ABSTRACT
BACKGROUND: The role of intestinal transporter regulation in optimising nutrient absorption has been studied extensively in rodent and cell line models but not in human subjects. AIMS: The aim of the present study was to investigate the response in vivo of zinc transporters in the human enterocyte to dietary zinc supplementation. SUBJECTS: Eighteen patients who had previously undergone ileostomy, all free of any symptoms of inflammatory bowel disease. METHODS: Subjects took a daily zinc supplement of 25 mg for 14 days in a double blind, placebo controlled, crossover trial. The effect of the supplement on expression in ileal biopsies of the zinc transporters SLC30A1, SLC30A4, SLC30A5, SLC39A1, SLC39A4, and metallothionein was measured by reverse transcription-polymerase chain reaction RT-PCR. Expression of SLC30A1, SLC30A5, and SLC39A4 was also examined by immunoblotting. RESULTS: The zinc supplement reduced SLC30A1 mRNA (1.4-fold) together with SLC30A1, SLC30A5, and SLC39A4 protein (1.8-fold, 3.7-fold, and to undetectable levels, respectively) in ileal mucosa and increased metallothionein mRNA (1.7-fold). The supplement had no effect on expression of SLC30A4 or SLC39A1 mRNA. Localisation of SLC30A5 at the apical human enterocyte/colonocyte membrane and also at the apical membrane of Caco-2 cells was demonstrated by immunohistochemistry. Commensurate with these observations in zinc supplemented human subjects, SLC30A1, SLC30A5, and SLC39A4 mRNA and protein were reduced in Caco-2 cells cultured at 200 muM compared with 100 muM zinc. CONCLUSIONS: These observations indicate that, in response to variations in dietary zinc intakes, regulated expression of plasma membrane zinc transporters in the human intestine contributes to maintenance of zinc status.
Subject(s)
Carrier Proteins/metabolism , Dietary Supplements , Gene Expression Regulation/drug effects , Ileum/metabolism , Zinc/pharmacology , Adult , Aged , Caco-2 Cells , Carrier Proteins/genetics , Cell Membrane/metabolism , Cross-Over Studies , Dose-Response Relationship, Drug , Double-Blind Method , Enterocytes/drug effects , Enterocytes/metabolism , Female , Homeostasis/drug effects , Humans , Male , Middle Aged , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction/methodsABSTRACT
Elevated plasma homocysteine is considered to be a risk factor for cardiovascular disease. The mechanisms for this effect are not fully understood but there is some evidence for a role for reactive oxygen species (ROS). This study was conducted to explore the effects of elevated plasma total homocysteine (tHcy) concentration on activity of antioxidant enzymes in the circulation. The study group consisted of 10 patients with inherited defects of homocysteine metabolism, from whom 41 blood samples were collected over a period of six months. Blood samples were also collected from 13 of their obligate heterozygous parents. For data analysis samples were classified as those with plasma tHcy < 20 microM or > 20 microM. The activity of erythrocyte superoxide dismutase (SOD) and plasma glutathione peroxidase (GSHPx) was elevated in samples with plasma tHcy > 20 microM. Moreover, a significant correlation was demonstrated between plasma GSHPx activity, plasma glutathione peroxidase protein and plasma tHcy. III vitro studies confirmed that this observation was not due to a simple chemical enhancement of enzyme activity. Homocysteine protected GSHPx from loss of activity following incubation at 37 degrees C. A similar effect was seen with another thiol-containing amino acid, cysteine. Results suggest that elevated plasma tHcy represents an oxidative stress, resulting in an adaptive increase in activity of antioxidant enzymes in the circulation.
