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B B Kahn

Publications and source records attributed to B B Kahn.

90 records · Page 5Linked to original sources
PubMed1991

High fat feeding causes insulin resistance and a marked decrease in the expression of glucose transporters (Glut 4) in fat cells of rats.

With the identification of two different glucose transporter species in adipose cells it is crucial to determine the role of these transporters in the alterations in glucose transport activity associated with different metabolic and nutritional states. In the present study we assess levels of expression of Glut 1 and Glut 4 transporters and basal and insulin-stimulated glucose transport activity in adipocytes from Sprague-Dawley rats fed standard chow (control), combined liquid diet and standard chow (overfed), high fat diet, or energy-restricted diet for 7 weeks. High fat feeding was associated with relative postprandial hypoglycemia (P less than 0.05) and hypoinsulinemia (P less than 0.05). Although the high fat fed animals had lower body weights (P less than 0.05) than control rats, their body compositions showed obesity, with 36% heavier epididymal fat pads (P less than 0.05) and a 47% increase in adipocyte volume (P less than 0.05). Fat feeding caused a 78% reduction in insulin-stimulated glucose transport per adipocyte (P less than 0.05). In parallel we found 92% and 94% reductions in Glut 4 protein and mRNA per adipocyte, respectively, (P less than 0.01) in fat-fed rats. Substantial reductions were also seen in Glut 1 protein and mRNA per fat cell in the same rats (62% and 76%, respectively; P less than 0.05). However, the changes in Glut 1 expression were of the same magnitude as changes in the cytoskeletal protein beta-actin, reflecting a decreased expression of several proteins in this nutritional state. Even though overfeeding and energy restriction brought about opposite changes in adiposity, no significant alterations were demonstrated in glucose transport rate or glucose transporter expression. The impaired insulin-stimulated glucose transport in adipose cells from high fat-fed rats occurs in the presence of a dramatic decrease in the expression of the major insulin-responsive glucose transporter (Glut 4). The reduced gene expression may be caused by chronic hypoinsulinemia and may contribute to the insulin resistance observed in this state.

Adipose Tissue↗
PubMed1991

Expression of the low Km GLUT-1 glucose transporter is turned on in perivenous hepatocytes of insulin-deficient diabetic rats.

In normal fed rats the low Km glucose transporter GLUT-1 is expressed only in one row of hepatocytes immediately surrounding a terminal hepatic venule, while the high Km GLUT-2 is expressed in every hepatocyte. Previously, we showed that additional perivenous hepatocytes express GLUT-1 in fasting animals. In diabetes, as in starvation, the liver functions to release glucose into the circulation, but unlike starvation, circulating extracellular glucose is high in diabetes. By immunofluorescence and Western blotting we studied whether glucose or insulin is the primary extracellular signal for inducing GLUT-1 expression in hepatocytes. We observed that streptozocin-induced diabetes causes induction of GLUT-1 expression in the plasma membrane of hepatocytes within four cell rows of a terminal hepatic venule; GLUT-2 expression is unaltered. Chronic insulin treatment of diabetic rats reduces the number of rows of hepatocytes expressing GLUT-1 from approximately four to approximately two. In contrast, chronic insulin infusion into nondiabetic rats does not affect the number of hepatocytes expressing GLUT-1. Thus, both fasting and diabetes induce GLUT-1 expression in specific hepatocytes that normally do not express this gene. This induction correlates with low insulin levels in the blood, and not with circulating glucose levels.

Animals↗
PubMed1990

Divergent molecular mechanisms for insulin-resistant glucose transport in muscle and adipose cells in vivo.

Glucose homeostasis depends on regulated changes in glucose transport in insulin-responsive tissues (e.g. muscle and adipose cells). This transport is mediated by at least two distinct glucose transporters: "adipose-muscle" and "erythrocyte-brain." To understand the molecular basis for in vivo insulin resistance we investigated the effects of fasting and refeeding on the expression of these two glucose transporters in adipose cells and skeletal muscle. In vivo insulin resistance seen with fasting and hyperresponsiveness seen with refeeding influence glucose transporter expression in a transporter-specific and tissue-specific manner. In adipose cells only the adipose-muscle glucose transporter mRNA and protein decrease dramatically with fasting and increase above control levels with refeeding, changes that parallel effects on insulin-stimulated glucose transport. In contrast, in muscle expression of both glucose transporters increase with fasting and return to control levels with refeeding, also in accordance with changes in glucose uptake in vitro. Although expression of the adipose-muscle glucose transporter predicts the physiological response at the tissue level, factors in the hormonal/metabolic milieu appear to override its increased expression in muscle resulting in insulin-resistant glucose uptake in this tissue in vivo.

Adipose Tissue↗
PubMed1990

Effect of endurance training on glucose transport capacity and glucose transporter expression in rat skeletal muscle.

The effect of 10 wk endurance swim training on 3-O-methylglucose (3-MG) uptake (at 40 mM 3-MG) in skeletal muscle was studied in the perfused rat hindquarter. Training resulted in an increase of approximately 33% for maximum insulin-stimulated 3-MG transport in fast-twitch red fibers and an increase of approximately 33% for contraction-stimulated transport in slow-twitch red fibers compared with nonexercised sedentary muscle. A fully additive effect of insulin and contractions was observed both in trained and untrained muscle. Compared with transport in control rats subjected to an almost exhaustive single exercise session the day before experiment both maximum insulin- and contraction-stimulated transport rates were increased in all muscle types in trained rats. Accordingly, the increased glucose transport capacity in trained muscle was not due to a residual effect of the last training session. Half-times for reversal of contraction-induced glucose transport were similar in trained and untrained muscles. The concentrations of mRNA for GLUT-1 (the erythrocyte-brain-Hep G2 glucose transporter) and GLUT-4 (the adipocyte-muscle glucose transporter) were increased approximately twofold by training in fast-twitch red muscle fibers. In parallel to this, Western blot demonstrated a approximately 47% increase in GLUT-1 protein and a approximately 31% increase in GLUT-4 protein. This indicates that the increases in maximum velocity for 3-MG transport in trained muscle is due to an increased number of glucose transporters.

3-O-Methylglucose↗
PubMed1990

Differential regulation of two glucose transporters in rat liver by fasting and refeeding and by diabetes and insulin treatment.

Two species of glucose transporter (GT) are present in the liver: the erythroid/brain GT and the newly characterized liver GT. No information is available regarding the functional role of these two species or whether their expression is regulated concordantly in states in which hepatic glucose uptake or output are markedly altered. In this study, we analyzed the effect of fasting and refeeding and streptozocin-induced diabetes and subsequent insulin treatment on the expression of the erythroid/brain and liver GT polypeptides and their mRNAs in rat liver. The erythroid/brain GT mRNA in livers of control rats corresponded to 1-3% of the amount of liver GT mRNA. After a 4-day fast, its level increased approximately 3-fold and represented 8-10% of the liver GT mRNA, whereas the corresponding protein increased 4-fold. In livers from diabetic rats, levels of the erythroid/brain GT mRNA increased up to 2.4-fold and gradually returned to normal with chronic insulin treatment. Levels of the corresponding protein were minimally altered. Levels of immunoreactive liver GTs were not significantly changed by 2 days of fasting, 7 or 14 days of diabetes, or subsequent insulin treatment for 3, 5, or 7 days but increased up to 75% with refeeding for 3-48 h. Liver GT mRNA levels minimally decreased in diabetic or insulin-treated rats, decreased 45% after a 2-day fast, and increased up to 5-fold on refeeding for 24 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Feed↗
PubMed1990

Evidence against altered expression of GLUT1 or GLUT4 in skeletal muscle of patients with obesity or NIDDM.

Studies of experimental diabetes in rodents induced by the beta-cell toxin streptozocin have shown that the insulin-resistant glucose transport of peripheral tissues (muscle and adipose) in these animals can be ascribed in part to a pretranslational reduction of the major insulin-sensitive glucose transporter (GLUT4) in these tissues. Because a central feature of non-insulin-dependent diabetes mellitus (NIDDM) is an imparied ability of insulin to enhance glucose disposal in skeletal muscle, we examined the hypothesis that reduced expression of GLUT4 is a characteristic finding in the skeletal muscle of subjects with NIDDM. Biopsies of skeletal muscles were obtained from 17 patients with NIDDM and 10 lean and 9 obese nondiabetic subjects. Among the diabetic subjects, 7 were newly diagnosed and untreated. Compared with age-matched and body-weight-matched healthy control subjects, there was no significant alteration in the level of GLUT4 mRNA demonstrated by Northern blot and slot blot or GLUT4 protein determined by immunoblotting muscle membranes. Neither GLUT4 mRNA nor protein concentration correlated with the degree of glycemic control, fasting plasma insulin or glucose, diabetes duration, body mass index, sex, or age. GLUT1 mRNA and protein levels were also not significantly different between diabetic and matched control subjects. Thus, unlike streptozocin-induced diabetes in rodents, there is no evidence that impaired expression of the major insulin-responsive glucose transporter is responsible for insulin-resistant glucose transport in the skeletal muscle of these lean and moderately obese NIDDM patients.

Animals↗
PubMed1990

Regulation of glucose-transporter gene expression in vitro and in vivo.

Understanding of the fundamental mechanisms underlying the complex regulation of glucose homeostasis has been dramatically transformed recently by the realization that glucose transport in mammalian tissues is mediated by a family of structurally related but genetically distinct glucose-transporter proteins. The regulatory factors and intracellular signaling pathways that influence expression of the genes encoding these proteins are just being identified. Factors that regulate glucose-transporter gene expression in vitro include oncogenes, growth factors, insulin, oral hypoglycemic agents, vanadate, glucocorticoids, ambient glucose levels, and the state of cellular differentiation. In vivo, glucose-transporter gene expression in adipose cells, skeletal muscle, and liver is markedly affected by various altered nutritional and metabolic states. Recent studies have demonstrated that two glucose transporters expressed in the same tissue may be regulated differently in response to the same metabolic perturbation. Furthermore, transporter regulation appears to be tissue specific. These observations lay the groundwork for future studies aimed at unraveling the functional roles of the individual transporter species in different tissues, the molecular processes involved in regulating the expression of these genes, and the impact of dysregulated glucose-transporter gene expression in the pathogenesis of insulin-resistant states such as diabetes.

Animals↗
PubMed1989

Regulation of glucose transporter-specific mRNA levels in rat adipose cells with fasting and refeeding. Implications for in vivo control of glucose transporter number.

Fasting in the rat is associated with a rapid and progressive decrease in insulin-stimulated glucose transport activity in adipose cells, which is not only restored to normal, but increased transiently to supranormal levels by refeeding. The mechanisms for these changes in glucose transport activity appear to involve alterations in both glucose transporter number and intrinsic activity (glucose turnover number). In this study, we use the human hepatoma Hep G2 glucose transporter complementary DNA clone to examine the molecular basis for these alterations. Extractable RNA per adipose cell is decreased 35% with 3 d of fasting and increased to 182% of control with 6 d of refeeding after 2 d of fasting. This parallels changes in adipose cell intracellular water, so that total RNA/water space remains relatively constant. When the changes in total RNA/cell are taken into account, Northern and slot blot analyses with quantitative densitometry reveal a 36% decrease in specific glucose transporter mRNA level in cells from the fasted rats. The mRNA level in cells from the fasted/refed rats is restored to normal. These observations correlate closely with previous measurements of glucose transporter number in adipose cell membrane fractions using cytochalasin B binding and Western blotting. The levels of specific mRNAs for tubulin and actin on a per cell basis show similar but more dramatic changes and mRNAs encoding several differentiation-dependent adipose cell proteins are also significantly affected. Thus, the levels of mRNA for multiple adipose cell genes are affected by fasting and refeeding. In particular, this demonstrates that in vivo metabolic alterations can influence the level of a glucose transporter mRNA in adipose cells. This may have implications for the regulation of glucose transporter number and glucose transport activity.

Adipose Tissue↗
PubMed1989

Differential regulation of two glucose transporters in adipose cells from diabetic and insulin-treated diabetic rats.

At least two genetically distinct glucose transporters (GTs) coexist in adipose cells, one cloned from human hepatoma cells and rat brain (HepG2/brain) and another from rat skeletal muscle, heart, and adipose cells (adipose cell/muscle). Here we demonstrate differential regulation of these two GTs in adipose cells of diabetic and insulin-treated diabetic rats and compare changes in the expression of each GT with marked alterations in insulin-stimulated glucose transport activity. Adipose cell/muscle GTs detected by immunoblotting with the monoclonal antiserum 1F8 (James, D. E., R. Brown, J. Navarro, and P. F. Pilch. 1988. Nature (Lond.). 333:183-185), which reacts with the protein product of the newly cloned adipose cell/muscle GT cDNA, decrease 87% with diabetes and increase to 8.5-fold diabetic levels with insulin treatment. These changes concur qualitatively with previous detection of GTs by cytochalasin B binding and with insulin-stimulated 3-O-methylglucose transport. Northern blotting reveals that the adipose/muscle GT mRNA decreases 50% with diabetes and increases to 6.8-fold control (13-fold diabetic) levels with insulin treatment. In contrast, GTs detected with antisera to the carboxyl terminus of the HepG2 GT or to the human erythrocyte GT show no significant change with diabetes or insulin treatment. The HepG2/brain GT mRNA is unchanged with diabetes and increases threefold with insulin treatment. These results suggest that (a) altered expression of the adipose cell/muscle GT forms the molecular basis for the dysregulated glucose transport response to insulin characteristic of diabetes, (b) the expression of two types of GTs in rat adipose cells is regulated independently, and (c) alterations in mRNA levels are only part of the mechanism for in vivo regulation of the expression of either GT species.

Actins↗
PubMed1988

Divergent mechanisms for the insulin resistant and hyperresponsive glucose transport in adipose cells from fasted and refed rats. Alterations in both glucose transporter number and intrinsic activity.

The effects of fasting and refeeding on the glucose transport response to insulin in isolated rat adipose cells have been examined using 3-O-methylglucose transport in intact cells and cytochalasin B binding and Western blotting in subcellular membrane fractions. After a 72-h fast, basal glucose transport activity decreases slightly and insulin-stimulated activity decreases greater than 85%. Following 48 h of fasting, insulin-stimulated glucose transport activity is diminished from 3.9 +/- 0.5 to 1.3 +/- 0.3 fmol/cell per min (mean +/- SEM). Similarly, the concentrations of glucose transporters are reduced with fasting in both the plasma membranes from insulin-stimulated cells from 38 +/- 5 to 18 +/- 3 pmol/mg of membrane protein and the low density microsomes from basal cells from 68 +/- 8 to 34 +/- 9 pmol/mg of membrane protein. Ad lib. refeeding for 6 d after a 48-h fast results in up to twofold greater maximally insulin-stimulated glucose transport activity compared with the control level (7.1 +/- 0.4 vs. 4.5 +/- 0.2 fmol/cell per min), before returning to baseline at 10 d. However, the corresponding concentration of glucose transporters in the plasma membranes is restored only to the control level (45 +/- 5 vs. 50 +/- 5 pmol/mg of membrane protein). Although the concentration of glucose transporters in the low density microsomes of basal cells remains decreased, the total number is restored to the control level due to an increase in low density microsomal protein. Thus, the insulin-resistant glucose transport in adipose cells from fasted rats can be explained by a decreased translocation of glucose transporters to the plasma membrane due to a depleted intracellular pool. In contrast, the insulin hyperresponsive glucose transport observed with refeeding appears to result from (a) a restored translocation of glucose transporters to the plasma membrane from a repleted intracellular pool and (b) enhanced plasma membrane glucose transporter intrinsic activity.

3-O-Methylglucose↗
PubMed1988

Factitious elevation of thyrotropin in a new ultrasensitive assay: implications for the use of monoclonal antibodies in "sandwich" immunoassay.

Three patients who had falsely elevated serum TSH concentrations (initial values, 30.5, 74, and greater than 50 mU/L) in a mouse monoclonal immunoradiometric assay are reported. Two patients were treated for hypothyroidism inappropriately, and one underwent unnecessary diagnostic testing. Immunoaffinity chromatography of serum from one patient indicated that the serum TSH level was truly low. Addition of mouse serum or immunoglobulin G (IgG) or absorption of patient serum with solid phase-bound mouse IgG-1 reduced the TSH content in the serum of the three patients to undetectable levels. Blocking studies revealed that all patients had antibodies directed at mouse IgG-1, the subclass of mouse antibody present in the assay kit. The serum of one patient who had autoimmune disease with elevated serum Igs had much broader species cross-reactivity than that of another patient who had known exposure to rats and mice. We hypothesize that such antimouse antibodies can arise either from endogenous autoimmunity or exogenous animal exposure. Serum TSH elevations also were found when the serum samples were tested in other mouse monoclonal immunoassays, underscoring the fact that antibody interference can potentially affect many assays used in endocrinology and other areas of medicine to make major diagnostic and therapeutic decisions. Clinicians must be aware of such interactions; relatively simple laboratory maneuvers can differentiate true from false results in assays of this type.

Adult↗
PubMed1987

Mechanism for markedly hyperresponsive insulin-stimulated glucose transport activity in adipose cells from insulin-treated streptozotocin diabetic rats. Evidence for increased glucose transporter intrinsic activity.

The effects of insulin therapy in streptozotocin diabetic rats on the glucose transport response to insulin in adipose cells have been examined. At sequential intervals during subcutaneous insulin infusion, isolated cells were prepared and incubated with or without insulin, and 3-O-methylglucose transport was measured. Insulin treatment not only reversed the insulin-resistant glucose transport associated with diabetes, but resulted in a progressive hyperresponsiveness, peaking with a 3-fold overshoot at 7-8 days (12.1 +/- 0.3 versus 3.4 +/- 0.1 fmol/cell/min, mean +/- S.E.) and remaining elevated for more than 3 weeks. During the peak overshoot, glucose transporters in subcellular membrane fractions were assessed by cytochalasin B binding. Insulin therapy restored glucose transporter concentration in the plasma membranes of insulin-stimulated cells from a 40% depleted level previously reported in the diabetic state to approximately 35% greater than control (38 +/- 4 versus 28 +/- 2 pmol/mg of membrane protein). Glucose transporter concentration in the low-density microsomes from basal cells was also restored from an approximately 45% depleted level back to normal (50 +/- 4 versus 50 +/- 6 pmol/mg of membrane protein), whereas total intracellular glucose transporters were further increased due to an approximately 2-fold increase in low-density microsomal membrane protein. However, these increases remained markedly less than the enhancement of insulin-stimulated glucose transport activity in the intact cell. Thus, insulin treatment of diabetic rats produces a marked and sustained hyperresponsive insulin-stimulated glucose transport activity in the adipose cell with little more than a restoration to the non-diabetic control level of glucose transporter translocation. Because this enhanced glucose transport activity occurs through an increase in Vmax, insulin therapy appears to be associated with a marked increase in glucose transporter intrinsic activity.

3-O-Methylglucose↗
PubMed1987

Mechanism for enhanced glucose transport response to insulin in adipose cells from chronically hyperinsulinemic rats. Increased translocation of glucose transporters from an enlarged intracellular pool.

The mechanism for the increased glucose transport response to insulin in adipose cells from chronically hyperinsulinemic rats was examined. Rats were infused with insulin s.c. for 2 wk. Isolated adipose cells were incubated with and without insulin, 3-O-methylglucose transport was measured, and glucose transporters in subcellular membrane fractions were assessed by cytochalasin B binding. Adipose cells from insulin-treated rats showed no change in basal but a 55% increase in insulin-stimulated glucose transport activity compared with those from control rats (7.1 +/- 0.8 vs. 4.6 +/- 0.5 fmol/cell per min, mean +/- SEM) and a corresponding increase in the concentration of glucose transporters in the plasma membranes (44 +/- 5 vs. 32 +/- 6 pmol/mg of membrane protein). In the low-density microsomes, glucose transporter concentrations in both basal and insulin-stimulated states were the same, but the total numbers were greater in cells from the insulin-treated rats because of a 39% increase in low-density microsomal protein. Therefore, chronic experimental hyperinsulinemia in the rat enhances the stimulatory action of insulin on glucose transport in the adipose cell by increasing the concentration of glucose transporters in the plasma membranes. This results from an enlarged intracellular pool due to increased intracellular protein and enhanced glucose transporter translocation in response to insulin.

3-O-Methylglucose↗
PubMed1987

Interference by anti-immunoglobulin G antibodies in immunoradiometric assays of thyrotropin involving mouse monoclonal antibodies.

"Sandwich"-type assays are subject to positive interference by the patient's "heterophile" antibodies. If present, these bind to the animal immunoglobulins in the assay reagents, forming artefactual sandwiches indistinguishable from those formed with the analyte itself. Immunoglobulins from non-immunized animals, added to the assay reagents, can diminish this effect by blocking the patient's antibodies. Elsewhere, we studied several patients with anti-mouse immunoglobulin activity, whose serum gave spuriously high results for thyrotropin (TSH) concentrations. Here we have studied this phenomenon by adding, to pooled zero-TSH serum, antibodies to mouse, goat, and horse immunoglobulins and then assaying TSH by several other sandwich-type assays involving mouse monoclonal antibodies. Assays not supplemented with blocking immunoglobulins from mice or other animals were more susceptible to this effect. When large amounts of antibody were added, the antibody excess diminished the interference. However, the presence of blocking immunoglobulins could reverse such antibody excess, actually enhancing, instead of diminishing, the positive interference. Users should be aware that blocking immunoglobulins may diminish but not necessarily eliminate this problem with such assays.

Animals↗
PubMed1985

Subcellular translocation of glucose transporters: role in insulin action and its perturbation in altered metabolic states.

In this article we have described the hypothesis that insulin stimulates glucose transport through glucose transporter translocation from an intracellular pool to the plasma membrane. In addition, we have shown that changes in the numbers and subcellular distributions of glucose transporters correlate with alterations in insulin-stimulated glucose transport activity in several experimental models of insulin resistance and hyperresponsiveness. However, in experiments with counterregulatory hormones and with hyperresponsive states induced by nutritional repletion following deprivation, changes in insulin responsiveness cannot be fully explained by such alterations in the numbers and/or subcellular distribution of glucose transporters. Thus, evidence has been presented for changes in glucose transporter intrinsic activity that both inhibit and augment insulin-stimulated glucose transport rates. Finally, we have discussed data suggesting that the translocation process is applicable to human tissue and that significant changes in adipose cell glucose transport activity have been correlated with total glucose disposal in various metabolic states in humans. Determining the physiologic factors involved in modulating these events at the cellular level is an important area for further investigation.

Adipose Tissue↗
PubMedDate not supplied

Determination of acetochlor in technical and formulated products by capillary gas chromatography: PVM 5:1999.

A peer-verified gas chromatographic (GC) method is presented for the weight percent (wt %) determination of acetochlor herbicide in technical and formulated products. During method development, the method was found to be rugged by the Youden Ruggedness Test. Two laboratories with experience in the wt % determination of acetochlor in various matrixes participated in this study. Each laboratory received 10 blind duplicate test samples of the following 5 matrixes: one acetochlor technical and 4, different, emulsifiable concentrate (EC) formulations--Harness EC (MON 5841), Harness Export/Fist (MON 8435), Surpass EC (HF), and Surpass EC (LF). Each participant was asked to make duplicate weighings of each of the test samples and to inject each test sample solution twice. All test samples were analyzed on the same day, and 8 data points (replicates) per matrix were obtained. The test samples were dissolved in acetone that contained dipentyl phthalate as an internal standard. They were analyzed by GC on a 15 m capillary column by using split injection and a flame ionization detector. Acetochlor (wt %) was determined by comparing the ratios of peak area of acetochlor/peak area of dipentyl phthalate internal standard obtained for the test sample and calibration solutions. Repeatability of the method, expressed as the within-laboratory (between replicates) relative standard deviation (RSDr), was found to be 0.09-0.77% for the 5 matrixes. Reproducibility of the method, expressed as the within-test sample relative standard deviation (RSDR), was found to be 0.18-0.78% for the

Algorithms↗