Insulin-stimulated Protein Synthesis Research Articles

261 Variations in Protein Deposition Might Be Associated with Insulin Resistance and Differences in Amino Acids Metabolism in Growing Gilts

Abstract Pigs with similar body weight (BW), same genetic background and sex vary in terms of protein deposition (PD) overgrowth. Insulin stimulates protein synthesis, and differences in PD could be associated with the variation among pigs in postprandial insulin secretion. Therefore, this study aimed to investigate if changes in preprandial and postprandial amino acids (AA) response and insulin resistance differ between pigs having Low or High PD. Seventy gilts (33 kg BW, ± 3.6; Alphagène) were assigned to a randomized complete block (BW) design and housed in the same pen equipped with automatic feeders. Pigs were fed ad libitum with a diet balanced to maximize PD. Dual X-Ray estimated body protein (BP) and lipid (BL) masses on d 1 and 21, and by difference, PD was calculated. On d 23, pigs were grouped in Low (157 g/d) and High (219 g/d) PD. Eight pigs per PD group were canulated in a jugular vein, and after 48h recovery receiving feed at 90% of the measured feed intake, they were fasted for eight hours to study preprandial metabolism (-5, -1 min before the meal). In sequence, all pigs received a 300 g meal and blood samples were collected at 10, 15, 20, 30, 40, 50, 60, 75, 90, 120, 150, 178, 178 min, relative to the meal, to measure AA, insulin and glucose concentrations. In addition, insulin resistance (HOMA-IR) was calculated based on fasting glucose and insulin concentrations. The AA response was modeled for each PD group based on the average group response using differential equations derived from the Erlang equation. Thus, the basal concentration, area under the curve (AUC) and AA flux could be compared among groups using the parameters obtained by the non-linear model. An F-test was used to compare the difference in the indexes between Low and High PD pigs. The basal concentration of methionine, valine and leucine were similar, whereas AUC was greater for Low PD pigs with no difference in flux, with the exception of methionine (P < 0.10; Table 1). Conversely, lysine (15%) and threonine (10%) basal concentrations were less in Low PD pigs. Lysine AUC was greater (8%), whereas others AA AUC were on average 27% less for High PD than for Low PD pigs. Lysine maximal concentration was 11% greater for High PD pigs and occurred later than for Low PD (1.15h vs 1.03h). Such differences in AA postprandial metabolism might be partially due to metabolic differences in response to anabolic hormones such as insulin. For instance, HOMA-IR differed (P < 0.05) among PD categories, with Low PD showing a 44% lower index for insulin resistance than high PD pigs. High PD pigs are less insulin-resistant and show differences in pre- and postprandial AA metabolism.

Iron deficiency attenuates protein synthesis stimulated by branched-chain amino acids and insulin in myotubes

Iron deficiency anemia indicates poor nutrition and is a public health problem. Iron deficiency is also associated with muscle weakness. However, the intracellular mechanisms by which iron deficiency induces muscle weakness are obscure. The purpose of the present study was to evaluate the effect of iron deficiency on protein synthesis in basal and branched-amino acids (BCAA)- and insulin-stimulated state in muscle cells. Differentiated C2C12 myotubes were incubated with an iron chelator, deferoxamine mesylate, and then stimulated with BCAA or insulin to activate protein synthesis. This iron deprivation resulted in a significant reduction in the abundance of iron-containing proteins, such as the mitochondrial complex 1 subunit protein, compared to control cells, but not of protein that does not contain iron, such as citrate synthase. Proteins involved in glucose utilization, such as glucose transpoter-1, hexokinase and AMP-activated protein kinase (AMPK), were upregulated under iron deficiency. Additionally, rates of BCAA- and insulin-stimulated protein synthesis, measured by puromycin incorporation, were lower in iron-deficient myotubes than in control cells. We suggest that low iron availability attenuates BCAA- and insulin-stimulated protein synthesis, possibly via activation of AMPK in myotubes. The present findings advance the understanding of the importance of iron to skeletal muscle protein synthesis and, thus, may contribute to the prevention of sarcopenia and frailty.

Insulin Stimulation of Protein Synthesis and mTOR Signaling in Chick Myotube Cultures.

Insulin stimulates protein synthesis in skeletal muscles. Protein synthesis is controlled by the mechanistic target of rapamycin (mTOR) signaling in skeletal muscles. This study was conducted to investigate the effect of insulin on protein synthesis and mTOR signaling in chick myotube cultures. Chick myotubes were incubated with insulin (1 µg/ml) for 1 h. Protein synthesis, measured using the surface sensing of translation method, was significantly increased by insulin. The phosphorylation of AKT (Thr308 and Ser473), p70 ribosomal S6 kinase 1 (S6K1, Thr389), S6 ribosomal protein (Ser235/236), and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1, Thr37/46) was also significantly increased by insulin. These results suggest that insulin stimulates protein synthesis via mTOR signaling (phosphorylation of AKT, S6K1, S6 ribosomal protein, and 4E-BP1) in chick myotube cultures.

Mechanisms involved in follistatin-induced hypertrophy and increased insulin action in skeletal muscle.

BackgroundSkeletal muscle wasting is often associated with insulin resistance. A major regulator of muscle mass is the transforming growth factor β (TGF‐β) superfamily, including activin A, which causes atrophy. TGF‐β superfamily ligands also negatively regulate insulin‐sensitive proteins, but whether this pathway contributes to insulin action remains to be determined.MethodsTo elucidate if TGF‐β superfamily ligands regulate insulin action, we used an adeno‐associated virus gene editing approach to overexpress an activin A inhibitor, follistatin (Fst288), in mouse muscle of lean and diet‐induced obese mice. We determined basal and insulin‐stimulated 2‐deoxy‐glucose uptake using isotopic tracers in vivo. Furthermore, to evaluate whether circulating Fst and activin A concentrations are associated with obesity, insulin resistance, and weight loss in humans, we analysed serum from morbidly obese subjects before, 1 week, and 1 year after Roux‐en‐Y gastric bypass (RYGB).ResultsFst288 muscle overexpression markedly increased in vivo insulin‐stimulated (but not basal) glucose uptake (+75%, P < 0.05) and increased protein expression and intracellular insulin signalling of AKT, TBC1D4, PAK1, pyruvate dehydrogenase‐E1α, and p70S6K, while decreasing TBC1D1 signaling (P < 0.05). Fst288 increased both basal and insulin‐stimulated protein synthesis, but no correlation was observed between the Fst288‐driven hypertrophy and the increase in insulin‐stimulated glucose uptake. Importantly, Fst288 completely normalized muscle glucose uptake in insulin‐resistant diet‐induced obese mice. RYGB surgery doubled circulating Fst and reduced activin A (−24%, P < 0.05) concentration 1 week after surgery before any significant weight loss in morbidly obese normoglycemic patients, while major weight loss after 1 year did not further change the concentrations.ConclusionsWe here present evidence that Fst is a potent regulator of insulin action in muscle, and in addition to AKT and p70S6K, we identify TBC1D1, TBC1D4, pyruvate dehydrogenase‐E1α, and PAK1 as Fst targets. Circulating Fst more than doubled post‐RYGB surgery, a treatment that markedly improved insulin sensitivity, suggesting a role for Fst in regulating glycaemic control. These findings demonstrate the therapeutic potential of inhibiting TGF‐β superfamily ligands to improve insulin action and Fst's relevance to muscle wasting‐associated insulin‐resistant conditions in mice and humans.

Hormonal control of growth in the wing imaginal disks of Junonia coenia: the relative contributions of insulin and ecdysone.

The wing imaginal disks of Lepidoptera can be grown in tissue culture, but require both insulin and ecdysone to grow normally. Here, we investigate the contributions the two hormones make to growth. Ecdysone is required to maintain mitoses, whereas in the presence of insulin alone mitoses stop. Both ecdysone and insulin stimulate protein synthesis, but only ecdysone stimulates DNA synthesis. Insulin stimulates primarily cytoplasmic growth and an increase in cell size, whereas ecdysone, by virtue of its stimulation of DNA synthesis and mitosis, stimulates growth by an increase in cell number. Although both hormones stimulate protein synthesis, they do so in different spatial patterns. Both hormones stimulate protein synthesis in the inter-vein regions, but ecdysone stimulates synthesis more strongly in the veins and in the margin of the wing disk. We propose that the balance of insulin and ecdysone signaling must be regulated to maintain normal growth, and when growth appears to be due primarily to an increase in cell number, or an increase in cell size, this may indicate growth occurred under conditions that favored a stronger role for ecdysone, or insulin, respectively.

GW28-e0155 Decrease Lean Mass and Increase Fat Mass of SGA Adult Rats Associated with Down-Regulation of the mTOR/S6K Pathway in Skeletal Muscle

Small for gestational age (SGA) is linked to increased risk of obesity and metabolic disorders in adulthood. Amino acids and insulin stimulate protein synthesis through mammalian target of rapamycin (mTOR) signaling pathway. Phosphorylation of ribosomal protein S6 kinase (S6K) is the effector of

Insulin Receptor β-Subunit Haploinsufficiency Impairs Hippocampal Late-Phase LTP and Recognition Memory

The insulin receptor (IR) is a protein tyrosine kinase playing a pivotal role in the regulation of peripheral glucose metabolism and energy homoeostasis. IRs are also abundantly distributed in the cerebral cortex and hippocampus, where they regulate synaptic activity required for learning and memory. As the major anabolic hormone in mammals, insulin stimulates protein synthesis partially through the activation of the PI3K/Akt/mTOR pathway, playing fundamental roles in neuronal development, synaptic plasticity and memory. Here, by means of a multidisciplinary approach, we report that long-term synaptic plasticity and recognition memory are impaired in IR β-subunit heterozygous mice. Since IR expression is diminished in type-2 diabetes as well as in Alzheimer's disease (AD) patients, these data may provide a mechanistic link between insulin resistance, impaired synaptic transmission and cognitive decline in humans with metabolic disorders.

Rag GTPases and AMPK/TSC2/Rheb mediate the differential regulation of mTORC1 signaling in response to alcohol and leucine

Leucine (Leu) and insulin both stimulate muscle protein synthesis, albeit at least in part via separate signaling pathways. While alcohol (EtOH) suppresses insulin-stimulated protein synthesis in cultured myocytes, its ability to disrupt Leu signaling and Rag GTPase activity has not been determined. Likewise, little is known regarding the interaction of EtOH and Leu on the AMPK/TSC2/Rheb pathway. Treatment of myocytes with EtOH (100 mM) decreased protein synthesis, whereas Leu (2 mM) increased synthesis. In combination, EtOH suppressed the anabolic effect of Leu. The effects of EtOH and Leu were associated with coordinate changes in the phosphorylation state of mTOR, raptor, and their downstream targets 4EBP1 and S6K1. As such, EtOH suppressed the ability of Leu to activate these signaling components. The Rag signaling pathway was activated by Leu but suppressed by EtOH, as evidenced by changes in the interaction of Rag proteins with mTOR and raptor. Overexpression of constitutively active (ca)RagA and caRagC increased mTORC1 activity, as determined by increased S6K1 phosphorylation. Furthermore, the caRagA-caRagC heterodimer blocked the inhibitory effect of EtOH. EtOH and Leu produced differential effects on AMPK signaling. EtOH enhanced AMPK activity, resulting in increased TSC2 (S1387) and eEF2 phosphorylation, whereas Leu had the opposite effect. EtOH also decreased the interaction of Rheb with mTOR, and this was prevented by Leu. Collectively, our results indicate that EtOH inhibits the anabolic effects that Leu has on protein synthesis and mTORC1 activity by modulating both Rag GTPase function and AMPK/TSC2/Rheb signaling.

Chromium Improves Protein Deposition Through Regulating the mRNA Levels of IGF-1, IGF-1R, and Ub in Rat Skeletal Muscle Cells

The aim of this study was to evaluate the impact of three different chromium forms--chromic chloride (CrCl3), chromium picolinate (CrPic), and a newly synthesized complex of chromium chelated with small peptides (CrSP)--on protein metabolism in vitro. In cultured skeletal muscle cells, CrSP was able to increase the basal and insulin-stimulated levels of protein deposition in skeletal muscles cells. CrCl3 and CrPic augmented insulin-stimulated protein synthesis. At the molecular level, insulin significantly increased the mRNA levels of insulin-like growth factor 1 and insulin-like growth factor 1 receptor. These impacts could be enhanced by the addition of chromium, especially CrSP. The mRNA levels of ubiquitin were significantly reduced when cells were cultured with chromium or/and insulin. Assuming that the mRNA level increase or decrease results in increased or decreased levels of these proteins, chromium would improve protein anabolism and reduce protein catabolism and then prove protein deposition in rat skeletal muscle cells.

Effect of thyroid hormone on insulin action in cultured arterial smooth muscle cells.

The effect of triiodothyronine (T3) on insulin action in cultured smooth muscle cells of the rat aorta were studied. Insulin binding to smooth muscle cells was increased by T3. The Scatchard analysis indicated an increase of insulin binding sites and an elevation of the affinity of insulin to its binding sites. T3 alone had no significant effect on DNA, RNA and protein synthesis in smooth muscle cells and did not increase the transport of 3-O-methylglucose into the cells. However, the insulin-induced 3-O-methylglucose transport into the cells was stimulated. The insulin-stimulated protein synthesis was not significantly altered by T3 treatment, whereas the effect of insulin on DNA and RNA synthesis was even reduced in the presence of T3. The study suggests, that T3 treatment produces predominantly a stimulation of processes which are necessary for the production of energy. The anabolic effect of insulin is decreased and must by inferior to the credit of energy production.

Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40

Insulin stimulates protein synthesis and cell growth by activation of the protein kinases Akt (also known as protein kinase B, PKB) and mammalian target of rapamycin (mTOR). It was reported that Akt activates mTOR by phosphorylation and inhibition of tuberous sclerosis complex 2 (TSC2). However, in recent studies the physiological requirement of Akt phosphorylation of TSC2 for mTOR activation has been questioned. Here, we identify PRAS40 (proline-rich Akt/PKB substrate 40 kDa) as a novel mTOR binding partner that mediates Akt signals to mTOR. PRAS40 binds the mTOR kinase domain and its interaction with mTOR is induced under conditions that inhibit mTOR signalling, such as nutrient or serum deprivation or mitochondrial metabolic inhibition. Binding of PRAS40 inhibits mTOR activity and suppresses constitutive activation of mTOR in cells lacking TSC2. PRAS40 silencing inactivates insulin-receptor substrate-1 (IRS-1) and Akt, and uncouples the response of mTOR to Akt signals. Furthermore, PRAS40 phosphorylation by Akt and association with 14-3-3, a cytosolic anchor protein, are crucial for insulin to stimulate mTOR. These findings identify PRAS40 as an important regulator of insulin sensitivity of the Akt-mTOR pathway and a potential target for the treatment of cancers, insulin resistance and hamartoma syndromes.

Activation of Mammalian Target of Rapamycin (mTOR) by Insulin Is Associated with Stimulation of 4EBP1 Binding to Dimeric mTOR Complex 1

Insulin stimulates protein synthesis by promoting phosphorylation of the eIF4E-binding protein, 4EBP1. This effect is rapamycin-sensitive and mediated by mammalian target of rapamycin (mTOR) complex 1 (mTORC1), a signaling complex containing mTOR, raptor, and mLST8. Here we demonstrate that insulin produces a stable increase in the kinase activity of mTORC1 in 3T3-L1 adipocytes. The response was associated with a marked increase in 4EBP1 binding to raptor in mTORC1, and it was abolished by disrupting the TOR signaling motif in 4EBP1. The stimulatory effects of insulin on both 4EBP1 kinase activity and binding occurred rapidly and at physiological concentrations of insulin, and both effects required an intact mTORC1. Results of experiments involving size exclusion chromatography and coimmunoprecipitation of epitope-tagged subunits provide evidence that the major insulin-responsive form is dimeric mTORC1, a structure containing two heterotrimers of mTOR, raptor, and mLST8.

MTOR-dependent stimulation of the association of eIF4G and eIF3 by insulin

Insulin stimulates protein synthesis by increasing translation initiation. This response is mediated by mTOR and is believed to result from 4EBP1 phosphorylation, which allows eIF4E to bind eIF4G. Here, we present evidence that mTOR interacts directly with eIF3 and that mTOR controls the association of eIF3 and eIF4G. Activating mTOR signaling with insulin increased by as much as five-fold the amount of eIF4G bound to eIF3. This novel effect was blocked by rapamycin and other inhibitors of mTOR, and it required neither eIF4E binding to eIF4G nor eIF3 binding to the 40S ribosomal subunit. The increase in eIF4G associated with eIF3 occurred rapidly and at physiological concentrations of insulin. Moreover, the magnitude of the response was similar to the increase in eIF4E binding to eIF4G produced by insulin. Thus, increasing eIF4G association with eIF3 represents a potentially important mechanism by which insulin, as well as amino acids and growth factors that activate mTOR, stimulate translation.

Differential p70S6k and 4E-BP1 regulation by insulin and amino acids in vascular endothelial and smooth muscle cells

Differential stimulation of vascular endothelial and smooth muscle cells proliferation is responsible for atherosclerotic lesions. Amino acids and insulin modulate p70S6k and 4E-BP1 activity, regulating cell growth and proliferation. We hypothesised that nutritional (amino acids) and hormonal (insulin) signals differently modulate protein anabolism in human vascular endothelial (HUVEC) and smooth muscle (HVSMC) cells. We evaluated p70S6kinase and 4E-BP1 phosphorylation in the two cell types, grown in amino acid-free medium with or without insulin (INS, 100 nM) or/and amino acids mixture (AA, 3 mM) and with the selective addition or deprivation of branched chain amino acids (BCAA, 0.5 mM). INS stimulated p70S6k and 4E-BP1 phosphorylation transiently in HUVEC and persistently in HVSMC. AA and INS+AA stimulated p70S6k and 4E-BP1 phosphorylation persistently in HUVEC and HVSMC. AA, but not BCAA alone or BCAA-deprived AA, induced p70S6k phosphorylation in HUVEC. BCAA deprivation decreased the p70S6k phosphorylation induced by AA with or without insulin in HVSMC. These results show that anabolic stimuli modulate p70S6k and 4E-BP1 activity differently in the two vascular cell types, suggesting that insulin stimulates protein synthesis for a longer time in HUSMC than in HUVEC. We speculate that hyperinsulinaemia frequently associated with atherosclerosis could induce a selective HVSMC proliferation.

Acute treatment with TNF-α attenuates insulin-stimulated protein synthesis in cultures of C2C12myotubes through a MEK1-sensitive mechanism

Insulin and TNF-alpha exert opposing effects on skeletal muscle protein synthesis that are mediated in part by the rapamycin-sensitive mammalian target of rapamycin (mTOR) pathway and the PD-98059-sensitive, extracellular signal-regulated kinase (ERK)1/2 pathway. The present study examined the separate and combined effects of insulin (INS), TNF, PD-98059, or dnMEK1 adenovirus on the translational control of protein synthesis in C(2)C(12) myotubes. Cultures were treated with INS, TNF, PD-98059, dnMEK1, or a combination of INS + TNF with PD-98059 or dnMEK1. INS stimulated protein synthesis, enhanced eIF4E.eIF4G association, and eIF4G phosphorylation and repressed eIF4E.4E-BP1 association vs. control. INS also promoted phosphorylation of ERK1/2, S6K1, and 4E-BP1 and dephosphorylation of eIF4E. TNF alone did not have an effect on protein synthesis (vs. control), eIF4E.eIF4G association, or the phosphorylation of eIF4G, S6K1, or 4E-BP1, although it transiently increased ERK1/2 and eIF4E phosphorylation. When myotubes were treated with TNF + INS, the cytokine blocked the insulin-induced stimulation of protein synthesis. This appeared to be due to an attenuation of insulin-stimulated eIF4E.eIF4G association, because other stimulatory effects of INS, e.g., phosphorylation of ERK1/2, 4E-BP1, S6K1, eIF4G, and eIF4E and eIF4E.4E-BP1 association, were unaffected. Finally, treatment of myotubes with PD-98059 or dnMEK1 adenovirus before TNF + INS addition resulted in a derepression of protein synthesis and the association of eIF4G with eIF4E. These findings suggest that TNF abrogates insulin-induced stimulation of protein synthesis in myotubes through a decrease in eIF4F complex assembly independently of S6K1 and 4E-BP1 signaling and dependently on a MEK1-sensitive signaling pathway.

Pancreastatin modulates insulin signaling in rat adipocytes: mechanisms of cross-talk.

Pancreastatin (PST), a chromogranin A-derived peptide, has counterregulatory effects on insulin in the hepatocyte and the adipocyte, suggesting a possible role in insulin resistance. The mechanism of PST action on glucose and lipid metabolism is typical of a calcium-mobilizing hormone and involves a receptor Gq/11 protein-phospholipase C (PLC)-beta pathway. In the rat adipocyte, PST inhibits insulin-mediated glucose transport, glucose utilization, and lipid synthesis, and it has a lipolytic effect but stimulates basal and insulin-stimulated protein synthesis. We have also recently studied the PST receptor-effector system in adipocyte membranes. To further investigate the mechanisms of PST effect on insulin action, we studied the cross-talk of PST with insulin signaling in the rat adipocyte. We found that PST inhibits insulin-stimulated GLUT4 translocation to the membrane, which may explain the reported inhibition of glucose transport. Tyrosine phosphorylation of the activated insulin receptor, insulin receptor substrate (IRS)-1, and p60-70 was also blunted, preventing their association with p85 phosphatidylinositol 3-kinase (PI3K) and their activity. The mechanism of this inhibition involves the activation of the "classical" protein kinase C isoforms and the serine phosphorylation of insulin receptor and IRS-1. On the other hand, PST activates the mitogen-activated protein kinase (MAPK) signaling module and enhances the effect of insulin. This pathway may account for the described effect of PST on protein synthesis. In conclusion, PST seems to inhibit the insulin-stimulated PI3K pathway in the adipocyte, whereas it activates the MAPK pathway. These data provide some clues to the PST cross-talk with insulin signaling that may explain the PST effects on glucose metabolism and protein synthesis.

Glucocorticoid-induced insulin resistance of protein synthesis is independent of the rapamycin-sensitive pathways in rat skeletal muscle.

This study was designed to evaluate the role of p70 S6 kinase (p70(S6K) ), p90 S6 kinase (p90(RSK)) and mitogen-activated protein (MAP) kinase pathways in the insulin resistance of muscle protein synthesis observed during glucocorticoid treatment. Dexamethasone treatment decreased the effect of insulin on protein synthesis (-35. 2%) in epitrochlearis muscle incubated in vitro. This resistance is associated with a total blockage of the stimulation of p70(S6K) by insulin without any significant decrease in the amount of the kinase. However, the effect of rapamycin (inhibitor of several intracellular pathways including p70(S6K) pathways) on muscle protein synthesis was not modified by dexamethasone in rat muscles. This suggested that 'rapamycin-sensitive pathways' associated with the insulin stimulation of protein synthesis were not altered by glucocorticoids and thus are not responsible for the insulin resistance observed. As incubation of muscles with a MAP kinase inhibitor (PD98059) did not modify the stimulation of protein synthesis by insulin and as glucocorticoids did not alter the effect of insulin on p90(RSK )activity, our results provide evidence that glucocorticoid-induced alterations in muscle protein synthesis regulation by insulin do not involve factors or kinases that are dependent on MAP kinase and/or p90(RSK).

Role of ERK1/ERK2 and p70 S6K pathway in insulin signalling of protein synthesis

The signalling pathways by which insulin triggers protein synthesis were studied using an antisense strategy to deplete ERK1/ERK2 and rapamycin to inhibit the p70 S6K pathway. The results indicated that ERK1/ERK2 principally regulated the amount of the protein synthesis machinery available in the cell while the p70 S6K pathway contributed to modulating its activation in response to insulin. ERK1/ERK2 also mediated in a small proportion of insulin-stimulated protein synthesis which included the induction of c-fos protein. When c-fos induction was blocked the majority of insulin-stimulated protein synthesis still occurred and thus did not require transcriptional regulation of c-fos or its targets.

Insulin stimulates phenylalanine uptake across the hind limb in fed lambs

Although insulin has been shown to stimulate muscle protein accretion in nonruminants (e.g., Wray-Cahen et al., 1998) evidence for this same effect in ruminants has been equivocal (e.g., Oddy et al., 1987; Wolff et al., 1989). Moreover, when insulin has been reported to have an anabolic effect in ruminants, the animal has been in the fasted state. In addition, Garlick and Grant (1988) have shown that branched-chain amino acids (BCAA) enhanced insulin-stimulated protein synthesis in fasted rats. The primary aim of the present study was to ascertain whether insulin increases phenylalanine (Phe) uptake (used as an index of skeletal muscle protein anabolism) across the hind limb of fed lambs. A secondary aim was to determine if infusion of BCAA, alone or in combination with insulin, would stimulate Phe uptake greater than insulin alone.

Pancreastatin inhibits insulin action in rat adipocytes.

Pancreastatin (PST), a regulatory peptide with a general inhibitory effect on secretion, is derived from chromogranin A, a glycoprotein present throughout the neuroendocrine system. We have previously demonstrated the counterregulatory role of PST on insulin action in rat hepatocytes. Here, we are reporting the PST effects on rat adipocytes. PST dose dependently inhibits basal and insulin-stimulated glucose transport, lactate production, and lipogenesis, impairing the main metabolic actions of insulin in adipocytes. These effects were observed in a wide range of insulin concentrations, leading to a shift to the right in the dose-response curve. Maximal effect was observed at 10 nM PST, and the IC50 value was approximately 1 nM. Moreover, PST has a lipolytic effect in rat adipocytes (ED50 0.1 nM), although it was completely inhibited by insulin. In contrast, PST dose dependently stimulated protein synthesis and enhanced insulin-stimulated protein synthesis. In summary, these data show the lipokinetic effect of PST and the inhibitory effect of PST on insulin metabolic action within a range of physiological concentrations. Therefore, these results give new pathophysiological basis for the association of PST with insulin resistance.