Biomedical subjects

A Bonen

Publications and source records attributed to A Bonen.

At least 37 records · Page 2Linked to original sources

PubMed2001

Tissue-specific and isoform-specific changes in MCT1 and MCT4 in heart and soleus muscle during a 1-yr period.

We examined the postnatal changes (days 10, 36, 84, 160, 365) of monocarboxylate transporters (MCT)1 and MCT4 in rat heart and soleus muscle. In the heart, MCT1 protein and mRNA remained unaltered from day 10 until 1 yr of age. Both MCT4 protein and mRNA in the heart were detected at 10 days of age, but the MCT4 protein and transcript were not detected thereafter. In the soleus muscle, MCT1 protein (+38%) and mRNA (+136%) increased during the first 84 days and remained stable until 1 yr of age. In contrast, soleus MCT4 protein decreased by 90% over the course of 1 yr, with the most rapid decrease (-60%) occurring by day 84 (P < 0.05). At the same time, MCT4 mRNA was increased by 74% from days 10 to 84 (P < 0.05), remaining stable thereafter. In conclusion, developmental changes in MCT transport proteins are tissue specific and isoform specific. Furthermore, it appears that MCT1 expression in the heart and MCT1 and MCT4 expression in the soleus are regulated by pretranslational processes, whereas posttranscriptional processes regulate MCT4 expression in the soleus muscle.

Animals↗

PubMed2000

Acute regulation of fatty acid uptake involves the cellular redistribution of fatty acid translocase.

We used muscle contraction, which increases fatty acid oxidation, as a model to determine whether fatty acid transport is acutely regulated by fatty acid translocase (FAT/CD36). Palmitate uptake by giant vesicles, obtained from skeletal muscle, was increased by muscle contraction. Kinetic studies indicated that muscle contraction increased V(max), but K(m) remained unaltered. Sulfo-N-succinimidyl oleate, a specific inhibitor of FAT/CD36, fully blocked the contraction-induced increase in palmitate uptake. In giant vesicles from contracting muscles, plasma membrane FAT/CD36 was also increased in parallel with the increase in long chain fatty acid uptake. Further studies showed that like GLUT-4, FAT/CD36 is located in both the plasma membrane and intracellularly (endosomally). With muscle contraction, FAT/CD36 at the surface of the muscle was increased, while concomitantly, FAT/CD36 in the intracellular pool was reduced. Similar responses were observed for GLUT-4. We conclude that fatty acid uptake is subject to short term regulation by muscle contraction and involves the translocation of FAT/CD36 from intracellular stores to the sarcolemma, analogous to the regulation of glucose uptake by GLUT-4.

Animals↗

PubMed2000

Muscle palmitate uptake and binding are saturable and inhibited by antibodies to FABP(PM).

Studies show that uptake of long-chain fatty acids (LCFA) across the plasma membranes (PM) may occur partly via a carrier-mediated process and that the plasma membrane fatty acid-binding protein (FABP(PM)) may be a component of this system. To test the hypothesis that FABP(PM) is involved in transsarcolemmal transport of LCFA in muscle, we measured palmitate uptake in giant sarcolemmal vesicles and palmitate binding to PM proteins in rat muscles, (1) in the presence of increasing amounts of unbound palmitate and (2) in the absence or presence of antibody to FABP(PM). Both palmitate uptake and binding were found to be saturable functions of the unbound palmitate concentration with calculated Vmax values of 10.5 +/- 1.2 pmol/mg protein/15 sec and 45.6 +/- 2.9 nmol/mg protein/15 min and Km values of 12.8 +/- 3.8 and 18.4 +/- 1.8 nmol/L, respectively. The Vmax values for both palmitate uptake and binding were significantly decreased by 75-79% in the presence of a polyclonal antibody to the rat hepatic FABP(PM). Antibody inhibition was found to be dose-dependent and specific to LCFA. Glucose uptake was not affected by the presence of the antibody to FABP(PM). Palmitate uptake and binding were also inhibited in the presence of trypsin and phloretin. These results support the hypothesis that transsarcolemmal LCFA transport occurs in part by a carrier-mediated process and that FABP(PM) is a component of this process in muscle.

Animals↗

PubMed2000

Denervation provokes greater reductions in insulin-stimulated glucose transport in muscle than severe diabetes.

We have examined the independent and combined effects of insulin insufficiency (streptozotocin (STZ)-induced diabetes, 85 mg/kg i.p.) and reduced muscle activity (denervation) (7 days) on basal, insulin-stimulated and contraction-stimulated glucose transport in rat muscles (soleus, red and white gastrocnemius). There were four treatments: control, denervated, diabetic, and denervated + diabetic muscles. Contraction-stimulated glucose transport was lowered (approximately 50%) (p < 0.05) to the same extent in all experimental groups. In contrast, there was a much smaller reduction insulin-stimulated glucose transport in muscles from diabetic animals (18-24% reduction, p < 0.05) than in denervated muscles (40-60% reduction, p < 0.05) and in denervated + diabetic muscles (40-60% reduction, p < 0.05). GLUT-4 mRNA reduction was greatest in denervated + diabetic muscles (approximately -75%, p < 0.05). GLUT-4 protein was decreased (p < 0.05) to a similar extent in all three experimental conditions (approximately -30-40%). In conclusion, (1) muscle inactivity (denervation) and STZ-induced diabetes had similar effects on reducing contraction-stimulated glucose transport, but (2) muscle inactivity (denervation), rather than severe diabetes, produced a 2-fold greater impairment in skeletal muscle insulin-stimulated glucose transport.

Animals↗

PubMed2000

Lactate transporters (MCT proteins) in heart and skeletal muscles.

Lactate traverses the cell membranes of many tissues, including the heart and skeletal muscle via a facilitated monocarboxylate transport system that functions as a proton symport and is stereoselective for L-lactate. In the past few years, seven monocarboxylate transporters have been cloned. Monocarboxylate transporters are ubiquitously distributed among many tissues, and the transcripts of several monocarboxylate transporters are present within many of the same tissues. This complicates the identification of their metabolic function. There is also evidence that that there is some species specificity, with differences in MCT tissue distributions in hamsters, rats, and humans. MCT1 and MCT3-M/MCT4 are present in rat and human muscles, and MCT1 expression is highly correlated with the oxidative capacity of skeletal muscles and with their capacity to take up lactate from the circulation. MCT1 is also present in heart and is located on the plasma membrane (in subdomains), T-tubules, and in caveolae. With training, MCT1 is increased in rat and human muscle, and in rat hearts, resulting in an increased uptake of lactate from the buffers perfused through these tissues and an increase in lactate efflux out of purified vesicles. In humans, the training-induced increases in MCT1 are associated with an increased lactate efflux out of muscle. MCT3-M/MCT4 is not correlated with the muscles' oxidative capacities but is equally abundant in Type IIa and IIb muscles, whereas it is markedly lower in slow-twitch (Type I) muscles. Clearly, we are at the threshold of a new era in understanding the regulation of lactate movement into and out of skeletal muscle and cardiac cells.

Animals↗

PubMed2000

Endurance training increases FFA oxidation and reduces triacylglycerol utilization in contracting rat soleus.

We examined the effects of 8 wk of intense endurance training on free fatty acid (FFA) transporters and metabolism in resting and contracting soleus muscle using pulse-chase procedures. Endurance training increased maximal citrate synthase activity in red muscles (+54 to +91%; P </= 0.05) but failed to increase cytosolic fatty acid binding protein content, mRNA for fatty acyl-CoA synthase, and the putative FFA transporters or transport of palmitic acid into giant sarcolemmal vesicles. At rest, only triacylglycerol (TG) synthesis was significantly increased by training (+100.9 +/- 8.7 vs. +66.6 +/- 6.7 nmol/g wet wt; P </= 0.05). Muscle contraction increased TG synthesis (+46%; P </= 0.05) and palmitate oxidation (+115%; P </= 0.05) in untrained rats. Endurance training further enhanced synthesis of monoacylglycerol (MG), diacylglycerol (DG) and TG during contraction (+36, +69 and +71%, respectively; P </= 0.05), as well as exogenous palmitate oxidation (+41%; P </= 0.05) relative to untrained rats. Compared with those in untrained rats, TG breakdown and oxidation during contraction were reduced after training by 49 and 30%, respectively (P </= 0.05). In conclusion, endurance training 1) increases FFA oxidation and incorporation into endogenous lipid pools during contraction and 2) reduces the rate of intramuscular TG utilization during contraction when exogenous FFA availability is adequate. The enhanced FFA uptake subsequent to training appears to be independent of altered maximal transport rates of FFA into the muscle cell.

Animals↗

PubMed2000

Abundance and subcellular distribution of MCT1 and MCT4 in heart and fast-twitch skeletal muscles.

The expression of two monocarboxylate transporters (MCTs) was examined in muscle and heart. MCT1 and MCT4 proteins are coexpressed in rat skeletal muscles, but only MCT1 is expressed in rat hearts. Among six rat fast-twitch muscles (red and white gastrocnemius, plantaris, extensor digitorum longus, red and white tibialis anterior) there was an inverse relationship between MCT1 and MCT4 (r = -0.94). MCT1 protein was correlated with MCT1 mRNA (r = 0.94). There was no relationship between MCT4 mRNA and MCT4 protein. MCT1 (r = -0.97) and MCT4 (r = 0.88) protein contents were correlated with percent fast-twitch glycolytic fiber. When normalized for their mRNAs, MCT1 but not MCT4 was still correlated with the percent fast-twitch glycolytic fiber composition of rat muscles (r = -0.98). MCT1 and MCT4 were also measured in plasma membranes (PM), triads (TR), T tubules (TT), sarcoplasmic reticulum (SR), and intracellular membranes (IM). There was an intracellular pool of MCT4 but not of MCT1. The MCT1 subcellular distribution was as follows: PM (100%) > TR (31.6%) > SR (15%) = TT (14%) > IM (1.7%). The MCT4 subcellular distribution was considerably different [PM (100%) > TR (66.5%) > TT (36%) = SR (43%) > IM (24%)]. These studies have shown that 1) the mechanisms regulating the expression of MCT1 (transcriptional and posttranscriptional) and MCT4 (posttranscriptional) are different and 2) differences in MCT1 and MCT4 expression among muscles, as well as in their subcellular locations, suggest that they may have different roles in muscle.

Animals↗

PubMed2000

Reduced insulin-stimulated glucose transport in denervated muscle is associated with impaired Akt-alpha activation.

Insulin signaling was examined in muscle made insulin resistant by short-term (24-h) denervation. Insulin-stimulated glucose transport in vitro was reduced by 28% (P < 0.05) in denervated muscle (DEN). In control muscle (SHAM), insulin increased levels of surface-detectable GLUT-4 (i.e., translocated GLUT-4) 1.8-fold (P < 0.05), whereas DEN surface GLUT-4 was not increased by insulin (P > 0.05). Insulin treatment in vivo induced a rapid appearance of phospho[Ser(473)]Akt-alpha in SHAM 3 min after insulin injection. In DEN, phospho[Ser(473)]Akt-alpha also appeared at 3 min, but Ser(473)-phosphorylated Akt-alpha was 36% lower than in SHAM (P < 0. 05). In addition, total Akt-alpha protein in DEN was 37% lower than in SHAM (P < 0.05). Akt-alpha kinase activity was lower in DEN at two insulin levels tested: 0.1 U insulin/rat (-22%, P < 0.05) and 1 U insulin/rat (-26%, P < 0.01). These data indicate that short-term (24-h) denervation, which lowers insulin-stimulated glucose transport, is associated with decreased Akt-alpha activation and impaired insulin-stimulated GLUT-4 appearance at the muscle surface.

Animals↗

PubMed2000

Isoform-specific regulation of the lactate transporters MCT1 and MCT4 by contractile activity.

We examined the isoform-specific regulation of monocarboxylate transporter (MCT)1 and MCT4 expression by contractile activity in red and white tibialis anterior muscles. After 1 and 3 wk of chronic muscle stimulation (24 h/day), MCT1 protein expression was increased in the red muscles (+78%, P < 0.05). In the white muscles, MCT1 was increased after 1 wk (+191%) and then was decreased after 3 wk. In the red muscle, MCT1 mRNA accumulation was increased only after 3 wk (+21%; P < 0.05). In the white muscle, MCT1 mRNA was increased after 1 wk (+30%; P < 0.05) and 3 wk (+15%; P < 0.05). MCT4 protein was not altered in either the red or white muscles after 1 or 3 wk. MCT4 mRNA was transiently lowered (approximately 15%) in both muscles in the 1st wk, but MCT4 mRNA levels were back to control levels after 3 wk. In conclusion, chronic contractile activity induces the expression of MCT1 but not MCT4. This increase in MCT1 alone was sufficient to increase lactate uptake from the circulation.

Animals↗

PubMed2000

The transferrin receptor defines two distinct contraction-responsive GLUT4 vesicle populations in skeletal muscle.

Insulin and contraction increase glucose transport in an additive fashion in skeletal muscle. However, it is still unclear whether they do so by inducing the recruitment of GLUT4 transporters from the same or distinct intracellular compartments to the plasma membrane and the T-tubules. Using the transferrin receptor as a recognized marker of recycling endosomes, we have examined whether insulin and/or contraction recruit GLUT4 from this pool to either the plasma membranes or T-tubules, isolated by subcellular fractionation of perfused hindlimb muscles. Either stimulus independently increased GLUT4 translocation from an intracellular fraction to both the plasma membrane and T-tubules. The combination of insulin and contraction induced a marked (approximately threefold) and almost fully additive increase in GLUT4 content, but only in the plasma membrane. Insulin did not stimulate transferrin receptor recruitment from the GLUT4-containing intracellular fraction to either the plasma membrane or the T-tubules. In contrast, contraction stimulated the recruitment of the transferrin receptor from the same GLUT4-containing intracellular fraction to the plasma membrane but not to the T-tubules. Contraction-induced recruitment of the transferrin receptor was also observed from immunopurified GLUT4 vesicles. It is concluded that muscle contraction stimulates translocation of GLUT4 from two distinct intracellular compartments: 1) a population of recycling endosomes that is selectively recruited to the plasma membrane and 2) from GLUT4 storage vesicles that are also insulin-responsive and recruited to both the plasma membrane and the T-tubules. The lack of additive translocation of GLUT4 to the T-tubules may be linked to the failure of GLUT4-containing recycling endosomes to be recruited to these structures.

Animals↗

PubMed2000

Fatty acid transport proteins facilitate fatty acid uptake in skeletal muscle.

In view of the importance of long chain fatty acids (LCFAs) to many cellular processes, it may be desirable to regulate the LCFA disposition in the cell. Such regulation may be present at the level of the plasma membrane, since a number of putative LCFA transport proteins have been cloned. The development of a model system of giant vesicles has proven to be important in identifying the metabolic role of the LCFA transport system. LCFA transport rates and transporters (FABPpm and FAT/CD36) are scaled with oxidative capacity of heart and muscle. FAT/CD36 is a critical LCFA transport protein in muscle. With chronic contraction the increase in this protein also results in an increase in LCFA transport. Most importantly, LCFA transport is also increased acutely by muscle contraction, involving the translocation of FAT/CD36 from intracellular depots to the surface of the muscle cell. The acute (minutes) and chronic (days) regulation of LCFA transporters and transport by muscle may be an important mechanism for LCFA utilization during exercise and adaptable with training and with a metabolic disease such as type 2 diabetes.

Animals↗

PubMed1999

Muscle-specific overexpression of FAT/CD36 enhances fatty acid oxidation by contracting muscle, reduces plasma triglycerides and fatty acids, and increases plasma glucose and insulin.

Increasing evidence has implicated the membrane protein CD36 (FAT) in binding and transport of long chain fatty acids (FA). To determine the physiological role of CD36, we examined effects of its overexpression in muscle, a tissue that depends on FA for its energy needs and is responsible for clearing a major fraction of circulating FA. Mice with CD36 overexpression in muscle were generated using the promoter of the muscle creatine kinase gene (MCK). Transgenic (MCK-CD36) mice had a slightly lower body weight than control litter mates. This reflected a leaner body mass with less overall adipose tissue, as evidenced by magnetic resonance spectroscopy. Soleus muscles from transgenic animals exhibited a greatly enhanced ability to oxidize fatty acids in response to stimulation/contraction. This increased oxidative ability was not associated with significant alterations in histological appearance of muscle fibers. Transgenic mice had lower blood levels of triglycerides and fatty acids and a reduced triglyceride content of very low density lipoproteins. Blood cholesterol levels were slightly lower, but no significant decrease in the cholesterol content of major lipoprotein fractions was measured. Blood glucose was significantly increased, while insulin levels were similar in the fed state and higher in the fasted state. However, glucose tolerance curves, determined at 20 weeks of age, were similar in control and transgenic mice. In summary, the study documented, in vivo, the role of CD36 to facilitate cellular FA uptake. It also illustrated importance of the uptake process in muscle to overall FA metabolism and glucose utilization.

Adipose Tissue↗

PubMed1999

Cellular fatty acid transport in heart and skeletal muscle as facilitated by proteins.

Despite the importance of long-chain fatty acids (FA) as fuels for heart and skeletal muscles, the mechanism of their cellular uptake has not yet been clarified. There is dispute as to whether FA are taken up by the muscle cells via passive diffusion and/or carrier-mediated transport. Kinetic studies of FA uptake by cardiac myocytes and the use of membrane protein-modifying agents have suggested the bulk of FA uptake is due to a protein component. Three membrane-associated FA-binding proteins were proposed to play a role in FA uptake, a 40-kDa plasma membrane FA-binding protein (FABPpm), an 88-kDa FA translocase (FAT/CD36), and a 60-kDa FA transport protein (FATP). In cardiac and skeletal myocytes the intracellular carrier for FA is cytoplasmic heart-type FA-binding protein (H-FABP), which likely transports FA from the sarcolemma to their intracellular sites of metabolism. A scenario is discussed in which FABPpm, FAT/CD36, and H-FABP, probably assisted by an albumin-binding protein, cooperate in the translocation of FA across the sarcolemma.

Animals↗

PubMed1999

Preferential inhibition of lactate oxidation relative to glucose oxidation in the rat heart following diabetes.

OBJECTIVE: Alterations in myocardial metabolism occur early after the onset of diabetes suggesting that they may play a role in the development of cardiac dysfunction. Inhibition of myocardial pyruvate dehydrogenase (PDH), glucose transport and glycolysis have all been reported following diabetes. In vivo lactate is also a potential source of energy for the heart and its oxidation should not be affected by changes in glucose transport and glycolysis. Therefore, the objective of this study, was to test the hypothesis that following diabetes the inhibition of glucose oxidation would be greater than the inhibition of lactate oxidation. METHODS: Hearts from control and one-week-old diabetic rats were perfused with [1-13C]glucose (11 mmol/l) alone, [1-13C]glucose plus lactate (0.5 mmol/l) or glucose plus [3-13C]lactate (0.5 or 1.0 mmol/l) as substrates. Glucose and lactate oxidation rates were determined by combining 13C-NMR glutamate isotopomer analysis of tissue extracts with measurements of oxygen consumption. RESULTS: In diabetic hearts perfused with glucose alone, glucose oxidation was decreased compared to controls (0.31 +/- 0.08 vs. 0.71 +/- 0.11 mumoles/min/g wet weight; p < 0.05). Surprisingly, in hearts perfused with glucose plus 0.5 mmol/l lactate, there was no difference in glucose oxidation between control and diabetic groups (0.20 +/- 0.05 vs. 0.16 +/- 0.04 mumoles/min/g wet weight respectively). However, under these conditions lactate oxidation was markedly reduced in the diabetic group (0.89 +/- 0.18 vs. 0.24 +/- 0.05 mumoles/min/g wet weight; p < 0.05). At 1.0 mmol/l lactate oxidation was still significantly depressed in the diabetic group. CONCLUSION: There was a greater decrease in lactate oxidation relative to glucose oxidation in hearts from diabetic animals. These results demonstrate that diabetes leads to a specific inhibition of lactate oxidation independent of its effects on pyruvate dehydrogenase.

Analysis of Variance↗

PubMed1999

Fatty acid transporters (FABPpm, FAT, FATP) in human muscle.

Recently, a number of putative LCFA transporters have been identified: fatty acid binding protein (FABPpm), fatty acid translocase (FAT/CD36), and fatty acid transport protein (FATP). We have demonstrated, for the first time, that transcripts of all three putative LCFA transporters (FAT mRNA, FATP mRNA, and mAspAT/FABPpm mRNA) are present in human skeletal muscle.

Adult↗

PubMed1999

Muscle contractile activity increases fatty acid metabolism and transport and FAT/CD36.

We have examined whether 1) fatty acid (FA) uptake, 2) FA transporter expression, and 3) FA metabolism are increased when the oxidative capacity of skeletal muscle is increased. The oxidative capacities of red and white tibialis anterior and extensor digitorum longus muscles were increased via chronic stimulation (10 Hz, 24 h/day for 7 days). The contralateral muscles served as controls. After 7 days of increased muscle activity 1) palmitate uptake by giant sarcolemmal vesicles was increased twofold (P < 0.05), 2) the expression of FA translocase (FAT)/CD36 was increased at both the mRNA (3.2- to 10-fold) and protein (3.4-fold) levels, and 3) palmitate oxidation and esterification into triacylglycerols and phospholipids were increased 1.5-, 2.7-, and 1.7-fold, respectively (P < 0.05). These data show that when the oxidative capacity of muscle is increased, there is a parallel increase in the rate of FA transport and FA transporters at the sarcolemmal membrane, which is associated with the enhanced expression of the membrane transporter FAT/CD36.

Animals↗

PubMed1999

Protein-mediated palmitate uptake and expression of fatty acid transport proteins in heart giant vesicles.

Giant sarcolemmal vesicles were isolated from rat heart and hindlimb muscles for a) characterization of long-chain fatty acid transport in the absence of metabolism and b) comparison of fatty acid transport protein expression with fatty acid transport. Giant vesicles contained cytosolic fatty acid binding protein. Palmitate uptake was completely divorced from its metabolism. All palmitate taken up was recovered in the intravesicular cytosol as unesterified FA. Palmitate uptake by heart vesicles exhibited a K m of 9.7 nm, similar to that of muscle (K m = 9.7 nm). Vmax (2.7 pmol/mg protein/s) in heart was 8-fold higher than in muscle (0.34 pmol/mg protein/s). Palmitate uptake was inhibited in heart (55-80%) and muscle (31-50%) by trypsin, phloretin, sulfo-N-succinimidyloleate (SSO), or a polyclonal antiserum against the 40 kDa plasma membrane fatty acid binding protein (FABPpm). Palmitate uptake by heart and by red and white muscle vesicles correlated well with the expression of fatty acid translocase (FAT/CD36) and fatty acid binding protein FABPpm, which may act in concert. The expression of fatty acid transport protein (FATP), was 10-fold lower in heart vesicles than in white muscle vesicles. It is concluded that long-chain fatty acid uptake by heart and muscle vesicles is largely protein-mediated, involving FAT/CD36 and FABPpm. The role of FATP in muscle and heart remains uncertain.

Animals↗

PubMed1998

Lactic acid efflux from white skeletal muscle is catalyzed by the monocarboxylate transporter isoform MCT3.

The newly cloned proton-linked monocarboxylate transporter MCT3 was shown by Western blotting and immunofluorescence confocal microscopy to be expressed in all muscle fibers. In contrast, MCT1 is expressed most abundantly in oxidative fibers but is almost totally absent in fast-twitch glycolytic fibers. Thus MCT3 appears to be the major MCT isoform responsible for efflux of glycolytically derived lactic acid from white skeletal muscle. MCT3 is also expressed in several other tissues requiring rapid lactic acid efflux. The expression of both MCT3 and MCT1 was decreased by 40-60% 3 weeks after denervation of rat hind limb muscles, whereas chronic stimulation of the muscles for 7 days increased expression of MCT1 2-3-fold but had no effect on MCT3 expression. The kinetics and substrate and inhibitor specificities of monocarboxylate transport into cell lines expressing only MCT3 or MCT1 have been determined. Differences in the properties of MCT1 and MCT3 are relatively modest, suggesting that the significance of the two isoforms may be related to their regulation rather than their intrinsic properties.

Animals↗