Intestinal Alkaline Phosphatase Research Articles

Xylooligosaccharides benefits the growth, digestive functions and TOR signaling in Megalobrama amblycephala fed diets with fish meal replaced by rice protein concentrate

This study was conducted to examine the effects of fish meal (FM) replacement by rice protein concentrate (RPC) supplemented with xylooligosaccharides (XOS) on the growth, intestinal digestive and absorptive capabilities and amino acid metabolism of blunt snout bream. Fish were randomly divided into six groups, and were fed with the control diet (containing 10% FM), the RPC diet (FM entirely replaced by RPC) and the RPC diet supplemented with 0.5, 1.5, 2.3 and 3% XOS, respectively, for 8 weeks. Fish fed the RPC diet obtained significantly lower values in daily growth index, feed efficiency ratio, nitrogen and energy retention and plasma concentrations of methionine, leucine, phenylalanine, lysine and histidine compared to the control. However, the supplement of 1.5% XOS restored these parameters to the levels similar to the control group. Fish fed the RPC diet showed no statistical difference in intestinal lipase, alkaline phosphatase and Na+K+-ATPase activities compared to the control group, but exhibited lower protease, carboxypeptidase A, aminopeptidase N and γ-glutamyl transpeptidase activities. However, these effects were reserved with the administration of 1.5% XOS. Also, the transmission electron microscopy analysis showed a significantly lower microvilli length in the mid-intestine of fish fed the RPC diet compared to the control, but it improved significantly with the supplementation of XOS. Hepatic activities of alanine transaminase, aspartate aminotransferase and glutamate dehydrogenase were significantly lower in fish fed the RPC diet compared to the control, but they improved with the supplementation of 1.5% XOS. Similarly, fish fed the RPC diet obtained relatively low expression levels of the target of rapamycin (TOR) and ribosomal protein S6 kinase 1 but higher expressions of eukaryotic translation initiation factor 4E-binding protein 2 in the intestine, liver and muscle. Opposite trend was observed in fish fed the RPC diet supplemented with 1.5% XOS. Overall, these results showed that RPC supplemented with 1.5% XOS could completely replace FM in the feed of blunt snout bream without any adverse effect on the growth performance, intestinal enzymes activities, gut morphology, amino acid metabolism and TOR signaling.

The effect of the branched-chain amino acids on the in-vitro activity of bovine intestinal alkaline phosphatase.

Branched-chain amino acids (BCAA) are used as nutritional support for patients with a range of conditions including liver cirrhosis and in-born errors of amino acid metabolism, and they are commonly used "sports" or exercise supplements. The effects of the BCAA on the in-vitro activity of calf intestinal alkaline phosphatase (EC. 3.1.3.1) were studied. All three BCAA were found to be uncompetitive inhibitors of the enzyme with L-leucine being the most potent ( = 24.9 mmol/L) and L-valine, the least potent ( = 37 mmol/L). Mixed BCAA are able to act in combination to inhibit the enzyme. Given the important role of intestinal alkaline phosphatase in gut homeostasis, these findings have potential implications for those taking high levels of BCAA as supplements.

Supplemented Alkaline Phosphatase Supports the Immune Response in Patients Undergoing Cardiac Surgery: Clinical and Computational Evidence

Alkaline phosphatase (AP) is an enzyme that exhibits anti-inflammatory effects by dephosphorylating inflammation triggering moieties (ITMs) like bacterial lipopolysaccharides and extracellular nucleotides. AP administration aims to prevent and treat peri- and post-surgical ischemia reperfusion injury in cardiothoracic surgery patients. Recent studies reported that intravenous bolus administration and continuous infusion of AP in patients undergoing coronary artery bypass grafting with cardiac valve surgery induce an increased release of liver-type “tissue non-specific alkaline phosphatase” (TNAP) into the bloodstream. The release of liver-type TNAP into circulation could be the body's way of strengthening its defense against a massive ischemic insult. However, the underlying mechanism behind the induction of TNAP is still unclear. To obtain a deeper insight into the role of AP during surgery, we developed a mathematical model of systemic inflammation that clarifies the relation between supplemented AP and TNAP and describes a plausible induction mechanism of TNAP in patients undergoing cardiothoracic surgery. The model was validated against clinical data from patients treated with bovine Intestinal AP (bIAP treatment) or without AP (placebo treatment), in addition to standard care procedures. We performed additional in-silico experiments adding a secondary source of ITMs after surgery, as observed in some patients with complications, and predicted the response to different AP treatment regimens. Our results show a strong protective effect of supplemented AP for patients with complications. The model provides evidence of the existence of an induction mechanism of liver-type tissue non-specific alkaline phosphatase, triggered by the supplementation of AP in patients undergoing cardiac surgery. To the best of our knowledge this is the first time that a quantitative and validated numerical model of systemic inflammation under clinical treatment conditions is presented.

Ameliorative effect of silymarin against linezolid-induced hepatotoxicity in methicillin-resistant Staphylococcus aureus (MRSA) infected Wistar rats.

Linezolid has a better choice for eradication of methicillin-resistant Staphylococcus aureus (MRSA) infections, but its use is limited because of linezolid-induced hepatotoxicity, myelosuppression, and lactic acidosis. This research elucidated the role of silymarin against hepatoxicity of linezolid therapy in MRSA infected Wistar rats. The rats were rendered neutropenic by an intraperitoneal injection of cyclophosphamide injection. The neutropenic rats were injected subcutaneously with 106 CFU/ml of MRSA. The rats were divided into 6 groups. Normal control, Infected, Infected animals treated with linezolid 50 mg/kg/twice/day and Infected animals treated with linezolid and different dose of silymarin 25, 50, and 100 mg/kg/twice/day for 14 days. On the 15th day, the blood, liver, kidney, and bone marrow were collected for biochemical and histopathological examination. The MRSA was confirmed by PCR assay. The minimum inhibitory concentration of linezolid was 0.5-2 μg/ml. The linezolid induced liver damage was confirmed by elevation of marker enzymes alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), Lactate dehydrogenase (LDH) levels, serum bilirubin, lactate, and histopathological studies of the liver. The linezolid treated rats also showed myelosuppression, lactic acidosis, oxidative stress and decreased intestinal alkaline phosphatase (IAP). The silymarin administration exhibited marked hepatoprotective effect by significantly lowering the liver marker enzymes, serum parameters, and cytological findings reflect the hepatoprotection. Additionally, Silymarin showed protection against myelosuppression and lactic acidosis evidenced by bone marrow smear and serum lactate estimation. Antioxidant effect of silymarin was confirmed by decreased levels of lipid peroxidation, restored the enzymatic and non-enzymatic antioxidants of the liver nearer to normal. The present study indicates that the silymarin could be a better herbal therapeutic agent which protects against the linezolid induced hepatotoxicity in MRSA infected rats.

Hybrid compounds from chalcone and 1,2-benzothiazine pharmacophores as selective inhibitors of alkaline phosphatase isozymes

Chalcones and 1,2-benzothiazines are two important classes of bioactive compounds, each scaffold endowed with diverse pharmacological activities. Combining both of these pharmacophores in a single molecule was aimed to yield multi-modal agents. Herein, we report a series of hybrid compounds 3a–3o derived from chalcones and 1,2-benzothiazine cores. They were synthesized from commercially available sodium saccharin, and the resulting 1,2-benzothiazine-derived ketone was then condensed with aromatic aldehydes in an aldol condensation to obtain the respective chalcones. The compounds were characterized using different analytical techniques including FT-IR, NMR spectroscopy, mass spectrometry and X-ray crystallography. Some synthesized chalcones revealed potent and/or selective inhibitory properties towards alkaline phosphatase isozymes transiently expressed in COS-7 cells. A detailed structure-activity and selectivity study was carried out with regard to the effect of different substituents at ortho-, meta- and para-positions of the phenyl residue. Compound 3c was the most effective human intestinal alkaline phosphatase (h-IAP) inhibitor (IC50 value of 1.04 μM), while it was not active against human tissue non-specific alkaline phosphatase (h-TNAP) isozyme. In contrast, 3i was a selective inhibitor of h-TNAP with IC50 values of 0.25 ± 0.01 μM. The possible binding interactions of the most effective inhibitors of h-TNAP and h-IAP were obtained from molecular docking studies.

Self-emulsifying drug delivery systems changing their zeta potential via a flip-flop mechanism

To overcome the mucus layer and cell membrane barrier, self-emulsifying drug delivery systems (SEDDS) exhibiting negative zeta potential, switching to positive values when having reached the cell membrane is a promising approach. Accordingly, a novel conjugate was synthesized by covalent attachment of phosphotyrosine to octadecylamine, which was incorporated into SEDDS. Generated system presented an average diameter of 32 nm and zeta potential of around −12 mV when being diluted 1:100 in 100 mM HEPES buffer pH 7.5 containing 5 mM MgCl2 and 0.2 mM ZnCl2. Incubation of SEDDS with isolated intestinal alkaline phosphatase (IAP) resulting in enzymatic cleavage of phosphate ester moiety caused a shift in zeta potential up to +5.3 mV. As non-toxicity of the developed SEDDS diluted 1:1000 in 25 mM HEPES buffer pH 7.5 containing 5% glucose was observed on Caco-2 cells by employing resazurin assay, this system may provide an inspiring strategy for future zeta potential changing drug delivery systems to master the mucus and membrane barrier.

Loss of MYO5B Leads to Reductions in Na+ Absorption With Maintenance of CFTR-Dependent Cl– Secretion in Enterocytes

Inactivating mutations in MYO5B cause microvillus inclusion disease (MVID), but the physiological cause of the diarrhea associated with this disease is unclear. We investigated whether loss of MYO5B results inaberrant expression of apical enterocyte transporters. We studied alterations in apical membrane transporters in MYO5B-knockout mice, as well as mice with tamoxifen-inducible, intestine-specific disruption of Myo5b (VilCreERT2;Myo5bflox/flox mice) or those not given tamoxifen (controls). Intestinal tissues were collected from mice and analyzed by immunostaining, immunoelectron microscopy, or cultured enteroids were derived. Functions of brush border transporters in intestinal mucosa were measured in Ussing chambers. We obtained duodenal biopsy specimens from individuals with MVID and individuals without MVID (controls) and compared transporter distribution by immunocytochemistry. Compared to intestinal tissues from littermate controls, intestinal tissues from MYO5B-knockout mice had decreased apical localization of SLC9A3 (also called NHE3), SLC5A1 (also called SGLT1), aquaporin (AQP) 7, and sucrase isomaltase, and subapical localization of intestinal alkaline phosphatase and CDC42. However, CFTR was present on apical membranes ofenterocytes from MYO5B knockout and control mice. Intestinal biopsies from patients with MVID had subapical localization of NHE3, SGLT1, and AQP7, but maintained apical CFTR. After tamoxifen administration, VilCreERT2;Myo5bflox/flox mice lost apical NHE3, SGLT1, DRA, and AQP7, similar to germline MYO5B knockout mice. Intestinal tissues from VilCreERT2;Myo5bflox/flox mice had increased CFTR in crypts andCFTR localized to the apical membranes of enterocytes. Intestinal mucosa from VilCreERT2;Myo5bflox/flox mice given tamoxifen did not have an intestinal barrier defect, based on Ussing chamber analysis, but did have decreased SGLT1 activity and increased CFTR activity. Although trafficking of many apical transporters is regulated by MYO5B, trafficking of CFTR is largely independent of MYO5B. Decreased apical localization of NHE3, SGLT1, DRA, and AQP7 might be responsible for dysfunctional water absorption in enterocytes of patients with MVID. Maintenance of apical CFTR might exacerbate water loss by active secretion of chloride into the intestinal lumen.

UHPLC-Q-TOF/MS detection of UV-induced TpT dimeric lesions in genomic DNA

Ultraviolet (UV) radiation induces mutagenicity and cytotoxicity in human cells by the formation of DNA lesions, including cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts (6-4PPs), mainly on thymine-thymine (TpT) dinucleotides. Here, we firstly synthesized the two TpT dimeric lesions with satisfactory yields using a unique UV-irradiated water droplet approach followed by HPLC purification. By the use of purified TpT lesions as standards, we further developed and optimized a quantitative UHPLC-Q-TOF/MS method for the detection of CPDs and 6-4PPs. After the optimization of the enzyme composition and the pH values of hydrolysis solution, a combination of snake venom phosphodiesterase, nuclease P1, and calf intestine alkaline phosphatase can be used for one-step enzymatic digestion to efficiently release the dimeric lesions (CPDs and 6-4PPs) from the genomic DNA. By the use of the one-step digestion and UHPLC-Q-TOF/MS assay for scanning all dimeric lesions, we demonstrate that only are TpT dimeric lesions detectable in genomic DNA of HCT116 cells upon UVC irradiation. The estimated frequency of the CPD of TpT increases from 28.7 to 409 per 106 bases with increasing UVC dosage from 40 J/m2 to 1200 J/m2, while the 6-4PP of TpT increases from 3.7 to 54 per 106 bases. The proposed UHPLC-Q-TOF/MS method is promising for accurate identification and quantitative detection of UV-induced dimeric lesions in cellular DNA.

Excess of nutrient-induced morphofunctional adaptation and inflammation degree in a Caco2/HT-29 in vitro intestinal co-culture.

The intestinal cell function can be modulated by the type and quantity of nutrients. The aim of this study was to evaluate the effects of an excess of nutrients on intestinal morphofunctional features and a possible association of inflammation in a 70/30 Caco2/HT-29 intestinal in vitro co-culture. An excess of nutrients (EX) was obtained by progressively increasing the medium change frequency with respect to standard cell growth conditions (ST) from confluence (T0) to 15 d after confluence (T15). In comparison with the ST group, the EX group revealed a maintenance in the number of microvilli, an increase in follicle like-structures and mucus production, and a decrease in the number of tight junction. The specific activity of markers of intestinal differentiation, alkaline phosphatase and aminopeptidase N, and of the enterocyte differentiation specific marker, dipeptidyl peptidase-IV, were progressively raised. The transepithelial electrical resistance, indicative of the co-culture barrier properties, decreased, whereas Lucifer yellow Papp evaluation, an index of the paracellular permeability to large molecules, showed an increase. Reactive oxygen species and nitric oxide production, indicative of an oxidative status, together with interleukin-6, interleukin-8, indicative of a low-grade inflammation, and peptide YY secretion were higher in the EX group than in the ST group. The differences between ST and EX were particularly evident at T15. These data support the suitability of our in vitro gut model for obesity studies at the molecular level and the necessity to standardize the medium frequency change in intestinal culture.

Loss of Intestinal Alkaline Phosphatase Leads to Distinct Chronic Changes in Bone Phenotype

BackgroundThe gut is becoming increasingly recognized as the source of various systemic diseases, and recently, it has been linked to bone metabolism via the so-called gut-bone axis. The microbiome and gut-derived mediators are thought to impact upon bone metabolism, and administration of probiotics has been shown to have beneficial effects in bone. The gut brush border enzyme intestinal alkaline phosphatase (IAP) plays an important role in controlling calcium absorption, inhibiting lipopolysaccharides, and other inflammatory mediators responsible for endotoxemia and appears to preserve the normal gut microbiota. Interestingly, IAP-deficient mice (AKP3−/−) also display a significant decrease in fecal Lactobacillus, the genus shown to be beneficial to bone. Materials and methodsIAP mRNA levels in mouse bone were measured using quantitative real-time polymerase chain reaction. Femurs of IAP-knockout (KO) and wild-type (WT) mice were analyzed by microcomputed tomography and histopathology. Serum levels of alkaline phosphatase, calcium, and phosphorus were measured. Target cell response upon exposure to serum from IAP-KO and WT mice was quantified using primary bone marrow macrophages. ResultsIAP was not significantly expressed in bones of WT or KO animals. IAP (alkaline phosphatase 3) expression in bone was vanishingly low compared to the duodenum (bone versus duodenum, 56.9 ± 17.7 versus 25,430.3 ± 10,884.5 relative expression, P = 0.01). Bone histology of younger IAP-KO and WT animals was indistinguishable, whereas older IAP-deficient mice showed a distinctly altered phenotype on histology and computed tomography scan. Younger KO mice did not display any abnormal levels in blood chemistry. Older IAP-KO animals showed an isolated increase in serum alkaline phosphatase levels reflecting an environment of active bone formation (IAP-WT versus IAP-KO, 80 ± 27.4 U/I versus 453 ± 107.5 U/I, P = 0.004). There was no significant difference in serum calcium or phosphorus levels between KO and WT mice. Serum from IAP-KO mice induced a significantly higher inflammatory target cell response. ConclusionsThrough its multiple functions, IAP seems to play a crucial role in connecting the gut to the bone. IAP deficiency leads to chronic changes in bone formation, most likely through dysbiosis and systemic dissemination of proinflammatory mediators.

Ontogenetic development of digestive tract and digestive enzymatic activities inSqualiobarbus curriculuslarvae

Squaliobarbus curriculus is an economically important freshwater fish. The ontogenetic development of the digestive system of S. curriculus larvae was studied histologically and enzymatically from hatching to 30 days posthatching (DPH). Amylase, lipase, alkaline phosphatase and pepsin activities were detected from the hatching stage, indicating that these enzymes were genetically preprogrammed. Marked increases in intestinal amylase, trypsin and alkaline phosphatase activities between 10 and 20 DPH corresponded to feed acquisition and transformation. Larval development in S. curriculus could be divided into three phases: phase I (endotrophic period): 1–3 DPH; phase II (endo-exotrophic period): 4–5 DPH; and phase III (exclusively exotrophic period): from 6 DPH onward. At hatching, the digestive tract of the larvae was an undifferentiated straight tube. On 3 DPH, the digestive tract differentiated into the mouth cavity, oesophagus and intestine. On 6 DPH, feeding was totally exotrophic and the yolk sac was completely exhausted. During the growth of S. curriculus larvae, the intestinal mucosa formed and the number of goblet cells and microvilli increased, demonstrating maturation of the digestive system. The study about the digestive development of S. curriculus larvae will contribute to better larval-rearing strategies.

Adiposity and metabolic health in mice deficient in intestinal alkaline phosphatase

ABSTRACTIntestinal alkaline phosphatase 3 (AKP3) is an enzyme that was reported to play a role in lipid metabolism and to prevent high fat diet-induced metabolic syndrome in mice.To investigate a potential functional role of AKP3 in diet-induced adiposity and metabolic health, we have kept male and female wild-type or AKP3 deficient mice on a high fat diet for 15 weeks to induce obesity and compared those with mice kept on standard fat diet.Body weight as well as adipose tissue mass were statistically significantly higher upon high fat diet feeding for mice of both genders and genotypes. Female mice of either genotype kept on high fat diet gained less weight, resulting in smaller adipose tissue depots with smaller adipocytes. However, AKP3 deficiency had no significant effect on body weight gain or adipose tissue mass and did not affect adipocyte size or density. Gene expression analysis revealed no effect of the genotype on inflammatory parameters in adipose tissue, except for tumor necrosis factor alpha, which was higher in mesenteric adipose tissue of female obese mice. Plasma glucose and insulin levels were also not affected in obese AKP3 deficient mice.Overall, our data do not support a functional role of AKP3 in adipose tissue development, or insulin sensitivity.

Intestine-specific expression of human chimeric intestinal alkaline phosphatase attenuates Western diet-induced barrier dysfunction and glucose intolerance.

Intestinal epithelial cell derived alkaline phosphatase (IAP) dephosphorylates/detoxifies bacterial endotoxin lipopolysaccharide (LPS) in the gut lumen. We have earlier demonstrated that consumption of high‐fat high‐cholesterol containing western type‐diet (WD) significantly reduces IAP activity, increases intestinal permeability leading to increased plasma levels of LPS and glucose intolerance. Furthermore, oral supplementation with curcumin that increased IAP activity improved intestinal barrier function as well as glucose tolerance. To directly test the hypothesis that targeted increase in IAP would protect against WD‐induced metabolic consequences, we developed intestine‐specific IAP transgenic mice where expression of human chimeric IAP is under the control of intestine‐specific villin promoter. This chimeric human IAP contains domains from human IAP and human placental alkaline phosphatase, has a higher turnover number, narrower substrate specificity, and selectivity for bacterial LPS. Chimeric IAP was specifically and uniformly overexpressed in these IAP transgenic (IAPTg) mice along the entire length of the intestine. While IAP activity reduced from proximal P1 segment to distal P9 segment in wild‐type (WT) mice, this activity was maintained in the IAPTg mice. Dietary challenge with WD impaired glucose tolerance in WT mice and this intolerance was attenuated in IAPTg mice. Significant decrease in fecal zonulin, a marker for intestinal barrier dysfunction, in WD fed IAPTg mice and a corresponding decrease in translocation of orally administered nonabsorbable 4 kDa FITC dextran to plasma suggests that IAP overexpression improves intestinal barrier function. Thus, targeted increase in IAP activity represents a novel strategy to improve WD‐induced intestinal barrier dysfunction and glucose intolerance.

Synthesis of sulfadiazinyl acyl/aryl thiourea derivatives as calf intestinal alkaline phosphatase inhibitors, pharmacokinetic properties, lead optimization, Lineweaver-Burk plot evaluation and binding analysis

To seek the new medicinal potential of sulfadiazine drug, the free amino group of sulfadiazine was exploited to obtain acyl/aryl thioureas using simple and straightforward protocol. Acyl/aryl thioureas are well recognized bioactive pharmacophore containing moieties. A new series (4a–4j) of sulfadiazine derived acyl/aryl thioureas was synthesized and characterized through spectroscopic and elemental analysis. The synthesized derivatives 4a–4j were subjected to calf intestinal alkaline phosphatase (CIAP) activity. The derivative 4a–4j showed better inhibition potential compared to standard monopotassium phosphate (MKP). The compound 4c exhibited higher potential in the series with IC50 0.251 ± 0.012 µM (standard KH2PO4 4.317 ± 0.201 µM). Lineweaver-Burk plots revealed that most potent derivative 4c inhibition CIAP via mixed type pathway. Pharmacological investigations showed that synthesized compounds 4a–4j obey Lipinsk’s rule. ADMET parameters evaluation predicted that these molecule show significant lead like properties with minimum possible toxicity and can serve as templates in drug designing. The synthetic compounds show none mutagenic and irritant behavior. Molecular docking analysis showed that compound 4c interacts with Asp273, His317 and Arg166 amino acid residues.

Deazapurine Analogues Bearing a 1H‐Pyrazolo[3,4‐b]pyridin‐3(2H)‐one Core: Synthesis and Biological Activity

A new methodology for the synthesis of fluorinated and non‐fluorinated 1H‐pyrazolo[3,4‐b]pyridin‐3‐ones was developed. The reactions proceed by cyclization of electron‐rich 3‐amino‐1H‐pyrazol‐5(4H)‐ones with 1,3‐diketones and the products were obtained in good to excellent yields and with excellent regioselectivity. The products, which can be regarded as deazapurine analogues, were tested for the alkaline phosphatase (h‐TNAP and h‐IAP) and nucleotide pyrophosphatase/phosphodiesterase (h‐NPP1 and h‐NPP3) inhibition profile. Most of the derivatives exhibited selective and potent inhibition on tissue non‐specific and intestinal alkaline phosphatase and nucleotide pyrophosphatase (NPP1 and NPP3) enzymes. Possible binding modes of the most potent inhibitors were studied by molecular docking analysis.

Vertical Ultrafiltration-Facilitated DNA Digestion for Rapid and Sensitive UHPLC-MS/MS Detection of DNA Modifications.

LC-MS/MS technologies provide important and powerful analytical tools for chemical structure-dependent identification and quantification of epigenetically crucial DNA modifications. To perform LC-MS/MS analysis, it is better to convert DNA to 2'-deoxynucleosides through enzymatic digestion. Here, we observed that inorganic cations Na+ and K+ and phosphate buffers, which were often found in various DNA solutions, significantly inhibited DNA digestion as catalyzed by typical set of DNase I, snake venom phosphodiesterase, and calf intestine alkaline phosphatase, leading to poor or varying performance on UHPLC-MS/MS analysis. We then developed an efficient and unique vertical-ultrafiltration approach, enabling usto remove these inorganic salts without DNA loss. Consequently, the removal of inorganic salts by ultrafiltration facilitated the followed DNA digestion and thus enhanced the final UHPLC-MS/MS detection. Benefiting from the developed vertical-ultrafiltration approach, it is also feasible to integrate the desalting step with the other two steps of DNA digestion and protein removal. By investigating the time course of DNA digestion, we observed a differential release rate of 2'-deoxycytidine and 5-methyl-2'-deoxycytidine causing a measurement bias on the methylation frequency. We further exploited Mg2+ to eliminate this bias by stimulating DNase set-based DNA digestion. These innovative approaches enable us to perform rapid, sensitive, and robust UHPLC-MS/MS analysis of methylated DNA 2'-deoxycytidine, demethylation intermediates, and probably other DNA modifications.

Pressure Effect on Thermal Inactivation of Calf Intestine Alkaline Phosphatase

Pressure Effect on Thermal Inactivation of Calf Intestine Alkaline Phosphatase

Correction: Human alkaline phosphatase dephosphorylates microbial products and is elevated in preterm neonates with a history of late-onset sepsis.

[This corrects the article DOI: 10.1371/journal.pone.0175936.].

An Enzyme-Responsive "Turn-on" Fluorescence Polymeric Superamphiphile as a Potential Visualizable Phosphate Prodrug Delivery Vehicle.

The development of inexpensive and highly efficient enzyme-responsive polymers has significantly contributed to targeted drug delivery systems. Here, a superamphiphile with a capability of fluorescent dissociation sensing is designed. It is constructed with negatively charged adenosine 5'-triphosphate (ATP) and negatively charged fluorescein diphosphate (FDP), which are used as fluorescence detection, and a cationic diblock copolymer methoxy-poly(ethylene glycol)113 -b-poly(2-dimethyl-aminoethyl methacrylate)70 . Upon addition of calf intestinal alkaline phosphatase, the superamphiphile disintegrates, presumably due to the enzymatic hydrolysis of ATP. This process is accompanied by an increase in the fluorescence emission intensity of fluorescein owing to the hydrolysis of FDP. The in vitro application of the superamphiphile is also proven. Thus, the "turn-on" fluorescence of the superamphiphile serves as a real-time module for detection of the disintegration of superamphiphile.

Factors Known to Influence the Development of Necrotizing Enterocolitis to Modify Expression and Activity of Intestinal Alkaline Phosphatase in a Newborn Neonatal Rat Model.

Prematurity, formula feeding, and early weaning strongly influence enterocyte differentiation. Intestinal alkaline phosphatase (IAP), an endogenous protein expressed in the intestines, is one enzyme that is affected by these factors. IAP supplementation decreases the severity of necrotizing enterocolitis (NEC) injury. We, therefore, hypothesized that prematurity predisposes this population to NEC due to IAP deficiency and investigated IAP expression and function in a neonatal rat model. Pre- and full-term newborn Sprague-Dawley rat pups were sacrificed on consecutive days of life both after vaginal or cesarean birth and following either breast or formula feeding. Compared with controls, cesarean delivery and formula feeding are associated with lower levels of IAP. The formula-fed pups continued to have low baseline IAP activity. Neither prematurity nor formula feeding led to differences of intestinal injury. Prematurity and formula feeding are associated with inhibition of IAP expression and activity. Both may increase the risk of NEC and early enteral supplementation of IAP to newborns at risk of NEC may be of therapeutic benefit.