Intestinal Alkaline Phosphatase Activity Research Articles

The Intestinal Epithelial Cell Differentiation Marker Intestinal Alkaline Phosphatase (ALPi) Is Selectively Induced by Histone Deacetylase Inhibitors (HDACi) in Colon Cancer Cells in a Kruppel-like Factor 5 (KLF5)-dependent Manner

The histone deacetylase inhibitor (HDACi) sodium butyrate promotes differentiation of colon cancer cells as evidenced by induced expression and enzyme activity of the differentiation marker intestinal alkaline phosphatase (ALPi). Screening of a panel of 33 colon cancer cell lines identified cell lines sensitive (42%) and resistant (58%) to butyrate induction of ALP activity. This differential sensitivity was similarly evident following treatment with the structurally distinct HDACi, MS-275. Resistant cell lines were significantly enriched for those harboring the CpG island methylator phenotype (p = 0.036, Chi square test), and resistant cell lines harbored methylation of the ALPi promoter, particularly of a CpG site within a critical KLF/Sp regulatory element required for butyrate induction of ALPi promoter activity. However, butyrate induction of an exogenous ALPi promoter-reporter paralleled up-regulation of endogenous ALPi expression across the cell lines, suggesting the presence or absence of a key transcriptional regulator is the major determinant of ALPi induction. Through microarray profiling of sensitive and resistant cell lines, we identified KLF5 to be both basally more highly expressed as well as preferentially induced by butyrate in sensitive cell lines. KLF5 overexpression induced ALPi promoter-reporter activity in resistant cell lines, KLF5 knockdown attenuated butyrate induction of ALPi expression in sensitive lines, and butyrate selectively enhanced KLF5 binding to the ALPi promoter in sensitive cells. These findings demonstrate that butyrate induction of the cell differentiation marker ALPi is mediated through KLF5 and identifies subsets of colon cancer cell lines responsive and refractory to this effect.

커피박 첨가가 육계의 소장 효소 활성도, 생화학 지표 및 장내 미생물 균총에 미치는 영향

Animal Products Research and Developmental Division, National Institute of Science, Suwon 441-706, KoreaABSTRACT The current study was performed to investigate the effects of dietary supplementation of dried coffee meal (CM) on growth performance, intestinal and blood biochemical index, intestinal enzymes, and cecal microbial populations. A total of 162, 3-day-old male broiler chicks were randomly allocated into three dietary groups: control group (CON), basal diet added with 0.5% CM (CM I), and basal diet added with 1.0% CM (CM II). Dietary supplementation of CM did not change bird performance and the relative weight of intestinal mucosal tissues. The birds fed the diet supplemented with CM (0.5 and 1.0%) significantly decreased mucosal glucose concentration (P<0.05) without affecting blood glucose level compared with those fed control diet. The level of blood aspartate aminotransferase (AST) significantly increased in CM II group (P<0.05) without affecting γ-glutamyl transpeptidase (γ-GTP) compared with that in the CON group. The specific activity of intestinal maltase, leucine aminopeptidase (LAP) and alkaline phosphatase (ALP) were not affected by dietary supplementation of CM, whereas sucrase activity in birds fed the diet supplemented with CM was decreased (P<0.05) compared to that in the control birds. The colony forming units (CFU) of E. coli in the cecum of CM-fed birds was significantly decreased (P<0.05) compared with that of control birds without changing the CFU of Lactobacillus. In conclusion, dietary supplementation of lower level of CM (0.5%) can be used as a beneficial feed resource without liver toxicity in broiler chicks.(Key words: coffee meal, broiler, biochemical profiles, intestinal enzymes, antimicrobial activity)

Yu Ping Feng San, an ancient Chinese herbal decoction, regulates the expression of inducible nitric oxide synthase and cyclooxygenase-2 and the activity of intestinal alkaline phosphatase in cultures.

Yu Ping Feng San (YPFS), a Chinese herbal decoction comprising Astragali Radix (AR; Huangqi), Atractylodis Macrocephalae Rhizoma (AMR; Baizhu), and Saposhnikoviae Radix (SR; Fangfeng), has been used clinically to treat inflammatory bowel diseases (IBD). Previously, we demonstrated a dual role of YPFS in regulating cytokine release in cultured macrophages. In this study, we elucidated the anti-inflammatory effect of YPFS that is mediated through modulating the expression of three key enzymes involved in IBD: inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and intestinal alkaline phosphatase (IALP). In a lipopolysaccharide (LPS)-induced chronic-inflammation model of cultured murine macrophages, YPFS treatment suppressed the activation of iNOS and COX-2 expression in a dose-dependent manner. Conversely, application of YPFS in cultured small intestinal enterocytes markedly induced the expression of IALP in a time-dependent manner, which might strengthen the intestinal detoxification system. A duality of YPFS in modulating the expression of iNOS and COX-2 was determined here. The expression of iNOS and COX-2 in macrophages was induced by YPFS, and this activation was partially blocked by the NF-κB-specific inhibitor BAY 11-7082, indicating a role of NF-κB signaling. These YPFS-induced changes in gene regulation strongly suggest that the anti-inflammatory effects of YPFS are mediated through the regulation of inflammatory enzymes.

Regulation of intestinal calcium absorption by luminal calcium content: role of intestinal alkaline phosphatase.

Intestinal alkaline phosphatase is a brush border enzyme that is stimulated by calcium. Inhibition of intestinal alkaline phosphatase increases intestinal calcium absorption. We hypothesized that intestinal alkaline phosphatase acts as a minute-to-minute regulatory mechanism of calcium entry. The aim of this study was to evaluate the mechanism by which intestinal luminal calcium controls intestinal calcium absorption. We performed kinetic studies with purified intestinal alkaline phosphatase and everted duodenal sacs and showed that intestinal alkaline phosphatase modifies the luminal pH as a function of enzyme concentration and calcium luminal content. A decrease in pH occurred simultaneously with a decrease in calcium absorption. The inhibition of intestinal alkaline phosphatase by l-phenylalanine caused an increase in calcium absorption. This effect was also confirmed in calcium uptake experiments with isolated duodenal cells. Changes in luminal pH arising from intestinal alkaline phosphatase activity induced by luminal calcium concentration modulate intestinal calcium absorption.

Lactoferrin up-regulates intestinal gene expression of brain-derived neurotrophic factors BDNF, UCHL1 and alkaline phosphatase activity to alleviate early weaning diarrhea in postnatal piglets

The molecular mechanisms underlying how dietary lactoferrin (Lf) impacts gut development and maturation and protects against early weaning diarrhea are not well understood. In this study, we supplemented postnatal piglets with an Lf at a dose level of 155 and 285 mg/kg/day from 3 to 38 days following birth. Our findings show that the high dose of Lf up-regulated messenger RNA expression levels of genes encoding brain-derived neurotrophic factor (BDNF) and ubiquitin carboxy-terminal hydrolase L1 (ubiquitin thiolesterase (UCHL1) and, to a lesser extent, glial cell line-derived neurotrophic factor, in the duodenum (P<.05). Piglets in the high and low Lf group had 30% and 7% larger jejunal crypts compared with the control group (P<.05). Escherichia coli 16S rRNA copy number per gram of ascending colon contents was significantly reduced (P=.001), while the copy number of Bifidobacteria and Lactobacillus spp. was not affected. In addition, Lf increased intestinal alkaline phosphatase activity (P<.05) and delayed the onset of food transitional diarrhea, reducing its frequency and duration (P<.05). The incidence of diarrhea in the high and low Lf groups was decreased 54% and 15%, respectively, compared with the control group (P=.035). In summary, these findings provide new evidence that dietary Lf supplementation up-regulated gene expression of BDNF and UCHL1, decreased the colon microbiota of E. coli, improved gut maturation and reduced early weaning diarrhea in piglets. The molecular basis underlying these findings suggests that Lf may enhance gut development and immune function by providing new insight into the gut–brain–microbe axis that has not been previously reported.

Luminal ATP: the missing link between intestinal alkaline phosphatase, the gut microbiota, and inflammation?

gut dysbiosis is a common feature to many inflammatory diseases but the cause-and-effect relationships between the gut microbiota and host inflammation are poorly understood. For metabolic disorders and obesity, however, bacterial lipopolysaccharide (LPS), an emblematic member of pathogen-associated

Utilization of real‐time electrospray ionization mass spectrometry to gain further insight into the course of nucleotide degradation by intestinal alkaline phosphatase

Related with its ability to degrade nucleotides, intestinal alkaline phosphatase (iAP) is an important participant in intestinal pH regulation and inflammatory processes. However, its activity has been investigated mainly by using artificial non-nucleotide substrates to enable the utilization of conventional colorimetric methods. To capture the degradation of the physiological nucleotide substrate of the enzyme along with arising intermediates and the final product, the enzymatic assay was adapted to mass spectrometric detection. Therewith, the drawbacks associated with colorimetric methods could be overcome. Enzymatic activity was comparatively investigated with a conventional colorimetric malachite green method and a single quadrupole mass spectrometer with an electrospray ionization source using the physiological nucleotide substrates ATP, ADP or AMP and three different pH-values in either methodological approach. By this means the enzymatic activity was assessed on the one hand by detecting the phosphate release spectrometrically at defined time points of enzymatic reaction or on the other by continuous monitoring with mass spectrometric detection. Adaption of the enzymatic assay to mass spectrometric detection disclosed the entire course of all reaction components--substrate, intermediates and product--resulting from the degradation of substrate, thereby pointing out a stepwise removal of phosphate groups. By calculating enzymatic substrate conversion rates a distinctively slower degradation of AMP compared to ADP or ATP was revealed together with the finding of a substrate competition between ATP and ADP at alkaline pH. The comparison of colorimetric and mass spectrometric methods to elucidate enzyme kinetics and specificity clearly underlines the advantages of mass spectrometric detection for the investigation of complex multi-component enzymatic assays. The entire course of enzymatic substrate degradation was revealed with different nucleotide substrates, thus allowing a specific monitoring of intestinal alkaline phosphatase activity.

Intestinal alkaline phosphatase is protective to the preterm rat pup intestine

BackgroundNecrotizing enterocolitis (NEC) is the most common surgical emergency in neonates, with a mortality rate between 10 and 50%. The onset of necrotizing enterocolitis is highly variable and associated with numerous risk factors. Prior research has shown that enteral supplementation with intestinal alkaline phosphatase (IAP) decreases the severity of NEC. The aim of this study is to investigate whether IAP is protective to the preterm intestine in the presence of formula feeding and in the absence of NEC. MethodsPreterm rat pups were fed formula with or without supplementation with IAP, and intestine was obtained on day of life 3 for analysis of IAP activity, mRNA expression of TNFα, IL-6 and iNOS and permeability and cytokine expression after LPS exposure. ResultsThere was no difference in the absolute and intestine specific alkaline phosphatase activity in both groups. Rat pups fed IAP had decreased mRNA expression of the inflammatory cytokines TNFα, IL-6 and iNOS. Pups supplemented with IAP had decreased permeability and inflammatory cytokine expression after exposure to LPS ex vivo when compared to formula fed controls. ConclusionsOur results support that IAP is beneficial to preterm intestine and decreases intestinal injury and inflammation caused by LPS.

Exacerbation of intestinal brush border enzyme activities and oxidative stress in streptozotocin-induced diabetic rats by monocrotophos

The present study was undertaken to investigate the potential of monocrotophos (MCP), one of the widely used broad spectrum systemic organophosphorus insecticides (OPI) in India, to alter small intestinal structure and function. Further, its potential to exacerbate diabetes induced alterations in intestinal structure and function was also studied in experimentally induced diabetic rats. Rats were rendered diabetic with an acute dose of streptozotocin (60mg/kgb.w.). MCP was orally administered at a sublethal dose (1/20 LD50 i.e. 0.9mg/kgb.w./d) for 15days to both normal and diabetic rats. MCP significantly increased unit weight of intestine in diabetic rats. MCP alone increased (up to 57%) the activities of intestinal brush border disaccharidases in normal rats and further augmented the enzyme activities in diabetic rats. Similar results were found with intestinal alkaline phosphatase activity. In addition, Na+/K+-ATPase activity was found to be aggravated in diabetic rats by MCP treatment. Oxidative stress markers showed similar degree of change in both MCP and diabetic rats while MCP aggravated oxidative stress condition in diabetic rats. Scanning electron microscopy and histological analysis of the small intestine revealed increased length of villi, congestion, goblet cell hyperplasia and infiltration of inflammatory cells in MCP and diabetic rats while MCP also induced necrotic lesions in diabetic rats. Collectively, our findings provide evidence that multiple doses of MCP has the propensity to augment diabetes associated intestinal dysfunctions in rats.

Interplay between intestinal alkaline phosphatase, diet, gut microbes and immunity.

Intestinal alkaline phosphatase (IAP) plays an essential role in intestinal homeostasis and health through interactions with the resident microbiota, diet and the gut. IAP's role in the intestine is to dephosphorylate toxic microbial ligands such as lipopolysaccharides, unmethylated cytosine-guanosine dinucleotides and flagellin as well as extracellular nucleotides such as uridine diphosphate. IAP's ability to detoxify these ligands is essential in protecting the host from sepsis during acute inflammation and chronic inflammatory conditions such as inflammatory bowel disease. Also important in these complications is IAP's ability to regulate the microbial ecosystem by forming a complex relationship between microbiota, diet and the intestinal mucosal surface. Evidence reveals that diet alters IAP expression and activity and this in turn can influence the gut microbiota and homeostasis. IAP's ability to maintain a healthy gastrointestinal tract has accelerated research on its potential use as a therapeutic agent against a multitude of diseases. Exogenous IAP has been shown to have beneficial effects when administered during ulcerative colitis, coronary bypass surgery and sepsis. There are currently a handful of human clinical trials underway investigating the effects of exogenous IAP during sepsis, rheumatoid arthritis and heart surgery. In light of these findings IAP has been marked as a novel agent to help treat a variety of other inflammatory and infectious diseases. The purpose of this review is to highlight the essential characteristics of IAP in protection and maintenance of intestinal homeostasis while addressing the intricate interplay between IAP, diet, microbiota and the intestinal epithelium.

Intestinal alkaline phosphatase: novel functions and protective effects

Important protective roles of intestinal alkaline phosphatase (IAP)--including regulation of intestinal surface pH, absorption of lipids, detoxification of free nucleotides and bacterial lipopolysaccharide, attenuation of intestinal inflammation, and possible modulation of the gut microbiota--have been reviewed recently. IAP is modulated by numerous nutritional factors. The present review highlights new findings on the properties of IAP and extends the list of its protective functions. Critical assessment of data suggests that some IAP properties are a direct result of dephosphorylation of proinflammatory moieties, while others (e.g., gut barrier protection and microbiota shaping) may be secondary to IAP-mediated downregulation of inflammation. IAP and tissue-nonspecific alkaline phosphatase isoforms characterize the small intestine and the colon, respectively. Gastrointestinal administration of exogenous IAP ameliorates gut inflammation and favors gut tissue regeneration, whereas enteral and systemic IAP administration attenuates systemic inflammation only. Finally, the IAP gene family has a strong evolutionary link to food-driven changes in gastrointestinal tract anatomy and microbiota composition. Therefore, stimulation of IAP activity by dietary intervention is a goal for preserving gut homeostasis and health by minimizing low-grade inflammation.

Performance, organ zinc concentration, jejunal brush border membrane enzyme activities and mRNA expression in piglets fed with different levels of dietary zinc

This study aimed at investigating the effect of dietary zinc on performance, jejunal brush border membrane enzyme activities and mRNA levels of enzymes and two zinc transporters in piglets. A total of 126 piglets were weaned at 26 ±1 days of age and randomly allocated into three groups fed with diets 50, 150 and 2500 mg zinc/kg. Performance was recorded and at weekly intervals, eight piglets per group were killed. The activities of isolated brush border membrane enzymes including lactase, maltase, sucrase, aminopeptidase-N and intestinal alkaline phosphatase (IAP), and the relative transcript abundance of aminopeptidase-N (APN), sucrase–isomaltase (SUC), IAP and the two zinc transporters SLC39A4 (ZIP4) and SLC30A1 (ZnT1) were investigated in the jejunum. Feeding pharmacological zinc levels increased weight gain (p < 0.001) during the first week, but performance was lower (p < 0.05) in the third week. Organ zinc concentrations were increased by high dietary zinc level. The activity of IAP was higher (p < 0.05) with the highest dietary zinc level, no effects were determined for other enzymes. Dietary zinc level had no effect on transcript abundance of digestive enzymes. The mRNA levels decreased (p < 0.001) for ZIP4, and increased for ZnT1 (p < 0.05) with pharmacological zinc levels. In conclusion, pharmacological zinc levels improved performance in the short-term. Intestinal mRNA level of zinc transporters changed with high zinc supply, but this did not prevent zinc accumulation in tissues, suggesting hampered homoeostatic regulation. This might cause impaired performance during longer supply.

Effects of corn replacement by sorghum in broiler diets on performance and intestinal mucosa integrity

The effect of replacing corn with low-tannin sorghum on broiler performance, carcass yield, integrity of mucosa of small intestine segments, and activity of membrane enzymes of the jejunum is investigated. A total of 594 male Cobb-500 broiler chicks were randomly assigned to 3 dietary treatments: 100% corn (control), 50% corn replacement with low-tannin sorghum (low sorghum), and 100% corn replacement with low-tannin sorghum (high sorghum). Body weight gain, feed consumption, feed conversion, and carcass yield were determined at 7, 21, and 42 d, and segments of the small intestine were collected. Feed conversion and weight gain were impaired at d 42 in broilers fed the high-sorghum diet, but no differences were observed for carcass yield among the treatments (P > 0.05). Crypt cell mitotic index of the jejunum and ileum at d 21 and 42 was lower in broilers fed the control diet than in those fed low- and high-sorghum diets (P < 0.05). Aminopeptidase activity was higher in broilers fed the control diet than in those fed low- and high-sorghum diets irrespective of age (P < 0.05). Conversely, intestinal alkaline phosphatase activity in the small intestine did not differ among the dietary treatments (P > 0.05). Our results indicate that 50% corn replacement with low-tannin sorghum is suitable for broiler diets, whereas 100% corn replacement with low-tannin sorghum had negative effects on the intestinal mucosa and performance of broilers at 42 d.

NFAT5 represses canonical Wnt signaling via inhibition of β -catenin acetylation and participates in regulating intestinal cell differentiation

The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis, which is regulated by multiple signaling pathways. The Wnt/β-catenin pathway has a critical role in this process. Previously, we have shown that the calcineurin-dependent nuclear factor of activated T cell (NFAT) is involved in the regulation of intestinal cell differentiation, as noted by the alteration of brush-border enzyme intestinal alkaline phosphatase (IAP) activity. Here, we show that calcineurin-independent NFAT5 interacts with β-catenin to repress Wnt signaling. We found that overexpression of NFAT5 inhibits, whereas knockdown of NFAT5 increases, TOPflash reporter activity and the expression of Wnt/β-catenin target genes, suggesting that NFAT5 inhibits Wnt signaling. In addition, we demonstrated that NFAT5 directly interacts with the C-terminal transactivation domain (TAD) of β-catenin, inhibits CBP interaction with β-catenin, and inhibits CBP-mediated β-catenin acetylation. Moreover, NFAT5 is expressed in the mucosa of human intestine, with the most pronounced staining in the most differentiated region near the epithelial surface. Knockdown of NFAT5 attenuated sodium butyrate (NaBT)-mediated induction of IAP and sucrase activities; overexpression of NFAT5 induced IAP promoter activity. In summary, we provide evidence showing that NFAT5 is a regulator of Wnt signaling. Importantly, our results suggest that NFAT5 regulation of intestinal cell differentiation may be through inhibition of Wnt/β-catenin signaling.

Sa1731 Neurotensin-HIF-1α-MicroRNA 210 Axis Orchestrates Hypoxia, Colonic Inflammation and Intestinal Angiogenesis

G A A b st ra ct s or constructs overexpressing HA-tagged 14-3-3σ plasmids. 14-3-3σ, Rictor (an mTORC2 protein), MUC2, cleaved Notch1, p-Akt(S473), Akt1 and Akt2 protein expression was determined by Western blotting. Enterocyte differentiation was induced by treatment of HT29 cells with sodium butyrate (NaBT) and assayed by measuring intestinal alkaline phosphatase (IAP) activity, an enterocyte differentiation marker. The expression of the goblet cell differentiation marker MUC2 and the Notch1 target gene HES1 was also determined by RT-PCR. The interaction between Akt andmTORC2was determined by immunoprecipitation followed by Western blot analysis. RESULTS. Knockdown of 14-3-3 σ did not alter basal or NaBT-induced of IAP activity. However, knockdown of 14-3-3 σ significantly increased MUC2 expression associated with the inhibition of Notch1 signaling as shown by decreased cleaved Notch1 protein and decreased HES1 mRNA expression. These results suggested that 14-3-3σ plays an important role in the regulation of goblet cell differentiation without impacting enterocyte differentiation. In addition, overexpression of 14-3-3σ decreased Akt (Ser473) phosphorylation and knockdown of either Akt1 or Akt2 decreased MUC2 expression. Moreover, overexpression of 14-3-3σ decreased the interaction between Akt and mTORC2. These results suggest that 14-3-3σmay regulate Akt activity through the inhibition of mTORC2-mediated Akt (Ser473) phosphorylation. CONCLUSIONS. Our results suggest a role for 14-3-3σ in goblet cell differentiation through the alteration of TSC2, Notch and Akt signaling pathways. Importantly, our data demonstrate that 14-3-3 σ regulation of Akt phosphorylation is mediated through the altered interaction between mTORC2 and Akt.

699 NFAT5 Represses Canonical Wnt Signaling via Inhibition of β-Catenin Acetylation and Participates in Regulating Intestinal Cell Differentiation

The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis, which is regulated by multiple signaling pathways. The Wnt/b-catenin pathway has a critical role in this process. Previously, we have shown that the calcineurin-dependent nuclear factor of activated T cell (NFAT) is involved in the regulation of intestinal cell differentiation, as noted by the alteration of brush-border enzyme intestinal alkaline phosphatase (IAP) activity. Here, we show that calcineurinindependent NFAT5 interacts with b-catenin to repress Wnt signaling. We found that overexpression of NFAT5 inhibits, whereas knockdown of NFAT5 increases, TOPflash reporter activity and the expression of Wnt/b-catenin target genes, suggesting that NFAT5 inhibits Wnt signaling. In addition, we demonstrated that NFAT5 directly interacts with the C-terminal transactivation domain (TAD) of b-catenin, inhibits CBP interaction with b-catenin, and inhibits CBP-mediated b-catenin acetylation. Moreover, NFAT5 is expressed in the mucosa of human intestine, with the most pronounced staining in the most differentiated region near the epithelial surface. Knockdown of NFAT5 attenuated sodium butyrate (NaBT)-mediated induction of IAP and sucrase activities; overexpression of NFAT5 induced IAP promoter activity. In summary, we provide evidence showing that NFAT5 is a regulator ofWntsignaling.Importantly, ourresults suggest thatNFAT5regulationofintestinalcelldifferentiation maybethrough inhibition of Wnt/b-catenin signaling.

Influence of different probiotic lactic Acid bacteria on microbiota and metabolism of rats with dysbiosis.

Lactic acid bacteria (LAB) are often used for prevention and treatment of dysbiosis. However, the action of various strains of LAB on metabolism and digestion under these conditions are poorly understood. The purpose of this study was to investigate the influence of probiotic LAB on metabolism, digestion and microbiota in animals with dysbiosis. After administration of ampicillin and metronidazole male Wistar rats, were fed products containing Enterococcus faecium L3 (E.f.), Lactobacillus fermentum Z (L.f.) or milk (control 1). Animals in control group 2 were fed milk, after water instead of antibiotics. Dyspeptic symptoms disappeared after administration of probiotic compared with control 1. At the end of the experiment, an increase in the content of enterococci and lactobacilli in the proximal part of the small intestine was found in the animals treated with E.f. and L.f., respectively. After the introduction of probiotic enterococci, the quantity of lactobacilli and bifidobacteria in the intestines of rats increased, and the content of Klebsiella spp. and Escherichia coli decreased in comparison with the control group 1 and the group fed lactobacilli. The activity of alkaline phosphatase and aspartate transaminase was greater in blood serum of rats with dysbiosis receiving milk and lactobacilli. Intestinal alkaline phosphatase activity increased in the epithelium and chyme in the jejunum of the animals treated with L. f. and in the chyme only in the animals treated with E. f. Thus, the specific effects of different strains of probiotic LAB on the microbiota, and on metabolism and digestion of various nutrients were demonstrated.

The small intestinal apical hydrolase activities are decreased in the piglet with bowel inflammation induced by dextran sodium sulfate1

Inflammatory bowel disease (IBD) is characterized by cramping, abdominal pain, bloating, constipation, and diarrhea. We tested the hypothesis that compromised activities of the major small intestinal apical hydrolases contribute to the symptoms of IBD. Changes in hydrolytic kinetics, target protein abundances, and mRNA expression of intestinal alkaline phosphatase (IAP), lactase, maltase, sucrase-isomaltase (SI), maltase-glucoamylase (MGA), and aminopeptidase N (APN) in piglets with colonic inflammation chemically induced by dextran sodium sulfate (DSS) were investigated. Yorkshire piglets at 5 d of age, with an average initial BW of about 3 kg, were fitted with intragastric catheters and were divided into control (CON; n = 6) and treatment groups (DSS; n = 5). Both groups were infused with an equal volume of either saline or 1.25 g of DSS · kg BW(-1) · d(-1) in saline, respectively, for 10 d. Enzyme kinetic experiments for IAP, lactase, maltase, SI, MGA, and APN were measured at 37°C with isolated proximal jejunal apical membrane. Target hydrolase protein abundances in the apical membrane were analyzed by Western blotting and their mRNA abundances in the jejunum were measured by quantitative real-time reverse transcription (RT-) PCR with β-actin as the housekeeping gene. Expressed as percentage of the CON, DSS treatment decreased (P < 0.05) the maximal specific activities of IAP (53%), lactase (78%), maltase (56%), SI (72%), MGA (29%), and APN (22%) as well as the target hydrolase protein abundances of IAP (39%), lactase (35%), SI (36%), and APN (54%), respectively. Decreases (P < 0.05) in the mRNA abundances (% of the CON) for lactase (25%), SI (52%), MGA (75%), and APN (39%) were observed in the DSS group. However, DSS treatment increased (P < 0.05) the jejunal IAP mRNA abundance by 3.5 fold. We conclude that decreases in the small intestinal apical activities of these examined hydrolases likely contribute to overgrowth of pathogenic bacterial populations in the distal small intestine and the colon, leading to the pathogenesis of IBD.

Ontogeny of the digestive enzymes in butter catfish Ompok bimaculatus (Bloch) larvae

The digestive physiology of butter catfish was studied by assessing the activity of different pancreatic (trypsin, chymotrypsin, α-amylase and lipase), gastric (pepsin) and intestinal (alkaline phosphatase) enzymes from hatching until the juvenile stage (30dph). Larvae were reared at 27°C and fed with Artemia nauplii from 2days post hatching (dph) until 10dph, from 7–10dph with Artemia nauplii and zooplankton (Cyclopoida) and from 10dph onwards only with zooplankton. The assessment of the activity of digestive enzymes showed that enzymes involved in the digestion of proteins, lipids and carbohydrates were present in butter catfish larvae before mouth opening and increased after the onset of exogenous feeding, coinciding with the histological organization of the exocrine pancreas. The specific activity of most of the pancreatic enzymes increased until 15dph and decreased thereafter coinciding with the increase of pepsin. A progressive shift in activity from alkaline (trypsin and chymotrypsin) to acid (pepsin) proteases indicated a change in the digestive physiology of the specimen, as alkaline proteases were no longer the main digestive enzymes involved in protein digestion after the onset of acidic digestion between 15 and 21dph. The maturation of the intestine and the achievement of a juvenile-like mode of digestion were demonstrated by changes in enzyme activities from the exocrine pancreas and stomach that coincided with alterations in enzyme production occurring in the intestine (e.g. alkaline phosphatase). Considering the ontogenetic development of the digestive enzymes from the pancreas, stomach and intestine, butter catfish larvae might be weaned between 15 and 21dph, as larvae have achieved the complete maturation of their digestive capacities. These results contradict previous recommendations, which were based solely on the histological organization of the gastric gland and histochemical properties of mucous cells from the stomach, to wean butter catfish larvae at earlier ages. These findings on the functional development of the digestive system in butter catfish would be useful to improve the actual larval rearing techniques for this promising catfish species from the Indian sub-continent.

The role of intestinal alkaline phosphatase in pediatric inflammatory bowel and celiac diseases

Intestinal alkaline phosphatase enzyme plays a pivotal role in the maintenance of intestinal mucosal barrier integrity with the detoxification capacity of lipopolysaccharide, the ligand of Toll-like receptor 4. The inappropriate immune responses and the damage of the mucosal barrier may contribute to the initiation of inflammatory bowel and celiac diseases. In the inflamed colonic mucosa of children with inflammatory bowel disease and in the duodenal mucosa of newly diagnosed children with celiac disease, the decreased intestinal alkaline phosphatase and increased Toll-like receptor 4 protein expression may generate enhanced lipopolysaccharide activity, which may strengthen tissue damaging processes. The enhancement of intestinal alkaline phosphatase activity in an animal model of colitis and in therapy resistant, adult patients with ulcerative colitis reduced the symptoms of intestinal inflammation. In accordance with these results, the targeted intestinal administration of the enzyme in the two examined disorders may be a supplemental therapeutic option in the future.