Uptake and glutathione conjugation of ethacrynic acid and efflux of the glutathione adduct by periportal and perivenous rat hepatocytes

We assessed the impact of zonal factors on the hepatic reduced glutathione (GSH) conjugation of ethacrynic acid (EA). Uptake of EA by enriched periportal (PP) and perivenous (PV) rat hepatocytes was characterized by both saturable (V(max)(uptake) = 3.4 +/- 1.7 and 3. 2 +/- 0.8 nmol/min/mg protein and K(m)(uptake) = 51 +/- 13 and 44 +/- 15 microM) and nonsaturable (12 +/- 5 and 12 +/- 3 microl/min/mg protein) components. Values for the overall GSH conjugation rates of EA (200 microM) were similar among the zonal hepatocytes and resembled those for the influx transport rates. In the absence of the hepatocyte membrane, GSH conjugation in PV and PP hepatocyte cytosol was similar, but a higher perivenous GSH conjugation activity toward EA (PV/PP of 2.4) that mirrored the higher PV/PP ratios of immunodetectable GSTs Ya (1.7) and Yb2 (2.5) was found in cell lysates obtained by the dual-digitonin-pulse perfusion technique. The GSH conjugation rates in the subcellular fragments were, however, much greater than those observed for intact hepatocytes. Efflux rates of the glutathione conjugate EA-SG from zonal hepatocytes were similar, as were levels of the immunodetectable multidrug-resistance protein 2/canalicular multispecific organic anion transporter (Mrp2/cMoat) in the 100,000g pellets. The composite results suggest that the GSTs responsible for EA metabolism are more abundant in the PV region, albeit that the gradient of enzymatic activities is shallow. Despite the existence of zonal metabolic activity, the overall GSH conjugation rate of EA is homogeneous among cells because the reaction is rate limited by uptake, which occurs evenly. Results on EA-SG efflux suggest the acinar homogeneity in Mrp2/cMoat function for canalicular transport.     

Effect of chronic ethanol feeding on rat hepatocytic glutathione. Compartmentation, efflux, and response to incubation with ethanol.

Hepatocytes from rats that were fed ethanol chronically for 6-8 wk were found to have a modest decrease in cytosolic GSH (24%) and a marked decrease in mitochondrial GSH (65%) as compared with pair-fed controls. Incubation of hepatocytes from ethanol-fed rats for 4 h in modified Fisher's medium revealed a greater absolute and fractional GSH efflux rate than controls with maintenance of constant cellular GSH, indicating increased net GSH synthesis. Inhibition of gamma-glutamyltransferase had no effect on these results, which indicates that no degradation of GSH had occurred during these studies. Enhanced fractional efflux was also noted in the perfused livers from ethanol-fed rats. Incubation of hepatocytes in medium containing up to 50 mM ethanol had no effect on cellular GSH, accumulation of GSH in the medium, or cell viability. Thus, chronic ethanol feeding causes a modest fall in cytosolic and a marked fall in mitochondrial GSH. Fractional GSH efflux and therefore synthesis are increased under basal conditions by chronic ethanol feeding, whereas the cellular concentration of GSH drops to a lower steady state level. Incubation of hepatocytes with ethanol indicates that it has no direct, acute effect on hepatic GSH homeostasis.     

Effect of phenobarbital on the distribution of drug metabolizing enzymes between periportal and perivenous rat hepatocytes prepared by digitonin-collagenase liver perfusion

Intact periportal (pp) or perivenous (pv) hepatocytes were prepared by digitonin-collagenase liver perfusion. The degree of separation was indicated by significant differences between the pp and pv cells in their activity of the pp markers, alanine aminotransferase (pp/pv = 2.1), gamma-glutamyltranspeptidase (3.4) and lactate dehydrogenase (1.3), and of the pv markers, glutamate dehydrogenase (0.73) and pyruvate kinase (0.81). This pattern was not altered by a 3-day pretreatment with phenobarbital (PB). The hepatocytes isolated from the pv area contained higher activities of microsomal NADPH-cytochrome c reductase, 7-ethoxycoumarin O-deethylase, 7-ethoxyresorufin O-deethylase and benzo(a)pyrene hydroxylase, and of cytosolic glutathione transferase. Cytochrome P-450 and UDP-glucuronosyltransferase were slightly higher in pv cells. Treatment with PB induced NADPH-cytochrome c reductase, glutathione transferase, cytochrome P-450 and UDP-glucuronosyltransferase but the degree of induction was found to be at least as strong in pp cells as in pv cells. The induction of 7-ethoxyresorufin O-deethylase and 7-ethoxycoumarin O-deethylase was clearly more prominent in pp cells. On the other hand, PB reduced the activities of benzo(a)pyrene hydroxylase and alcohol dehydrogenase in both cell types. These results demonstrate by direct enzyme assay of separated cells the dominance of the pv-region for metabolizing drugs in the normal liver. Contrary to several other studies, however, our data indicate that induction by PB occurs panacinarily, i.e., relatively more in the pp region, thus diminishing rather than exaggerating the original pv dominance.     

Effects of chronic ethanol feeding on rat hepatocytic glutathione. Relationship of cytosolic glutathione to efflux and mitochondrial sequestration.

Chronic ethanol feeding to rats increases the sinusoidal component of hepatic glutathione (GSH) efflux, despite a lower steady-state GSH pool size. In the present studies, no increase of biliary GSH efflux in vivo was found in chronic ethanol-fed cells. Studies were performed on ethanol-fed and pair-fed cells to identify the kinetic parameters of cellular GSH concentration-dependent efflux. The relationship between cytosolic GSH and the rate of efflux was modeled by the Hill equation, revealing a similar Vmax, 0.22 +/- 0.013 vs. 0.20 +/- 0.014 nmol/min per 10(6) cells for ethanol-fed and pair-fed cells, respectively, whereas the Km was significantly decreased (25.3 +/- 2.3 vs. 33.5 +/- 1.4 nmol/10(6) cells) in ethanol-fed cells. The difference in Km was larger when the data were corrected for the increased water content in ethanol-fed cells. We found a direct correlation between mitochondria and cytosolic GSH, revealing that mitochondria from ethanol-fed cells have less GSH at all cytosolic GSH values. The rate of resynthesis in depleted ethanol-fed cells in the presence of methionine and serine was similar to control cells and gamma-glutamylcysteine synthetase remained unaffected by chronic ethanol. However, the reaccumulation of mitochondrial GSH as the cytosolic pool increased was impaired in the ethanol cells. The earliest time change in GSH regulation was a 50% decrease in the mitochondrial GSH at 2 wk.     

Efcts of chronic ethanol feeding on serum lipoprotein metabolism in the rat

In rats, chronic ethanol feeding was found to enhance the postprandial hyperlipemia and to increase the incorporation of dietary palmitic acid-³H and intravenously injected L-lysine-¹⁴C into serum lipoproteins. The main increases of total amount, labeling, and specific activity of lipid and protein occurred in the d < 1.019 lipoprotein fraction. Fat absorption and the clearance of injected chylomicrons were not affected by ethanol feeding. Blocking of lipoprotein and chylomicron removal with Triton did not prevent the action of ethanol on serum lipids, indicating that the ethanol effect is not likely due to defective removal of lipids from the circulation. Ethanol enhanced the incorporation of chylomicron fatty acids into newly synthetized very low density lipoproteins, as shown by an increased reappearance of the fatty acid label into the lipids of this fraction after injection of palmitate-¹⁴C/glycerol-³H doubly labeled chylomicrons. These results indicate that alcoholic hyperlipemia is due, at least in part, to an increase in newly synthetized lipoproteins. The hyperlipemia produced by ethanol was accompanied by hepatic steatosis. The simultaneous production of fatty liver and hyperlipemia makes it unlikely that defective lipoprotein synthesis or secretion is a primary mechanism for the pathogenesis of the alcoholic fatty liver.     

Decrease in microviscosity and cholesterol content of rat liver plasma membranes after chronic ethanol feeding.

This investigation was performed to determine whether chronic ethanol feeding alters the lipid composition or the fluidity of liver plasma membranes. Male Sprague-Dawley rats were pair-fed nutritionally adequate liquid diets containing ethanol as 36% of energy or an isocaloric amount of carbohydrate for 4-5 wk. Contrasting with other membranes, chronic ethanol feeding resulted in an increase in hepatic plasma membrane fluidity as assessed by fluorescence anisotropy. This alteration was associated with a decrease in plasma membrane cholesterol content.     

Effect of chronic ethanol ingestion on alpha-mannosidase isoenzymes in rat liver

Identification of biochemical changes induced by ethanol ingestion would aid in the diagnosis and management of many alcohol-related problems in man. In this paper we identify a pH 5.5 alpha-mannosidase activity in the rat which is affected by chronic ethanol consumption. Chronic (16 wk) ingestion of alcohol (36% of calories) causes the activity of this alpha-mannosidase (thought to be the cytosolic alpha-mannosidase) in liver to decrease by 50%. We hypothesize that this deficiency of (pH 5.5) alpha-mannosidase activity may account for the reduced rate of secretion of glycoproteins by livers of alcohol-fed rats reported by other investigators (Volentine et al, Hepatology 1987;7:490-495).     

Thiol-disulfide effects on hepatic glutathione transport. Studies in cultured rat hepatocytes and perfused livers.

In cultured rat hepatocytes, cystine led to an inhibition of GSH efflux by lowering the Vmax by approximately 35% without affecting the Km. The cystine-mediated inhibition of GSH efflux was rapid in onset (< 1 h), with near maximum effect at 0.1 mM. Inhibition was still observed when cystine uptake was prevented. Cystine and sulfobromophthalein-GSH, a selective inhibitor of sinusoidal transport of GSH, did not exhibit additive inhibitory effects on GSH efflux. Depletion of ATP or membrane depolarization after cystine treatment were excluded as potential mechanisms. DTT not only reversed the cystine-mediated inhibition of GSH efflux, it stimulated GSH efflux up to 400-500%. The DTT effect was immediate in onset, reaching maximum after 30 min, and was partially reversed by cystine, suggesting that the two share a common site(s) of action. DTT treatment did not alter cellular ATP levels or change the membrane potential. In cultured hepatocytes, DTT treatment increased the Vmax of GSH efflux by approximately 500% without affecting the Km. Inhibition of microtubular function and vesicular acidification did not affect basal or DTT stimulated efflux. Both cystine and DTT effects on sinusoidal GSH efflux were confirmed in perfused livers. In summary, the capacity of the sinusoidal GSH transporter is markedly influenced by thiol-disulfide status.     

Effect of chronic ethanol diet on expression of gonadotropin genes in the male rat.

The present study elucidates further the defect in the function of hypothalamic-pituitary-gonadal axis in the male rat after a 6% ethanol diet for 5 weeks. The body, testis, prostate and seminal vesicle weights of the ethanol-treated animals did not differ from those of the pair-fed controls. However, the weights of the ethanol and pair-fed groups were significantly lower than in untreated controls (P less than .001). The pituitary luteinizing hormone beta mRNA level of ethanol-treated rats was significantly higher (P less than .05) but the follicle-stimulating hormone beta mRNA level was lower (P less than .05) than those of the diet control group. Additionally, the follicle-stimulating hormone beta mRNA level of the diet controls was higher than that of the untreated controls (P less than .05). There were no significant differences in the levels of common-alpha subunit or prolactin mRNA between the groups. The serum and testicular testosterone concentrations of the ethanol and diet control groups did not differ, but both were significantly lower than in untreated controls (P less than .01). The serum luteinizing hormone of ethanol rats was significantly higher than that of diet controls (P less than .01), but the pituitary luteinizing hormone, the serum and pituitary follicle-stimulating hormone and the prolactin of ethanol and diet control animals did not differ.     

Evidence for potentiation of lipid peroxidation in the rat liver after chronic ethanol feeding

Hepatic steatosis was induced in rats by feeding nutritionally adequate liquid diet containing ethanol as 36% of energy for 4-5 weeks. After 24 h fasting and withdrawal from ethanol, liver ischaemia for 30 min followed by 2 h reperfusion resulted in a significant increase in microsomal lipid peroxide content and a decrease in reduced glutathione content as well as in protein synthesis with a rapid accumulation of triglyceride in the liver. In rats fed a non-ethanol diet or those fed a high-cholesterol diet with hepatic steatosis, however, similar phenomena were not found. These findings suggest that chronic ethanol feeding potentiates hepatic lipid peroxidation.     

Iron mediates production of a neutrophil chemoattractant by rat hepatocytes metabolizing ethanol.

Ethanol metabolism in hepatocytes is accompanied by release of a potent lipid chemoattractant for neutrophils. Production of the factor may initiate the inflammation associated with alcoholic hepatitis. In previous studies with a cytosol system from liver, production was blocked by iron chelators as well as by catalase and superoxide dismutase, suggesting the involvement of oxyradicals in formation of the chemoattractant. These studies have examined the role of iron in intact hepatocytes using cells from rats fed an iron-deficient diet, a control diet or a diet containing 3% carbonyl iron. The iron content averaged 1.4 nmol/mg protein in iron-deficient cells, 6.3 in controls and 135.3 in iron-loaded cells. Hepatocytes from all groups were established in primary culture and incubated with ethanol (10 mM); the medium was assayed for chemoattractant activity for human neutrophils. Cultures from chow-fed or iron-loaded animals produced chemoattractant as previously reported. By contrast, chemoattractant production was undetectable in the iron-deficient cultures. Addition of ferric citrate (10 microM) restored chemoattractant production while increasing cellular iron in the deficient cells less than 50% (to 2.3 nmol/mg protein). Addition of desferrioxamine mesylate to cultures of iron-loaded cells ablated chemoattractant production. The data provide evidence for the importance of hepatocellular iron in production of this alcohol-related lipid chemoattractant and suggest that a small intracellular pool of "free" iron plays a critical role.     

Stereoselective glutathione conjugation and amidase-catalyzed hydrolysis of alpha-bromoisovalerylurea enantiomers in isolated rat hepatocytes

alpha-Bromoisovalerylurea (BIU) is used as model substrate for studies on the pharmacokinetics of glutathione conjugation in vivo. Its metabolism in isolated rat hepatocytes is presently studied. A major part of the substrate was conjugated with glutathione, but also amidase-catalyzed hydrolysis occurred, resulting in the products urea and alpha-bromoisovaleric acid (BI). The amidase activity was located in the microsomal fraction of the rat liver. The product of hydrolysis, BI, also was conjugated efficiently with glutathione. In glutathione-depleted hepatocytes, no glutathione conjugates but only urea and BI were formed. A pronounced stereoselectivity in the metabolism of the BIU enantiomers was observed: (R)-BIU was conjugated with glutathione much faster than (S)-BIU. (S)-BIU was hydrolyzed substantially in the cells and the glutathione conjugate of the hydrolytic product, (S)-BI, could be detected. At high BIU concentrations (500 microM of the racemate) intracellular glutathione was seriously depleted; then, the cosubstrate availability most likely was the rate-limiting factor in the conjugation of BIU with glutathione. More urea was formed from (racemic) BIU in isolated rat hepatocytes in the present study than in the perfused liver and the intact rat in previous studies. This in vivo-in vitro difference is tentatively assigned to differences in glutathione availability in these systems. The results suggest that BI may also be a useful model substrate to study the kinetics of glutathione conjugation in vivo and in vitro.     

Hepatic cholesterol synthesis and esterification in rats after chronic ethanol feeding

1. Chronic (5 weeks) alcohol-fed and isocaloric glucose pair-fed control rats had similar body weights, liver weights and liver protein contents. 2. Hepatic esterified cholesterol and triacylglycerol levels were two- to three-fold higher in alcohol-fed rats than in controls. 3. Hepatic cholesterol synthesis rates measured in vivo with 3H2O were significantly reduced in alcohol-fed rats. 4. Hepatic 3-hydroxy-3-methylglutaryl-CoA reductase (NADPH) (EC 1.1.1.34) activity was increased and the apparent Km for 3-hydroxymethyl-3-glutaryl-CoA was decreased in alcohol-fed rats. 5. Hepatic acyl-CoA:cholesterol acyltransferase (cholesterol acyltransferase; EC 2.3.1.26) activity was significantly increased in alcohol-fed rats. 6. These results indicate that there is no direct relationship between 3-hydroxy-3-methylglutaryl-CoA reductase activity and sterol synthesis in liver of alcohol-fed rats.     

Insulin and glucagon regulation of glutathione S-transferase expression in primary cultured rat hepatocytes

Diabetes is a major cause of morbidity and mortality, and complications resulting from diabetes have been attributed in part to increased oxidative stress. Glutathione S-transferases (GSTs) constitute a major protective mechanism against oxidative stress. Studies of the expression and activity of GSTs during diabetes are inconclusive, with both increased and decreased GST expression being reported in vivo. Insulin and glucagon effects on GST expression and the signaling pathway involved in the glucagon regulation of GST expression were examined in primary cultured rat hepatocytes. The addition of insulin resulted in the elevation of alpha-class GST protein levels, whereas alpha- and pi-class GST protein levels were markedly decreased in hepatocytes cultured with glucagon. In contrast, mu-class GST protein expression was unaffected by insulin or glucagon treatment. Insulin concentrations >/=1 nM resulted in increased GST activities and alpha-class GST protein levels, whereas glucagon concentrations >/=20 nM decreased alpha- and pi-class protein levels and activity. Treatment of cells with 8-bromo-cAMP or dibutyryl-cAMP also resulted in decreased alpha- and pi-class GST protein levels. Pretreatment with N-[2-(4-bromocinnamylamino)ethyl]-5-isoquinoline sulfonamide (H89), a selective inhibitor of protein kinase A, before glucagon addition markedly attenuated the glucagon effect. This study demonstrates that insulin and glucagon regulate, in an opposing manner, the expression of alpha-class GSTs and that glucagon completely inhibits pi-class GST expression in vitro, suggesting that hepatic GST expression may be decreased during diabetes. Furthermore, the present study implicates cAMP and protein kinase A in mediating the inhibitory effect of glucagon on GST expression.     

Effect of bicarbonate feeding on immobilization osteoporosis in the rat.

A time course study of immobilization osteoporosis was carried out in an experimental model in which the hind half of the rat was paralyzed by spinal section between the 3rd and 4th lumbar vertebrae. The earliest changes in the femure were a reduction in water content and in bone volume (3 days after operation). A significant reduction in both hydroxyproline and phosphate content was observed 8 days after immobilization. Calcium, magnesium, sodium, citrate, and carbonate were found significantly decreased at 15 days. Bone potassium remained unchanged. The rate of calcium accretion and the size of the exchangeable pools, when expressed per unit of bone mass, were not significantly affected by treatment. Sham-operated rats fed a bicarbonate-enriched diet, showed increased rates of calcium accretion and bone calcium content. The latter was not accompanied by increases in phosphate and carbonate contents. Operated animals fed the same diet showed an increased rate of bone resorption attributed to an assumed higher rate of parathyroid hormone secretion induced by bicarbonate feeding.     

Mechanisms behind the inhibitory effect of ethanol on the conjugation of morphine in rat hepatocytes

Liver microsomes were isolated by calcium aggregation, and isolated hepatocytes from male Wistar rats were prepared according to a two-step Ca++-free collagenase perfusion method. With the hepatocytes maximal inhibition of glucuronidation (about 40%) was reached at 10 mM ethanol after incubation at 37 degrees C for 60 min. UDP-glucuronic acid concentration and energy charge in the hepatocytes also did decrease maximally (about 90 and 50%, respectively) and the amount of UDP-glucose was tripled in the presence of 10 mM and higher concentrations of ethanol. The alcohol dehydrogenase inhibitor 4-methylpyrazole abolished ethanol-induced inhibition of morphine glucuronidation in the hepatocytes. Acetaldehyde (250-50 microM) and the pH decrease induced by ethanol did not reduce morphine-3-glucuronide formation by the cells. Cellular uptake of morphine and excretion of morphine metabolites were similar in the absence and presence of ethanol. Ethanol (60 mM) did not affect the glucuronidation of morphine (1.7 mM added) during a 30-min incubation at 37 degrees C with the microsomes (UDP-glucuronic acid, 5 mM). When the concentration of UDP-glucuronic acid in the microsomes was lowered from 1 to 0.1 mM, the decrease in morphine-3-glucuronide formation was similar to that observed in cells. The data indicate that the inhibition by ethanol of morphine glucuronidation was due to decreased levels of UDP-glucuronic acid. The mechanism is likely to be inhibition of UDP-glucose dehydrogenase activity by ethanol from increased intracellular NADH/NAD ratio accompanying ethanol oxidation.     

Structural and functional integrity of rat liver perfused in backward and forward directions

The purpose of this study was to assess if reversal of the direction of isolated rat liver perfusion would cause significant alterations in hepatic functions and structure. Five isolated rat livers were perfused forward and another five backward with oxygenated Ringer's solution for up to 90 min (hydrostatic pressure: less than or equal to 13 cm H2O; flow rate: forward 3.88 +/- 0.34 ml/min per gram and backward 3.76 +/- 0.34 ml/min per gram). At the end of the experiment, livers were perfusion-fixed for morphological examination. The following results were obtained: No significant differences were noted between the forward and backward perfusions with respect to oxygen uptake, mean bile flow (forward 0.57 +/- 0.12; backward 0.60 +/- 0.14 ml/min per gram), average bile acid excretion (forward 2.39 +/- 1.11; backward 2.83 +/- 0.94 nmol/min per gram), hydroxylation pattern of bile acids, urea synthesis, release of lactic dehydrogenase, glucose secretion, and redox ratios. Light and electron microscopy, including morphometry of parenchymal and sinusoidal areas, revealed that the backward perfusion caused a greater degree of sinusoidal distension, but no other noteworthy differences. Hepatic ultrastructure was well preserved. We conclude that reversing the direction of perfusion does not alter structure and major hepatic functions significantly.     

Insulin and glucocorticoid dependence of hepatic gamma-glutamylcysteine synthetase and glutathione synthesis in the rat. Studies in cultured hepatocytes and in vivo.

We reported that glucagon and phenylephrine decrease hepatocyte GSH by inhibiting gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in GSH synthesis (Lu, S.C., J. Kuhlenkamp, C. Garcia-Ruiz, and N. Kaplowitz. 1991. J. Clin. Invest. 88:260-269). In contrast, we have found that insulin (In, 1 microgram/ml) and hydrocortisone (HC, 50 nM) increased GSH of cultured hepatocytes up to 50-70% (earliest significant change at 6 h) with either methionine or cystine alone as the sole sulfur amino acid in the medium. The effect of In occurred independent of glucose concentration in the medium. Changes in steady-state cellular cysteine levels, cell volume, GSH efflux, or expression of gamma-glutamyl transpeptidase were excluded as possible mechanisms. Both hormones are known to induce cystine/glutamate transport, but this was excluded as the predominant mechanism since the induction in cystine uptake required a lag period of greater than 6 h, and the increase in cell GSH still occurred when cystine uptake was blocked. Assay of GSH synthesis in extracts of detergent-treated cells revealed that In and HC increased the activity of GCS by 45-65% (earliest significant change at 4 h) but not GSH synthetase. In and HC treatment increased the Vmax of GCS by 31-43% with no change in Km. Both the hormone-mediated increase in cell GSH and GCS activity were blocked with either cycloheximide or actinomycin D. Finally, when studied in vivo, streptozotocin-treated diabetic and adrenalectomized rats exhibited lower hepatic GSH levels and GCS activities than respective controls. Both of these abnormalities were prevented with hormone replacement. Thus, both in vitro and in vivo, In and glucocorticoids are required for normal expression of GCS.     

微囊化培养对肝细胞增殖生长和功能表达的影响

背景:前期研究证实,肝细胞在微囊化培养过程中结构形态发生变化,细胞的骨架会发生重排.目的:在前期研究基础上,进一步考察微囊化培养体系对细胞生长和功能表达的影响.方法:采用海藻酸钠-多聚赖氨酸-海藻酸钠微囊化人肝癌细胞系HepG2细胞,通过显微观察、苏木精-伊红染色、MTT实验、Realtime RT-PCR与Elisa实验,检测微囊内细胞的生长形态、细胞团的形态结构、细胞的活性、以及细胞的功能表达情况.结果与结论:HepG2细胞在微囊内聚集成团,以三维方式生长,细胞间连接紧密,与传统的平面培养相比,微囊化细胞的生长速率减慢,但微囊化细胞的功能基因表达水平及白蛋白的分泌量增高,说明海藻酸钠-多聚赖氨酸-海藻酸钠微胶囊所提供的微环境有利于肝细胞类组织的体外构建.结果也提示海藻酸钠-多聚赖氨酸-海藻酸钠微胶囊能够实现肝细胞的体外类组织化培养,有望成为一种更具前景的三维培养模式,应用于癌症治疗、高通量药物筛选以及肝组织工程的研究.     

Restricted expression of the erythroid/brain glucose transporter isoform to perivenous hepatocytes in rats. Modulation by glucose.

The "erythroid/brain" glucose transporter (GT) isoform is expressed only in a subset of hepatocytes, those forming the first row around the terminal hepatic venules, while the "liver" GT is expressed in all hepatocytes. After 3 d of starvation, a three- to fourfold elevation of expression of the erythroid/brain GT mRNA and protein is detected in the liver as a whole; this correlates with the expression of this GT in more hepatocytes, those forming the first three to four rows around the hepatic venules. Starvation-dependent expression of the erythroid/brain GT on the plasma membrane of these additional hepatocytes is lost within 3 h of glucose refeeding; however, by immunoblotting we show that the protein is still present. Its loss from the surface is possibly explained by internalization.