Reduction of Fructose-induced Hypertriglyceridemia and Fatty Liver in Rats by 4-(4′-Chlorobenzyloxy)benzyl Nicotinate (KCD-232)

The effect of KCD-232, a new compound with a structure of 4-(4′-chlorobenzyloxy)benzyl nicotinate, on fructose-induced hypertriglyceridemia and fatty liver was studied in rats refed experimental diets for 3 days after having been starved for 2 days.

In younger or older rats refed 7% corn oil-containing diets, both the serum and liver triglyceride (TG) levels were higher in the fructose (F) group than in the glucose (G) group, and those of the F group were higher in the older rats than the younger rats. KCD-232 effectively inhibited fructose-induced hypertriglyceridemia and fatty liver in both the younger and older rats. In older rats refed 7% corn oil diets, an increase in hepatic fatty acid (FA) synthesis and decreases in the activities of hepatic TG lipase (HTGL) and adipose tissue lipoprotein lipase (LPL) were found in the F group, while the serum free FA level and hepatic FA oxidative activity of the F group were equal to those of the G group. KCD-232 strongly inhibited the fructose-enhanced FA synthesis, slightly repressed the fructose-suppressed HTGL activity and had no effect on LPL activity and FA oxidation.

These results suggest that fructose-induced hypertriglyceridemia and fatty liver are due to an enhanced hepatic FA synthesis and decreased hydrolytic activities required for TG clearance from the circulation system and that KCD-232 prevents them by inhibiting enhanced FA synthesis.     

Endothelin-1 modulates angiotensin II in the development of hypertension in fructose-fed rats

Two of the most potent vasoconstrictors, endothelin-1 (ET-1) and angiotensin II (Ang II), are upregulated in fructose hypertensive rats. It is unknown whether an interrelationship exists between these peptides that may contribute to the development of fructose-induced hypertension. The objective of this study was to investigate the existence of an interaction between the endothelin and renin angiotensin systems that may play a role in the development of fructose-induced hypertension. High fructose feeding and treatment with either bosentan, a dual endothelin receptor antagonist, or with L-158,809, an angiotensin type 1 receptor antagonist, were initiated simultaneously in male Wistar rats. Systolic blood pressure, fasted plasma parameters, insulin sensitivity, plasma Ang II, and vascular ET-1-immunoreactivity were determined following 6 weeks of high fructose feeding. Rats fed with a high fructose diet exhibited insulin resistance, hyperinsulinemia, hypertriglyceridemia, hypertension, and elevated plasma Ang II. Treatment with either bosentan or L-158,809 significantly attenuated the rise in blood pressure with no effect on insulin levels or insulin sensitivity in fructose-fed rats. Bosentan treatment significantly reduced plasma Ang II levels, while L-158,809 treatment significantly increased vascular ET-1-immunoreactivity in fructose-fed rats. Thus, treatment with the endothelin receptor antagonist prevented the development of fructose-induced hypertension and decreased plasma Ang II levels. These data suggest that ET-1 contributes to the development of fructose-induced hypertension through modulation of Ang II levels.     

Dissociation of inositol-requiring enzyme (IRE1α)-mediated c-Jun N-terminal kinase activation from hepatic insulin resistance in conditional X-box-binding protein-1 (XBP1) knock-out mice

Hepatic insulin resistance has been attributed to both increased endoplasmic reticulum (ER) stress and accumulation of intracellular lipids, specifically diacylglycerol (DAG). The ER stress response protein, X-box-binding protein-1 (XBP1), was recently shown to regulate hepatic lipogenesis, suggesting that hepatic insulin resistance in models of ER stress may result from defective lipid storage, as opposed to ER-specific stress signals. Studies were designed to dissociate liver lipid accumulation and activation of ER stress signaling pathways, which would allow us to delineate the individual contributions of ER stress and hepatic lipid content to the pathogenesis of hepatic insulin resistance. Conditional XBP1 knock-out (XBP1Δ) and control mice were fed fructose chow for 1 week. Determinants of whole-body energy balance, weight, and composition were determined. Hepatic lipids including triglyceride, DAGs, and ceramide were measured, alongside markers of ER stress. Whole-body and tissue-specific insulin sensitivity were determined by hyperinsulinemic-euglycemic clamp studies. Hepatic ER stress signaling was increased in fructose chow-fed XBP1Δ mice as reflected by increased phosphorylated eIF2α, HSPA5 mRNA, and a 2-fold increase in hepatic JNK activity. Despite JNK activation, XBP1Δ displayed increased hepatic insulin sensitivity during hyperinsulinemic-euglycemic clamp studies, which was associated with increased insulin-stimulated IRS2 tyrosine phosphorylation, reduced hepatic DAG content, and reduced PKCε activity. These studies demonstrate that ER stress and IRE1α-mediated JNK activation can be disassociated from hepatic insulin resistance and support the hypothesis that hepatic insulin resistance in models of ER stress may be secondary to ER stress modulation of hepatic lipogenesis.     

Salicylic acid reverses phorbol 12-myristate-13-acetate (PMA)- and tumor necrosis factor alpha (TNFalpha)-induced insulin receptor substrate 1 (IRS1) serine 307 phosphorylation and insulin resistance in human embryonic kidney 293 (HEK293) cells

Salicylates, including aspirin, have been shown to improve insulin sensitivity both in human and animal models. Although it has been suggested that salicylates sensitize insulin action by inhibiting IkappaB kinase beta (IKKbeta), the detailed mechanisms remain unclear. Protein kinase C isoforms and tumor necrosis factor alpha (TNFalpha) signaling pathways are well described mediators of insulin resistance; they are implicated in the activation of IKKbeta and the subsequent inhibition of proximal insulin signaling via insulin receptor substrate 1 (IRS1) and Akt. This study investigated the effect of salicylic acid on phorbol 12-myristate 13-acetate (PMA)- and TNFalpha-induced insulin resistance in a human embryonic kidney 293 (HEK293) cell line stably expressing recombinant human IRS1. The results showed that both PMA and TNFalpha inhibited insulin-induced Akt phosphorylation and promoted IRS1 phosphorylation on Ser-307. Salicylic acid pretreatment completely reversed the effects of PMA and TNFalpha on both Akt and IRS1. Whereas PMA activated protein kinase C isoforms and IKKbeta, TNFalpha activated neither. On the other hand, both PMA and TNFalpha activated the c-Jun N-terminal kinase (JNK), which has been reported to directly phosphorylate IRS1 Ser-307. SP600125, a JNK inhibitor, prevented PMA and TNFalpha-induced IRS1 Ser-307 phosphorylation. Finally, salicylic acid inhibited JNK activation induced by both PMA and TNFalpha. Taken together, these observations suggest that salicylic acid can reverse the inhibitory effects of TNFalpha on insulin signaling via an IKKbeta-independent mechanism(s), potentially involving the inhibition of JNK activation. The role of JNK in salicylic acid-mediated insulin sensitization, however, requires further validation because the JNK inhibitor SP600125 appears to have other nonspecific activity in addition to inhibiting JNK activity.     

Resistin promotes cardiac hypertrophy via the AMP-activated protein kinase/mammalian target of rapamycin (AMPK/mTOR) and c-Jun N-terminal kinase/insulin receptor substrate 1 (JNK/IRS1) pathways

Resistin has been suggested to be involved in the development of diabetes and insulin resistance. We recently reported that resistin is expressed in diabetic hearts and promotes cardiac hypertrophy; however, the mechanisms underlying this process are currently unknown. Therefore, we wanted to elucidate the mechanisms associated with resistin-induced cardiac hypertrophy and myocardial insulin resistance. Overexpression of resistin using adenoviral vector in neonatal rat ventricular myocytes was associated with inhibition of AMP-activated protein kinase (AMPK) activity, activation of tuberous sclerosis complex 2/mammalian target of rapamycin (mTOR) pathway, and increased cell size, [(3)H]leucine incorporation (i.e. protein synthesis) and mRNA expression of the hypertrophic marker genes, atrial natriuretic factor, brain natriuretic peptide, and β-myosin heavy chain. Activation of AMPK with 5-aminoimidazole-4-carbozamide-1-β-D-ribifuranoside or inhibition of mTOR with rapamycin or mTOR siRNA attenuated these resistin-induced changes. Furthermore, resistin increased serine phosphorylation of insulin receptor substrate (IRS1) through the activation of the apoptosis signal-regulating kinase 1/c-Jun N-terminal Kinase (JNK) pathway, a module known to stimulate insulin resistance. Inhibition of JNK (with JNK inhibitor SP600125 or using dominant-negative JNK) reduced serine 307 phosphorylation of IRS1. Resistin also stimulated the activation of p70(S6K), a downstream kinase target of mTOR, and increased phosphorylation of the IRS1 serine 636/639 residues, whereas treatment with rapamycin reduced the phosphorylation of these residues. Interestingly, these in vitro signaling pathways were also operative in vivo in ventricular tissues from adult rat hearts overexpressing resistin. These data demonstrate that resistin induces cardiac hypertrophy and myocardial insulin resistance, possibly via the AMPK/mTOR/p70(S6K) and apoptosis signal-regulating kinase 1/JNK/IRS1 pathways.     

Mammalian target of rapamycin regulates IRS-1 serine 307 phosphorylation

Insulin signaling can be negatively regulated by phosphorylation of serine 307 of the insulin receptor substrate (IRS)-1. Rapamycin, an inhibitor of the kinase mTOR, can prevent serine 307 phosphorylation and the development of insulin resistance. We further investigated the role of mTOR in regulating serine 307 phosphorylation, demonstrating that serine 307 phosphorylation in response to insulin, anisomycin, or tumor necrosis factor was quantitatively and temporally associated with activation of mTOR and could be inhibited by rapamycin. Amino acid stimulation activated mTOR and resulted in IRS-1 serine 307 phosphorylation without activating PKB or JNK. Okadaic acid, an inhibitor of the phosphatase PP2A, activated mTOR and stimulated the phosphorylation of serine 307 in a rapamycin-sensitive manner, indicating serine 307 phosphorylation requires mTOR activity but not PP2A, suggesting that mTOR itself may be responsible for phosphorylating serine 307. Finally, we demonstrated that serine 307 phosphorylated IRS-1 is detected primarily in the cytosolic fraction.     

c-Jun N-terminal kinase (JNK) mediates feedback inhibition of the insulin signaling cascade

Activation of the c-Jun N-terminal kinase (JNK) by proinflammatory cytokines inhibits insulin signaling, at least in part, by stimulating phosphorylation of rat/mouse insulin receptor substrate 1 (Irs1) at Ser(307) (Ser(312) in human IRS1). Here we show that JNK mediated feedback inhibition of the insulin signal in mouse embryo fibroblasts, 3T3-L1 adipocytes, and 32D(IR) cells. Insulin stimulation of JNK activity required phosphatidylinositol 3-kinase and Grb2 signaling. Moreover, activation of JNK by insulin was inhibited by a cell-permeable peptide that disrupted the interaction of JNK with cellular proteins. However, the direct binding of JNK to Irs1 was not required for its activation by insulin, whereas direct binding was required for Ser(307) phosphorylation of Irs1. Insulin-stimulated Ser(307) phosphorylation was reduced 80% in cells lacking JNK1 and JNK2 or in cells expressing a mutant Irs1 protein lacking the JNK binding site. Reduced Ser(307) phosphorylation was directly related to increased insulin-stimulated tyrosine phosphorylation, Akt phosphorylation, and glucose uptake. These results support the hypothesis that JNK is a negative feedback regulator of insulin action by phosphorylating Ser(307) in Irs1.     

Lipoic acid prevents liver metabolic changes induced by administration of a fructose-rich diet

Background

To evaluate whether co-administration of R/S-α-lipoic acid can prevent the development of oxidative stress and metabolic changes induced by a fructose-rich diet (F).

Methods

We assessed glycemia in the fasting state and during an oral glucose tolerance test, triglyceridemia and insulinemia in rats fed with standard diet (control) and fructose without or with R/S-α-lipoic acid. Insulin resistance and hepatic insulin sensitivity were also calculated. In liver, we measured reduced glutathione, protein carbonyl groups, antioxidant capacity by ABTS assay, antioxidant enzymes (catalase and superoxide dismutase 1 and 2), uncoupling protein 2, PPARδ and PPARγ protein expressions, SREBP-1c, fatty acid synthase and glycerol-3-phosphate acyltransferase-1 gene expression, and glucokinase activity.

Results

R/S-α-lipoic acid co-administration to F-fed rats a) prevented hyperinsulinemia, hypertriglyceridemia and insulin resistance, b) improved hepatic insulin sensitivity and glucose tolerance, c) decreased liver oxidative stress and increased antioxidant capacity and antioxidant enzymes expression, d) decreased uncoupling protein 2 and PPARδ protein expression and increased PPARγ levels, e) restored the basal gene expression of PPARδ, SREBP-1c and the lipogenic genes fatty acid synthase and glycerol-3-phosphate acyltransferase, and f) decreased the fructose-mediated enhancement of glucokinase activity.

Conclusions

Our results suggest that fructose-induced oxidative stress is an early phenomenon associated with compensatory hepatic metabolic mechanisms, and that treatment with an antioxidant prevented the development of such changes.

General significance

This knowledge would help to better understand the mechanisms involved in liver adaptation to fructose-induced oxidative stress and to develop effective strategies to prevent and treat, at early stages, obesity and type 2 diabetes mellitus.     

Enhanced prereceptor glucocorticoid metabolism and lipogenesis impair insulin signaling in the liver of fructose-fed rats

Overconsumption of fructose, as a highly lipogenic sugar, may profoundly affect hepatic metabolism and has been associated with many components of the metabolic syndrome, particularly with insulin resistance and Type 2 diabetes. In this study, we proposed that high fructose diet may enhance lipogenesis and decrease insulin sensitivity in the liver through dysregulation of glucocorticoid signaling. Therefore, we examined the effects of long-term consumption of 10% fructose solution on triglyceridemia, liver histology and intracellular corticosterone level, as well as on 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) and hexose-6-phosphate dehydrogenase (H6PDH) mRNA and protein levels in the rat liver. Glucocorticoid action was assessed by glucocorticoid receptor (GR) expression and intracellular redistribution. We also analyzed the expression of enzymes involved in gluconeogenesis and lipogenesis, phosphoenolpyruvate carboxykinase (PEPCK) and lipin-1. The results have shown that fructose-rich diet led to increase in 11βHSD1 and H6PDH protein levels, while hepatic corticosterone concentration remained unchanged. Concomitantly, GR was increasingly accumulated in the cytoplasm, whereas its nuclear level was unchanged and accompanied by diminished PEPCK mRNA level. Elevation of lipin-1 in the liver microsomes suggested that fructose diet led to an increase in lipogenesis and consequently to hypertriglyceridemia. The observed increase of insulin receptor supstrate-1 phosphorylation on Ser³⁰⁷ represents a hallmark of impaired insulin signaling in the liver of fructose-fed rat and probably is a consequence of the alterations in 11βHSD1 and lipin-1 levels. Overall, our findings suggest that fructose-rich diet may perturb hepatic prereceptor glucocorticoid metabolism and lipogenesis, resulting in hypertriglyceridemia and attenuated hepatic insulin sensitivity.     

(−)-Epicatechin mitigates high-fructose-associated insulin resistance by modulating redox signaling and endoplasmic reticulum stress

We investigated the capacity of dietary (−)-epicatechin (EC) to mitigate insulin resistance through the modulation of redox-regulated mechanisms in a rat model of metabolic syndrome. Adolescent rats were fed a regular chow diet without or with high fructose (HFr; 10% w/v) in drinking water for 8 weeks, and a group of HFr-fed rats was supplemented with EC in the diet. HFr-fed rats developed insulin resistance, which was mitigated by EC supplementation. Accordingly, the activation of components of the insulin signaling cascade (insulin receptor, IRS1, Akt, and ERK1/2) was impaired, whereas negative regulators (PKC, IKK, JNK, and PTP1B) were upregulated in the liver and adipose tissue of HFr rats. These alterations were partially or totally prevented by EC supplementation. In addition, EC inhibited events that contribute to insulin resistance: HFr-associated increased expression and activity of NADPH oxidase, activation of redox-sensitive signals, expression of NF-κB-regulated proinflammatory cytokines and chemokines, and some sub-arms of endoplasmic reticulum stress signaling. Collectively, these findings indicate that EC supplementation can mitigate HFr-induced insulin resistance and are relevant for defining interventions that can prevent/mitigate MetS-associated insulin resistance.     

Use of a monoclonal antibody to distinguish between precursor and mature forms of human lysosomal alpha-glucosidase

The levels of functional mRNA encoding glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) were examined in hepatocytes from fasted and fasted/carbohydrate-refed rats and in hepatocytes inoculated into primary culture. Functional G6PDH mRNA was assessed in a cell-free protein synthesis system in vitro. We observed that hepatocytes from fasted/carbohydrate-refed rats had a 12-fold higher level of mRNA than did hepatocytes from fasted rats. The possibility that the adrenal glucocorticoids and insulin were responsible for the increase in G6PDH mRNA in refed rats was examined by studying the effect of insulin and the synthetic glucocorticoid, dexamethasone, on the level of functional G6PDH mRNA in primary cultures of rat hepatocytes maintained in a chemically defined medium. Hepatocytes from fasted rats were inoculated into primary culture and maintained for 48 h either in the absence of hormones or in the presence of insulin alone, dexamethasone alone or both hormones together. We observed that dexamethasone alone caused a fourfold increase in G6PDH mRNA while insulin caused about a twofold increase. Both hormones together elicited an increase that was additive. A comparison of functional G6PDH mRNA levels with the effect of the hormones on G6PDH activity and relative rate of enzyme synthesis suggests that the glucocorticoid elevates the level of G6PDH mRNA within the cell without causing a concommitant increase in the rate of synthesis of the enzyme or the level of G6PDH activity. The results obtained with the primary cultures of hepatocytes indicate that insulin and the glucocorticoids are probably involved with the regulation of hepatic G6PDH mRNA. However, involvement of other hormones, such as thyroid hormone, seems likely since the induced levels of G6PDH mRNA in hepatocytes in culture was one-third of that observed in refed rats.     

Sustained Action of Ceramide on the Insulin Signaling Pathway in Muscle Cells: IMPLICATION OF THE DOUBLE-STRANDED RNA-ACTIVATED PROTEIN KINASE*

In vivo, ectopic accumulation of fatty acids in muscles leads to alterations in insulin signaling at both the IRS1 and Akt steps. However, in vitro treatments with saturated fatty acids or their derivative ceramide demonstrate an effect only at the Akt step. In this study, we adapted our experimental procedures to mimic the in vivo situation and show that the double-stranded RNA-dependent protein kinase (PKR) is involved in the long-term effects of saturated fatty acids on IRS1. C2C12 or human muscle cells were incubated with palmitate or directly with ceramide for short or long periods, and insulin signaling pathway activity was evaluated. PKR involvement was assessed through pharmacological and genetic studies. Short-term treatments of myotubes with palmitate, a ceramide precursor, or directly with ceramide induce an inhibition of Akt, whereas prolonged periods of treatment show an additive inhibition of insulin signaling through increased IRS1 serine 307 phosphorylation. PKR mRNA, protein, and phosphorylation are increased in insulin-resistant muscles. When PKR activity is reduced (siRNA or a pharmacological inhibitor), serine phosphorylation of IRS1 is reduced, and insulin-induced phosphorylation of Akt is improved. Finally, we show that JNK mediates ceramide-activated PKR inhibitory action on IRS1. Together, in the long term, our results show that ceramide acts at two distinct levels of the insulin signaling pathway (IRS1 and Akt). PKR, which is induced by both inflammation signals and ceramide, could play a major role in the development of insulin resistance in muscle cells.     

The HISS story overview: a novel hepatic neurohumoral regulation of peripheral insulin sensitivity in health and diabetes.

Data are reviewed that are consistent with the following working hypothesis that proposes a novel mechanism regulating insulin sensitivity, which when nonfunctional, leads to severe insulin resistance. Postprandial elevation in insulin levels activates a hepatic parasympathetic reflex release of a putative hepatic insulin-sensitizing substance (HISS), which activates glucose uptake at skeletal muscle. Insulin causes HISS release in fed but not fasted animals. The reflex is mediated by acetylcholine and involves release of nitric oxide in the liver. Interruption of the release of HISS is achieved by surgical denervation of the anterior hepatic nerve plexus, muscarinic receptor blockade, or nitric oxide synthase antagonism and leads to immediate severe insulin resistance. The nitric oxide donor, SIN-1, reverses L-NAME-induced insulin resistance. Denervation-induced insulin resistance is reversed by intraportal but not intravenous administration of acetylcholine or SIN-1. Liver disease is often associated with insulin resistance; the bile duct ligation model of liver disease results in parasympathetic neuropathy and insulin resistance that is reversed by intraportal acetylcholine. Possible relevance of this HISS-dependent control of insulin action to insulin resistance in diabetes, liver disease, and obesity is discussed.     

Molecular mechanism(s) of burn-induced insulin resistance in murine skeletal muscle: role of IRS phosphorylation

Hyperglycemia, glucose intolerance and elevated insulin levels frequently occur in burned patients; however, the mechanism(s) for this insulin resistance has not been fully elucidated. One possible mechanism could involve alterations in the phosphorylation of serine 307 of the insulin receptor substrate-1 (IRS-1) via activation of stress kinase enzymes, including SAPK/JNK. In the present study we examined the time course of the effect of burn injury to mice on: levels of IRS-1 protein, phosphorylation of serine 307 of IRS-1, SAPK/JNK kinase levels and activity and Akt kinase activity in hind limb skeletal muscle. Burn injury produced a reduction in hind limb muscle mass 24 h after injury, and, which persisted for 168 h. At 24 h after injury, there was a dramatic ( approximately 9-fold) increase in phosphorylation of IRS-1 serine 307 followed by a more moderate elevation thereafter. Total IRS-1 protein was slightly elevated at 24 h after injury and decreased to levels below sham treated animals at the later times. Burn injury did not appear to change total SAPK/JNK protein content, however, enzyme activity was increased for 7 days after injury. Akt kinase activity was decreased in skeletal muscle following burn injury; providing a biochemical basis for burn-induced insulin resistance. These findings are consistent with the hypothesis that burn-induced insulin resistance may be related, at least in part, to alterations in the phosphorylation of key proteins in the insulin signaling cascade, including IRS-1, and that changes in stress kinases, such as SAPK/JNK produced by burn injury, may be responsible for these changes in phosphorylation.     

Role of tumor necrosis factor α (TNFα) in the onset of fructose-induced nonalcoholic fatty liver disease in mice

Tumor necrosis factor α (TNFα) is known to be involved in dysregulation of hepatic lipid metabolism and insulin signaling. However, whether TNFα also plays a casual role in the onset of fructose-induced nonalcoholic fatty liver disease (NAFLD) has not yet been determined. Therefore, wild-type and TNFα receptor 1 (TNFR1)−/− mice were fed with either 30% fructose solution or plain tap water. Hepatic triglycerides, markers of inflammation and ATP concentration as well as plasma ALT levels were determined. Hepatic PAI-1, SREBP-1, FAS mRNA expression was assessed by real-time RT-PCR. Furthermore, lipid peroxidation and indices of insulin resistance were determined in liver tissue and plasma. In comparison to water controls, chronic intake of 30% fructose solution caused a significant ∼5-fold increase in triglyceride accumulation and neutrophil infiltration in livers of wild-type mice and a ∼8-fold increase in plasma ALT levels. In TNFR1−/− mice, hepatic steatosis was attenuated and neutrophil infiltration in the liver as well as plasma ALT levels was similar to water controls. The protective effect of the TNFR1 deletion against the onset of fructose-induced steatosis was associated with increased phospho AMPK and Akt levels, decreased SREBP-1 and FAS expression in the liver and decreased RBP4 plasma levels, whereas hepatic lipid peroxidation, iNOS protein and ATP levels were similar between wild-type and TNFR1−/− mice fed fructose. Taken together, these data suggest that TNFα plays a casual role in the onset of fructose-induced liver damage as well as insulin resistance in mice through signaling cascades downstream of TNFR1.     

IRS1 phosphorylation does not mediate mTORC1-induced insulin resistance

Increased mammalian target of rapamycin complex 1 (mTORC1) activity has been suggested to play important roles in development of insulin resistance in obesity. mTORC1 hyperactivity also increases endoplasmic reticulum (ER) stress, which in turn contributes to development of insulin resistance and glucose intolerance. Increased IRS1 phosphorylation at Ser307 in vitro is correlated with mTORC1- and ER stress-induced insulin resistance. This phosphorylation site correlates strongly with impaired insulin receptor signaling in diabetic mice and humans. In contrast, evidence from knock-in mice suggests that phosphorylation of IRS1 at Ser307 is actually required to maintain insulin sensitivity. To study the involvement of IRS1^Ser307 phosphorylation in mTORC1-mediated glucose intolerance and insulin sensitivity in vivo, we investigated the effects of liver specific TSC1 depletion in IRS1^Ser307Ala mice and controls. Our results demonstrate that blockade of IRS1^Ser307 phosphorylation in vivo does not prevent mTORC1-mediated glucose intolerance and insulin resistance.     

Vitamin A status and its metabolism contribute to the regulation of hepatic genes during the cycle of fasting and refeeding in rats

Vitamin A (VA) status and its metabolism affect hepatic metabolic homeostasis. We investigated if VA status and metabolism contribute to energy metabolism and expression of hepatic genes in the cycle of fasting and refeeding. Zucker lean rats with VA sufficient (VAS) or VA deficient (VAD) status were respectively grouped as: ad libitum (VAS-AD or VAD-AD), 48-h fasted (VAS-Fasted or VAD-Fasted), 48-h fasted and refed a VAS diet (VAS-Refed-VAS or VAD-Refed-VAS), or refed a VAD diet (VAS-Refed-VAD or VAD-Refed-VAD) for 6 h. Respiratory exchange ratio (RER) of rats fed the VAS or VAD diet was monitored for 6 weeks. From week four, rats fed the VAS diet had higher RER than those fed the VAD diet. VAS-Refed rats had higher plasma levels of glucose, triglyceride, insulin and leptin than VAD-Refed rats. The mRNA and protein levels of hepatic genes for fuel metabolism in the fasting and refeeding cycle were determined using real-time polymerase chain reaction and immunoblot, respectively. The mRNA levels of glucokinase (Gck), sterol regulatory element-binding protein 1c (Srebp-1c), and fatty acid synthase (Fas) were lowered in VAS-Fasted and VAD-Fasted rats, and increased in VAS-Refed-VAS, VAS-Refed-VAD and VAD-Refed-VAS, but not VAD-Refed-VAD, rats. The ACL and FAS protein levels only dropped in VAS-Fasted rats and increased in VAS-Refed-VAS rats. The GK protein level decreased only in VAS-Fasted rats, and increased in VAS-Refed-VAS, VAS-Refed-VAD and VAD-Refed-VAS (but not VAD-Refed-VAD) rats. We conclude that VA status and its metabolism in the fasting and refeeding cycle contribute to the regulation of hepatic gene expression in rats.     

Fructose augments infection-impaired net hepatic glucose uptake during TPN administration

During chronic total parenteral nutrition (TPN), net hepatic glucose uptake (NHGU) and net hepatic lactate release (NHLR) are markedly reduced (downward arrow approximately 45 and approximately 65%, respectively) with infection. Because small quantities of fructose are known to augment hepatic glucose uptake and lactate release in normal fasted animals, the aim of this work was to determine whether acute fructose infusion with TPN could correct the impairments in NHGU and NHLR during infection. Chronically catheterized conscious dogs received TPN for 5 days via the inferior vena cava at a rate designed to match daily basal energy requirements. On the third day of TPN administration, a sterile (SHAM, n = 12) or Escherichia coli-containing (INF, n = 11) fibrin clot was implanted in the peritoneal cavity. Forty-two hours later, somatostatin was infused with intraportal replacement of insulin (12 +/- 2 vs. 24 +/- 2 microU/ml, SHAM vs. INF, respectively) and glucagon (24 +/- 4 vs. 92 +/- 5 pg/ml) to match concentrations previously observed in sham and infected animals. After a 120-min basal period, animals received either saline (Sham+S, n = 6; Inf+S, n = 6) or intraportal fructose (0.7 mg x kg(-1) x min(-1); Sham+F, n = 6; Inf+F, n = 5) infusion for 180 min. Isoglycemia of 120 mg/dl was maintained with a variable glucose infusion. Combined tracer and arteriovenous difference techniques were used to assess hepatic glucose metabolism. Acute fructose infusion with TPN augmented NHGU by 2.9 +/- 0.4 and 2.5 +/- 0.3 mg x kg(-1) x min(-1) in Sham+F and Inf+F, respectively. The majority of liver glucose uptake was stored as glycogen, and NHLR did not increase substantially. Therefore, despite an infection-induced impairment in NHGU and different hormonal environments, small amounts of fructose enhanced NHGU similarly in sham and infected animals. Glycogen storage, not lactate release, was the preferential fate of the fructose-induced increase in hepatic glucose disposal in animals adapted to TPN.     

Mechanism of rapid-phase insulin response to elevation of portal glucose concentration

The hepatoportal region is important for glucose sensing; however, the relationship between the hepatoportal glucose-sensing system and the postprandial rapid phase of the insulin response has been unclear. We examined whether a rapid-phase insulin response to low amounts of intraportal glucose infusion would occur, compared that with the response to intrajugular glucose infusion in conscious rats, and assessed whether this sensing system was associated with autonomic nerve activity. The increases in plasma glucose concentration did not differ between the two infusions at 3 min, but the rapid-phase insulin response was detected only in the intraportal infusion. A sharp and rapid insulin response was observed at 3 min after intraportal infusion of a small amount of glucose but not after intrajugular infusion. Furthermore, this insulin response was also induced by intraportal fructose infusion but not by nonmetabolizable sugars. The rapid-phase insulin response at 3 min during intraportal infusion did not differ between rats that had undergone hepatic vagotomy or chemical sympathectomy with 6-hydroxydopamine compared with control rats, but this response disappeared in rats that had undergone chemical vagotomy with atropine. We conclude that the elevation of glucose concentration in the hepatoportal region induced afferent signals from undetectable sensors and that these signals stimulate pancreas to induce the rapid-phase insulin response via cholinergic nerve action.