Expression of the PEPT1, CAT, SOD2 and GPX1 genes in the zebrafish intestine supplemented with methionine dipeptide under predation risk.

This study evaluated the effect of methionine supplementation, predation risk and their interaction on gut histology, whole-body cortisol levels, and intestinal gene expression in zebrafish. A total of 360 one-year-old animals were maintained under two environmental conditions and fed diets containing different methionine sources. Fish were fed either a control diet (CTL, without methionine supplementation), a diet supplemented with dl-methionine (DLM), or a diet supplemented with methionine dipeptide (MM) in the absence (AP) of a predator or in the presence of the predator (PP) for 48 h or 20 days. Predator-induced stress for 20 days resulted in lower body weight. Zebrafish fed methionine-supplemented diets had higher weight gain than control fish. We found no effect of predation stress or methionine supplementation on cortisol level. Predation risk and methionine supplementation showed no interaction effect on dipeptide transporter gene expression. After 48 h of predation pressure, zebrafish had higher mRNA expression of SOD2, CAT and GPX1 in the gut. After 20 days of exposure to the predator, zebrafish fed methionine-supplemented diets had lower expression of GPX1, SOD2 and CAT than those diet CTL. Methionine dipeptide and free methionine supplementation improved growth, intestinal health and survivability of zebrafish both conditions.

Enrichment of the amnion with threonine in chicken embryos affects the small intestine development, ileal gene expression and performance of broilers between 1 and 21 days of age.

The effects of in ovo feeding with threonine (Thr) on intestinal morphology, ileal gene expression and performance of broiler chicken between 1 and 21 d of age (d) were assessed. On day 17.5 of incubation, fertile eggs were randomly allotted to 5 treatments of Thr injection in the amniotic fluid (0; 1.75; 3.5; 5.25; 7%, corresponding to 17.5; 35; 52.5 and 70 mg Thr/mL). After hatch, chicks were given a commercial corn-soybean diet up to 21 d. Daily feed intake (FI), body weight (BW), and food conversion ratio (FCR) were measured from 1 to 7, 14, and 21 d of age. The ileal gene expression of mucin (MUC2), peptide transporter (PepT1), and aminopeptidase enzyme (APN) were evaluated on day of hatch and at 21 d, as well as intestinal morphometric traits. In ovo feeding with threonine significantly increased final weight (FI) and weight gain (WG) and decreased FCR in the period from 1 to 21 d. Threonine levels affected beneficially the villus height, vilo: crypt ratio and villus area on day of hatch and at 21 d. At hatch, all Thr levels increased the expression of MUC2 and PepT1 compared to the control group. APN expression also increased, but for the lowest and the highest threonine levels (1.75 and 7%). At 21 d, there was no effect of threonine on the expression of MUC2, PepT1, and APN. In conclusion, in ovo threonine feeding beneficially affected the morphological and functional development of the intestinal mucosa, which ensured improved performance of chicks at hatch and at 21 d.

Maternal protein restriction during pregnancy affects gene expression and immunolocalization of intestinal nutrient transporters in rats.

Intrauterine dietary restriction may cause changes in the functioning of offspring organs and systems later in life, an effect known as fetal programming. The present study evaluated mRNA abundance and immunolocalization of nutrient transporters as well as enterocytes proliferation in the proximal, median and distal segments of small intestine of rats born to protein-restricted dams. Pregnant rats were fed hypoproteic (6% protein) or control (17% protein) diets, and offspring rats were evaluated at 3 and 16 weeks of age. The presence of SGLT1 (sodium-glucose co-transporter 1), GLUT2 (glucose transporter 2), PEPT1 (peptide transporter 1) and the intestinal proliferation were evaluated by immunohistochemical techniques and the abundance of specific mRNA for SGLT1, GLUT2 and PEPT1 was assessed by the real-time PCR technique. Rats born to protein-restricted dams showed higher cell proliferation in all intestinal segments and higher gene expression of SGLT1 and PEPT1 in the duodenum. Moreover, in adult animals born to protein-restricted dams the immunoreactivity of SGLT1, GLUT2 and PEPT1 in the duodenum was more intense than in control rats. Taken together, the results indicate that changes in the small intestine observed in adulthood can be programmed during the gestation. In addition, they show that this response is caused by both up-regulation in transporter gene expression, a specific adaptation mechanism, and intestinal proliferation, an unspecific adaptation mechanism.

Study of the mechanisms of uptake of 5-aminolevulinic acid derivatives by PEPT1 and PEPT2 transporters as a tool to improve photodynamic therapy of tumours.

Endogenous porphyrin accumulation after administration of 5-aminolevulinic acid is employed in photodynamic therapy of tumours. Due to its low membrane permeability, esterified 5-aminolevulinic acid derivatives less hydrophilic than the parental compound are under investigation. Knowledge of the mechanisms of 5-aminolevulinic acid derivatives uptake into target cells is essential to understand and improve photodynamic therapy and useful in the design of new derivatives with better affinity and with higher selectivity for tumour cells in specific tissues. The aim of this work was to assess the interaction of 5-aminolevulinic acid derivatives with the intestinal PEPT1 and renal transporter PEPT2 expressed in Pichia pastoris yeasts. We found that Undecanoyl, Hexyl, Methyl and 2-(hydroxymethyl)tetrahydropyranyl 5-aminolevulinic acid esters and the dendron 3m-ALA inhibited (14)C-5-aminolevulinic acid uptake by PEPT2. However, only the Undecanoyl ester inhibited 5-aminolevulinic acid uptake by PEPT1. We have also found through a new developed colorimetric method, that Hexyl and 2-(hydroxymethyl)tetrahydropyranyl 5-aminolevulinic acid esters display more affinity than 5-aminolevulinic acid for PEPT2 whereas none of the compounds surpass 5-aminolevulinic acid affinity for PEPT1. In addition, the Undecanoyl ester binds with high affinity to the membranes of PEPT2 and PEPT1-expressing yeasts and to the control yeasts. The main finding of this work was that some derivatives have the potential to improve 5-aminolevulinic acid-based photodynamic therapy by increased efficiency of transport into cells expressing PEPT2 such as kidney, mammary gland, brain or lung whereas in tissues expressing exclusively PEPT1 the parent 5-aminolevulinic acid remains the compound of choice.

Alterations in hexose, amino acid and peptide transporter expression in intestinal epithelial cells during Nippostrongylus brasiliensis infection in the rat.

Infection with the nematode Nippostrongylus brasiliensis induces various types of cytological alterations in the intestinal villus epithelium. The aim of this study was to analyse the expression of hexose, peptide and amino acid transporters in the small intestinal epithelium after infection. Brown-Norway rats were infected with 2000 N. brasiliensis L3 larvae and villus epithelial cells were isolated at various time points after infection. Expression of hexose transporters Na(+)/glucose cotransporter SGLT1 and glucose transporter GLUT-1, -2 and -5, a peptide transporter (PepT1) and an amino acid transporter (LAT2) was examined by reverse transcription-PCR, Western blotting or immunohistochemistry. Semi-quantitative reverse transcription-PCR studies of separated jejunal epithelial cells showed that expression levels of GLUT5, PepT1 and LAT2 were significantly decreased 7 and 14 days after infection, while these changes were not observed in the ileal epithelium. Although the apical surface glucose transporter SGLT1 showed no significant alteration in mRNA expression, Western blotting analyses of jejunal epithelial cell lysate showed a marked decrease. Contrary to SGLT1, GLUT5, PepT1 and LAT2, expression of GLUT1, which is essential in maintaining high rates of glucose influx, was significantly up-regulated in the jejunal epithelium 7 and 14 days after infection in reverse transcription-PCR as in Western blotting analyses. Immunohistochemical studies showed that GLUT1 immunoreactivity was localised to the basolateral membrane of intestinal epithelial cells 7 days after infection. These results show that N. brasiliensis infection results in an increase in GLUT1 and a decrease in various hexose, amino acid and peptide transporter expression in jejunal epithelial cells. Up-regulation of GLUT1 might be a compensatory response in injured epithelial cells.