angiopoietin family members and, especially, the elevated expression of VEGF 120 observed in parthenogenetic porcine placentas are related to the early miscarriage of parthenogenetic embryos in pigs.

Integrated QSAR study for inhibitors of Hedgehog Signal Pathway against multiple cell lines:a collaborative filtering method.

BACKGROUND: The Hedgehog Signaling Pathway is one of signaling pathways that are very important to embryonic development. The participation of inhibitors in the Hedgehog Signal Pathway can control cell growth and death, and searching novel inhibitors to the functioning of the pathway are in a great demand. As the matter of fact, effective inhibitors could provide efficient therapies for a wide range of malignancies, and targeting such pathway in cells represents a promising new paradigm for cell growth and death control. Current research mainly focuses on the syntheses of the inhibitors of cyclopamine derivatives, which bind specifically to the Smo protein, and can be used for cancer therapy. While quantitatively structure-activity relationship (QSAR) studies have been performed for these compounds among different cell lines, none of them have achieved acceptable results in the prediction of activity values of new compounds. In this study, we proposed a novel collaborative QSAR model for inhibitors of the Hedgehog Signaling Pathway by integration the information from multiple cell lines. Such a model is expected to substantially improve the QSAR ability from single cell lines, and provide useful clues in developing clinically effective inhibitors and modifications of parent lead compounds for target on the Hedgehog Signaling Pathway. RESULTS: In this study, we have presented: (1) a collaborative QSAR model, which is used to integrate information among multiple cell lines to boost the QSAR results, rather than only a single cell line QSAR modeling. Our experiments have shown that the performance of our model is significantly better than single cell line QSAR methods; and (2) an efficient feature selection strategy under such collaborative environment, which can derive the commonly important features related to the entire given cell lines, while simultaneously showing their specific contributions to a specific cell-line. Based on feature selection results, we have proposed several possible chemical modifications to improve the inhibitor affinity towards multiple targets in the Hedgehog Signaling Pathway. CONCLUSIONS: Our model with the feature selection strategy presented here is efficient, robust, and flexible, and can be easily extended to model large-scale multiple cell line/QSAR data. The data and scripts for collaborative QSAR modeling are available in the Additional file 1.

Decision tree and ensemble learning algorithms with their applications in bioinformatics.

Machine learning approaches have wide applications in bioinformatics, and decision tree is one of the successful approaches applied in this field. In this chapter, we briefly review decision tree and related ensemble algorithms and show the successful applications of such approaches on solving biological problems. We hope that by learning the algorithms of decision trees and ensemble classifiers, biologists can get the basic ideas of how machine learning algorithms work. On the other hand, by being exposed to the applications of decision trees and ensemble algorithms in bioinformatics, computer scientists can get better ideas of which bioinformatics topics they may work on in their future research directions. We aim to provide a platform to bridge the gap between biologists and computer scientists.