Mammalian cells utilize two transporters for the uptake of ascorbic acid (AA), Na-dependent vitamin C transporter SVCT-1 and SVCT-2. In the intestine, these transporters are involved in AA absorption and are expressed at the apical and basolateral membrane domains of the polarized epithelia, respectively. Little is known about the differential expression of these two transporters along the anterior-posterior axis of the intestinal tract and the molecular mechanism(s) that dictate this pattern of expression. We used mouse and human intestinal cDNAs to address these issues. The results showed a significantly lower rate of carrier-mediated AA uptake by mouse colon than jejunum. This was associated with a significantly lower level of expression of SVCT-1 and SVCT-2 at the protein, mRNA, and heterogeneous nuclear RNA (hnRNA) levels in the colon than the jejunum, implying the involvement of transcriptional mechanism(s). Similarly, expression levels of SVCT-1 and SVCT-2 mRNA and hnRNA were significantly lower in human colon. We also examined the levels of expression of hepatocyte nuclear factor 1α and specificity protein 1, which drive transcription of the and promoters, respectively, and found them to be markedly lower in the colon. Furthermore, significantly lower levels of the activating markers for histone (H3) modifications [H3 trimethylation of lysine 4 (H3K4me3) and H3 triacetylation of lysine 9 (H3K9ac)] were observed in the and promoters in the colon. These findings show, for the first time, that SVCT-1 and SVCT-2 are differentially expressed along the intestinal tract and that this pattern of expression is, at least in part, mediated via transcriptional/epigenetic mechanisms. Our findings show, for the first time, that transporters of the water-soluble vitamin ascorbic acid (i.e., the vitamin C transporters SVCT-1 and SVCT-2) are differentially expressed along the length of the intestinal tract and that the pattern of expression is mediated, at least in part, by transcriptional and epigenetic mechanism(s) affecting both and genes.
Thiamin is essential for normal metabolic activity of all mammalian cells, including those of the pancreas. Cells obtain thiamin from their surroundings and enzymatically convert it into thiamin pyrophosphate (TPP) in the cytoplasm; TPP is then taken up by mitochondria via a specific carrier the mitochondrial TPP transporter (MTPPT; product of the SLC25A19 gene). Chronic alcohol exposure negatively impacts the health of pancreatic acinar cells (PAC), but its effect on physiological/molecular parameters of MTPPT is not known. We addressed this issue using mouse pancreatic acinar tumor cell line 266-6 and primary PAC of wild-type and transgenic mice carrying the SLC25A19 promoter that were fed alcohol chronically. Chronic alcohol exposure of 266-6 cells (but not to its nonoxidative metabolites ethyl palmitate and ethyl oleate) led to a significant inhibition in mitochondrial TPP uptake, which was associated with a decreased expression of MTPPT protein, mRNA, and activity of the SLC25A19 promoter. Similarly, chronic alcohol feeding of mice led to a significant inhibition in expression of MTPPT protein, mRNA, heterogeneous nuclear RNA, as well as in activity of SLC25A19 promoter in PAC. While chronic alcohol exposure did not affect DNA methylation of the Slc25a19 promoter, a significant decrease in histone H3 euchromatin markers and an increase in H3 heterochromatin marker were observed. These findings show, for the first time, that chronic alcohol exposure negatively impacts pancreatic MTPPT, and that this effect is exerted, at least in part, at the level of Slc25a19 transcription and appears to involve epigenetic mechanism(s).
Thiamin is indispensable for the normal function of pancreatic acinar cells. These cells take up thiamin via specific carrier-mediated process that involves thiamin transporter-1 and -2 (THTR-1 and THTR-2; products of SLC19A2 and SLC19A3 genes, respectively). In this study we examined the effect of chronic exposure of pancreatic acinar cells in vitro (pancreatic acinar 266-6 cells) and in vivo (wild-type and transgenic mice carrying the SLC19A2 and SLC19A3 promoters) to the cigarette smoke component 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) on physiological and molecular parameters of the thiamin uptake process. The results show that chronic exposure of 266-6 cells to NNK (3 µM, 24 h) leads to a significant inhibition in thiamin uptake. The inhibition was associated with a significant decrease in the level of expression of THTR-1 and -2 at the protein and mRNA levels as well as in the activity of SLC19A2 and SLC19A3 promoters. Similarly chronic exposure of mice to NNK (IP 10 mg/100 g body weight, three times/week for 2 weeks) leads to a significant inhibition in thiamin uptake by freshly isolated pancreatic acinar cells, as well as in the level of expression of THTR-1 and -2 protein and mRNA. Furthermore, activity of the SLC19A2 and SLC19A3 promoters expressed in transgenic mice were significantly suppressed by chronic exposure to NNK. The effect of NNK on the activity of the SLC19A2 and SLC19A3 promoters was not mediated via changes in their methylation profile, rather it appears to be exerted via an SP1/GG and SP1/GC cis-regulatory elements in these promoters, respectively. These results demonstrate, for the first time, that chronic exposure of pancreatic acinar cells to NNK negatively impacts the physiological and molecular parameters of thiamin uptake by pancreatic acinar cells and that this effect is exerted, at least in part, at the level of transcription of the SLC19A2 and SLC19A3 genes.
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