Mechanism and regulation of folate uptake by pancreatic acinar cells: effect of chronic alcohol consumption

Abstract

Said HM, Mee L, Sekar VT, Ashokkumar B, Pandol SJ. Mechanism and regulation of folate uptake by pancreatic acinar cells: effect of chronic alcohol consumption. Am J Physiol Gastrointest Liver Physiol 298: G985-G993, 2010. First published April 1, 2010; doi:10.1152/ajpgi.00068.2010.-Folate plays an essential role in onecarbon metabolism, and a relationship exists between methyl group metabolism and pancreatic exocrine function. Little, however, is known about the mechanism(s) and regulation of folate uptake by pancreatic acinar cells and the effect of chronic alcohol use on the process. We addressed these issues using the rat-derived pancreatic acinar cell line AR42J and freshly isolated primary rat pancreatic acinar cells as models. We found [3 H]folic acid uptake to be 1) temperature and pH dependent with a higher uptake at acidic than at neutral/alkaline pH; 2) saturable as a function of substrate concentration at both buffer pH 7.4 and 6.0; 3) inhibited by folate structural analogs and by anion transport inhibitors at both buffer pH 7.4 and 6.0; 4) trans-stimulated by unlabeled folate; 5) adaptively regulated by the prevailing extracellular folate level, and 6) inhibited by modulators of the cAMP/PKA-mediated pathway. Both the reduced folate carrier (RFC) and the proton-coupled folate transporter (PCFT) were found to be expressed in AR42J and in primary pancreatic acinar cells, as well as in native human pancreas with expression of RFC being higher than PCFT. Chronic alcohol feeding of rats (4 wk; 36% of calories from ethanol) led to a significant decrease in folate uptake by freshly isolated primary pancreatic acinar cells compared with cells from pair-fed controls; this effect was associated with a parallel decrease in the level of expression of RFC and PCFT. These studies reveal that folate uptake by pancreatic acinar cells is via a regulated carrier-mediated process which may involve RFC and PCFT. In addition, chronic alcohol feeding leads to a marked inhibition in folate uptake by pancreatic acinar cells, an effect that is associated with reduction in level of expression of RFC and PCFT.RFC; PCFT; transporter; pancreas; chronic alcohol use FOLATE, A WATER-SOLUBLE VITAMIN, is essential for maintaining normal cellular function, growth, and development. The vitamin acts as a cofactor in a number of critical metabolic reactions that include synthesis of precursors of DNA and RNA, metabolism of certain amino acids (e.g., homocysteine), and other methylation reactions. Thus it is not surprising that folate deficiency leads to a variety of clinical abnormalities that range from megaloblastic anemia to neural tube defects.All mammalian cells, including those of the pancreatic acini, are unable to synthesize folate and thus must obtain the vitamin from the extracellular environment via transport across the plasma membrane. The pancreas maintains the second highest level of folate after the liver (3, 30), and folate is essential for its normal exocrine function and health. A reduction in amylase secretion, appeara...

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“…Mouse primary PAC were isolated from the pancreas of adult mice by the Worthington collagenase type IV (Lakewood, NJ) digestion method as described previously (42)(43)(44). Isolated cells were suspended in K-R buffer, and uptake was performed within 1 h of isolation.…”

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“…In addition to dietary inadequacy, alcohol may decrease internal folate levels through intestinal malabsorption, decreased hepatic storage and increased renal excretion (124,125). Folate homeostasis depends on transporter proteins, including reduced folate carrier, proton-coupled folate transporter, folate-binding protein, mitochondrial folate transporter and enzymes such as folylpolyglutamate synthetase (126)(127)(128)(129). Chronic ethanol exposure may downregulate gene expression, thus impairing the transportation of folate across membranes (126)(127)(128)(129).…”

“…Folate homeostasis depends on transporter proteins, including reduced folate carrier, proton-coupled folate transporter, folate-binding protein, mitochondrial folate transporter and enzymes such as folylpolyglutamate synthetase (126)(127)(128)(129). Chronic ethanol exposure may downregulate gene expression, thus impairing the transportation of folate across membranes (126)(127)(128)(129). With the exception of causing folate deficiency, ethanol intake interferes widely in folate-dependent intermediary metabolism by inhibiting enzymes in the one-carbon metabolism, particularly methionine synthase and its associated products of metabolism, thus disturbing the synthesis of nucleotides (130).…”

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“…Recent studies have delineated the mechanism of folate uptake by pancreatic acinar cells and show the involvement of a carrier-mediated mechanism at acidic and neutral/ alkaline pH levels (73). The reduced folate carrier (RFC) and the proton-coupled folate transporter (PCFT) are expressed in pancreatic acinar cells, with higher expression of RFC than PCFT (73); the contribution of these systems to folate uptake by pancreatic acinar cells is not clear but most likely depends on the prevailing extracellular pH. The pancreatic folate uptake process is adaptively regulated by the prevailing extracellular folate level and also appears to be under the regulation of an intracellular cAMP/PKA-mediated pathway (73).…”

“…Finally, chronic feeding of alcohol to animals was found to lead to a significant inhibition of pancreatic acinar folate uptake and suppression of the level of expression of RFC and PCFT (73). These findings may contribute to the deleterious effects of chronic alcohol exposure on pancreatic health [i.e., acute and chronic pancreatitis (35)], which is multifactorial and involves a reduction of pancreatic defense mechanisms (35,45,50).…”