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Endocrinology Vol. 138, No. 3 1267-1275
Copyright © 1997 by The Endocrine Society

ARTICLES

Changes in Facilitative Glucose Transporter Messenger Ribonucleic Acid Levels in the Diabetic Rat Kidney1

Edward Chin, A. Musa Zamah, Daniel Landau, Henning Gronboek, Allan Flyvbjerg, Derek LeRoith and Carolyn A. Bondy

Division of Endocrinology (E.C.), University of Alabama at Birmingham, and Veterans Administration Medical Center, Birmingham, Alabama; Developmental Endocrinology Branch (A.M.Z.), National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland; Diabetes Branch (Da.L., De.L.), National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland; Department of Nephrology (Da.L.), Children’s Hospital National Medical Center, Washington, DC; Institute of Experimental Clinical Research (H.G.), Aarhus Kommunehospital, DK-8000 Aarhus C., Denmark

Address all correspondence and requests for reprints to: C. A. Bondy, NIH-NICHHD-DEB, Building 10/Room 10N262, 10 Center Drive, MSC-1862, Bethesda, Maryland 20892-1862.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Facilitative glucose transporter (GLUTs 1, 2, 4, and 5) messenger RNAs (mRNAs) are differentially distributed in the rat nephron: GLUT1 is widely expressed, GLUT4 is selectively concentrated in thick ascending limbs, and GLUT2 and 5 are exclusively localized in proximal tubules, consistent with differential roles for these transporters in renal glucose handling. In the present study, quantitative in situ hybridization was used to evaluate changes in these mRNA levels during acute (2 and 7 days) and chronic (30, 90, and 180 days) streptozotocin-induced diabetes mellitus (STZ-DM). Medullary GLUT1 and GLUT4 mRNA levels were significantly increased during the acute phase but returned to normal after 1 week. Cortical GLUT1 mRNA levels, however, were decreased significantly from 7 days through 6 months of STZ-DM. Cortical GLUT2 mRNA was slightly increased acutely and increased 5-fold in chronic STZ-DM, with the largest increase focally concentrated in the convoluted portion of the proximal tubule. Proximal tubule GLUT5 mRNA levels also were increased significantly during chronic STZ-DM.

In summary, medullary GLUT1 and GLUT4 mRNA levels are acutely increased in STZ-DM, paralleling the increased renal epithelial metabolic activity accompanying early diabetes. Proximal tubular GLUT2 and 5 mRNA levels were increased in chronic STZ-DM, possibly adapting to the increased need for glucose transport out of these epithelial cells, whereas the concomitant decrease in cortical GLUT1 expression may reflect the decreased requirement for basolateral import of glucose into these same cells. Thus, renal GLUTs demonstrate complex, nephron segment-specific and duration-dependent responses to the effects of STZ-DM.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
THE FACILITATIVE glucose transporters (GLUTs 1–5) are a family of integral membrane proteins that allow the passive movement of glucose across cell membranes. The five known facilitative transporters have distinctive affinities for glucose, cell-specific distributions, and presumably, different roles in glucose metabolism (1–3, recent reviews). The kidney has a complex role in glucose homeostasis; it is responsible for reabsorption of filtered glucose; gluconeogenesis in hypoglycemic states and some nephron segments rely on glucose as a primary substrate for fuel metabolism (4, 5, 6). We have shown previously that GLUTs 1, 2, 4, and 5 (GLUT3 was not detected) demonstrate nephron segment-specific patterns of gene expression consistent with differential roles in renal glucose handling (7). For example, the distribution of GLUT1 parallels renal patterns of glycolysis, whereas the focal expression of GLUT4 in the thick ascending limb (TAL) reflects the intense oxidative metabolism in this segment (7). In contrast, GLUTs 2 and 5 are localized exclusively in the proximal tubule, presumably serving the basolateral efflux of reabsorbed or newly synthesized glucose from epithelial cells (7).

Diabetes has profound effects on the kidney and is the leading cause of end stage renal disease in the United States (8). Streptozotocin-induced diabetes mellitus (STZ-DM) in rats has been shown to cause renal hypertrophy (9), increase renal gluconeogenesis (10), increase renal glucose use (11), and produce tubular lesions as a result of glycogen accumulation (12). To investigate the role that alterations in facilitative GLUT expression might play in diabetic renal pathology, we used in situ hybridization to quantitate steady-state renal messenger RNA (mRNA) levels for GLUTs 1 2, 4, and 5 in both acute and chronic phases of STZ-DM.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Study design
Adult female Wistar rats (200 g, Mollegaards Breeding Center, Eiby, Denmark) were housed three per cage with 12:12 h (0600 to 1800 h) artificial light cycle, temperature 21 ± 1 C, and humidity 55 ± 2%. Animals were randomized into control, diabetic, and diabetic/insulin-treated groups. Controls received saline, and the other two groups received streptozotocin (STZ, 55 mg/kg BW, iv, Upjohn, Kalamazoo, MI). Insulin-treated rats received Ultralente Insulin (Novo-Nordisk, Bagsvaerd, Denmark) in an initial dose of 4–6 U, given for the first time approximately 24 h after the STZ treatment, i.e. on the morning of day 1, followed by 1–3 U daily thereafter, depending on blood glucose values. Tail blood glucose was measured by Haemoglucotest 1–44 and Reflolux II reflectance meter (Boehringer Mannheim, Mannheim, Germany). Urine was tested for glucose and ketone bodies by Neostix-4 (Ames, Stoke Poges, Slough, UK). Animals with blood glucose levels above 18 mmol/liter, urine glucose concentration more than 111 mmol/liter, and without ketonuria were included in the study. Animals had free access to standard rat chow (Altromin, no. 1324, Lage, Germany) and water. Animals were placed in metabolic cages 30, 60, and 90 days after injection for 24-h urine collections. Urine was stored at -80 C until assayed. Urinary albumin concentration was determined by RIA, as previously described (13).

Control rats from the day of injection (day zero, n = 6), diabetes/insulin-treated rats (2 and 7 days, n = 8, respectively), and rats 2, 7, 30, 90, and 180 days after the injections of STZ or placebo (6–8 in each arm of the age groups) were anesthetized with sodium barbital (50 mg/kg ip) and killed. The left kidney was rapidly removed, frozen over dry ice, stored at -70 C, and used for in situ hybridization. Frozen sections were cut at a thickness of 10 µ, thaw-mounted on poly-L-lysine coated slides, and stored at -70 C until use. Sections were cut longitudinally so that all sections included the papilla, outer medulla, and cortex. All sections for a given experiment were prepared, washed, exposed, and analyzed together.

Preparation of complementary RNA (cRNA) probes
The cDNAs used for probe synthesis in the present study were the rat GLUT1 (14), GLUT2 (15), GLUT4 (16), and GLUT5 (7), which have been previously described. High specific activity 35S-labeled cRNA probes were synthesized in 10-µl reactions containing 100 µCi cytidine 5' ({alpha} 35S) thiotriphosphate (Amersham SJ 40832), 100 µCi uridine 5' ({alpha} 35S) thiotriphosphate (Amersham SJ 40383), 10 mM NaCl, 6 mM MgCl2, 40 mM Tris (pH 7.5), 2 mM spermidine, 10 mM dithiothreitol, 500 µM each of unlabeled ATP and GTP, 25 µM each of unlabeled UTP and CTP, 0.5 µg linearized template, 15 U of the appropriate polymerase, and 15 U RNAsin (Promega, Madison, WI). The reaction was incubated at 42 C for 30 min; then an additional 15 U of the appropriate polymerase and 15 U RNAsin were added. The reaction was incubated at 42 C for 30 min, after which the DNA template was removed by digestion with DNase-I at 37 C for 10 min. Labeled cRNA was column purified (Bio-Spin 6, Bio-Rad) to separate unincorporated nucleotides. Labeled probe was precipitated with 5 µl transfer RNA (tRNA,) 10 µl 5 M NaCl, 10 µl DEPC H20 and 300 µl cold EtOH. Purified probe then underwent alkaline hydrolysis to produce fragments of an average length of 150 bases.

In situ hybridization
Tissue sections were prepared in the following manner. Before hybridization, sections were warmed to 25 C, fixed in 10% formaldehyde, and soaked for 10 min in 0.25% acetic anhydride/0.1 M triethanolamine hydrochloride/0.9% NaCl. Tissue was then dehydrated through an ethanol series, delipidated in chloroform, rehydrated, and air-dried. The 35S-labeled probes (107 dpm/ml or approximately 50 ng/ml) were added to hybridization buffer composed of 50% formamide, 0.3 M NaCl, 20 mM Tris HCl, pH 8, 5 mM EDTA, 500 µg tRNA, 10% dextran sulfate, 10 mM dithiothreitol, and 0.02% each of BSA, ficoll, and polyvinylpyrolidone. After the 35S-labeled probe in hybridization buffer was added to the sections, coverslips were placed over the sections, and the slides were incubated in humidified chambers overnight (14 h) at 55 C.

Slides were washed several times in 4 x SSC to remove cover slips and hybridization buffer, dehydrated and immersed in 0.3 M NaCl, 50% formamide, 20 mM Tris HCl, 1 mM EDTA at 60 C for 10 min. Sections were then treated with RNAse A (20 µg/ml) for 30 min at room temperature, followed by a 15-min wash in 0.1 x SSC at 55 C. Slides were air-dried and apposed to Hyperfilm-ßMax (Amersham) along with autoradiographic standards (ARC Inc., St. Louis, MO) for 1–4 days and then dipped in Kodak NTB2 nuclear emulsion, stored with desiccant at 4 C for 3–30 days, developed, and counterstained with Mayer’s hematoxylin and eosin for microscopic evaluation.

Quantitative densitometry
Quantification of mRNA level was done as follows. Film autoradiographic images, registered with a solid-state video camera (Sony XC-77, Sony Corp.) and a 55-mm MicroNikkor lens over a light box of variable intensity (Northern Light Precision 890, Imaging Research Inc., Toronto, Canada), were digitized to a 640 x 480 matrix with light transmittance coded in 256 equal grey levels (LG-3 frame grabber card, Scion Corp., Frederick, MD). The images were quantified using a Macintosh II-based image analysis program (Image 1.49, developed by W.S. Rasband, Research Services Branch, NIMH, Bethesda, MD), in which the gray scale values from the autoradiographic standard curve were used in a third-degree polynomial function curve fit. Anatomically matched, standardized areas of the tubulo-interstitial cortex, outer stripe of the outer medulla (OS/OM) and inner stripe of the outer medulla (IS/OM), inner medulla, and papilla were defined by cursor control and their transmittance measured. Two to three measurements were taken for each structure in each kidney, and an average value was determined. Data was normalized as a percentage of the age-matched control and expressed as mean ± SEM for each group. One-way ANOVA, followed by unpaired two tailed t tests, was used to evaluate significant differences between diabetic and control groups at the different time points.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Clinical data
Controls had a steady increase in body weight over the study period. STZ-DM rats weighed significantly less than the controls (201.8 ± 1.7 vs. 192.1 ± 2.6 g) as early as day 2. In contrast, STZ-DM kidneys are significantly heavier, and the renal fractional weight (KW/BW%) is significantly increased, compared with their respective control groups at all time points. Treatment with insulin normalizes the weight loss and the changes in renal weights associated with STZ-DM (Table 1Go). Blood glucose levels in the STZ-DM groups are increased 5-fold compared with controls and are normalized by insulin treatment (Table 1Go). All diabetic animals developed glycosuria but not ketonuria (data not shown). Urinary albumin excretion was markedly elevated in the STZ-DM rats by 30 days, compared with controls (864 ± 149 vs. 133 ± 34 µg/24 h; P < 0.001), and continued to increase through 180 days (1560 ± 202 vs. 418 ± 115; P < 0.001).


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  		Table 1. Body weight, kidney weight, kidney weight as a percentage of body weight (KW/BW%), blood glucose, and urinary albumin excretion in control, streptozotocin-induced diabetic, and insulin-treated rats

 
GLUT1 and GLUT4
GLUT1 mRNA is most abundant in the renal papilla and in the IS/OM (Fig. 1AGo). GLUT1 mRNA is also detected in the OS/OM, in medullary rays and in the cortex where we have previously shown that it is localized in epithelial cells of the proximal straight tubule, Henle’s loop, distal tubule and collecting duct (7). In the acute phase of STZ-DM, GLUT1 mRNA levels are significantly increased throughout the medulla, particularly in the papilla and the IS/OM; the increases in medullary GLUT1 mRNA levels are progressively normalized by 30 days and remain normal through the chronic period (Fig. 1Go). Cortical GLUT1 mRNA levels, however, begin to decrease by day 7 and remain significantly decreased up to 6 months after the onset of STZ-DM.



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  		Figure 1. GLUT1 mRNA in control (Con) and diabetic (DM) rat kidneys, as shown by in situ hybridization. A and B, Representative film autoradiographs of longitudinal sections from day zero control and day 2 diabetic kidneys hybridized to a GLUT1 cRNA probe. The boxes illustrate regions chosen for quantitative analysis. C, Effects of STZ-DM (DM) on renal GLUT1 mRNA levels determined by quantitative densitometry, as described in Materials and Methods. Levels are expressed as per cent of control, and each experimental group had its own control group. Results are given as means ± SEM; n = 6–8 animals for each group for this and each of the following figures. Pa, Papilla; IM, inner medulla; Cx, cortex. *, P < 0.05; **, P < 0.01. Bar = 0.93 mm.

 
GLUT4 mRNA is most abundant in the IS/OM (Fig. 2AGo), where it is localized in epithelial cells of the TAL, as previously described (7). Medullary GLUT4 mRNA levels are increased by almost 90% in the acute phase of STZ-DM but are similar to control levels by day 30 and thereafter (Fig. 2CGo). Insulin treatment had little effect on GLUT4 mRNA levels at the 2-day time point, but with 7 days of insulin treatment (Fig. 2CGo), GLUT4 mRNA levels were normalized in diabetic kidneys.



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  		Figure 2. GLUT4 mRNA in control, diabetic, and diabetic/insulin-treated kidneys. A and B, Representative film autoradiographs of longitudinal sections from day zero control and day 2 diabetic kidneys hybridized to a GLUT4 cRNA probe; C, quantitation of STZ-DM (DM)-associated changes in renal GLUT4 mRNA levels. See legend to Fig. 1Go for details. *, P < 0.05; **, P < 0.01. Bar = 0.93 mm.

 
GLUT2 and GLUT5
GLUT2 mRNA is somewhat more abundant in proximal straight tubules (PSTs, localized in the OS/OM), compared with proximal convoluted tubules (PCTs, localized in the cortex) in control kidneys (Fig. 3AGo), and increase by 30–50% in PSTs and by 400% in PCTs 1 month after induction of STZ-DM (Fig. 3Go, B–D).



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  		Figure 3. GLUT2 mRNA in control and diabetic kidneys. A and B, Representative film autoradiographs of longitudinal sections from 30-day control and diabetic kidneys hybridized to a GLUT2 cRNA probe. The boxes in Fig. 1BGo illustrate a region of the cortex and OS/OM chosen for quantitative analysis. C, Quantitation of STZ-DM (DM) associated changes in renal GLUT2 mRNA levels; D and E, paired bright and dark field photomicrographs showing the cellular localization of GLUT2 mRNA in the renal cortex. In the emulsion-coated sections, the mRNA hybridization signal appears as black grains in the bright field and as white grains in the dark field illumination. GLUT2 mRNA is abundant in the tubular epithelial cells of the PCT. GLUT2 mRNA is less abundant in the PST and not detected in the glomerulus (G). See legend to Fig. 1Go for details. Cx, Cortex. Bar = 0.93 mm (A and B) and 100 µm (D and E). *, P < 0.05; **, P < 0.01.

 
GLUT5 mRNA is concentrated in medullary rays and OS/OM (Fig. 4AGo), where it is localized in PSTs (7). STZ-DM had no detectable effect on this mRNA in the acute phase, but PST GLUT5 mRNA levels were increased significantly at 30 and 90 days after onset of diabetes, returning to normal thereafter.



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  		Figure 4. GLUT5 mRNA in control and diabetic kidneys. A and B, Representative film autoradiographs of longitudinal sections from control and 30-day diabetic kidneys hybridized to a GLUT5 cRNA probe. A region of the OS/OM chosen for quantitative analysis is illustrated by the box in Fig. 1BGo. C, Quantitation of STZ-DM (DM) associated changes in renal GLUT5 mRNA levels. See legend to Fig. 1Go for details. Arrowheads, medullary ray. Bar = 0.93 mm. *, P < 0.05; **, P < 0.01.

 
Effects of insulin treatment
Two days of insulin treatment had little effect on elevated medullary GLUT1 and 4 mRNA levels and was associated with an increase in cortical GLUT1 and 2 mRNA levels (Table 2Go). Interestingly, insulin demonstrates segment specific effects on GLUT2 mRNA, producing a normalization of elevated levels in the PST of the OS/OM, but a further increase GLUT2 mRNA in cortical PCTs. After 7 days of insulin, however, all GLUT mRNA levels were normalized, except for a persistent slight elevation in GLUT1 in the IS/OM. No groups were treated with insulin for longer than 7 days; and, thus, there is no data on effects of insulin on the cortical changes in GLUT1, 2, and 5 mRNAs seen in chronic diabetes.


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  		Table 2. Effects of insulin treatment on renal glucose transporter (GT) 1, 4 and 2 mRNA expression, 2 and 7 days after STZ-DM

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
The present study shows that STZ-DM is associated with complex, nephron segment-specific alterations in facilitative transporter mRNA levels, which differ in acute and chronic phases of the disease. In the acute phase, from 2–7 days after STZ treatment, medullary GLUT1 and 4 mRNA levels are increased significantly. Whereas expression of these transporters normalizes in the medulla after 1 week, cortical GLUT1 levels decrease and GLUT2 and GLUT5 mRNA levels increase in the chronic phase of diabetes. These changes cannot be ascribed to morphological changes associated with diabetes, i.e. renal hypertrophy, because the hybrid signal was measured in a standard-sized area for each region, and because all sections are of equal thickness (10 µ), hypertrophy of the kidney, as a whole, or of specific regions is not expected to influence mean signal intensity/area in the different groups.

STZ-DM has been shown to increase glomerular filtration rate, tubular sodium transport, and Na+/K+ ATPase activity (17, 18). The distribution of renal GLUT1 expression is correlated with renal tubular glycolysis in support of active transport mechanisms (5). Thus, the increase in GLUT1 mRNA levels throughout the medulla in acute STZ-DM may be related to the increased metabolic demands of early diabetes. The present, data showing decreases in cortical GLUT1 mRNA levels during chronic diabetes, confirm the results of a recent study that evaluated GLUT1 protein and mRNA levels in proximal tubules fractionated from the renal cortex in STZ-DM (19). The reduction in GLUT1 expression in proximal tubules during prolonged diabetes may be caused by the saturation of these tubular cells with glucose reabsorbed from the filtrate; and thus, expression of the high affinity GLUT1 (which normally facilitates the passage of glucose from the circulation across the basolateral membranes into these cells) would be gratuitous. Interestingly, cortical GLUT1 mRNA levels were stable early in diabetes when medullary levels were increased and were decreased later in diabetes, by which time medullary levels had normalized, suggesting that local tubular factors are more important in regulation of renal GLUT1 expression than are systemic factors.

The present study demonstrates that proximal tubule GLUT2 mRNA levels increase dramatically in chronic diabetes, with maximal elevation of almost 5 times normal from 1–3 months and continued elevation after 6 months of diabetes. A previous study also reported increased renal GLUT2 mRNA and protein in chronic STZ-DM (19). Interestingly, the use of in situ hybridization in our study reveals that the balance of proximal tubule GLUT2 gene expression shifts from the straight portion in normal rats to the convoluted segment in the diabetic rats. The adaptive significance of this increase in proximal tubule GLUT2 expression seems clear; these cells actively reabsorb a large load of filtered glucose in diabetes via a Na+-GLUT. The glucose that accumulates in these epithelial cells must be transferred back to the circulation across the basolateral membranes largely via the low-affinity, high-capacity GLUT2 transporter, which is localized on these membranes (20, 21). Hence, the increase in proximal tubule GLUT2 may serve to accommodate the increased glucose transport load. The significance of the selective increase in PCT GLUT2 mRNA in STZ-DM is not clear, but it is noteworthy that insulin treatment also produced striking segment-specific effects, with a marked increase in PCT and decrease or normalization in PST GLUT2 mRNA levels. The present data suggests that regulation of renal GLUT2 is similar to the small intestine, where GLUT2 mRNA and protein both are increased after 45 days of STZ-DM (22). This is in contrast to the pancreatic ß cell, however, where GLUT2 is decreased in experimental and spontaneous diabetes and increased during hyperglycemic glucose infusion (23).

This is the first study to describe GLUT5 mRNA levels in the diabetic kidney. GLUT5 is exclusively localized in PSTs (7) and may serve to transport fructose and glucose (24). Our study shows that renal GLUT5 mRNA levels are increased in chronic diabetes in correlation with the sharp increase in GLUT2 mRNA levels. This observation agrees with a recent study of increased GLUT5 mRNA and protein in the small intestine of 45-day diabetic rats (22). Renal fructose metabolism has not been well characterized, but the enzymes necessary for fructose metabolism are localized in proximal tubules (4, 10) and are linked to renal gluconeogenesis. Gluconeogenesis is elevated in ketotic animals (4, 10), and it is possible that GLUT5 serves to transport fructose into proximal tubule cells for entry into gluconeogenic pathways. On the other hand, fructose is a product of the sorbitol pathway, which is increased in the chronic diabetic rat kidney (10, 25), and thus, it is possible that GLUT5 may serve to transport fructose out of the cell.

This also is the first study to evaluate renal tubular GLUT4 mRNA levels in diabetes. A recent study (26) used RT-PCR to quantitate GLUT4 mRNA levels in dissected glomeruli and microvessels from rat kidneys after 1 week of STZ-DM and reported a decrease of approximately 70%. GLUT4 immunoreactivity and glucose transport also were decreased in this microdissected tissue. GLUT1 was not investigated in this previous study, though the present data, showing significantly decreased GLUT1 mRNA levels in the renal cortex, suggests that decreases in GLUT1 may contribute to the effects seen in the Marcus et al. study. Furthermore, muscle and adipocyte GLUT4 levels generally are decreased in STZ-DM and increased with insulin treatment (reviewed in Ref.23); thus, our finding that GLUT4 mRNA levels are transiently increased in early diabetes, demonstrates a renal tubular specific pattern of regulation of GLUT4 expression in STZ-DM. We have shown previously that GLUT4 mRNA and protein are focally localized in TALs in the IS/OM (7). The TALs have the highest level of Na+/K+ ATPase activity of any segment of the nephron and preferentially use glucose to fuel their pump activity (4, 27). Renal sodium reabsorption is increased in diabetes (18), and the timing of GLUT4 mRNA elevation in the diabetic TAL correlates with increases in Na+/K+ ATPase activity observed in TALs after 2 days of STZ-DM (28). Thus, it seems that TAL-specific increases in GLUT4 expression may serve to increase glucose uptake to fuel the heightened Na+/K+ ATPase in this segment. We have shown previously that VP positively regulates renal GLUT4 mRNA levels (7), and VP is increased in uncontrolled diabetes (29); hence, VP may stimulate the increases in TAL GLUT4 mRNA levels in early STZ-DM.

The observation that GLUT 1 and 4 gene expression did not normalize at the 2-day point in the insulin-treated group is probably caused by the fact that the animals had only received a single injection of insulin at this point and had not yet metabolically stabilized. In support of this view, the fact that the early changes in GLUTs 1 and 4 are totally normalized with insulin treatment by day 7 is a strong argument against a general toxic effect of STZ. Furthermore, if the changes in gene expression were caused by a toxic effect of STZ, it would be difficult to explain why only GLUTs 1 and 4 were affected immediately after exposure, whereas GLUTs 2 and 5 were affected weeks later.

In summary, this study has shown that STZ-DM is accompanied by complex, nephron segment-specific, temporally dependent changes in steady-state mRNA levels for each of the four facilitative GLUTs found in the rat kidney. The high-affinity transporters, GLUTs 1 and 4, demonstrate parallel increases in mRNA levels in the metabolically highly active renal medulla during the first week of diabetes, presumably facilitating basolateral epithelial glucose import for use as substrate to fuel increased active transport functions. The low-affinity transporters, GLUTs 2 and 5, demonstrate parallel increases in steady-state mRNA levels in proximal tubules during the chronic phase of diabetes, presumably to support the increased requirements for basolateral epithelial glucose export. At the same time, in the same proximal tubules, GLUT1 mRNA levels are reduced, illustrating the complexity of facilitative GLUT regulation.


   Acknowledgments
 
We thank Ricardo Dreyfuss for expert photography and Drs. Morris Birnbaum, Graeme Bell, and Harvey Lodish for providing the cDNAs used for riboprobe synthesis. We are grateful to Kirsten Nyborg and Karen Mathiesen for excellent technical assistance.


   Footnotes
 
1 This work was supported by the Clarence Rice Fellowship of the Washington, D.C. Affiliate of the American Diabetes Association (D.L.) and by grants from the Danish Diabetes Association, the Danish Medical Research Council, the Ruth König Petersen Foundation, and the Aage Louis-Hansen Memorial Foundation. Back

Received September 25, 1996.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Mueckler M 1990 Family of glucose-transporter genes implications for glucose homeostasis and diabetes. Diabetes 39:6–11[Abstract]
  2. Gould G, Bell G 1990 Facilitative glucose transporters: an expanding family. TIBS 15:18–23
  3. Thorens B, Charron M, Lodish H 1990 Molecular physiology of glucose transporters. Diabetes Care 13:219–227[Abstract]
  4. Silva P 1990 Energy and fuel substrate metabolism in the kidney. Semin Nephrol 10:432–444[Medline]
  5. Ross B, Espinal J, Silva P 1986 Glucose metabolism in renal function. Kidney Int 29:54–67[Medline]
  6. Silverman M, Turner R 1992 Glucose transport in the renal proximal tubule. In: Windhager E (ed) Handbook of Physiology, Section 8: Renal Physiology. Oxford University Press, New York, vol 2:2017–2038
  7. Chin E, Zhou J, Bondy CA 1993 Anatomical and developmental patterns of facilitative glucose transporter gene expression in the rat kidney. J Clin Invest 91:1810–1815
  8. Nathan DM 1993 Long-term complications of diabetes mellitus. N Engl J Med 328:1676–1685[Free Full Text]
  9. Seyer-Hansen K 1983 Renal hypertrophy in experimental diabetes. Kidney Int 23:643–646[Medline]
  10. Guder WG, Schmolke M, Wirthensohn G 1992 Carbohydrate and lipid metabolism of the renal tubule in diabetes mellitus. Eur J Clin Chemi Clin Biochem 30:669–674
  11. Steer KA, Sochaor M, McLean P 1985 Renal hypertrophy in experimental diabetes changes in pentose phosphate pathway activity. Diabetes 34:485–490[Abstract]
  12. Holck P, Rasch R 1993 Structure and localization of glycogen in the diabetic rat kidney. Diabetes 42:891–900[Abstract]
  13. Christensen CK, Orskov H 1984 Rapid screening PEG radioimmunoassay for quantitation of pathological microalbuminuria. Diabetic Nephrol 3:92–94
  14. Werner H, Adamo M, Lowe Jr WL, Roberts Jr CT, LeRoith D 1989 Developmental regulation of rat brain/Hep G2 glucose transporter gene expression. Mol Endocrinol 3:273–279[Abstract]
  15. Thorens B, Sarkar H, Kaback H, Lodish H 1988 Cloning and functional expression in bacteria of a novel glucose transporter present in liver, intestine, kidney, and ß-pancreatic islet cells. Cell 55:281–290[CrossRef][Medline]
  16. Birnbaum MJ 1989 Identification of a novel gene encoding an insulin-responsive glucose transporter protein. Cell 57:305–315[CrossRef][Medline]
  17. Körner A, Eklöf A, Celsi G, Aperia A 1994 Increased renal metabolism in diabetes mechanism and functional implications. Diabetes 43:629–633[Abstract]
  18. Wald H, Popovtzer MM 1984 The effect of streptozotocin-induced diabetes mellitus on urinary excretion of sodium and renal Na+-K+-ATPase activity. Pflugers Arch. 401:97–100
  19. Dominguez JH, Camp K, Maianu L, Feister H, Garvey WT 1994 Molecular adaptations of GLUT1 and GLUT2 in renal proximal tubules of diabetic rats. American Journal of Physiology 266 (Renal Fluid Electrolyte Physiol. 35):F283–F290
  20. Thorens B, Lodish H, Brown D 1990 Differential localization of two glucose transporter isoforms in the rat kidney. Am J Physiol 259:C286–C294
  21. Cramer S, Pardridge W, Hirayama B 1992 Colocalization of GLUT2 glucose transporter, sodium/glucose cotransporter and {gamma}-glutamyl transpeptidase in rat kidney with double-peroxidase immunocytochemistry. Diabetes 41:766–770[Abstract]
  22. Burant CF, Flink S, DePaoli AM, Chen J, Lee W-S, Hediger MA, Buse JB, Chang EB 1994 Small intestine hexose transport in experimental diabetes. J Clin Invest 93:578–585
  23. Kahn BB 1992 Facilitative glucose transporters: regulatory mechanisms and dysregulation in diabetes. J Clin Invest 89:1367–1374
  24. Rand EB, Depaoli AM, Davidson NO, Bell GI, Burant CF 1993 Sequence, tissue distribution and functional characterization of the rat fructose transporter GLUT5. Amer J Physiol 264:G1169–G1176
  25. Beyers-Mears A, Ku L, Cohen MP 1980 Glomerular polyol accumulation in diabetes and its prevention by oral sorbi nil. Diabetes 33:604–607[Abstract]
  26. Marcus RG, England R, Nguyen K, Charron MJ, Briggs JP, Brosius III FC 1994 Altered renal expression of the insulin-responsive glucose transporter GLUT4 in experimental diabetes mellitus. Am J Physiol 267:F816–F824
  27. Knepper M, Rector Jr F 1991 Urinary concentration and dilution. In: Brenner BM, Rector Jr FC (ed) The Kidney. W. B. Saunders Company, Philadelphia, vol 2:445–482
  28. Khadouri C, Barlet-Bas C, Doucet A 1987 Mechanism of increased tubular Na+-K+-ATPase during streptozotocin-induced diabetes. Pflugers Arch 409:296–301[CrossRef][Medline]
  29. Vaamonde CA, Pérez GO 1990 Tubular function in diabetes mellitus. Semin Nephrol 10:203–218[Medline]



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Na+-Glucose Transporter-2 Messenger Ribonucleic Acid Expression in Kidney of Diabetic Rats Correlates with Glycemic Levels: Involvement of Hepatocyte Nuclear Factor-1{alpha} Expression and Activity
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 Am. J. Physiol. Renal Physiol.Home page
C. Schmidt, K. Hocherl, and M. Bucher
Regulation of renal glucose transporters during severe inflammation
Am J Physiol Renal Physiol, February 1, 2007; 292(2): F804 - F811.
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 Mol. Endocrinol.Home page
C. Keembiyehetty, R. Augustin, M. O. Carayannopoulos, S. Steer, A. Manolescu, C. I. Cheeseman, and K. H. Moley
Mouse Glucose Transporter 9 Splice Variants Are Expressed in Adult Liver and Kidney and Are Up-Regulated in Diabetes
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 DiabetesHome page
H. Rahmoune, P. W. Thompson, J. M. Ward, C. D. Smith, G. Hong, and J. Brown
Glucose Transporters in Human Renal Proximal Tubular Cells Isolated From the Urine of Patients With Non-Insulin-Dependent Diabetes
Diabetes, December 1, 2005; 54(12): 3427 - 3434.
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 J. Pharmacol. Exp. Ther.Home page
B. Grover, D. Buckley, A. R. Buckley, and W. Cacini
Reduced Expression of Organic Cation Transporters rOCT1 and rOCT2 in Experimental Diabetes
J. Pharmacol. Exp. Ther., March 1, 2004; 308(3): 949 - 956.
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 IOVSHome page
B. R. Merriman-Smith, A. Krushinsky, J. Kistler, and P. J. Donaldson
Expression Patterns for Glucose Transporters GLUT1 and GLUT3 in the Normal Rat Lens and in Models of Diabetic Cataract
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 DiabetesHome page
P. Odetti, C. Pesce, N. Traverso, S. Menini, E. P. Maineri, L. Cosso, S. Valentini, S. Patriarca, D. Cottalasso, U. M. Marinari, et al.
Comparative Trial of N-Acetyl-Cysteine, Taurine, and Oxerutin on Skin and Kidney Damage in Long-Term Experimental Diabetes
Diabetes, February 1, 2003; 52(2): 499 - 505.
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 Nephrol Dial TransplantHome page
A. Mogyorosi and F. N. Ziyadeh
GLUT1 and TGF-{beta}: the link between hyperglycaemia and diabetic nephropathy
Nephrol. Dial. Transplant., December 1, 1999; 14(12): 2827 - 2829.
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