Sepsis and endotoxin (LPS) have been demonstrated to impair insulin-mediated glucose uptake in sk... more Sepsis and endotoxin (LPS) have been demonstrated to impair insulin-mediated glucose uptake in skeletal muscle. However, the intracellular mechanism responsible for this defect is not fully defined. The purpose of the present study was to determine whether specific elements of the insulin receptor (IR) signaling pathway in skeletal muscle are altered by LPS. In vivo injection of Escherichia coli LPS resulted in a 44% reduction in whole body glucose disposal under euglycemic hyperinsulinemic conditions, which was largely accounted for by a decreased rate of glycogen synthesis. Scatchard analysis indicated that the number and affinity of the high-affinity insulin binding sites in muscle were similar between control and LPS-treated rats. Western blot analysis indicated that under basal conditions, the levels of total and phosphorylated IR, insulin receptor substrate (IRS)-1, and mitogen-activated protein (MAP) kinase were not significantly different between control and endotoxic rats. In control animals, muscle obtained 2 min after intravenous injection of a maximally stimulating dose of insulin demonstrated a marked increase in the amount of phosphorylated IR (approximately 5-fold), IRS-1 (approximately 10-fold), and MAP kinase (approximately 10-fold). Insulin-stimulated phosphorylation of IR, IRS-1, and MAP kinase was markedly diminished (approximately 75%, 90%, and 78%, respectively) in LPS-treated rats. However, there was no concomitant reduction in the total abundance of these proteins under hyperinsulinemic conditions. These data demonstrate that LPS alters multiple steps in the insulin signal transduction pathway, but not insulin binding, in skeletal muscle that may mediate the observed impairment in glucose uptake.
Small molecule nonpeptidyl molecules are potentially attractive drug candidates as adjunct therap... more Small molecule nonpeptidyl molecules are potentially attractive drug candidates as adjunct therapies in the treatment of sepsis-induced metabolic complications. As such, the current study investigates the use of aurintricarboxylic acid (ATA), which stimulates insulinlike growth factor 1 receptor and AKT signaling, for its ability to ameliorate the protein metabolic effects of endotoxin (lipopolysaccharide [LPS]) + interferon + (IFN-+) in C2C12 myotubes and sepsis in skeletal muscle. Aurintricarboxylic acid dose-and time-dependently increases mTOR (mammalian or mechanistic target of rapamycin)Ydependent protein synthesis. Pretreatment with ATA prevents the LPS/IFN-+Yinduced decrease in protein synthesis at least in part by maintaining mTOR kinase activity, whereas posttreatment with ATA is able to increase protein synthesis when added up to 6 h after LPS/IFN-+. Aurintricarboxylic acid also reverses the amino acid resistance, which is detected in response to nutrient deprivation. Conversely, ATA decreases the basal rate of protein degradation and prevents the LPS/IFN-+ increase in proteolysis, and the latter change is associated reduced atrogin 1 and MuRF1 mRNA. The ability of ATA to antagonize LPS/IFN-+Yinduced changes in protein metabolism was associated with its ability to prevent the increases in interleukin 6 and nitric oxide synthase 2 and decreases in insulinlike growth factor 1. In vivo studies indicate ATA acutely increases skeletal muscle, but not cardiac, protein synthesis and attenuates the loss of lean body mass over 5 days. These data suggest ATA and other small molecule agonists of endogenous anabolic hormones may prove beneficial in treating sepsis by decreasing the inflammatory response and improving muscle protein balance.
Excitatory amino acids (EAAs) are the principal mediators of fast excitatory neurotransmission in... more Excitatory amino acids (EAAs) are the principal mediators of fast excitatory neurotransmission in the mammalian central nervous system. Previous studies have demonstrated that N-methyl-D-aspartate (NMDA), an EAA agonist, produces a stress response that mimics that observed in animals receiving lipopolysaccharide (LPS). The present investigation determined the role that NMDA receptors play in the hemodynamic, metabolic, and hormonal changes induced by LPS. Chronically catheterized fasted rats received LPS with or without prior injection of MK 801, an NMDA receptor antagonist. LPS produced a classical stress response characterized by hypotension, tachycardia, increased glucose flux, and elevated plasma levels of glucagon, corticosterone, and catecholamines. MK 801 (intravenously) prevented the tachycardia in response to LPS, but did not consistently alter the fall in arterial blood pressure. The NMDA receptor antagonist also blunted the early elevation in plasma epinephrine and norepinephrine levels seen in LPS-injected rats, and this was associated with a smaller increment in plasma glucose and lactate concentrations and glucose flux. To confirm that MK 801 was functioning by antagonizing NMDA receptors within the brain, a second group of rats received an intracerebroventricular injection of MK 801 prior to LPS. The central administration of MK 801 also attenuated the increase in heart rate. These results indicate that central NMDA receptor stimulation mediates the LPS-induced tachycardia and suggest that the partial inhibition of the glucose metabolic response to LPS by MK 801 resulted from the smaller increment in plasma catecholamines.
The purpose of the present study was to determine whether the intracerebroventricular (ICV) injec... more The purpose of the present study was to determine whether the intracerebroventricular (ICV) injection of carbachol enhances whole body glucose utilization and, if so, how this cholinergic agonist influences in vivo regional glucose uptake. An ICV cannula and vascular catheters were placed in rats prior to the experiment. Whole body glucose flux was assessed in overnight-fasted conscious unrestrained rats using [3-3H]glucose. Hyperglycemia was elicited 30 min after carbachol (50 nmol), and resulted from an increased rate of hepatic glucose production (135%) that exceeded an elevated rate of peripheral glucose uptake (105%). The glucose metabolic clearance rate was not altered by carbachol. Despite the hyperglycemia, no compensatory increase in plasma insulin levels were observed. Carbachol, however, did increase glucagon (64-164%), catecholamines (3.5- to 15-fold), and corticosterone (62-160%). Complete alpha- and beta-adrenergic blockade prevented the carbachol-induced changes in glucose flux. In vivo glucose uptake (Rg) by individual tissues was determined at the peak of the carbachol-induced hyperglycemia, using [2-14C]deoxyglucose. In addition, a separate group of control rats received an intravenous hormone infusion that increased plasma glucose, glucagon, and catecholamine levels to the same extent seen in carbachol-treated rats. The Rg in liver, spleen, and lung was elevated to a similar extent in carbachol-treated (60, 47, and 48%, respectively) and hormone-infused (60, 53, and 70%, respectively) rats. In contrast, whereas the hormone infusion increased Rg by ileum, skin, and kidney (80, 67, and 110%, respectively), no change was observed in these tissues from rats injected with carbachol.(ABSTRACT TRUNCATED AT 250 WORDS)
Premature rupture of the membranes (PROM) is still an unsolved problem in obstetrics. Neither pre... more Premature rupture of the membranes (PROM) is still an unsolved problem in obstetrics. Neither prevention nor diagnosis nor therapy is readily available. The amniotic sac is stabilized mechanically by a dense network of collagenous fibres; in the event of PROM, the collagenous mesh is dissolved and the thickness of the amniotic sac is reduced. In a prospective study involving 32 patients, we measured the thickness of the membranes in vitro by high-frequency ultrasound and by light microscopy of histological sections, between gestational weeks 28 and 32. We compared 18 patients with premature rupture of the amnio-chorionic membrane with 14 patients following induction of labour, with intact membranes at the same gestational age in vitro. We were able to show that the membrane in the PROM group (0.54 +/- 0.9 mm) was markedly thinner with fewer collagenous fibres than in the control group (0.74 +/- 1.01 mm). The application of high-frequency ultrasound in vivo in PROM may become helpful for the prediction of PROM; further studies are therefore desirable.
Premature rupture of the membranes (PROM) accounts for approximately 30 per cent of all preterm d... more Premature rupture of the membranes (PROM) accounts for approximately 30 per cent of all preterm deliveries. PROM is thought to be mainly due to a decrease in membrane integrity. The aim of our investigation was to determine, post-partum after 28 normal deliveries, the thickness of the amniochorionic membrane using a 20 MHz high-frequency ultrasound. The data obtained were compared with histological sections for measurement accuracy using a linear regression analysis method. The membrane thickness of the total study group was 0.83 +/- 0.11 mm (0.72-1.08 mm). Based on a statistical comparison with the histological sections, the high-frequency ultrasound examination was shown to be highly reliable, with a correlation coefficient of r = 0.96 (P < 0.0001). High-frequency ultrasonographic examinations of membrane thickness are an objective and reliable method and may be a gain to prenatal diagnostics once this method can be used in vivo.
Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, howeve... more Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however, they increase in obesity and elevations appear to be prognostic of diabetes. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1-14 C]-leucine metabolism, tissue-specific protein synthesis and branched-chain keto-acid (BCKA) dehydrogenase complex (BCKDC) activities. Male obese Zucker rats (11weeks old) had increased body weight (BW, 53%), liver (107%) and fat (,300%), but lower plantaris and gastrocnemius masses (221-24%). Plasma BCAAs and BCKAs were elevated 45-69% and ,100%, respectively, in obese rats. Processes facilitating these rises appeared to include increased dietary intake (23%), leucine (Leu) turnover and proteolysis [35% per g fat free mass (FFM), urinary markers of proteolysis: 3-methylhistidine (183%) and 4-hydroxyproline (766%)] and decreased BCKDC per g kidney, heart, gastrocnemius and liver (247-66%). A process disposing of circulating BCAAs, protein synthesis, was increased 23-29% by obesity in whole-body (FFM corrected), gastrocnemius and liver. Despite the observed decreases in BCKDC activities per gm tissue, rates of whole-body Leu oxidation in obese rats were 22% and 59% higher normalized to BW and FFM, respectively. Consistently, urinary concentrations of eight BCAA catabolism-derived acylcarnitines were also elevated. The unexpected increase in BCAA oxidation may be due to a substrate effect in liver. Supporting this idea, BCKAs were elevated more in liver (193-418%) than plasma or muscle, and per g losses of hepatic BCKDC activities were completely offset by increased liver mass, in contrast to other tissues. In summary, our results indicate that plasma BCKAs may represent a more sensitive metabolic signature for obesity than BCAAs. Processes supporting elevated BCAA]BCKAs in the obese Zucker rat include increased dietary intake, Leu and protein turnover along with impaired BCKDC activity. Elevated BCAAs/BCKAs may contribute to observed elevations in protein synthesis and BCAA oxidation.
The present study determined whether the muscle atrophy produced by colitis is associated with al... more The present study determined whether the muscle atrophy produced by colitis is associated with altered rates of muscle protein synthesis or degradation, as well as the potential role of the local (e.g., muscle) insulin-like growth factor (IGF) system and muscle-specific ubiquitin E3 ligases atrogin-1 and MuRF1 in mediating altered muscle protein balance. Colitis was induced in C57BL/6 mice by intra-rectal administration of trinitrobenzne sulfonic acid (TNBS), and blood and tissues collected on day 10. Mice with inflammatory bowel disease (IBD) demonstrated reduced skeletal muscle mass and protein content, whereas colonic segment weight and gross damage score were both increased in mice with colitis, compared to time-matched control values. There was no change in muscle protein synthesis in mice with IBD, but there was an increased protein breakdown (45%), proteasome activity (85%), and mRNA expression for atrogin-1 and MuRF1 (200-300%) in muscle. These changes were associated with a reduction in liver (but not muscle) IGF-I mRNA as well as a reduction in both total and free IGF-I in the blood. Colitis decreased the hepatic content of IGF binding protein (IGFBP)-3 mRNA by 40% and increased IGFBP-1 mRNA by 100%. In contrast, colitis did alter IGFBP mRNAs in muscle. The TNFα, IL-6 and NOS2 mRNA content of both liver and skeletal muscle was increased in TNBS-treated mice, and plasma TNFα and IL-6 concentrations were also elevated. These data suggest TNBS-induced colitis is independent of a change in muscle protein synthesis but dependent on stimulation of protein degradation via increased expression of muscle-specific atrogenes, which may be mediated in part by the reduction in circulating concentration of IGF-I and the concomitant increase in inflammatory mediators observed in the blood and muscle per se.
Glucose turnover is increased during shock and in acute sepsis, but relatively little information... more Glucose turnover is increased during shock and in acute sepsis, but relatively little information is available concerning the regulation of carbohydrate metabolism during the hypermetabolic phase of sepsis. In these studies peritoneal sepsis was induced in rats, following chronic vascular catheterization, by intraperitoneal administration of a pooled fecal inoculum. The resultant peritonitis has been shown to produce a sustained hypermetabolic state during the first three days of infection. Glucose and lactate kinetics were studied using a constant infusion of radiolabeled tracers during the peak of the hypermetabolic phase (day 2). The septic animals exhibited a 42% increase in glucose turnover and a 63% increase in the metabolic clearance rate of glucose, as compared to time-matched control rats. Hepatic glycogenolysis could only contribute 1% to 2% to the increased rate of glucose appearance. A major portion of the elevated glucose turnover was accounted for by a 93% increase in glucose recycling, indicating an enhancement of gluconeogenesis from glucose-derived gluconeogenic precursors. The increased importance of lactate as a precursor for gluconeogenesis in sepsis was indicated by the elevated lactate turnover (34%) and the increased percentage of 14C-glucose derived from 14C-lactate. The insulin to glucagon ratio was decreased in the septic animals as a result of a reduction in the plasma insulin concentration (56%) and an increased glucagon concentration (67%). We conclude that during the hypermetabolic phase of sepsis, the increased peripheral glucose uptake generated more gluconeogenic precursors but did not appear to have a major direct contribution to the increased aerobic metabolism.
Gram-negative hypermetabolic sepsis has been previously reported to produce whole body insulin re... more Gram-negative hypermetabolic sepsis has been previously reported to produce whole body insulin resistance. The present study was performed to determine in vivo which tissues are responsible for the sepsis-induced decrease in insulin-mediated glucose uptake (IMGU). and whether that decrease was related to a change in regional blood flow. Vascular catheters were placed in rats and sepsis was induced by subcutaneous injections of Escherichia coli. Insulin action was assessed 20 hours after the first injection of bacteria by the combined use of the euglycemic hyperinsulinemic clamp and the tracer 2-deoxyglucose IdGlc) technique. Insulin was infused at various rates in separate groups of septic and nonseptic rats for 3 hours to produce steady-state insulin levels between 70 and 20,000 pU/mL. Rats were injected with [U-'4C]-dGlc 140 minutes after the start of the euglycemic hyperinsulinemic clamp for determination of the glucose metabolic rate (Rg) in selected tissues. The maximal response to insulin was decreased 30% to 40% in the gastrocnemius, and in the red and white quadriceps. The former two muscles also showed a decrease in insulin sensitivity. However, the insulin resistance seen in hindlimb muscles was not evident in all muscles of the body, since tMGU by abdominal muscle, diaphragm, and heart wes not impaired by sepsis. The basal Rg by skin, spleen, ileum, and lung was increased by sepsis, and was higher than the insulin-stimulated increases in Rg by these tissues in nonseptic animals. Cardiac output was similar in septic and nonseptic rats and did not change during the infusion of insulin. Under basal conditions, sepsis appeared to redistribute blood flow away from the red quadriceps and skin, and increased flow to the liver (arterial), lung, and small intestine. When plasma insulin levels were elevated, hepatic arterial blood flow was increased, and flow to the red quadriceps and skin was decreased in nonseptic animals. Hyperinsulinemia did not produce any consistent change in regional blood flow in septic animals. The results of this study indicate that a decrease rate of IMGU in muscle is primarily responsible for the whole body insulin resistance seen during hypermetabolic sepsis, and that the impairment of insulin action in skeletal muscle is not
The purpose of the present study was to determine how a high dose of endotoxin (lipopolysaccharid... more The purpose of the present study was to determine how a high dose of endotoxin (lipopolysaccharide [LPS]), which produces hypoglycemia, alters in vivo glucose uptake by individual tissues. Catheterized conscious fasted rats were injected intravenously (i.v.) with either saline, LPS (1 mg/100 g body weight [BW], lethal dose [LD] 100), or 3-mercaptopicolinic acid (3-MP), an inhibitor of gluconeogenesis. In the latter two groups, blood glucose levels were clamped at either 6 mmol/L (euglycemia) or 3 mmol/L (hypoglycemia). In the first series of experiments, whole-body glucose flux was determined using [3-3H]glucose, and in the second study in vivo glucose uptake (Rg) by individual tissues was estimated by the tracer [U-14C]-2-deoxyglucose technique. The relative contribution of hypoglycemia per se to the LPS effect was determined by comparing the values from LPS- versus 3-MP-treated animals. There was no difference in the rate of whole-body glucose utilization (Rd) between saline-infused control rats and LPS-treated animals that were hypoglycemic. However, Rg by diaphragm, spleen, liver, and lung was increased in hypoglycemic LPS-treated rats. The increased Rg in these tissues was not observed in 3-MP-treated rats with a comparable hypoglycemia. Only the gastrocnemius muscle showed a reduction in Rg under hypoglycemic conditions, and the decrease was similar in both LPS- and 3-MP-treated animals. When sufficient glucose was infused into LPS-injected rats to maintain euglycemia, whole-body glucose Rd was increased compared with that in hypoglycemic LPS-treated rats.(ABSTRACT TRUNCATED AT 250 WORDS)
Glycogen repletion rates in liver and skeletal muscle were quantitated, and the contribution of g... more Glycogen repletion rates in liver and skeletal muscle were quantitated, and the contribution of gluconeogenesis to hepatic glycogen repletion and glucose output were determined during glucose infusion in hemodynamically stable endotoxemic
The increased glucose turnover seen during the hypermetabolic, hyperdynamic phase of sepsis is pa... more The increased glucose turnover seen during the hypermetabolic, hyperdynamic phase of sepsis is part of the body's defense mechanisms. In contrast, the metabolism of ethanol (ETOH] is known to compromise hepatic gluconeogenesis under certain conditions. This study tested the hypothesis that acute infusion of ETOH inhibits the elevated glucose production that is manifested during infection and thereby alters the normal responses to sepsis. In catheterized conscious rats, ETOH or saline infusion was started 24 hours before the induction of sepsis. and continued throughout the experiment.
The purpose of the present study was to determine how hypoglycemia alters glucose uptake by indiv... more The purpose of the present study was to determine how hypoglycemia alters glucose uptake by individual tissues and whether this response is altered by gram-negative infection. A hypermetabolic septic state was produced in catheterized rats by subcutaneous injections of live Escherichia co/i. The next morning, animals were infused with saline, somatostatin to produce a euglycemic insulinopenic state (6 mmol/L glucose, 5 plJ/mL insulin), or 3-mercaptopicolinate (3-MP) to inhibit gluconeogenesis and produce a hypoglycemic insulinopenic (4.5 or 2 mmol/L glucose, 5 pU/mL insulin) condition. After 140 minutes, ['YE]2-deoxyglucose was injected intravenously (IV) to determine in vivo glucose uptake by individual tissues. Sepsis increased whole body glucose disposal (Rd) by 53% under basal euglycemic conditions and this increase resulted from an enhanced rate of glucose removal by liver, spleen, lung, ileum, and skin. Under euglycemic insulinopenic conditions, total glucose Rd decreased in both septic and nonseptic rats as a result of a decreased rate of glucose uptake by muscle. However, because the absolute rate of glucose uptake was still elevated by sepsis, the rate of non-insulin-mediated glucose uptake (NIMGU) was46% higher in septic rats than in nonseptic animals. Severe hypoglycemia (2 mmol/L) produced a relative insulin deficiency and decreased whole body Rd in both septic and nonseptic animals by 53% to 56%. compared with euglycemic insulinopenic animals. The decrease in blood glucose decreased glucose uptake by all tissues examined, except brain and heart. However, sepsis still increased glucose uptake by liver, spleen, lung, ileum, and skin (25% to SO%), compared with hypoglycemic nonseptic rats. These results indicate that gram-negative sepsis increases NIMGU under basal conditions due to an increased glucose uptake by macrophage-rich tissues, and that this enhanced rate is maintained during hypoglycemia. Since both septic and nonseptic rats were insulinopenic, the increased rate of glucose uptake by liver, spleen, lung, ileum, and skin must have occurred predominantly by insulin-independent mechanisms.
Sepsis and endotoxin (LPS) have been demonstrated to impair insulin-mediated glucose uptake in sk... more Sepsis and endotoxin (LPS) have been demonstrated to impair insulin-mediated glucose uptake in skeletal muscle. However, the intracellular mechanism responsible for this defect is not fully defined. The purpose of the present study was to determine whether specific elements of the insulin receptor (IR) signaling pathway in skeletal muscle are altered by LPS. In vivo injection of Escherichia coli LPS resulted in a 44% reduction in whole body glucose disposal under euglycemic hyperinsulinemic conditions, which was largely accounted for by a decreased rate of glycogen synthesis. Scatchard analysis indicated that the number and affinity of the high-affinity insulin binding sites in muscle were similar between control and LPS-treated rats. Western blot analysis indicated that under basal conditions, the levels of total and phosphorylated IR, insulin receptor substrate (IRS)-1, and mitogen-activated protein (MAP) kinase were not significantly different between control and endotoxic rats. In control animals, muscle obtained 2 min after intravenous injection of a maximally stimulating dose of insulin demonstrated a marked increase in the amount of phosphorylated IR (approximately 5-fold), IRS-1 (approximately 10-fold), and MAP kinase (approximately 10-fold). Insulin-stimulated phosphorylation of IR, IRS-1, and MAP kinase was markedly diminished (approximately 75%, 90%, and 78%, respectively) in LPS-treated rats. However, there was no concomitant reduction in the total abundance of these proteins under hyperinsulinemic conditions. These data demonstrate that LPS alters multiple steps in the insulin signal transduction pathway, but not insulin binding, in skeletal muscle that may mediate the observed impairment in glucose uptake.
Small molecule nonpeptidyl molecules are potentially attractive drug candidates as adjunct therap... more Small molecule nonpeptidyl molecules are potentially attractive drug candidates as adjunct therapies in the treatment of sepsis-induced metabolic complications. As such, the current study investigates the use of aurintricarboxylic acid (ATA), which stimulates insulinlike growth factor 1 receptor and AKT signaling, for its ability to ameliorate the protein metabolic effects of endotoxin (lipopolysaccharide [LPS]) + interferon + (IFN-+) in C2C12 myotubes and sepsis in skeletal muscle. Aurintricarboxylic acid dose-and time-dependently increases mTOR (mammalian or mechanistic target of rapamycin)Ydependent protein synthesis. Pretreatment with ATA prevents the LPS/IFN-+Yinduced decrease in protein synthesis at least in part by maintaining mTOR kinase activity, whereas posttreatment with ATA is able to increase protein synthesis when added up to 6 h after LPS/IFN-+. Aurintricarboxylic acid also reverses the amino acid resistance, which is detected in response to nutrient deprivation. Conversely, ATA decreases the basal rate of protein degradation and prevents the LPS/IFN-+ increase in proteolysis, and the latter change is associated reduced atrogin 1 and MuRF1 mRNA. The ability of ATA to antagonize LPS/IFN-+Yinduced changes in protein metabolism was associated with its ability to prevent the increases in interleukin 6 and nitric oxide synthase 2 and decreases in insulinlike growth factor 1. In vivo studies indicate ATA acutely increases skeletal muscle, but not cardiac, protein synthesis and attenuates the loss of lean body mass over 5 days. These data suggest ATA and other small molecule agonists of endogenous anabolic hormones may prove beneficial in treating sepsis by decreasing the inflammatory response and improving muscle protein balance.
Excitatory amino acids (EAAs) are the principal mediators of fast excitatory neurotransmission in... more Excitatory amino acids (EAAs) are the principal mediators of fast excitatory neurotransmission in the mammalian central nervous system. Previous studies have demonstrated that N-methyl-D-aspartate (NMDA), an EAA agonist, produces a stress response that mimics that observed in animals receiving lipopolysaccharide (LPS). The present investigation determined the role that NMDA receptors play in the hemodynamic, metabolic, and hormonal changes induced by LPS. Chronically catheterized fasted rats received LPS with or without prior injection of MK 801, an NMDA receptor antagonist. LPS produced a classical stress response characterized by hypotension, tachycardia, increased glucose flux, and elevated plasma levels of glucagon, corticosterone, and catecholamines. MK 801 (intravenously) prevented the tachycardia in response to LPS, but did not consistently alter the fall in arterial blood pressure. The NMDA receptor antagonist also blunted the early elevation in plasma epinephrine and norepinephrine levels seen in LPS-injected rats, and this was associated with a smaller increment in plasma glucose and lactate concentrations and glucose flux. To confirm that MK 801 was functioning by antagonizing NMDA receptors within the brain, a second group of rats received an intracerebroventricular injection of MK 801 prior to LPS. The central administration of MK 801 also attenuated the increase in heart rate. These results indicate that central NMDA receptor stimulation mediates the LPS-induced tachycardia and suggest that the partial inhibition of the glucose metabolic response to LPS by MK 801 resulted from the smaller increment in plasma catecholamines.
The purpose of the present study was to determine whether the intracerebroventricular (ICV) injec... more The purpose of the present study was to determine whether the intracerebroventricular (ICV) injection of carbachol enhances whole body glucose utilization and, if so, how this cholinergic agonist influences in vivo regional glucose uptake. An ICV cannula and vascular catheters were placed in rats prior to the experiment. Whole body glucose flux was assessed in overnight-fasted conscious unrestrained rats using [3-3H]glucose. Hyperglycemia was elicited 30 min after carbachol (50 nmol), and resulted from an increased rate of hepatic glucose production (135%) that exceeded an elevated rate of peripheral glucose uptake (105%). The glucose metabolic clearance rate was not altered by carbachol. Despite the hyperglycemia, no compensatory increase in plasma insulin levels were observed. Carbachol, however, did increase glucagon (64-164%), catecholamines (3.5- to 15-fold), and corticosterone (62-160%). Complete alpha- and beta-adrenergic blockade prevented the carbachol-induced changes in glucose flux. In vivo glucose uptake (Rg) by individual tissues was determined at the peak of the carbachol-induced hyperglycemia, using [2-14C]deoxyglucose. In addition, a separate group of control rats received an intravenous hormone infusion that increased plasma glucose, glucagon, and catecholamine levels to the same extent seen in carbachol-treated rats. The Rg in liver, spleen, and lung was elevated to a similar extent in carbachol-treated (60, 47, and 48%, respectively) and hormone-infused (60, 53, and 70%, respectively) rats. In contrast, whereas the hormone infusion increased Rg by ileum, skin, and kidney (80, 67, and 110%, respectively), no change was observed in these tissues from rats injected with carbachol.(ABSTRACT TRUNCATED AT 250 WORDS)
Premature rupture of the membranes (PROM) is still an unsolved problem in obstetrics. Neither pre... more Premature rupture of the membranes (PROM) is still an unsolved problem in obstetrics. Neither prevention nor diagnosis nor therapy is readily available. The amniotic sac is stabilized mechanically by a dense network of collagenous fibres; in the event of PROM, the collagenous mesh is dissolved and the thickness of the amniotic sac is reduced. In a prospective study involving 32 patients, we measured the thickness of the membranes in vitro by high-frequency ultrasound and by light microscopy of histological sections, between gestational weeks 28 and 32. We compared 18 patients with premature rupture of the amnio-chorionic membrane with 14 patients following induction of labour, with intact membranes at the same gestational age in vitro. We were able to show that the membrane in the PROM group (0.54 +/- 0.9 mm) was markedly thinner with fewer collagenous fibres than in the control group (0.74 +/- 1.01 mm). The application of high-frequency ultrasound in vivo in PROM may become helpful for the prediction of PROM; further studies are therefore desirable.
Premature rupture of the membranes (PROM) accounts for approximately 30 per cent of all preterm d... more Premature rupture of the membranes (PROM) accounts for approximately 30 per cent of all preterm deliveries. PROM is thought to be mainly due to a decrease in membrane integrity. The aim of our investigation was to determine, post-partum after 28 normal deliveries, the thickness of the amniochorionic membrane using a 20 MHz high-frequency ultrasound. The data obtained were compared with histological sections for measurement accuracy using a linear regression analysis method. The membrane thickness of the total study group was 0.83 +/- 0.11 mm (0.72-1.08 mm). Based on a statistical comparison with the histological sections, the high-frequency ultrasound examination was shown to be highly reliable, with a correlation coefficient of r = 0.96 (P < 0.0001). High-frequency ultrasonographic examinations of membrane thickness are an objective and reliable method and may be a gain to prenatal diagnostics once this method can be used in vivo.
Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, howeve... more Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however, they increase in obesity and elevations appear to be prognostic of diabetes. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1-14 C]-leucine metabolism, tissue-specific protein synthesis and branched-chain keto-acid (BCKA) dehydrogenase complex (BCKDC) activities. Male obese Zucker rats (11weeks old) had increased body weight (BW, 53%), liver (107%) and fat (,300%), but lower plantaris and gastrocnemius masses (221-24%). Plasma BCAAs and BCKAs were elevated 45-69% and ,100%, respectively, in obese rats. Processes facilitating these rises appeared to include increased dietary intake (23%), leucine (Leu) turnover and proteolysis [35% per g fat free mass (FFM), urinary markers of proteolysis: 3-methylhistidine (183%) and 4-hydroxyproline (766%)] and decreased BCKDC per g kidney, heart, gastrocnemius and liver (247-66%). A process disposing of circulating BCAAs, protein synthesis, was increased 23-29% by obesity in whole-body (FFM corrected), gastrocnemius and liver. Despite the observed decreases in BCKDC activities per gm tissue, rates of whole-body Leu oxidation in obese rats were 22% and 59% higher normalized to BW and FFM, respectively. Consistently, urinary concentrations of eight BCAA catabolism-derived acylcarnitines were also elevated. The unexpected increase in BCAA oxidation may be due to a substrate effect in liver. Supporting this idea, BCKAs were elevated more in liver (193-418%) than plasma or muscle, and per g losses of hepatic BCKDC activities were completely offset by increased liver mass, in contrast to other tissues. In summary, our results indicate that plasma BCKAs may represent a more sensitive metabolic signature for obesity than BCAAs. Processes supporting elevated BCAA]BCKAs in the obese Zucker rat include increased dietary intake, Leu and protein turnover along with impaired BCKDC activity. Elevated BCAAs/BCKAs may contribute to observed elevations in protein synthesis and BCAA oxidation.
The present study determined whether the muscle atrophy produced by colitis is associated with al... more The present study determined whether the muscle atrophy produced by colitis is associated with altered rates of muscle protein synthesis or degradation, as well as the potential role of the local (e.g., muscle) insulin-like growth factor (IGF) system and muscle-specific ubiquitin E3 ligases atrogin-1 and MuRF1 in mediating altered muscle protein balance. Colitis was induced in C57BL/6 mice by intra-rectal administration of trinitrobenzne sulfonic acid (TNBS), and blood and tissues collected on day 10. Mice with inflammatory bowel disease (IBD) demonstrated reduced skeletal muscle mass and protein content, whereas colonic segment weight and gross damage score were both increased in mice with colitis, compared to time-matched control values. There was no change in muscle protein synthesis in mice with IBD, but there was an increased protein breakdown (45%), proteasome activity (85%), and mRNA expression for atrogin-1 and MuRF1 (200-300%) in muscle. These changes were associated with a reduction in liver (but not muscle) IGF-I mRNA as well as a reduction in both total and free IGF-I in the blood. Colitis decreased the hepatic content of IGF binding protein (IGFBP)-3 mRNA by 40% and increased IGFBP-1 mRNA by 100%. In contrast, colitis did alter IGFBP mRNAs in muscle. The TNFα, IL-6 and NOS2 mRNA content of both liver and skeletal muscle was increased in TNBS-treated mice, and plasma TNFα and IL-6 concentrations were also elevated. These data suggest TNBS-induced colitis is independent of a change in muscle protein synthesis but dependent on stimulation of protein degradation via increased expression of muscle-specific atrogenes, which may be mediated in part by the reduction in circulating concentration of IGF-I and the concomitant increase in inflammatory mediators observed in the blood and muscle per se.
Glucose turnover is increased during shock and in acute sepsis, but relatively little information... more Glucose turnover is increased during shock and in acute sepsis, but relatively little information is available concerning the regulation of carbohydrate metabolism during the hypermetabolic phase of sepsis. In these studies peritoneal sepsis was induced in rats, following chronic vascular catheterization, by intraperitoneal administration of a pooled fecal inoculum. The resultant peritonitis has been shown to produce a sustained hypermetabolic state during the first three days of infection. Glucose and lactate kinetics were studied using a constant infusion of radiolabeled tracers during the peak of the hypermetabolic phase (day 2). The septic animals exhibited a 42% increase in glucose turnover and a 63% increase in the metabolic clearance rate of glucose, as compared to time-matched control rats. Hepatic glycogenolysis could only contribute 1% to 2% to the increased rate of glucose appearance. A major portion of the elevated glucose turnover was accounted for by a 93% increase in glucose recycling, indicating an enhancement of gluconeogenesis from glucose-derived gluconeogenic precursors. The increased importance of lactate as a precursor for gluconeogenesis in sepsis was indicated by the elevated lactate turnover (34%) and the increased percentage of 14C-glucose derived from 14C-lactate. The insulin to glucagon ratio was decreased in the septic animals as a result of a reduction in the plasma insulin concentration (56%) and an increased glucagon concentration (67%). We conclude that during the hypermetabolic phase of sepsis, the increased peripheral glucose uptake generated more gluconeogenic precursors but did not appear to have a major direct contribution to the increased aerobic metabolism.
Gram-negative hypermetabolic sepsis has been previously reported to produce whole body insulin re... more Gram-negative hypermetabolic sepsis has been previously reported to produce whole body insulin resistance. The present study was performed to determine in vivo which tissues are responsible for the sepsis-induced decrease in insulin-mediated glucose uptake (IMGU). and whether that decrease was related to a change in regional blood flow. Vascular catheters were placed in rats and sepsis was induced by subcutaneous injections of Escherichia coli. Insulin action was assessed 20 hours after the first injection of bacteria by the combined use of the euglycemic hyperinsulinemic clamp and the tracer 2-deoxyglucose IdGlc) technique. Insulin was infused at various rates in separate groups of septic and nonseptic rats for 3 hours to produce steady-state insulin levels between 70 and 20,000 pU/mL. Rats were injected with [U-'4C]-dGlc 140 minutes after the start of the euglycemic hyperinsulinemic clamp for determination of the glucose metabolic rate (Rg) in selected tissues. The maximal response to insulin was decreased 30% to 40% in the gastrocnemius, and in the red and white quadriceps. The former two muscles also showed a decrease in insulin sensitivity. However, the insulin resistance seen in hindlimb muscles was not evident in all muscles of the body, since tMGU by abdominal muscle, diaphragm, and heart wes not impaired by sepsis. The basal Rg by skin, spleen, ileum, and lung was increased by sepsis, and was higher than the insulin-stimulated increases in Rg by these tissues in nonseptic animals. Cardiac output was similar in septic and nonseptic rats and did not change during the infusion of insulin. Under basal conditions, sepsis appeared to redistribute blood flow away from the red quadriceps and skin, and increased flow to the liver (arterial), lung, and small intestine. When plasma insulin levels were elevated, hepatic arterial blood flow was increased, and flow to the red quadriceps and skin was decreased in nonseptic animals. Hyperinsulinemia did not produce any consistent change in regional blood flow in septic animals. The results of this study indicate that a decrease rate of IMGU in muscle is primarily responsible for the whole body insulin resistance seen during hypermetabolic sepsis, and that the impairment of insulin action in skeletal muscle is not
The purpose of the present study was to determine how a high dose of endotoxin (lipopolysaccharid... more The purpose of the present study was to determine how a high dose of endotoxin (lipopolysaccharide [LPS]), which produces hypoglycemia, alters in vivo glucose uptake by individual tissues. Catheterized conscious fasted rats were injected intravenously (i.v.) with either saline, LPS (1 mg/100 g body weight [BW], lethal dose [LD] 100), or 3-mercaptopicolinic acid (3-MP), an inhibitor of gluconeogenesis. In the latter two groups, blood glucose levels were clamped at either 6 mmol/L (euglycemia) or 3 mmol/L (hypoglycemia). In the first series of experiments, whole-body glucose flux was determined using [3-3H]glucose, and in the second study in vivo glucose uptake (Rg) by individual tissues was estimated by the tracer [U-14C]-2-deoxyglucose technique. The relative contribution of hypoglycemia per se to the LPS effect was determined by comparing the values from LPS- versus 3-MP-treated animals. There was no difference in the rate of whole-body glucose utilization (Rd) between saline-infused control rats and LPS-treated animals that were hypoglycemic. However, Rg by diaphragm, spleen, liver, and lung was increased in hypoglycemic LPS-treated rats. The increased Rg in these tissues was not observed in 3-MP-treated rats with a comparable hypoglycemia. Only the gastrocnemius muscle showed a reduction in Rg under hypoglycemic conditions, and the decrease was similar in both LPS- and 3-MP-treated animals. When sufficient glucose was infused into LPS-injected rats to maintain euglycemia, whole-body glucose Rd was increased compared with that in hypoglycemic LPS-treated rats.(ABSTRACT TRUNCATED AT 250 WORDS)
Glycogen repletion rates in liver and skeletal muscle were quantitated, and the contribution of g... more Glycogen repletion rates in liver and skeletal muscle were quantitated, and the contribution of gluconeogenesis to hepatic glycogen repletion and glucose output were determined during glucose infusion in hemodynamically stable endotoxemic
The increased glucose turnover seen during the hypermetabolic, hyperdynamic phase of sepsis is pa... more The increased glucose turnover seen during the hypermetabolic, hyperdynamic phase of sepsis is part of the body's defense mechanisms. In contrast, the metabolism of ethanol (ETOH] is known to compromise hepatic gluconeogenesis under certain conditions. This study tested the hypothesis that acute infusion of ETOH inhibits the elevated glucose production that is manifested during infection and thereby alters the normal responses to sepsis. In catheterized conscious rats, ETOH or saline infusion was started 24 hours before the induction of sepsis. and continued throughout the experiment.
The purpose of the present study was to determine how hypoglycemia alters glucose uptake by indiv... more The purpose of the present study was to determine how hypoglycemia alters glucose uptake by individual tissues and whether this response is altered by gram-negative infection. A hypermetabolic septic state was produced in catheterized rats by subcutaneous injections of live Escherichia co/i. The next morning, animals were infused with saline, somatostatin to produce a euglycemic insulinopenic state (6 mmol/L glucose, 5 plJ/mL insulin), or 3-mercaptopicolinate (3-MP) to inhibit gluconeogenesis and produce a hypoglycemic insulinopenic (4.5 or 2 mmol/L glucose, 5 pU/mL insulin) condition. After 140 minutes, ['YE]2-deoxyglucose was injected intravenously (IV) to determine in vivo glucose uptake by individual tissues. Sepsis increased whole body glucose disposal (Rd) by 53% under basal euglycemic conditions and this increase resulted from an enhanced rate of glucose removal by liver, spleen, lung, ileum, and skin. Under euglycemic insulinopenic conditions, total glucose Rd decreased in both septic and nonseptic rats as a result of a decreased rate of glucose uptake by muscle. However, because the absolute rate of glucose uptake was still elevated by sepsis, the rate of non-insulin-mediated glucose uptake (NIMGU) was46% higher in septic rats than in nonseptic animals. Severe hypoglycemia (2 mmol/L) produced a relative insulin deficiency and decreased whole body Rd in both septic and nonseptic animals by 53% to 56%. compared with euglycemic insulinopenic animals. The decrease in blood glucose decreased glucose uptake by all tissues examined, except brain and heart. However, sepsis still increased glucose uptake by liver, spleen, lung, ileum, and skin (25% to SO%), compared with hypoglycemic nonseptic rats. These results indicate that gram-negative sepsis increases NIMGU under basal conditions due to an increased glucose uptake by macrophage-rich tissues, and that this enhanced rate is maintained during hypoglycemia. Since both septic and nonseptic rats were insulinopenic, the increased rate of glucose uptake by liver, spleen, lung, ileum, and skin must have occurred predominantly by insulin-independent mechanisms.
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