Effects of raising muscle glycogen synthesis rate on skeletal muscle ATP turnover rate in type 2 diabetes

Mitochondrial dysfunction has been implicated in the pathogenesis of type 2 diabetes. We hypothesized that any impairment in insulin-stimulated muscle ATP production could merely reflect the lower rates of muscle glucose uptake and glycogen synthesis, rather than cause it. If this is correct, muscle ATP turnover rates in type 2 diabetes could be increased if glycogen synthesis rates were normalized by the mass-action effect of hyperglycemia. Isoglycemic- and hyperglycemic-hyperinsulinemic clamps were performed on type 2 diabetic subjects and matched controls, with muscle ATP turnover and glycogen synthesis rates measured using (31)P- and (13)C-magnetic resonance spectroscopy, respectively. In diabetic subjects, hyperglycemia increased muscle glycogen synthesis rates to the level observed in controls at isoglycemia [from 19 ± 9 to 41 ± 12 μmol·l(-1)·min(-1) (P = 0.012) vs. 40 ± 7 μmol·l(-1)·min(-1) in controls]. This was accompanied by a modest increase in muscle ATP turnover rates (7.1 ± 0.5 vs. 8.6 ± 0.7 μmol·l(-1)·min(-1), P = 0.04). In controls, hyperglycemia brought about a 2.5-fold increase in glycogen synthesis rates (100 ± 24 vs. 40 ± 7 μmol·l(-1)·min(-1), P = 0.028) and a 23% increase in ATP turnover rates (8.1 ± 0.9 vs. 10.0 ± 0.9 μmol·l(-1)·min(-1), P = 0.025) from basal state. Muscle ATP turnover rates correlated positively with glycogen synthesis rates (r(s) = 0.46, P = 0.005). Changing the rate of muscle glucose metabolism in type 2 diabetic subjects alters demand for ATP synthesis at rest. In type 2 diabetes, skeletal muscle ATP turnover rates reflect the rate of glucose uptake and glycogen synthesis, rather than any primary mitochondrial defect.     

Muscle cellular properties in the ice hockey player: a model for investigating overtraining?

In this study, we hypothesized that athletes involved in 5-6 months of sprint-type training would display higher levels of proteins and processes involved in muscle energy supply and utilization. Tissue was sampled from the vastus lateralis of 13 elite ice hockey players (peak oxygen consumption = 51.8 ± 1.3 mL·kg(-1)·min(-1); mean ± standard error) at the end of a season (POST) and compared with samples from 8 controls (peak oxygen consumption = 45.5 ± 1.4 mL·kg(-1)·min(-1)) (CON). Compared with CON, higher activities were observed in POST (p < 0.05) only for succinic dehydrogenase (3.32 ± 0.16 mol·(mg protein)(-1)·min(-1) vs. 4.10 ± 0.11 mol·(mg protein)(-1)·min(-1)) and hexokinase (0.73 ± 0.05 mol·(mg protein)(-1)·min(-1) vs. 0.90 ± 0.05mol·(mg protein)(-1)·min(-1)) but not for phosphorylase, phosphofructokinase, and creatine phosphokinase. No differences were found in Na(+),K(+)-ATPase concentration (β(max): 262 ± 36 pmol·(g wet weight)(-1) vs. 275 ± 27 pmol·(g wet weight)(-1)) and the maximal activity of the sarcoplasmic reticulum Ca(2+)-ATPase (98.1 ± 6.1 μmol·(g protein)(-1)·min(-1) vs. 102 ± 3.3 μmol·(g protein)(-1)·min(-1)). Cross-sectional area was lower (p < 0.05) in POST but only for the type IIA fibres (6312 ± 684 μm(2) vs. 5512 ± 335 μm(2)), while the number of capillary counts per fibre and the capillary to fibre area ratio were generally higher (p < 0.05). These findings suggest that elite trained ice hockey players display elevations only in support of glucose-based aerobic metabolism that occur in the absence of alterations in excitation-contraction processes.     

Effect of hyperinsulinemia on amino acid utilization and oxidation independent of glucose metabolism in the ovine fetus

We studied the effect of acute hyperinsulinemia on amino acid (AA) utilization and oxidation rates independent of insulin-enhanced glucose metabolism in fetal sheep. Metabolic studies were conducted in each fetus (n = 11) under three experimental periods. After control period (C) study, a fetal hyperinsulinemic-euglycemic-euaminoacidemic (HI-euG-euAA) clamp was established, followed by a hyperinsulinemic-hypoglycemic-euaminoacidemic (HI-hypoG-euAA) clamp to decrease glucose metabolic rates toward C values. Infusions of (3)H(2)0, L-[1-(13)C]leucine, and [(14)C(U)]glucose were administered to measure blood flow, leucine oxidation, and fetal glucose uptake, utilization, and oxidation in each period. Fetal glucose utilization rate increased 1.7-fold with hyperinsulinemia (C 5.8 +/- 0.8 mg.kg(-1).min(-1), HI-euG-euAA 10 +/- 1.3 mg.kg(-1).min(-1), P < 0.0001), returning to rates not different from C with hypoglycemia (HI-hypoG-euAA 7.1 +/- 0.9 mg.kg(-1).min(-1) vs. C value, P = 0.15). Fetal glucose oxidation rate increased 1.7-fold with hyperinsulinemia (C 3.1 +/- 0.2 mg.kg(-1).min(-1), HI-euG-euAA 5.4 +/- 0.4 mg.kg(-1).min(-1), P < 0.0001) and decreased to near control rates with hypoglycemia (4.0 +/- 0.3 HI-hypoG-euAA vs. C value, P = 0.006). AA utilization rates increased with hyperinsulinemia for all essential and most nonessential AAs (P < 0.001) and did not change when insulin-induced increases in glucose utilization returned to control rates. Leucine oxidation rate increased 1.7-fold with hyperinsulinemia (C 1.0 +/- 0.3 micromol.min(-1).kg(-1), HI-euG-euAA 1.7 +/- 0.3 micromol.min(-1).kg(-1), P < 0.002) and did not change when glucose oxidation rate was decreased with hypoglycemia. These results demonstrate that, in fetal sheep, insulin promotes AA utilization and oxidation independent of its simultaneous effects on glucose metabolism. In acute hyperinsulinemic conditions, AA oxidation does not change when insulin-induced glucose utilization is prevented.     

Cardiac function following prolonged exercise: influence of age

This study sought to determine the influence of age on the left ventricular (LV) response to prolonged exercise (PE; 150 min). LV systolic and diastolic performance was assessed using echocardiography (ECHO) before (pre) and 60 min following (post) exercise performed at 80% maximal aerobic power in young (28 ± 4.5 years; n = 18; mean ± SD) and middle-aged (52 ± 3.9 years; n = 18) participants. LV performance was assessed using two-dimensional ECHO, including speckle-tracking imaging, to determine LV strain (LV S) and LV S rate (LV SR), in addition to Doppler measures of diastolic function. We observed a postexercise elevation in LV S (young: -19.5 ± 2.1% vs. -21.6 ± 2.1%; middle-aged: -19.9 ± 2.3% vs. -20.8 ± 2.1%; P < 0.05) and LV SR (young: -1.19 ± 0.1 vs. -1.37 ± 0.2; middle-aged: -1.20 ± 0.2 vs. -1.38 ± 0.2; P < 0.05) during recovery in both groups. Diastolic function was reduced during recovery, including the LV SR ratio of early-to-late atrial diastolic filling (SR(e/a)), in young (2.35 ± 0.7 vs. 1.89 ± 0.5; P < 0.01) and middle-aged (1.51 ± 0.5 vs. 1.05 ± 0.2; P < 0.01) participants, as were conventional indices including the E/A ratio. Dobutamine stress ECHO revealed a postexercise depression in LV S in response to increasing dobutamine dose, which was similar in both young (pre-exercise dobutamine 0 vs. 20 μg·kg(-1)·min(-1): -19.5 ± 2.1 vs. -27.2 ± 2.2%; postexercise dobutamine 0 vs. 20 μg·kg(-1)·min(-1): -21.6 ± 2.1 vs. -23.7 ± 2.2%; P < 0.05) and middle-aged participants (pre: -19.9 ± 2.3 vs. -25.3 ± 2.7%; post: -20.8 ± 2.1 vs. -23.5 ± 2.7; P < 0.05). This was despite higher noradrenaline concentrations immediately postexercise in the middle-aged participants compared with young (4.26 ± 2.7 nmol/L vs. 3.00 ± 1.4 nmol/L; P = 0.12). These data indicate that LV dysfunction is observed following PE and that advancing age does not increase the magnitude of this response.     

Evidence that renal arginine transport is impaired in spontaneously hypertensive rats

Low renal nitric oxide (NO) bioavailability contributes to the development and maintenance of chronic hypertension. We investigated whether impaired l-arginine transport contributes to low renal NO bioavailability in hypertension. Responses of renal medullary perfusion and NO concentration to renal arterial infusions of the l-arginine transport inhibitor l-lysine (10 μmol·kg(-1)·min(-1); 30 min) and subsequent superimposition of l-arginine (100 μmol·kg(-1)·min(-1); 30 min), the NO synthase inhibitor N(G)-nitro-l-arginine (2.4 mg/kg; iv bolus), and the NO donor sodium nitroprusside (0.24 μg·kg(-1)·min(-1)) were examined in Sprague-Dawley rats (SD) and spontaneously hypertensive rats (SHR). Renal medullary perfusion and NO concentration were measured by laser-Doppler flowmetry and polarographically, respectively, 5.5 mm below the kidney surface. Renal medullary NO concentration was less in SHR (53 ± 3 nM) compared with SD rats (108 ± 12 nM; P = 0.004). l-Lysine tended to reduce medullary perfusion (-15 ± 7%; P = 0.07) and reduced medullary NO concentration (-9 ± 3%; P = 0.03) while subsequent superimposition of l-arginine reversed these effects of l-lysine in SD rats. In SHR, l-lysine and subsequent superimposition of l-arginine did not significantly alter medullary perfusion or NO concentration. Collectively, these data suggest that renal l-arginine transport is impaired in SHR. Renal l-[(3)H]arginine transport was less in SHR compared with SD rats (P = 0.01). Accordingly, we conclude that impaired arginine transport contributes to low renal NO bioavailability observed in the SHR kidney.     

Glucose uptake during centrally induced stress is insulin independent and enhanced by adrenergic blockade.

Glucose utilization increases markedly in the normal dog during stress induced by the intracerebroventricular (ICV) injection of carbachol. To determine the extent to which insulin, glucagon, and selective (alpha/beta)-adrenergic activation mediate the increment in glucose metabolic clearance rate (MCR) and glucose production (R(a)), we used five groups of normal mongrel dogs: 1) pancreatic clamp (PC; n = 7) with peripheral somatostatin (0.8 microg x kg(-1) x min(-1)) and intraportal replacement of insulin (1,482 +/- 84 pmol x kg(-1) x min(-1)) and glucagon (0.65 ng x kg(-1) x min(-1)) infusions; 2) PC plus combined alpha (phentolamine)- and beta (propranolol)-blockade (7 and 5 microg x kg(-1) x min(-1), respectively; alpha+beta; n = 5); 3) PC plus alpha-blockade (alpha; n = 6); 4) PC plus beta-blockade (beta; n = 5); and 5) a carbachol control group without PC (Con; n = 10). During ICV carbachol stress (0-120 min), catecholamines, ACTH, and cortisol increased in all groups. Baseline insulin and glucagon levels were maintained in all groups except Con, where glucagon rose 33%, and alpha, where insulin increased slightly but significantly. Stress increased (P < 0.05) plasma glucose in Con, PC, and alpha but decreased it in beta and alpha+beta. The MCR increment was greater (P < 0.05) in beta and alpha+beta than in Con, PC, and alpha. R(a) increased (P < 0.05) in all groups but was attenuated in alpha+beta. Stress-induced lipolysis was abolished in beta (P < 0.05). The marked rise in lactate in Con, PC, and alpha was abolished in alpha+beta and beta. We conclude that the stress-induced increase in MCR is largely independent of changes in insulin, markedly augmented by beta-blockade, and related, at least in part, to inhibition of lipolysis and glycogenolysis, and that R(a) is augmented by glucagon and alpha- and beta-catecholamine effects.     

Two weeks of muscle immobilization impairs functional sympatholysis but increases exercise hyperemia and the vasodilatory responsiveness to infused ATP

During exercise, contracting muscles can override sympathetic vasoconstrictor activity (functional sympatholysis). ATP and adenosine have been proposed to play a role in skeletal muscle blood flow regulation. However, little is known about the role of muscle training status on functional sympatholysis and ATP- and adenosine-induced vasodilation. Eight male subjects (22 ± 2 yr, Vo(2max): 49 ± 2 ml O(2)·min(-1)·kg(-1)) were studied before and after 5 wk of one-legged knee-extensor training (3-4 times/wk) and 2 wk of immobilization of the other leg. Leg hemodynamics were measured at rest, during exercise (24 ± 4 watts), and during arterial ATP (0.94 ± 0.03 μmol/min) and adenosine (5.61 ± 0.03 μmol/min) infusion with and without coinfusion of tyramine (11.11 μmol/min). During exercise, leg blood flow (LBF) was lower in the trained leg (2.5 ± 0.1 l/min) compared with the control leg (2.6 ± 0.2 l/min; P < 0.05), and it was higher in the immobilized leg (2.9 ± 0.2 l/min; P < 0.05). Tyramine infusion lowers LBF similarly at rest, but, when tyramine was infused during exercise, LBF was blunted in the immobilized leg (2.5 ± 0.2 l/min; P < 0.05), whereas it was unchanged in the control and trained leg. Mean arterial pressure was lower during exercise with the trained leg compared with the immobilized leg (P < 0.05), and leg vascular conductance was similar. During ATP infusion, the LBF response was higher after immobilization (3.9 ± 0.3 and 4.5 ± 0.6 l/min in the control and immobilized leg, respectively; P < 0.05), whereas it did not change after training. When tyramine was coinfused with ATP, LBF was reduced in the immobilized leg (P < 0.05) but remained similar in the control and trained leg. Training increased skeletal muscle P2Y2 receptor content (P < 0.05), whereas it did not change with immobilization. These results suggest that muscle inactivity impairs functional sympatholysis and that the magnitude of hyperemia and blood pressure response to exercise is dependent on the training status of the muscle. Immobilization also increases the vasodilatory response to infused ATP.     

Mesenchymal stem cell transplantation for the infarcted heart: a role in minimizing abnormalities in cardiac-specific energy metabolism

Intense interest has been focused on cell-based therapy for the infarcted heart given that stem cells have exhibited the ability to reduce infarct size and mitigate cardiac dysfunction. Despite this, it is unknown whether mesenchymal stem cell (MSC) therapy can prevent metabolic remodeling following a myocardial infarction (MI). This study examines the ability of MSCs to rescue the infarcted heart from perturbed substrate uptake in vivo. C57BL/6 mice underwent chronic ligation of the left anterior descending coronary artery to induce a MI. Echocardiography was performed on conscious mice at baseline as well as 7 and 23 days post-MI. Twenty-eight days following the ligation procedure, hyperinsulinemic euglycemic clamps assessed in vivo insulin sensitivity. Isotopic tracer administration evaluated whole body, peripheral tissue, and cardiac-specific glucose and fatty acid utilization. To gain insight into the mechanisms by which MSCs modulate metabolism, mitochondrial function was assessed by high-resolution respirometry using permeabilized cardiac fibers. Data show that MSC transplantation preserves insulin-stimulated fatty acid uptake in the peri-infarct region (4.25 ± 0.64 vs. 2.57 ± 0.34 vs. 3.89 ± 0.54 μmol·100 g(-1)·min(-1), SHAM vs. MI + PBS vs. MI + MSC; P < 0.05) and prevents increases in glucose uptake in the remote left ventricle (3.11 ± 0.43 vs. 3.81 ± 0.79 vs. 6.36 ± 1.08 μmol·100 g(-1)·min(-1), SHAM vs. MI + PBS vs. MI + MSC; P < 0.05). This was associated with an enhanced efficiency of mitochondrial oxidative phosphorylation with a respiratory control ratio of 3.36 ± 0.18 in MSC-treated cardiac fibers vs. 2.57 ± 0.14 in the infarct-only fibers (P < 0.05). In conclusion, MSC therapy exhibits the potential to rescue the heart from metabolic aberrations following a MI. Restoration of metabolic flexibility is important given the metabolic demands of the heart and the role of energetics in the progression to heart failure.     

Effect of hyperinsulinemia on amino acid utilization in the ovine fetus

We studied the effect of an acute 4-h period of hyperinsulinemia (H) on net utilization rates (AAUR(net)) of 21 amino acids (AA) in 17 studies performed in 13 late-gestation fetal sheep by use of a novel fetal hyperinsulinemic-euglycemic-euaminoacidemic clamp. During H [84 +/- 12 (SE) microU/ml H, 15 +/- 2 microU/ml control (C), P < 0. 00001], euglycemia was maintained by glucose clamp (19 +/- 0.05 micromol/ml H, 1.19 +/- 0.04 micromol/ml C), and euaminoacidemia (mean 4.1 +/- 3.3% increase for all amino acid concentrations [AA], nonsignificantly different from zero) was maintained with a mixed amino acid solution adjusted to keep lysine concentration constant and other [AA] near C values. H produced a 63.7% increase in AAUR(net) (3.29 +/- 0.66 micromol. min(-1). kg(-1) H, 2.01 +/- 0.55 micromol. min(-1). kg(-1) C, P < 0.001), accounting for a 60.1% increase in fetal nitrogen uptake rate (2,064 +/- 108 mg. day(-1). kg(-1) H, 1,289 +/- 73 mg. day(-1). kg(-1) C, P < 0.001). Mean AA clearance rate (AAUR(net)/[AA]) increased by 64.5 +/- 18.9% (P < 0. 001). Thus acute physiological H increases net amino acid and nitrogen utilization rates in the ovine fetus independent of plasma glucose and [AA].     

Transaldolase exchange and its effects on measurements of gluconeogenesis in humans

The deuterated water method is used extensively to measure gluconeogenesis in humans. This method assumes negligible exchange of the lower three carbons of fructose 6-phsophate via transaldolase exchange since this exchange will result in enrichment of carbon 5 of glucose in the absence of net gluconeogenesis. The present studies tested this assumption. 2H?O and acetaminophen were ingested and [1-13C]acetate infused in 11 nondiabetic subjects after a 16-h fast. Plasma and urinary glucuronide enrichments were measured using nuclear magnetic resonance spectroscopy before and during a 0.35 mU·kg FFM?1·min?1 insulin infusion. Rates of endogenous glucose production measured with [3-3H]- and [6,6-2H?]glucose did not differ either before (14.0 ± 0.7 vs. 13.8 ± 0.7 μmol·kg?1·min?1) or during the clamp (10.4 ± 0.9 vs. 10.9 ± 0.7 μmol·kg?1·min?1), consistent with equilibration and quantitative removal of tritium during triose isomerase exchange. Plasma [3-13C] glucose-to-[4-13C]glucose and urinary [3-13C] glucuronide-to-[4-13C]glucuronide ratios were <1.0 (P < 0.001) in all subjects both before (0.66 ± 0.04 and 0.60 ± 0.04) and during (059 ± 0.05 and 0.56 ± 0.06) the insulin infusion, respectively, indicating that ～35-45% of the labeling of the 5th carbon of glucose by deuterium was due to transaldolase exchange rather than gluconeogenesis. When corrected for transaldolase exchange, rates of gluconeogenesis were lower (P < 0.001) and glycogenolysis higher (P < 0.001) than uncorrected rates both before and during the insulin infusion. In conclusion, assuming negligible dilution by glycerol and near-complete triose isomerase equilibration, these data provide strong experimental evidence that transaldolase exchange occurs in humans, resulting in an overestimate of gluconeogenesis and an underestimate of glycogenolysis when measured with the 2H?O method. Use of appropriate 13C tracers provides a means of correcting for transaldolase exchange.     

Osmotic regulation of prolactin secretion in the fetal sheep

The functions of prolactin in the fetus remain speculative. No obvious physiological role has been found for the prolactin present in the fetal or maternal plasma and amniotic fluid compartments. The aim of the present study was to investigate changes in fetal plasma prolactin following intracerebroventricular (i.c.r.) administration to the fetus of artificial cerebrospinal fluid of different tonicities. A lateral ventricle catheter was placed in 11 fetuses at 107-128 days of gestation. Either isotonic artificial cerebrospinal fluid (300 mOsm.1(-1);n = 9), 15% polyethylene glycol (340 mOsm.1(-1);n = 5), or 7% distilled water in isotonic artificial cerebrospinal fluid (270 mOsm.1(-1);n = 9) was infused i.c.v. at 35 mu1.min-1 for 240 min. At 180 min thyrotropin releasing hormone (TRH) was administered through a fetal a fetal jugular catheter. Fetal carotid arterial blood gases, plasma osmolality and concentrations of prolactin, vasopressin (AVP), and norepinephrine (NE) were measured. Administration of hypotonic artificial cerebrospinal fluid produced an increase in fetal plasma prolactin from 46.6 +/- 36 ng.ml-1 at 0 min to 83.3 +/- 49 ng.ml-1 at 180 min (mean +/- SEM; P less than 0.05). No changes in either AVP or NE were observed. Administration of hypertonic artificial cerebrospinal fluid produced a decrease in plasma prolactin from 85 +/- 57 at time 0 to 49 +/- 35 at 180 min (P less than 0.05). No changes in either AVP or NE were observed. Fetal plasma prolactin, AVP, and NE did not change during control infusion of isotonic artificial cerebrospinal fluid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of renal caveolin-1 reduces natriuresis and produces hypertension in sodium-loaded rats

Renal dopamine receptor function and ion transport inhibition are impaired in essential hypertension. We recently reported that caveolin-1 (CAV1) and lipid rafts are necessary for normal D(1)-like receptor-dependent internalization of Na-K-ATPase in human proximal tubule cells. We now hypothesize that CAV1 is necessary for the regulation of urine sodium (Na(+)) excretion (U(Na)V) and mean arterial blood pressure (MAP) in vivo. Acute renal interstitial (RI) infusion into Sprague-Dawley rats of 1 μg·kg?1·min?1 fenoldopam (FEN; D(1)-like receptor agonist) caused a 0.46 ± 0.15-μmol/min increase in U(Na)V (over baseline of 0.29 ± 0.04 μmol/min; P < 0.01). This increase was seen in Na(+)-loaded rats, but not in those under a normal-sodium load. Coinfusion with β-methyl cyclodextrin (βMCD; lipid raft disrupter, 200 μg·kg?1·min?1) completely blocked this FEN-induced natriuresis (P < 0.001). Long-term (3 day) lipid raft disruption via continuous RI infusion of 80 μg·kg?1·min?1 βMCD decreased renal cortical CAV1 expression (47.3 ± 6.4%; P < 0.01) and increased MAP (32.4 ± 6.6 mmHg; P < 0.001) compared with vehicle-infused animals. To determine whether the MAP rise was due to a CAV1-dependent lipid raft-mediated disruption, Na(+)-loaded rats were given a bolus RI infusion of CAV1 siRNA. Two days postinfusion, cortical CAV1 expression was decreased by 73.6 ± 8.2% (P < 0.001) and the animals showed an increase in MAP by 17.4 ± 2.9 mmHg (P < 0.01) compared with animals receiving scrambled control siRNA. In summary, acute kidney-specific lipid raft disruption decreases CAV1 expression and blocks D(1)-like receptor-induced natriuresis. Furthermore, chronic disruption of lipid rafts or CAV1 protein expression in the kidney induces hypertension.     

Validity of triple- and dual-tracer techniques to estimate glucose appearance

The triple-tracer (TT) dilution technique has been proposed to be the gold standard method to measure postprandial glucose appearance. However, validation against an independent standard has been missing. We addressed this issue and also validated the simpler dual-tracer (DT) technique. Sixteen young subjects with type 1 diabetes (age 19.5 ± 3.8 yr, BMI 23.4 ± 1.5 kg/m(2), HbA(1c) 8.7 ± 1.7%, diabetes duration 9.0 ± 6.9 yr, total daily insulin 0.9 ± 0.2 U·kg(-1)·day(-1), mean ± SD) received a variable intravenous 20% dextrose infusion enriched with [U-(13)C]glucose over 8 h to achieve postprandial-resembling glucose excursions while intravenous insulin was administered to achieve postprandial-resembling levels of plasma insulin. Primed [6,6-(2)H(2)]glucose was infused in a manner that mimicked the expected endogenous glucose production and [U-(13)C; 1,2,3,4,5,6,6-(2)H(7)]glucose was infused in a manner that mimicked the expected glucose appearance from a standard meal. Plasma glucose enrichment was measured by gas chromatography-mass spectrometry. The intravenous dextrose infusion served as an independent standard and was reconstructed using the TT and DT techniques with the two-compartment Radziuk/Mari model and an advanced stochastic computational method. The difference between the infused and reconstructed dextrose profile was similar for the two methods (root mean square error 6.6 ± 1.9 vs. 8.0 ± 3.5 μmol·kg(-1)·min(-1), TT vs. DT, P = NS, paired t-test). The TT technique was more accurate in recovering the overall dextrose infusion (100 ± 9 and 92 ± 12%; P = 0.02). The root mean square error associated with the mean dextrose infusion profile was 2.5 and 3.3 μmol·kg(-1)·min(-1) for the TT and DT techniques, respectively. We conclude that the TT and DT techniques combined with the advanced computational method can measure accurately exogenous glucose appearance. The TT technique tends to outperform slightly the DT technique, but the latter benefits from reduced experimental and computational complexity.     

Acute exercise and training alter blood lipid and lipoprotein profiles differently in overweight and obese men and women

Our purpose was to elucidate effects of acute exercise and training on blood lipids-lipoproteins, and high-sensitivity C-reactive protein (hsCRP) in overweight/obese men (n = 10) and women (n = 8); age, BMI, body fat percentage, and VO(2)max were (mean ± SEM): 45 ± 2.5 years, 31.9 ± 1.4 kg·m(-2), 41.1 ± 1.5%, and 25.2 ± 1.3 mlO(2)·kg(-1)·min(-1). Before exercise training subjects performed an acute exercise session on a treadmill (70% VO(2)max, 400 kcal energy expenditure), followed by 12 weeks of endurance exercise training (land-based or aquatic-based treadmill): 3 sessions·week(-1), progressing to 500 kcal·session(-1) during which subjects maintained accustomed dietary habits. After training, the acute exercise session was repeated. Blood samples, obtained immediately before and 24 h after acute exercise sessions, were analyzed for serum lipids, lipoproteins, and hsCRP adjusted for plasma volume shifts. Exercise training increased VO(2)max (+3.67 mlO(2)·kg(-1)·min(-1), P < 0.001) and reduced body weight (-2.7 kg, P < 0.01). Training increased high-density lipoprotein (HDL) and HDL(2b)-cholesterol (HDL-C) concentrations (+3.7 and +2.4 mg·dl(-1), P < 0.05) and particle numbers (+588 and +206 nmol·l(-1), P < 0.05) in men. In women despite no change in total HDL-C, subfractions shifted from HDL(3)-C (-3.2, P < 0.01) to HDL(2b)-C (+3.5, P < 0.05) and HDL(2a)-C (+2.2 mg·dl(-1), P < 0.05), with increased HDL(2b) particle number (+313 nmol·l(-1), P < 0.05). Training reduced LDL(3) concentration and particle number in women (-1.6 mg·dl(-1) and -16 nmol·l(-1), P < 0.05). Acute exercise reduced the total cholesterol (TC): HDL-C ratio in men (-0.16, P < 0.01) and increased hsCRP in all subjects (+0.05 mg·dl(-1), P < 0.05), regardless of training. Training did not affect acute exercise responses. Our data support the efficacy of endurance training, without dietary intervention, to elicit beneficial changes in blood lipids-lipoproteins in obese men and women.     

Coenzyme Q(1) as a probe for mitochondrial complex I activity in the intact perfused hyperoxia-exposed wild-type and Nqo1-null mouse lung

Previous studies showed that coenzyme Q(1) (CoQ(1)) reduction on passage through the rat pulmonary circulation was catalyzed by NAD(P)H:quinone oxidoreductase 1 (NQO1) and mitochondrial complex I, but that NQO1 genotype was not a factor in CoQ(1) reduction on passage through the mouse lung. The aim of the present study was to evaluate the complex I contribution to CoQ(1) reduction in the isolated perfused wild-type (NQO1(+/+)) and Nqo1-null (NQO1(-)/(-)) mouse lung. CoQ(1) reduction was measured as the steady-state pulmonary venous CoQ(1) hydroquinone (CoQ(1)H(2)) efflux rate during infusion of CoQ(1) into the pulmonary arterial inflow. CoQ(1)H(2) efflux rates during infusion of 50 μM CoQ(1) were not significantly different for NQO1(+/+) and NQO1(-/-) lungs (0.80 ± 0.03 and 0.68 ± 0.07 μmol·min(-1)·g lung dry wt(-1), respectively, P > 0.05). The mitochondrial complex I inhibitor rotenone depressed CoQ(1)H(2) efflux rates for both genotypes (0.19 ± 0.08 and 0.08 ± 0.04 μmol·min(-1)·g lung dry wt(-1) for NQO1(+/+) and NQO1(-/-), respectively, P < 0.05). Exposure of mice to 100% O(2) for 48 h also depressed CoQ(1)H(2) efflux rates in NQO1(+/+) and NQO1(-/-) lungs (0.43 ± 0.03 and 0.11 ± 0.04 μmol·min(-1)·g lung dry wt(-1), respectively, P < 0.05 by ANOVA). The impact of rotenone or hyperoxia on CoQ(1) redox metabolism could not be attributed to effects on lung wet-to-dry weight ratios, perfusion pressures, perfused surface areas, or total venous effluent CoQ(1) recoveries, the latter measured by spectrophotometry or mass spectrometry. Complex I activity in mitochondria-enriched lung fractions was depressed in hyperoxia-exposed lungs for both genotypes. This study provides new evidence for the potential utility of CoQ(1) as a nondestructive indicator of the impact of pharmacological or pathological exposures on complex I activity in the intact perfused mouse lung.     

Ursodeoxycholate modulates bile flow and bile salt pool independently from the cystic fibrosis transmembrane regulator (Cftr) in mice

Cystic fibrosis liver disease (CFLD) is treated with ursodeoxycholate (UDCA). Our aim was to evaluate, in cystic fibrosis transmembrane regulator knockout (Cftr(-/-)) mice and wild-type controls, whether the supposed therapeutic action of UDCA is mediated via choleretic activity or effects on bile salt metabolism. Cftr(-/-) mice and controls, under general anesthesia, were intravenously infused with tauroursodeoxycholate (TUDCA) in increasing dosage or were fed either standard or UDCA-enriched chow (0.5% wt/wt) for 3 wk. Bile flow and bile composition were characterized. In chow-fed mice, we analyzed bile salt synthesis and pool size of cholate (CA). In both Cftr(-/-) and controls intravenous TUDCA stimulated bile flow by ～250% and dietary UDCA by ～500%, compared with untreated animals (P < 0.05). In non-UDCA-treated Cftr(-/-) mice, the proportion of CA in bile was higher compared with that in controls (61 ± 4 vs. 46 ± 4%; P < 0.05), accompanied by an increased CA synthesis [16 ± 1 vs. 10 ± 2 μmol·h(-1)·100 g body wt (BW)(-1); P < 0.05] and CA pool size (28 ± 3 vs. 19 ± 1 μmol/100 g BW; P < 0.05). In both Cftr(-/-) and controls, UDCA treatment drastically reduced the proportion of CA in bile below 5% and diminished CA synthesis (2.3 ± 0.3 vs. 2.2 ± 0.4 μmol·day(-1)·100 g BW(-1); nonsignificant) and CA pool size (3.6 ± 0.6 vs. 1.5 ± 0.3 μmol/100 g BW; P < 0.05). Acute TUDCA infusion and chronic UDCA treatment both stimulate bile flow in cystic fibrosis conditions independently from Cftr function. Chronic UDCA treatment reduces the hydrophobicity of the bile salt pool in Cftr(-/-) mice. These results support a potential beneficial effect of UDCA on bile flow and bile salt metabolism in cystic fibrosis conditions.     

Glomerular filtration rate determinations in conscious type II diabetic mice

Diabetic nephropathy is a major cause of end-stage renal disease worldwide. The current studies were performed to determine the later stages of the progression of renal disease in type II diabetic mice (BKS; db/db). Methodology was developed for determining glomerular filtration rate (GFR) in conscious, chronically instrumented mice using continuous intravenous infusion of FITC-labeled inulin to achieve a steady-state plasma inulin concentration. Obese diabetic mice exhibited increased GFR compared with control mice. GFR averaged 0.313 ± 0.018 and 0.278 ± 0.007 ml/min in 18-wk-old obese diabetic (n = 11) and control (n = 13) mice, respectively (P < 0.05). In 28-wk-old obese diabetic (n = 10) and control (n = 15) mice, GFR averaged 0.348 ± 0.030 and 0.279 ± 0.009 ml/min, respectively (P < 0.05). GFR expressed per gram BW was significantly reduced in 18- and 28-wk-old obese diabetic compared with control mice (5.9 ± 0.3 vs. 9.0 ± 0.3; 6.6 ± 0.6 vs. 7.8 ± 0.3 μl·min(-1)·g body wt(-1)), respectively (P < 0.05). However, older nonobese type II diabetic mice had significantly reduced GFR (0.179 ± 0.023 ml/min; n = 6) and elevated urinary albumin excretion (811 ± 127 μg/day) compared with obese diabetic and control mice (514 ± 54, 171 ± 18 μg/day), which are consistent with the advanced stages of renal disease. These studies suggest that hyperfiltration contributes to the progression of renal disease in type II diabetic mice.     

Impact of protein coingestion on muscle protein synthesis during continuous endurance type exercise

This study investigates the impact of protein coingestion with carbohydrate on muscle protein synthesis during endurance type exercise. Twelve healthy male cyclists were studied during 2 h of fasted rest followed by 2 h of continuous cycling at 55% W(max). During exercise, subjects received either 1.0 g·kg(-1)·h(-1) carbohydrate (CHO) or 0.8 g·kg(-1)·h(-1) carbohydrate with 0.2 g·kg(-1)·h(-1) protein hydrolysate (CHO+PRO). Continuous intravenous infusions with l-[ring-(13)C(6)]phenylalanine and l-[ring-(2)H(2)]tyrosine were applied, and blood and muscle biopsies were collected to assess whole body protein turnover and muscle protein synthesis rates at rest and during exercise conditions. Protein coingestion stimulated whole body protein synthesis and oxidation rates during exercise by 22 ± 3 and 70 ± 17%, respectively (P < 0.01). Whole body protein breakdown rates did not differ between experiments. As a consequence, whole body net protein balance was slightly negative in CHO and positive in the CHO+PRO treatment (-4.9 ± 0.3 vs. 8.0 ± 0.3 μmol Phe·kg(-1)·h(-1), respectively, P < 0.01). Mixed muscle protein fractional synthetic rates (FSR) were higher during exercise compared with resting conditions (0.058 ± 0.006 vs. 0.035 ± 0.006%/h in CHO and 0.070 ± 0.011 vs. 0.038 ± 0.005%/h in the CHO+PRO treatment, respectively, P < 0.05). FSR during exercise did not differ between experiments (P = 0.46). We conclude that muscle protein synthesis is stimulated during continuous endurance type exercise activities when carbohydrate with or without protein is ingested. Protein coingestion does not further increase muscle protein synthesis rates during continuous endurance type exercise.     

Metabolic pathways promoting intrahepatic fatty acid accumulation in methionine and choline deficiency: implications for the pathogenesis of steatohepatitis

The pathological mechanisms that distinguish simple steatosis from steatohepatitis (or NASH, with consequent risk of cirrhosis and hepatocellular cancer) remain incompletely defined. Whereas both a methionine- and choline-deficient diet (MCDD) and a choline-deficient diet (CDD) lead to hepatic triglyceride accumulation, MCDD alone is associated with hepatic insulin resistance and inflammation (steatohepatitis). We used metabolic tracer techniques, including stable isotope ([13C?]palmitate) dilution and mass isotopomer distribution analysis (MIDA) of [13C?]acetate, to define differences in intrahepatic fatty acid metabolism that could explain the contrasting effect of MCDD and CDD on NASH in C57Bl6 mice. Compared with control-supplemented (CS) diet, liver triglyceride pool sizes were similarly elevated in CDD and MCDD groups (24.37 ± 2.4, 45.94 ± 3.9, and 43.30 ± 3.5 μmol/liver for CS, CDD, and MCDD, respectively), but intrahepatic neutrophil infiltration and plasma alanine aminotransferase (31 ± 3, 48 ± 4, 231 ± 79 U/l, P < 0.05) were elevated only in MCDD mice. However, despite loss of peripheral fat in MCDD mice, neither the rate of appearance of palmitate (27.2 ± 3.5, 26.3 ± 2.3, and 28.3 ± 3.5 μmol·kg?1·min?1) nor the contribution of circulating fatty acids to the liver triglyceride pool differed between groups. Unlike CDD, MCDD had a defect in hepatic triglyceride export that was confirmed using intravenous tyloxapol (142 ± 21, 122 ± 15, and 80 ± 7 mg·kg?1·h?1, P < 0.05). Moreover, hepatic de novo lipogenesis was significantly elevated in the MCDD group only (1.4 ± 0.3, 2.3 ± 0.4, and 3.4 ± 0.4 μmol/day, P < 0.01). These findings suggest that important alterations in hepatic fatty acid metabolism may promote the development of steatohepatitis. Similar mechanisms may predispose to hepatocyte damage in human NASH.     

Amino acid supplementation alters bone metabolism during simulated weightlessness.

High-protein and acidogenic diets induce hypercalciuria. Foods or supplements with excess sulfur-containing amino acids increase endogenous sulfuric acid production and therefore have the potential to increase calcium excretion and alter bone metabolism. In this study, effects of an amino acid/carbohydrate supplement on bone resorption were examined during bed rest. Thirteen subjects were divided at random into two groups: a control group (Con, n = 6) and an amino acid-supplemented group (AA, n = 7) who consumed an extra 49.5 g essential amino acids and 90 g carbohydrate per day for 28 days. Urine was collected for n-telopeptide (NTX), deoxypyridinoline (DPD), calcium, and pH determinations. Bone mineral content was determined and potential renal acid load was calculated. Bone-specific alkaline phosphatase was measured in serum samples collected on day 1 (immediately before bed rest) and on day 28. Potential renal acid load was higher in the AA group than in the Con group during bed rest (P < 0.05). For all subjects, during bed rest urinary NTX and DPD concentrations were greater than pre-bed rest levels (P < 0.05). Urinary NTX and DPD tended to be higher in the AA group (P = 0.073 and P = 0.056, respectively). During bed rest, urinary calcium was greater than baseline levels (P < 0.05) in the AA group but not the Con group. Total bone mineral content was lower after bed rest than before bed rest in the AA group but not the Con group (P < 0.05). During bed rest, urinary pH decreased (P < 0.05), and it was lower in the AA group than the Con group. These data suggest that bone resorption increased, without changes in bone formation, in the AA group.