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© 2006 American Society for Nutrition J. Nutr. 136:2308-2313, September 2006

Nutrient Physiology, Metabolism, and Nutrient-Nutrient Interactions

Vanadate but Not Tungstate Prevents the Fructose-Induced Increase in GLUT5 Expression and Fructose Uptake by Neonatal Rat Intestine1,2

Séverine Kirchner, Edward Kwon, Anjali Muduli, Carla Cerqueira, Xue-Lin Cui and Ronaldo P. Ferraris*

Department of Pharmacology and Physiology, University of Medicine and Dentistry of New Jersey (UMDNJ)—New Jersey Medical School, Newark, NJ 07103-2714

* To whom correspondence should be addressed. E-mail: ferraris{at}umdnj.edu.


   ABSTRACT
TOP
 ABSTRACT
Introduction
 Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
Intermediary signals, precociously enhancing GLUT5 transcription in response to perfusion of its substrate, fructose, in the small intestine of neonatal rats, are not known. Because glucose-6-phosphatase (G6Pase), glucose-6-phosphate translocase (G6PT), and fructose-1,6-bisphosphatase (FBPase) expression increases parallel to or precedes that of GLUT5, we investigated the link between these gluconeogenic genes and GLUT5 by using vanadate or tungstate, potent inhibitors of gluconeogenesis. Small intestinal perfusions of 20-d–old rats were performed with fructose alone, fructose + vanadate or tungstate, glucose alone, and glucose + vanadate or tungstate. As expected, fructose, but not glucose nor glucose + inhibitor perfusion, increased GLUT5 mRNA abundance and fructose transport. Fructose perfusion dramatically increased G6Pase mRNA abundance but had no effect on G6Pase activity. In sharp contrast, fructose perfusion did not increase FBPase gene expression but stimulated FBPase activity. Both vanadate and tungstate significantly inhibited G6Pase activity but did not prevent the fructose-induced increases in G6Pase and G6PT gene expression. Perfusion with fructose + vanadate prevented the fructose-induced increases in fructose transport and GLUT5 mRNA abundance, whereas perfusion with fructose + tungstate did not. Interestingly, vanadate, but not tungstate, inhibited the fructose-induced increase in FBPase activity. Thus, vanadate inhibition of fructose-induced increases in FBPase activity paralleled exactly vanadate inhibition of fructose-induced increases in GLUT5 mRNA abundance and activity. Fructose-induced changes in FBPase activity may regulate changes in GLUT5 expression and activity in the small intestine of neonatal rats. The marked increases in intestinal G6Pase and GLUT5 mRNA abundance may be a parallel response to different factors released during fructose perfusion.


   Introduction
TOP
 ABSTRACT
 Introduction
Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
Diets high in fructose can result in metabolic abnormalities, such as insulin resistance and hypertriglyceridemia, caused by disturbances in hepatic and intestinal sugar uptake and metabolism (1). One proposed mechanism involves intestinal fructose transporter (GLUT5),3 which has a significantly higher expression in adipocytes of young obese rats than lean controls. As the rats age and become diabetic, GLUT5 abundance and activity is markedly reduced in adipose tissue of obese rats, causing more marked insulin resistance and indicating a possible role of this transporter in the pathology of the metabolic syndrome associated with fructose feeding and insulin resistance (2). Interestingly, per capita fructose consumption by young children has increased faster than that by the general population (3), but few studies have focused on the physiological adaptations or gene regulation mechanisms underlying early exposure to fructose in the intestine. Fructose absorption rates and GLUT5 mRNA abundance are very low during the suckling and early weaning stages of rat intestinal development. However, the consumption of a high-fructose (HF) diet, or intestinal perfusion in vivo with a HF solution for 2–4 h, precociously and markedly increases GLUT5 activity and mRNA abundance in rats as young as 16–18 d of age, whereas the brush border sodium-glucose cotransporter (SGLT1) is not affected (4).

We identified by microarray fructose-1,6-bisphosphatase (FBPase) and glucose-6-phosphatase (G6Pase) as fructose-responsive genes, paralleling fructose-induced changes in GLUT5 (5). Hence, we hypothesized that products of these enzymes may serve as key intermediary signals in the fructose-induced activation of the GLUT5 mRNA level and activity in neonatal rat small intestine. FBPase is a highly regulated enzyme that catalyzes the second-to-last step in gluconeogenesis, preceding G6Pase in the metabolic pathway. FBPase is acutely regulated by fructose-2,6-bisphosphate, a product of the bifunctional enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. Years ago, Korieh and Crouzoulon (6) found that a high fructose diet increases rat hepatic and intestinal FBPase activities. G6Pase is a multicomponent enzyme that is tightly associated with the endoplasmic reticulum (ER) membrane: glucose-6-phosphate (G6P) hydrolysis involves a glucose-6-phosphate translocase (G6PT) protein, which transports G6P from the cytosol across the ER, and a glucose-6-phosphatase catalytic subunit (G6Pase) located on the luminal side of the ER (7). A high fructose diet increases rat hepatic G6Pase gene expression and activities as well as G6Pase activities in the intestine (6,8,9). Fructose perfusion also increases intestinal G6Pase mRNA levels, which is an increase known to precede fructose-induced increases in GLUT5 expression (5).

In light of the increasing incidence of fructose-induced metabolic syndromes, the effect of fructose on intestinal G6Pase and FBPase may be critically important because intestinal gluconeogenesis, previously thought of as insignificant relative to renal and hepatic gluconeogenesis, may actually contribute to the total endogenous production of glucose (7). Moreover, G6Pase likely plays a key role in this process (10). Although the existence of intestinal gluconeogenesis remains controversial even in neonates, the expression of gluconeogenic enzymes in the intestine is still high at this stage of development (11). We hypothesize that the gluconeogenic enzymes G6Pase and/or FBPase may regulate GLUT5 gene expression and fructose transport in the small intestine of neonatal rats. To evaluate the role of gluconeogenesis in the fructose-induced increase in GLUT5 expression, we used 2 potent inhibitors of gluconeogenesis: sodium orthovanadate (vanadate, V) (12) and sodium tungstate (tungstate, T) (13).


   Materials and Methods
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 ABSTRACT
 Introduction
 Materials and Methods
Results
 Discussion
 LITERATURE CITED
 
    Animals and experimental design. Pregnant female Sprague-Dawley rats (Taconic), who were allowed to consume water and nonpurified diet ad libitum (Purina Mills), were monitored until pups were born (d 0). The pups were kept with their dams until they were 20- to 22-d–old and then they were randomly separated into 4 groups. Sodium orthovanadate and sodium tungstate (Sigma), cell-permeable, potent inhibitors of G6Pase catalytic subunit activity, were selected to investigate the regulatory role of G6Pase in the fructose-induced increase of GLUT5 expression in the neonatal rat small intestine. Mid-weaned rat pups were assigned to 2 different sets of experiments to test the effects of each inhibitor. G (glucose), G + I (glucose + inhibitor), F (fructose) and F + I (fructose + inhibitor) in KRB solutions were continuously perfused for 4 h, where the inhibitor (I) was either vanadate (Expt. A) or tungstate (Expt. B). Both inhibitors were soluble in aqueous solution. A series of bioassays (not shown) found that 2 mmol/L was the most effective concentration in blocking the fructose-induced increase in fructose uptake in the neonatal rat intestine for both vanadate and tungstate. Moreover, for both inhibitors, the chosen concentration had no effect on mortality rate during perfusion. All procedures conducted in this study were approved by the Institutional Animal Care and Use Committee, University of Medicine and Dentistry, New Jersey Medical School.

    Perfusion model. The rat intestinal perfusion procedure was conducted following the method of Jiang and Ferraris (4). The small intestine was continuously perfused with sugar solution (100 mmol/L fructose or glucose in KRB ± inhibitor) at a rate of 30 mL/h at 37°C for 4 h. Composition of the perfusion solution was as follows (in mmol/L): 78 NaCl, 4.7 KCl, 2.5 CaCl2·5H2O, 1.2 MgSO4, 19 NaHCO3, 2.2 KH2CO3, and 100 mmol/L glucose or fructose (± 2 mmol/L inhibitor). Rat pups were kept under continuous anesthesia. The perfusion duration and sugar concentration were based on previous findings (4,14). Rats were killed by a lethal euthasol injection into the heart. Jejunal samples were collected for determinations of sugar uptake rates. Mucosal scrapes of intestine were quickly frozen in liquid nitrogen and then stored at –80°C for subsequent determinations of enzyme activities and mRNA levels.

    Fructose and glucose uptake measurements. After perfusion, everted jejunum sleeves were prepared as previously described (4) and incubated at 37°C in an oxygenated, stirred (1,200 rpm) solution containing either [D-14C]-labeled glucose for 1 minute or [D-14C]-labeled fructose for 2 min (Perkin Elmer). [L-3H]-labeled glucose was also added to correct for adherent fluid and passive diffusion of glucose and fructose. Results were expressed as nmol/mg wet weight of intestine.

    Total RNA extraction and processing. Total RNA was extracted using the TRIzol reagent (GIBCO-BRL). Total RNA samples were treated by RQ1 RNase-Free DNase prior to RT-PCR (Promega), to avoid genomic DNA amplification. cDNA was generated from 1µg DNase-treated RNA using SuperScript III RNase H-Reverse Transcriptase (Invitrogen). The samples were then stored at –20°C for subsequent relative quantification of specific mRNA levels by real-time PCR.

    Gene expression levels by real-time PCR. Real-time PCR was performed in the MX 3000P (Stratagene). Quantitative PCRs were performed on 10 µL of the RT reaction mixture using the Brilliant SYBR Green QPCR Master Mix (Stratagene). The total volume of the PCR was 25 µL, and contained 100 nmol/L of primers chosen with the primer3 tool web site (15) (Supplemental Table 1). Thermal cycling was initiated with incubation at 95°C for 10 min for SureStart Taq DNA polymerase activation. Forty steps of PCR were performed; each step consisted of heating the mixture at 95°C for 30 s for denaturing, at 59°C for 30 s for annealing, and 1 min for extension. Following the final cycle of the PCR, melting curves were systematically monitored (55°C temperature gradient at 0.05°C/s from 55 to 95°C). Serial dilutions of cDNAs generated from selected samples that expressed target genes at a suitable level were used to generate a standard curve for each target gene and the chosen reference gene transcript [housekeeping gene {alpha}-elongation factor 1(EF1{alpha})] to evaluate PCR efficiency. Relative quantification of the target gene transcript (G6Pase, G6PT, FBPase, SGLT1 or GLUT5), in comparison with EF1{alpha} expression level in the same sample, was made following the method by Pfaffl et al. (16).

    Enzyme activities. The assay of intestinal G6Pase activity was based on the specific inactivation of the enzyme upon brief exposure to an acidic pH at 37°C (17). Rat intestine scrapes were homogenized in Hepes 10 mmol/L, DTT 1 mmol/L, and saccharose 0.25 mol/L. One-half of each homogenate was brought to pH 4.5 by the adding 1 mol/L HCl and incubating for 10 min at 37°C to assay for nonspecific phosphohydrolase activity. The pH was then neutralized to 7.0 by adding 1 mol/L NaOH. Treated and nontreated microsomes were prepared and G6Pase activities assayed as previously described (18). Nonspecific alkaline phosphatase activities were assayed using the same protocol and were found to be negligible. FBPase activity assays were performed on the cytosolic fraction as previously described (18). All enzyme activities were expressed as per mg of protein. Protein concentration was determined using a Bio Rad protein assay kit (Bio Rad) with bovine serum albumin as standard. One unit of the enzyme activity was defined as the amount of enzyme that catalyzed the hydrolysis of 1 µmol of substrate under the specified conditions (37°C).

    Statistical analyses. Data are presented as means ± SEM. A 2-way ANOVA was first used to determine the difference of relative mRNA abundance among groups with different treatments. If there was a significant difference, Fisher's paired least significant difference (LSD) test was used to determine the particular effect that caused that difference (STATVIEW, Abacus Concepts). Differences were considered significant, P < 0.05.


   Results
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 ABSTRACT
 Introduction
 Materials and Methods
 Results
Discussion
 LITERATURE CITED
 
    Effect of fructose, vanadate, and tungstate on G6Pase and G6PT. Compared with intestine perfused with HG, G6Pase mRNA abundance increased by >800% in rat small intestine perfused with HF (P < 0.001; Tables 1 and 2). There was a >100% increase in G6PT mRNA levels in HF-perfused intestines (P < 0.001). Perfusion of vanadate with fructose did not prevent the fructose-induced increase in G6Pase and G6PT mRNA abundance (Table 1). Vanadate also did not alter the low expression levels of G6Pase and G6PT in HG-perfused intestine. Interestingly, there was no significant fructose-induced increase in G6Pase activity, but vanadate reduced G6Pase activity in both glucose- and fructose-perfused intestines (P < 0.0001). In the tungstate experiment, we found fructose-induced increases in G6Pase (>900%) and G6PT (>100%) mRNA abundance (P < 0.001; Table 2). These fructose-induced increases were similar to those in the vanadate study. Tungstate also had no effect on mRNA abundance of G6Pase and G6PT in both HF- and HG-perfused intestines. Tungstate inhibited G6Pase activity in HG- and HF-perfused intestines (P < 0.001; Table 2).


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  		TABLE 1 Effect of vanadate perfusion of neonatal rat small intestine on glucose- and fructose-induced changes of G6Pase, FBPase, and SGLT1 expression and activity1

 

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  		TABLE 2 Effect of tungstate perfusion of neonatal rat small intestine on glucose- and fructose-induced changes of G6Pase, FBPase, and SGLT1 expression and activity in neonatal rat intestine1

 
    Effect of fructose, vanadate and tungstate on FBPase. In sharp contrast to those of G6Pase, FBPase mRNA abundance did not change with fructose perfusion in both the vanadate (Table 1) and tungstate (Table 2) experiments. Vanadate and tungstate did not effect mRNA abundance of FBPase in both HG- and HF-perfused intestines. In another important difference from those of G6Pase, FBPase activities increased from 70 to 90% with HF perfusion (P < 0.01) in both experiments. Vanadate prevented the fructose-induced increase in FBPase activity. This effect of vanadate is specific, because tungstate did not prevent the fructose-induced increase in FBPase activity, and vanadate did not affect FBPase activity in glucose-perfused intestine.

    Effect of vanadate and tungstate on SGLT1. The SGLT1 mRNA abundance between HG- and HF-perfused intestines did not differ (Tables 1 and 2). Vanadate and tungstate each had no effect on SGLT1 expression in both HG- and HF-perfused intestines. Glucose uptake rate between the HG- and HF-perfused intestine did not differ. Vanadate but not tungstate markedly reduced (P < 0.0001) glucose uptake in both HG- and HF-perfused intestine.

    Effect of vanadate and tungstate on GLUT5. Compared with HG-perfused intestine, fructose perfusion markedly increased GLUT5 mRNA abundance by >400% (P < 0.0001, Fig. 1A,C).Vanadate prevented the fructose-induced increase in GLUT5 expression, but had no effect on GLUT5 expression in glucose-perfused intestine. Tungstate had no effect on GLUT5 expression in both HF- and HG-perfused intestines. Fructose perfusion increased the rate of intestinal fructose uptake by ~50% (P < 0.05), and vanadate also prevented this fructose-induced increase in GLUT5 activity (Fig. 1B). The effect of vanadate on GLUT5 expression and activity is specific because the fructose-induced increase in intestinal fructose uptake and GLUT5 mRNA abundance was not prevented by tungstate (Fig. 1C,D). Moreover, vanadate did not decrease fructose uptake and GLUT5 mRNA abundance in intestines perfused with glucose.


Figure 1
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  		Figure 1  The effect of a 4 h vanadate (A, B) or tungstate (C, D) perfusion of neonatal rat small intestine, with or without 100 mmol/L glucose (HG) or fructose (HF) solutions, on relative GLUT5 mRNA abundance (A, C) and fructose uptake rate (B, D). Bars represent means + SEM, n = 4. *Significant effects (P < 0.05) were: (A, B) sugar, interaction; (C, D) sugar. *Different from other means in the panel, P < 0.05. (Fisher's LSD test was conducted when the interaction was significant.) Data for the HG treatment were set to 1.0.

 

   Discussion
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
LITERATURE CITED
 
The main finding of this study was that intestinal perfusion with fructose increased G6Pase mRNA abundance and FBPase activity, and that this fructose-mediated effect on FBPase activity might play a role in modulating the well-known effect of fructose on GLUT5 expression and activity. Intestinal fructose perfusion dramatically increased G6Pase expression but not activity, and it stimulated FBPase activity without changing mRNA abundance. Perturbing gluconeogenesis with the G6Pase inhibitor vanadate decreased intestinal glucose uptake, and simultaneously prevented the fructose-induced increases in FBPase activity, fructose uptake, and GLUT5 expression. Because tungstate, like vanadate, inhibited G6Pase but did not prevent fructose-induced increases in FBPase activity, fructose uptake, or GLUT5 expression, there may be a potential link between FBPase activity and GLUT5 regulation.

    Luminal fructose and intestinal G6Pase and FBPase. Despite the dramatic and rapid increase in G6Pase mRNA levels, there was paradoxically no subsequent increase in intestinal G6Pase activity. A similar disconcordance has been observed in the adult rat liver where G6Pase mRNA abundance but not activity increased with dietary sucrose (8). G6Pase activity may not change (among other reasons) because: the mRNA is not translated and therefore G6Pase protein abundance is low; G6Pase protein abundance is high, but activity is reduced by allosteric inhibition; and sufficient amounts of substrate are unavailable to G6Pase. This latter explanation is possible because fructose markedly enhanced G6Pase but only modestly increased G6PT expression (G6PT transports G6P from the cytosol to the endoplasmic reticulum for processing by G6Pase). Therefore, G6Pase catalytic activity may also have been inhibited because of a limited supply of its substrate G6P. In this study, we were unable to determine glucose-6-phosphate transport in microsomes or G6Pase protein abundance because of limited amounts of neonatal jejunal tissue. The consumption of fructose has been shown to increase G6Pase activity in the liver of adult rats (6), but in these early studies, G6Pase mRNA abundance was not measured. It is unclear why these findings differ from ours and those of Pagliasotti et al. (8).

In contrast to G6Pase, fructose perfusion stimulated intestinal FBPase activities independent of changes in the abundance of FBPase mRNA. A high fructose diet also increased FBPase activities in adult rat liver and intestine (6). The discrepancy between FBPase gene expression and enzyme activity is not surprising, because complex transcriptional and post-transcriptional mechanisms are involved in the regulation of this enzyme (19). In particular, alterations in FBPase activity do not always involve changes in FBPase mRNA abundance and are highly dependant on allosteric effectors (20), including fructose-2,6-bisphosphate produced by the bifunctional enzyme fructose-2,6-bisphosphatase/6-phosphofructokinase-2 (21), which was also recently found to be fructose-responsive in the neonatal rat intestine (5). Because FBPase precedes G6Pase in the gluconeogenic pathway, the possibility that the fructose-induced activation of FBPase activities might directly or indirectly stimulate increases in G6Pase mRNA, cannot be ruled out.

    Effect of inhibitors on intestinal G6Pase and FBPase. Vanadium and tungsten compounds are potential therapeutic agents for diabetes, because they normalize plasma glucose homeostasis, carbohydrates, and lipid metabolism by currently unknown mechanisms (22,23). They generally mimic most of the actions of insulin in various insulin-responsive cells and tissues (24,25) but may also exhibit noninsulin-like effects (26). In neonatal rats, both vanadate and tungstate dramatically reduced intestinal G6Pase activity whether glucose or fructose was perfused along with the inhibitor. This finding confirms the results of previous work (12,13,27). Surprisingly, mRNA levels of intestinal G6Pase catalytic and translocase subunits were both vanadate- and tungstate-insensitive. Unfortunately, to our knowledge, there have been no studies on the effect of these inhibitors on intestinal G6Pase mRNA abundance. In contrast, our data support the post-transcriptional effect of vanadate on intestinal G6Pase and FBPase.

The vanadate effect on the fructose-induced increase in FBPase activity was not accompanied by changes in FBPase mRNA levels. Vanadate is known to increase the levels of the FBPase-potent allosteric inhibitor, fructose-2,6-bisphosphate, in rat hepatocytes (28), heart (29), and intestine in diabetic models (30). Vanadate, by increasing fructose-2,6-bisphosphate concentrations, can therefore inhibit the fructose-induced increase in FBPase activity. Vanadate inhibition of the fructose-induced effect may be specific, because vanadate did not inhibit baseline FBPase activity in glucose-perfused intestines, and tungstate did not prevent the fructose-induced increase in FBPase activity. However, tungstate was also reported to require 3- to 5-fold higher concentrations than vanadate to raise fructose-2,6-bisphosphate concentrations (25). Because we used similar modest concentrations of vanadate and tungstate (2 mmol/L, proven to be effective against G6Pase), we postulate that vanadate but not tungstate inhibited the fructose-induced increase in FBPase activity because vanadate was more effective at these concentrations. Hepatic FBPase has also been shown to be less vulnerable to tungstate inhibition, especially in nondiabetic rats (31).

    The role of G6Pase and FBPase in GLUT5 regulation. Luminal fructose in the neonatal rat small intestine consistently enhances GLUT5 expression and activity and also increases G6Pase mRNA abundance and FBPase activity. Are these events causally linked, or are they parallel, coincidental responses to fructose perfusion? The divergent effect of vanadate and tungstate on FBPase activities was an important clue, because vanadate, but not tungstate, was able to prevent the fructose-induced activation of GLUT5 in the neonatal rat small intestine. The vanadate-sensitivity of GLUT5 expression was found in adult rat adipocytes (32). Vanadate inhibition of fructose-induced increases in FBPase activity paralleled exactly vanadate inhibition of fructose-induced increases in GLUT5 mRNA abundance and activity. Hence, fructose-induced changes in FBPase activity may play a role in increasing GLUT5 mRNA abundance and activity perhaps via metabolites produced by gluconeogenesis thought to occur in neonatal intestines (11). Future studies will determine whether the fructose-induced activation of FBPase activity is sufficient by itself to trigger the induction of GLUT5 in the small intestine of neonate rats, by specifically modulating FBPase activity.

It is unclear, even in neonates that exhibit high gluconeogenic enzyme expression, whether gluconeogenic enzymes in the intestine actually produce endogenous glucose or are related to other processes in the intestine (11). Because salts of vanadium interfere not only with a variety of phosphatases but also with other enzyme systems such as ATPases (33), an indirect or more complex mechanism potentially unrelated to gluconeogenic capacities could be responsible for the observed prevention of fructose-induced activation of GLUT5 by vanadate. Vanadate inhibits glucose uptake without modifying SGLT1 mRNA levels in both glucose and fructose-perfused intestine of nondiabetic neonatal rats. Inhibition of intestinal glucose absorption by vanadate is thought to be mediated by decreases in activity and expression of {alpha}-glucosidases as well as of GLUT2, a facilitative glucose transporter located in the basolateral membrane of intestinal epithelial cells (34). Vanadate was also shown to regulate Na+/K+-ATPase activity and GLUT4 translocation in diabetic rats (35). The Na+/K+-ATPase mechanism could not be invoked in this experiment because Na+-dependent proline uptakes were not modified by vanadate perfusions (S. Kirchner and R. Ferraris, unpublished observations). Our data suggested that the vanadate effect on GLUT5 was mediated by a transcriptional mechanism but GLUT5 protein levels and translocation to the brush border membrane might be similarly affected. These alternative mechanisms of action of vanadate will be investigated in future studies by quantifying {alpha}-glucosidase, GLUT2, and GLUT5 protein.

The coactivation of intestinal G6Pase and GLUT5 gene expression by luminal fructose could be unrelated for the following reasons. First, only G6Pase mRNA abundance increased with fructose, whereas G6Pase activity did not; in contrast, both GLUT5 mRNA abundance and activity increased with fructose. Second, the tungstate-induced inhibition of G6Pase activity does not lead to an inhibition of the fructose-induced induction of GLUT5 in the neonate rat small intestine. This hypothesis was reinforced by a comparison of the promoter regions of G6Pase, G6PT, and GLUT5. Using an analytical program, the Gene2Promoter (36), we identified 5 models, each consisting of 3 potential transcription-factor-binding sites that follow the same sequential order of appearance on the 3 putative promoters. Within 1000 base pairs upstream of the transcription start site, 2 of these models were not found on SGLT1 or NaPi2b, which are 2 transporters whose mRNA levels are not increased by luminal fructose (5). Such a high similarity of regulatory frameworks on the 3 promoter regions suggests that G6Pase, G6PT, and GLUT5 genes are more likely regulated in parallel by similar pathways and mechanisms related to fructose appearance. In contrast to G6Pase and G6PT, the GLUT5 promoter would additionally contain a response element that would be blocked by vanadate.


   FOOTNOTES
 
1 This work was supported by NSF Grants IBN-998808 and 235011 (R.P.F.). Back

2 A list of primers used for quantification of the target gene transcripts (Supplemental Table 1) is available with the online posting of this paper at jn.nutrition.org. Back

3 Abbreviations used: FBPase, fructose-1,6-bisphosphatase; G6P, glucose-6-phosphate; G6Pase, glucose-6-phosphatase; G6PT, glucose-6-phosphate translocase; GLUT5, intestinal fructose transporter, HF, high fructose; HG, high glucose; SGLT1, sodium-glucose cotransporter. Back

Manuscript received 12 March 2006. Initial review completed 1 May 2006. Revision accepted 28 June 2006.


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 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
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