Intestinal Alkaline Phosphatase Research Articles

New findings of intestinal alkaline phosphatase: effects on intestinal and organ health of piglets challenged with ETEC F4 (K88)

The aim of this study was to assess the addition of intestinal alkaline phosphatase (IAP) to diets on the count of bacterial populations, pH of digestive organ contents, histopathological description, proinflammatory markers, hepatic glycogen reserve, and diarrhoea incidence of piglets challenged with Escherichia coli. Sixty-four crossbred piglets (7.16±0.28 kg body weight, 25-days-old) were assigned to four treatments in a completely randomised block design: negative control (NC), NC + antibiotic (ANT), NC + 15 mg IAP, or NC + 30 [...]

PATHOBIOME TRANSFORMATION AND FEATURES OF ENTERAL INSUFFICIENCY IN PATIENTS WITH PERITONITIS DURING THE COVID-19 PANDEMIC

We studied the features of acute enteral failure in patients with peritonitis in the setting of COVID-19 by studying the properties of the enteral pathobiome, biochemical, functional, pro-inflammatory and anti-apoptotic markers in diffuse peritonitis. A retrospective randomized cohort controlled study was conducted; it included 84 patients divided into two groups. Stratified randomization was used. The species composition and properties of the enteral luminal pathobiome were studied. 699 inoculations were made, resulting in 3024 isolates. The enteral morphofunctional coefficient, the level of alkaline phosphatase and its intestinal isoform were dynamically studied. In immunohistochemical studies, we studied the expression of CD-3 and Bcl-2. Patients with COVID-19 in addition to a decrease in the level of saprophytic microflora, showed a significant increase in Klebsiella spp., Candida, Clostridium spp. and Citrobacter (p≤0.05). The rate of antibiotic resistance in the comparison group was 8.3%, in the main group - 36.9%. MRSA pathogens were noted in 86.4% of cases, VRSA strains were found in 20.6% of cases. Patients with COVID-19 had a deficiency of intestinal alkaline phosphatase, which persisted even on the day 5 of treatment. Macro- and microscopic studies showed pronounced inflammatory and apoptotic changes in the intestinal wall of patients in the main group. A specific enteral pathobiome that is multiresistant to antibiotics, pronounced inflammatory and apoptotic changes in the intestinal wall, and a predisposition to perforation of the small intestine are characteristics of enteral failure in diffuse peritonitis in the setting of COVID-19.

Extraction, purification, and in vitro biological activities of intestinal alkaline phosphatase from pig intestine mucous waste

Extraction, purification, and in vitro biological activities of intestinal alkaline phosphatase from pig intestine mucous waste

Chitosan – Polyphosphate nanoparticles for a targeted drug release at the absorption membrane

The aim of this study was to develop nanoparticles (NPs) providing a targeted drug release directly on the epithelium of the intestinal mucosa.NPs were prepared via ionic gelation between cationic chitosan (Cs) and anionic polyphosphate (PP). The resulting NPs were characterized by their size, polydispersity index (PDI) and zeta potential. Isolated and cell-associated intestinal alkaline phosphatase (IAP) was employed to trigger polyphosphate cleavage in Cs-PP NPs which was quantified via malachite green assay. In parallel, the shift in zeta potential was determined. In-vitro drug release studies were performed in Franz diffusion cells with Cs-PP NPs containing rhodamine 123 as model active ingredient. Furthermore, cytotoxicity of Cs-PP NPs was assessed via resazurin assay on Caco-2 cells as well as via hemolysis assay on red blood cells.Cs-PP NPs exhibited an average size of 144.17 ± 10.95 nm and zeta potential of -12.6 ± 0.50 mV. The encapsulation efficiency of rhodamine 123 by Cs-PP NPs was 86.8%. After incubation with isolated IAP for 3 h the polyphosphate of Cs-PP NPs was cleaved to monophosphate and zeta potential raised up to -2.3 ± 0.30 mV. Cs-PP NPs showed a non-toxic profile. Within 3 h, 62.0 ± 10.8% and 14.1 ± 2.2% of total rhodamine 123 was released from Cs-PP NPs upon incubation with isolated as well as porcine intestine derived intestinal alkaline phosphatase (IAP), respectively.According to these results, Cs-PP NPs are promising drug delivery systems to enable a drug targeted release at the absorption membrane.

The immature intestinal epithelial cells in preterm infants play a role in the necrotizing enterocolitis pathogenesis: A review

Intestinal stem cells (ISCs) possess the self-renewal capacity to produce progenitor cells that differentiate into Paneth cells, enterocytes, goblet cells, or enteroendocrine cells in the small intestine. The differences in functionality and population of intestinal epithelial cells (IECs) between neonates (including premature and term infants) and adults for both humans and rodents are discussed. The impact of gestational age and postnatal age on each IEC type's functionality is explored. The specific characteristics of Paneth cells, goblet cells, and enterocytes that make preterm infants more vulnerable to infectious diseases than term infants or adults are described. These characteristics in the preterm intestine include a reduced thickness of mucus, lower intestinal alkaline phosphatase secreted by enterocytes, and diminished secretion of lysozymes by Paneth cells compared to term infants, which increase their risk of inflammation, dysbiosis, and development of necrotizing enterocolitis (NEC). Further studies are needed to characterize the critical role of growth factors, antibodies, cytokines, and immune cells from breast milk to prevent NEC development in preterm infants by promoting their maturation of IECs.

Adsorption and catalytic properties of enzymes on the surface of silicalite-1

Microporous hydrophobic silicalite-1 was used as a carrier for immobilization of different enzymes, such as horseradish peroxidase, calf intestinal alkaline phosphatase and two β-galactosidases of different origin, to create heterogeneous biocatalytic systems. The peculiarities of enzyme adsorption on the surface of silicalite-1, as well as catalytic properties of the obtained systems compared to enzyme activity in solution and on the surface of other carriers, are discussed.

Characterization and Structure of Alternatively Spliced Transcript Variant of Human Intestinal Alkaline Phosphatase (ALPI) Gene.

Intestinal-type alkaline phosphatase (IAP) is expressed at a high concentration in the brush border membrane of intestinal epithelial cells and is known to be a gut mucosal defense factor. In humans, a single gene (ALPI) for IAP has been isolated, and its transcription produces two kinds of alternatively spliced mRNAs (aAug10 and bAug10). Recently, we discovered that vitamin D up-regulated the expression of both types of human IAP alternative splicing variants in Caco-2 cells. However, the functional difference of protein encoded by the mRNA variants has remained elusive. In the present study, we aimed to provide further insight into the characterization and structure of IAP isoforms. To analyze the protein translated from the ALPI gene, we constructed two kinds of cDNA expression plasmids (aAug10 and bAug10), and the transfected cells were homogenized and assayed for alkaline phosphatase (ALP) activity. We also designed the homology-modeled 3D structures of the protein encoded by the mRNA variants (ALPI-aAug10 and ALPI-bAug10). The levels of ALP activity of COS-1 cells transfected with the aAug10 plasmid were increased significantly, while cells transfected with the bAug10 plasmid had undetectable ALP activity. The homology-modeled 3D structures revealed that the variant bAug10 lacks the central N-terminal α-helix and residue corresponding to Asp-42 of ALPI-aAug10 near the active site. This is the first report on the characterization and structure of alternatively spliced transcript variants of the human ALPI gene. Further studies on the regulation of aAug10 and/or bAug10 mRNA expression may identify novel physiological functions of IAP.

The role of gut-derived oxidized lipids and bacterial lipopolysaccharide in systemic inflammation and atherosclerosis.

Purpose of reviewThis review explores mechanisms by which gut-derived bacterial lipopolysaccharide (LPS) and oxidized phospholipids contribute to chronic systemic inflammation and atherosclerosis.Recent findingsGut-derived LPS enters through the small intestine via two distinct pathways that involve high density lipoproteins (HDL) and chylomicrons. Gut-derived LPS can bind to the LPS-binding protein (LBP) and to HDL3 in the small intestine and travel through the portal vein to the liver where it does not elicit an inflammatory reaction, and is inactivated or it can bind to HDL2 and travel through the portal vein to the liver where it elicits an inflammatory reaction. Alternatively, in the small intestine, LPS can bind to LBP and chylomicrons and travel through the lymphatics to the systemic circulation and enhance inflammatory processes including atherosclerosis. Oxidized phospholipids formed in the small intestine regulate the levels and uptake of LPS in small intestine by regulating antimicrobial proteins such as intestinal alkaline phosphatase. Gut-derived LPS and oxidized phospholipids may be responsible for the persistent inflammation seen in some persons with human immunodeficiency virus on potent antiretroviral therapy with undetectable virus levels.SummaryBy targeting gut-derived oxidized phospholipids, the uptake of gut-derived LPS may be reduced to decrease systemic inflammation and atherosclerosis.

Study about the combination strategy of Bacillus subtilis wt55 with AiiO-AIO6 to improve the resistance of zebrafish to Aeromonas veronii infection

Disease problems will seriously restrict the sustainable development of aquaculture, and the environmental-friendly prevention strategies are urgently needed. Probiotics and quorum-quenching enzyme are innovative strategies to control bacterial diseases. Firstly, the bacteriostatic activity of Bacillus subtilis wt55 strain and quenching enzyme AiiO-AIO6 on the growth of Aeromonas veronii were tested in vitro, and the results showed wt55 inhibit the growth of A. veronii, but AiiO-AIO6 did not. Then, the synergistic effects of simple combination of B. subtilis wt55 and AiiO-AIO6 were evaluated next. The results showed this combination could improve the survival rate and significantly reduce the number of invasive A. veronii in gut after challenge compared to the other groups, corresponding to the lower intestinal alkaline phosphatase activity. One of its effect mechanisms is the combination could inhibit the growth of A. veronii in vitro; the other is direct immersion of germ-free zebrafish proved AiiO-AIO6 did not directly regulate the innate immune response of the host, but wt55 did it, and the simple combination group could significantly reduce the expression of nuclear factor kappa-B (NF-κB) and proinflammatory cytokine interleukin-1β (IL-1β), increase the expression of lysozyme gene; and the third is intestinal microbiota also plays a regulatory role: the gut microbiota from combination group could significantly inhibit the expression of IL-1β and NF-κB, and increased the expression of transforming growth factor-β (TGF-β) and lysozyme. Given the effectiveness of this simple combination, a B. subtilis quorum-quenching recombinant expression strain in which AiiO-AIO6 was surface displayed on the spores and secreted by vegetative cells was built. The results showed that the survival rate after challenge was lower than that of the group treated with AiiO-AIO6 or wt55 alone, and the expression of proinflammatory cytokine IL-1β and NF-κB were significantly higher. Our study demonstrated the effectiveness of B. subtilis and AiiO-AIO6 simple combination and established an efficient B. subtilis expression system.

Phosphate decorated lipid-based nanocarriers providing a prolonged mucosal residence time

The aim of this study was to develop phosphate decorated lipid-based nanocarriers including self-emulsifying drug delivery systems (SEDDS), solid lipid nanoparticles (SLN) and nanostructured lipid carriers (NLC) to extend their mucosal residence time. All nanocarriers contained tetradecyltrimethylammonium bromide (TTAB) and polyoxyethylene (9) nonylphenol monophosphate ester (PNPP) for surface decoration. Zeta potential, cytotoxicity, charge conversion and phosphate release studies using isolated intestinal alkaline phosphatase (IAP) and Caco-2 cells were performed. Moreover, the residence time of nanocarriers was determined on porcine intestinal mucosa. Results showed a shift from negative to positive zeta potential due to the addition of TTAB and charge conversion back to a negative zeta potential when also PNPP was added. Up to a concentration of 0.3 %, lipid-based nanocarriers were not toxic. Charge conversion studies with IAP revealed the highest zeta potential shift for NLCTTAB-PNPP with almost Δ22 mV. Phosphate release studies using isolated IAP as well as Caco-2 cells showed a fast phosphate release for SEDDSTTAB-PNPP, SLNTTAB-PNPP and NLCTTAB-PNPP. SLN TTAB-PNPP and NLC TTAB-PNPP provided the highest increase in mucosal residence time that was 4-fold more prolonged than that of blank formulations. In conclusion, phosphate modified lipid-based nanocarriers can essentially prolong the intestinal residence time of their payload.

Effects of Different Viscous Guar Gums on Growth, Apparent Nutrient Digestibility, Intestinal Development and Morphology in Juvenile Largemouth Bass, Micropterus salmoides.

An 8-week feeding trial was conducted to investigate the effects of different viscous guar gums on the growth performance, apparent nutrient digestibility, intestinal development and morphology of juvenile largemouth bass. Four isoproteic and isolipidic diets (crude protein 42.5%, crude lipid 13.7%) were formulated to contain 8% cellulose (Control group), 8% low viscous guar gum with 2,500 mPa s (Lvs-GG group), 8% medium viscous guar gum with 5,200 mPa s (Mvs-GG group) and 8% high viscous guar gum with 6,000 mPa s (Hvs-GG group), respectively. Each diet was fed to quadruplicate groups of 40 fish (6.00 ± 0.01 g) per repetition. Dietary guar gum inclusion significantly decreased the weight gain rate, specific growth rate, protein efficiency ratio, protein productive value and lipid deposition rate, and these parameters decreased considerably with increasing guar gum viscous and were lowest in the Hvs-GG group. Dietary guar gum inclusion significantly decreased the apparent digestibility of dry matter, crude protein and crude lipid, and these parameters decreased considerably with increasing guar gum viscous and were lowest in the Hvs-GG group. Intestinal protease, lipase and creatine kinase activities in the guar gum groups were significantly lower than those in the control group, and intestinal protease and lipase activities decreased considerably with increased guar gum viscous. Intestinal alkaline phosphatase activity in the Hvs-GG group and intestinal Na+/K+-ATPase activity in the Mvs-GG and Hvs-GG groups were significantly lower than those in the Lvs-GG and control groups. Serum high-density lipoprotein, total cholesterol and triglyceride concentrations and superoxide dismutase activity in the guar gum groups were significantly lower than those in the control group. Intestinal villus height and muscular thickness in the guar gum groups were considerably higher than those in the control group, whereas the goblet cell relative number in the Mvs-GG and Hvs-GG groups and the microvillus height in the Lvs-GG and Hvs-GG groups were significantly lower than those in the control group. The expression level of IGF-1 in the guar gum groups and the expression level of GLP-2 in the Mvs-GG and Hvs-GG groups were significantly higher than those in the control group. These results indicated that guar gum diets adversely affected intestinal morphology, decreased intestinal digestive and absorptive enzyme activities, and caused poor nutrient digestibility and growth performance in juvenile largemouth bass. Moreover, the adverse effects of guar gum are closely related to its viscous, and high viscous guar gum produces more extreme negative impacts on juvenile largemouth bass.

Electrophoretic mobility of individual molecules of alkaline phosphatase.

The electrophoretic mobilities and catalytic rates of individual molecules of bovine intestinal alkaline phosphatase were determined in CHES and borate buffers of identical pH using a capillary electrophoresis based method. Both properties were found to be heterogeneous. In the presence of CHES, the mobility and rate were found to be-1.9±0.2×10-9 m2V-1 s-1 and 9.8±7.4× 104 min-1 (N=38), respectively. In the presence of borate, the mobility and rate were found to be-6.9±0.5×10-9 m2V-1 s-1 and 2.0±1.3× 104 min-1 (N=41), respectively. The means and variances for both properties were found to differ significantly between the two buffers. The difference in average mobility was attributed to an increase in negative charge caused by borate complexing with the carbohydrate moieties attached to the enzyme. The difference in variance was attributed to heterogeneous complexation with borate due to heterogeneity in the glycosylation. The differences in mean values for the catalytic rate were attributed to the inhibitory effect of borate and the difference in variance may suggest that the KI of this binding may also be heterogeneous.

Charge converting nanostructured lipid carriers containing a cell-penetrating peptide for enhanced cellular uptake

HypothesisThe aim of this study was the development of nanostructured lipid carriers (NLCs) decorated with a polycationic cell-penetrating peptide (CPP). A coating with polyphosphates (PP) enables charge conversion at target cells being triggered by the membrane bound enzyme intestinal alkaline phosphatase (IAP). ExperimentsThe CPP, stearyl-nona-L-arginine (R9SA) was obtained by solid phase synthesis. Formed nanocarriers were characterized regarding size, polydispersity index, zeta potential and charge conversion in the presence of IAP and on Caco-2 cells. The BCS class IV drug saquinavir (SQV) was loaded into NLCs in different concentrations. Mucus diffusion ability of the NLCs was evaluated by the rotating tube method. Furthermore, cellular uptake was evaluated on Caco-2 cells and endosomal escape properties were investigated using erythrocytes. FindingsAll NLCs were obtained in a size range between 146 nm and 152 nm and a polydispersity index of 0.2. Incubation of PP coated PP-R9SA-NLCs with IAP led to a charge conversion from −41.8 mV to 6.4 mV (Δ48.2 mV). After four hours of incubation with IAP, phosphate release reached a plateau, indicating a faster polyphosphate cleavage than on Caco-2. Drug load and encapsulation efficiency of SQV was obtained up to 80.6% and 46.5 µg/mg. Mucus diffusion was increasing in the following rank order: R9SA-NLCs < blank NLCs < PP-R9SA-NLCs. R9SA-NLCs and PP-R9SA-NLCs increased the cellular uptake 15.6- and 13.2-fold, respectively, compared to the control NLCs. Erythrocytes interaction study revealed enhanced endosomal escape properties for R9SA-NLCs and PP-R9SA-NLCs when incubated with IAP.

Intestinal alkaline phosphatase (IAP, IAP Enhancer) attenuates intestinal inflammation and alleviates insulin resistance.

In this study, we investigated the effects of intestinal alkaline phosphatase (IAP) in controlled intestinal inflammation and alleviated associated insulin resistance (IR). We also explored the possible underlying molecular mechanisms, showed the preventive effect of IAP on IR in vivo, and verified the dephosphorylation of IAP for the inhibition of intestinal inflammation in vitro. Furthermore, we examined the preventive role of IAP in IR induced by a high-fat diet in mice. We found that an IAP + IAP enhancer significantly ameliorated blood glucose, insulin, low-density lipoprotein, gut barrier function, inflammatory markers, and lipopolysaccharide (LPS) in serum. IAP could dephosphorylate LPS and nucleoside triphosphate in a pH-dependent manner in vitro. Firstly, LPS is inactivated by IAP and IAP reduces LPS-induced inflammation. Secondly, adenosine, a dephosphorylated product of adenosine triphosphate, elicited anti-inflammatory effects by binding to the A2A receptor, which inhibits NF-κB, TNF, and PI3K-Akt signalling pathways. Hence, IAP can be used as a natural anti-inflammatory agent to reduce intestinal inflammation-induced IR.

Design of nanostructured lipid carriers and solid lipid nanoparticles for enhanced cellular uptake

In this study PEG-free and zeta potential changing lipid-based nanocarriers providing enhanced cellular uptake were developed. Nanostructured lipid carriers (NLC), consisting of paraffin wax, caprylic/ capric triglyceride, cetyltrimethylammoniumchloride and either soy lecithin or polyglycerol-4 laurate and solid lipid nanoparticles (SLN) with the same composition but without the liquid lipid content were developed. All formulations exposed a positive surface charge and were then coated with the polyphosphate Graham’s salt. Phosphate release from these formulations was evaluated by incubation with intestinal alkaline phosphatase as well as on a Caco-2 monolayer and zeta potentials were measured. Additionally, cellular uptake studies were performed.Within 5 h, a remarkable amount of phosphate was released from all formulations incubated with intestinal alkaline phosphatase. Enzymatically induced phosphate release with intestinal alkaline phosphatase led to a zeta potential shift up to Δ 26 mV. Results of phosphate release and zeta potential change were confirmed on Caco-2 cells. Cellular uptake studies on Caco-2 cells showed an up to 5.6-times higher uptake compared to cells with inhibited phosphatase. According to these results, polyphosphate coating is a powerful tool to obtain lipid-based nanocarriers for enhanced cellular uptake.

Magnetic multi-enzyme cascade combined with liquid chromatography tandem mass spectrometry for fast DNA digestion and quantitative analysis of 5-hydroxymethylcytosine in genome of human bladder cancer T24 cells induced by tetrachlorobenzoquinone

Long-term exposure to halobenzoquinones (HBQs) can induce genomic damages and abnormal epigenetic modifications. High-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) has shown unique advantages in identification and sensitive analysis of these structurally modified DNA lesions. Prior to MS analysis, genomic DNA needs to be fully digested into mono-nucleosides. Here, we prepared Supernuclease (SN)-, snake venom phosphodiesterase (SVP)- and calf intestinal alkaline phosphatase (CIP)- individually immobilized magnetic nanoparticles (MNPs), and combined them according to certain formula to construct a recyclable SN-SVP-CIP magnetic nanoparticles (SNSC-MNPs) cascade for rapid and efficient DNA digestion. The SNSC-MNPs cascade can fully digest genomic DNA into mono-nucleosides within 30 min. The SNSC-MNPs cascade coupled with HPLC-MS/MS method can accurately and sensitively detect 5-hydroxymethylcytosine (5hmC) changes in genome of human bladder cancer T24 cells induced by tetrachlorobenzoquinone. The immobilization of enzymes on MNPs can enhance the stability and enzymatic activity of the three enzymes, which guarantees the reusability and longtime preservation of the cascades. The relative digestive efficiencies are among 86% -106% up to ten times of reuse. The newly synthesized SNSC-MNPs cascade coupled with HPLC-MS/MS method is promising for fast identification and analysis of structural modifications in genomic DNA.

Effects of Butyrate Supplementation on Inflammation and Kidney Parameters in Type 1 Diabetes: A Randomized, Double-Blind, Placebo-Controlled Trial.

Type 1 diabetes is associated with increased intestinal inflammation and decreased abundance of butyrate-producing bacteria. We investigated the effect of butyrate on inflammation, kidney parameters, HbA1c, serum metabolites and gastrointestinal symptoms in persons with type 1 diabetes, albuminuria and intestinal inflammation. We conducted a randomized placebo-controlled, double-blind, parallel clinical study involving 53 participants randomized to 3.6 g sodium butyrate daily or placebo for 12 weeks. The primary endpoint was the change in fecal calprotectin. Additional endpoints were the change in fecal short chain fatty acids, intestinal alkaline phosphatase activity and immunoglobulins, serum lipopolysaccharide, CRP, albuminuria, kidney function, HbA1c, metabolites and gastrointestinal symptoms. The mean age was 54 ± 13 years, and the median [Q1:Q3] urinary albumin excretion was 46 [14:121] mg/g. The median fecal calprotectin in the butyrate group was 48 [26:100] μg/g at baseline, and the change was −1.0 [−20:10] μg/g; the median in the placebo group was 61 [25:139] μg/g at baseline, and the change was −12 [−95:1] μg/g. The difference between the groups was not significant (p = 0.24); neither did we find an effect of butyrate compared to placebo on the other inflammatory markers, kidney parameters, HbA1c, metabolites nor gastrointestinal symptoms. Twelve weeks of butyrate supplementation did not reduce intestinal inflammation in persons with type 1 diabetes, albuminuria and intestinal inflammation.

Oral supplementation of alkaline phosphatase in poultry and swine.

The importance of intestinal alkaline phosphatase (IAP) in maintaining gut health and intestinal homeostasis is well established. The objective of this study was to investigate the tolerance of poultry and swine to dietary supplementation of a novel microbial-derived alkaline phosphatase (AP; E.C. 3.1.3.1 produced by Paenibacillus lentus strain CMG3709). Studies were conducted on day-old Ross 308 chicken (n = 1,000; Study 1) and weaned piglets (n = 180; Study 2) for a duration of 42 d; and consisted of four treatment groups (TG) based on the concentration of microbial-derived AP supplemented in their diet at 0; 12,000; 20,000; and 200,000 U/kg of feed. Parameters such as animal survival, hematology, coagulation, and biochemical indices were assessed at the end of the study. The effect of microbial AP on nutrient absorption through skin pigmentation and intestinal permeability were also investigated in broilers (n = 600; Study 3). In poultry (Study 1), there were no statistically significant differences between control and TG for any of the hematological and biochemical parameters, except for a marginal increase (P < 0.05) in serum phosphorus at the highest dose. This variation was not dose-dependent, was well within the reference range, and was not associated with any clinical correlates. In swine (Study 2), hematological parameters such as leukocyte, basophil, and lymphocyte counts were lower (P < 0.05) for the two highest doses but were traced back to individual variations within the group. The biochemical indices in piglets showed no significant differences between control and supplemental groups except for glucose (P = 0.0005), which showed a high effect (P = 0.008) of the random blood collection order. Nonetheless, glucose was within the normal reference range, and were not related to in-feed supplementation of AP as they had no biological significance. The survival rate in all three studies was over 98%. Dietary supplementation of microbial-derived AP up to 16.7 times the intended use (12,000 U/kg feed) level had no negative effects in both poultry and swine. In-feed supplementation of microbial-derived AP for 28 d improved intestinal pigment absorption (P < 0.0001) and reduced intestinal paracellular permeability (P = 0.0001) in broilers (Study 3). Based on these results, it can be concluded that oral supplementation of microbial-derived AP is safe for poultry and swine and effective at improving gut health in poultry.

Interactions between intestinal morphology, digestion, inflammatory responses, and gut microbiota of juvenile channel catfish elicited by dietary enzymatic rice protein

The reduction of fishmeal in aquafeeds has been the concern of researchers. Replacing fishmeal with plant proteins affects intestinal function and inflammation, but the interaction between the intestinal responses and gut microbiota remains unclear. In this study, juvenile channel catfish (Ictalurus punctatus) was fed with four diets in which enzymatic rice protein (RP) replaced fishmeal at levels of 0 (FM), 2.5% (RP2.5), 5.0% (RP5.0), and 7.5% (RP7.5) for 8 weeks to solve the problem mentioned above. Quantification of intestinal morphology showed that 2.5% or 5.0% RP significantly increased villus length and goblet cell number, accompanied by higher activities of intestinal trypsin, alkaline phosphatase (AKP), and Na+/K+-ATPase (NKA) in RP2.5 group (P < 0.05). In contrast, 7.5% RP slightly damaged the intestinal mucosa and significantly reduced the activities of amylase, AKP, and NKA, as well as decreased serum complement 4 (C4) and immunoglobulin M (IgM). Noteworthy, RT-qPCR showed that 2.5% RP significantly down-regulated intestinal mRNA expression level of il8, while up-regulated mif, tlr4, tlr7, tgfβ3, and cldn2. In contrast, 7.5% RP up-regulated the mRNA expression levels of il1β, il8, and mif, while down-regulated cldn3d. Analysis of gut microbiota showed that 2.5% RP increased the relative abundance of Bacteroidetes and significantly activated potential functions of gut microbiota involved in carbohydrate metabolism. The 7.5% RP increased the diversity of the gut microbiota, accompanied by a significant increase in the relative abundance of conditionally pathogenic bacteria such as Vibrio, Serratia, and Aeromonas (classified as Proteobacteria). Notably, Vibrio was the biomarker species with the greatest difference between the FM and RP7.5 groups (genus level). Correlation analysis indicated that Vibrio may affect immunity through the C4 pathway and further lead to gut inflammation and digestive impairment. Taken above, these results indicated that RP could affect intestinal morphology, digestion, and inflammation, and interact with the composition and potential function of gut microbiota. The low RP supplement (2.5%) improved intestinal morphology and digestion, while high supplement (7.5%) disrupted gut microbiota homeostasis, resulting in damage to intestinal mucosa and inflammatory response.

Lactic Acid Bacterial Supplementation Ameliorated the Lipopolysaccharide-Induced Gut Inflammation and Dysbiosis in Mice.

Lipopolysaccharide (LPS), a gut-transmitted endotoxin from Gram-negative bacteria, causes inflammatory diseases leading to the loss of gut barrier integrity and has been identified as a major pathogenic stimulator in many dysfunctions. Hence, supplementation with probiotics is believed to be one of the most effective strategies for treating many inflammatory gut disorders. Although probiotics are known to have a variety of therapeutic characteristics and to play a beneficial role in host defense responses, the molecular mechanisms by which they achieve these beneficial effects are unknown due to species- and strain-specific behaviors. Therefore, in this study, the protective role of five indigenous lactic acid bacterial strains in ameliorating LPS-induced gut barrier impairment in the C57BL/6 mice model was elucidated. Lacticaseibacillus rhamnosus LAB3, Levilactobacillus brevis LAB20, and Lactiplantibacillus plantarum LAB31 were isolated from infant feces; Pediococcus acidilactici LAB8 from fermented food (Bekang); and Lactiplantibacillus plantarum LAB39 from beetroot. Intraperitoneal injection of LPS (10 mg/kg of body weight) increased the levels of lipocalin and serum markers TNF-α, IL-6, and IL-1β, and the overall disease activity index in the treated group. Furthermore, gene expression of NF-kB, IL-12, and Cox-2; mucin-producing genes Muc-2 and Muc-4; and intestinal alkaline phosphatase (IAP) was deleteriously altered in the ileum of LPS-treated mice. Furthermore, LPS also induced dysbiosis in gut microbiota where higher abundances of Klebsiella, Enterobacter, and Salmonella and decreased abundances of Lactobacillus, Bifidobacteria, Roseburia, and Akkermansia were observed. Western blotting results also suggested that LPS treatment causes the loss of gut barrier integrity relative to the pre-supplementation with LAB strains, which enhanced the expression of tight junction proteins and ameliorated the LPS-induced changes and inflammation. Taken together, the study suggested that LAB3 and LAB39 were more potent in ameliorating LPS-induced gut inflammation and dysbiosis.