G6Pase mRNA Research Articles

Identification of Isthmin-1 in common carp (Cyprinus carpio L.) and the effects on glucose metabolism in vivo and in vitro.

Isthmin-1 (Ism1) plays roles in glucose uptake in mammals as an adipokine. To investigate its role in the glucose metabolism of common carp (Cyprinus carpio. L), the Ism1 sequence was cloned, and its expression and distribution in tissues were detected. In addition, we prepared and purified the recombinant Ism1 protein using the E. coli expression system and assessed changes in the expression of key genes related to glucose metabolism through both in vivo injection experiments and primary hepatocyte experiments in vitro. The results revealed that the open reading frame of Ism1 was 1377 bp long, encoding 458 amino acids. Similarity analysis indicated that Ism1 exhibited a close evolutionary relationship with goldfish (Carassius auratus), sharing 98.35% amino acid similarity. Ism1 was expressed in all tissues of common carp, with the highest level observed in the heart, followed by the gill, head kidney, and hepatopancreas. Distinct patterns of Ism1 expression were identified during the oral glucose tolerance test and long-term high-carbohydrate and high-fat diet feeding experiments. In vivo studies demonstrated that the serum glucose concentration was reduced on treatment with Ism1, accompanied by a significant upregulation of mRNA levels for gk, hk, and pfk genes in hepatopancreas; conversely pepck and g6pase mRNA levels were significantly downregulated in the hepatopancreas under these conditions as well. Furthermore, our primary hepatocyte experiment confirmed that Ism1 could inhibit pepck and g6pase mRNA expression, while promoting gk, hk, and pfk mRNA expression levels. In conclusion, Ism1, in common carp, could participate in the glucose metabolism, which provides essential information for future studies on the function of Ism1.

Preparation and identification of hypoglycemic bioactive peptide from Amygdalus communis L. by multienzyme hydrolysis

Amygdalus communis L. is rich in high-quality protein and its natural peptides have potential in controlling and preventing diseases. The objective of this study was to evaluate the hypoglycemic activity of bioactive peptides obtained from Amygdalus communis L. protein by synergistic hydrolysis of trypsin and protamex. After ultrafiltration and purification, active peptide B4 with small molecular weight in four hypoglycemic active peptides showed the best inhibitory activities on α-amylase and α-glucosidase. Moreover, insulin resistance model was established with HepG2 cells to study the hypoglycemic mechanism of active peptide B4. The experimental results confirmed that the active peptide B4 was able to up-regulate GYS1 gene mRNA expression and down-regulate PEPCK, G6Pase and GP gene mRNA expression. Meanwhile, active peptide B4 significantly increased the glucose consumption and glycogen synthesis in IR-HepG2. This research enhances the additional value of nut proteins and contributes to the development of potential functional products to expand their application in the food field.

Regulation of plasma glucose levels by central dopamine D2 receptors is impaired in type 1 but not type 2 diabetic mouse models

Glucose metabolism is reported to be regulated by the central nervous system, but it is unclear whether this regulation is altered in diabetes. We investigated whether regulation of glucose metabolism by central dopamine D2 receptors is altered in type 1 and type 2 diabetic models. Intracerebroventricular injections of both the dopamine D2 receptor agonist quinpirole and the antagonist l-sulpiride induced hyperglycemia in control mice, but not in streptozotocin (STZ)-induced diabetic mice, a type 1 diabetic model. Hyperglycemia induced by quinpirole or l-sulpiride was diminished following fasting and these drugs did not affect hyperglycemia in the pyruvate tolerance test. In addition, both quinpirole and l-sulpiride increased hepatic glucose-6-phosphatase (G6Pase) mRNA. In STZ-induced diabetic mice, dopamine and dopamine D2 receptor mRNA in the hypothalamus, which regulates glucose homeostasis, were decreased. Hepatic glycogen and G6Pase mRNA were also decreased in STZ-induced diabetic mice. Neither quinpirole nor l-sulpiride increased hepatic G6Pase mRNA in STZ-induced diabetic mice. In diet-induced obesity mice, a type 2 diabetic model, both quinpirole and l-sulpiride induced hyperglycemia, and hypothalamic dopamine and dopamine D2 receptor mRNA were not altered. These results indicate that (i) stimulation or blockade of dopamine D2 receptors causes hyperglycemia by increasing hepatic glycogenolysis, and (ii) stimulation or blockade of dopamine D2 receptors does not affect glucose levels in type 1 but does so in type 2 diabetic models. Moreover, hypothalamic dopaminergic function and hepatic glycogenolysis are decreased in the type 1 diabetic model, which reduces hyperglycemia induced by stimulation or blockade of dopamine D2 receptors.

Vulgarin, a Sesquiterpene Lactone from Artemisia judaica, Improves the Antidiabetic Effectiveness of Glibenclamide in Streptozotocin-Induced Diabetic Rats via Modulation of PEPCK and G6Pase Genes Expression.

The current investigation assessed the effect of the eudesmanolid, Vulgarin (VGN), obtained from Artemisia judaica (A. judaica), on the antidiabetic potential of glibenclamide (GLB) using streptozotocin (STZ) to induce diabetes. Seven groups of rats were used in the study; the first group received the vehicle and served as normal control. The diabetic rats of the second to the fifth groups were treated with the vehicle (negative control), GLB at 5 mg/kg (positive control), VGN at 10 mg/kg (VGN-10) and VGN at 20 mg/kg (VGN-20), respectively. The diabetic rats of the sixth and seventh groups were administered combinations of GLB plus VGN-10 and GLB plus VGN-20, respectively. The diabetic rats treated with GLB plus VGN-20 combination showed marked improvement in the fasting blood glucose (FBG), insulin and glycated hemoglobin (HbA1c), as well as the lipid profile, compared with those treated with GLB alone. Further, the pancreatic tissues of the diabetic rats that received the GLB+VGN-20 combination showed superior improvements in lipid peroxidation and antioxidant parameters than those of GLB monotherapy. The insulin content of the β-cells was restored in all treatments, while the levels of glucagon and somatostatin of the α- and δ-endocrine cells were reduced in the pancreatic islets. In addition, the concurrent administration of GLB+VGN-20 was the most effective in restoring PEPCK and G6Pase mRNA expression in the liver. In conclusion, the results demonstrated that the GLB+VGN-20 combination led to greater glycemic improvement in diabetic rats compared with GLB monotherapy through its antioxidant effect and capability to modulate PEPCK and G6Pase gene expression in their livers.

Dietary Protein Modifies Hepatic Glycolipid Metabolism, Intestinal Immune Response, and Resistance to Streptococcus agalactiae of Genetically Improved Farmed Tilapia (GIFT: Oreochromis niloticus) Exposed to High Temperature

The present study investigates the effects of dietary protein levels on glucolipid metabolism, immune function, and resistance to Streptococcus agalactiae of genetically improved farmed tilapia (GIFT) exposed to high temperature. Six practical diets were prepared to feed 360 fish (initial weight 43.78 ± 0.12 g) with graded protein levels (26.45%, 29.28%, 31.69%, 33.68%, 36.18%, and 38.75% dry matter). The results showed that 26.45% dietary protein significantly improved glycolysis by increasing PK mRNA levels, while the 29.28% and 31.69% dietary protein levels promoted gluconeogenesis by increasing PEPCK and G6Pase mRNA levels. For lipid metabolism, 26.45% dietary protein enhanced lipid synthesis by increasing PPAR-γ, SREBP1c, and FAS mRNA levels, while 31.69% dietary protein enhanced the level of lipolysis by increasing the PPAR-α and CPT1 mRNA levels. The highest plasma TG and TC contents were observed in the 29.28% and 31.69% dietary protein groups, respectively. In terms of antioxidants and immunity, the 31.69% dietary protein level activated the expression levels of HSP90 mRNA, thus increasing the expression levels of antioxidant-related genes (CAT, SOD, and GPx), and upregulating the anti-inflammatory factor IL-10 mRNA levels. In addition, regarding the antioxidant enzymes, the highest GSH content was found in the 29.28% dietary protein group, while the 31.69% dietary protein group had the maximum GSH-Px activity. The lowest plasma ALT and AST activities were observed in the 31.69% dietary protein group. Ultimately, the survival rate of juvenile GIFT fed 31.69% dietary protein was highest after a Streptococcus agalactiae challenge. Overall, 29.28–31.69% dietary protein was recommended in the diet of GIFT in a high-temperature environment.

Targeting IRE1α and PERK in the endoplasmic reticulum stress pathway attenuates fatty acid-induced insulin resistance in bovine hepatocytes

Endoplasmic reticulum (ER) stress can be induced by various stimuli and triggers the unfolded protein response to activate intracellular signaling pathways that are mediated by 3 ER-resident sensors: inositol requiring protein-1α (IRE1α), PKR-like ER kinase (PERK), and activating transcription factor-6 (ATF6). In nonruminants, ER stress plays a critical role in hepatic insulin resistance. However, whether ER stress plays a role in nonesterified fatty acid (NEFA)-induced hepatic insulin resistance in dairy cows is still unknown. Experiments were conducted using primary bovine hepatocytes isolated from 5 healthy calves (body weight: 30-40 kg; 1 d old). First, hepatocytes were treated with NEFA (1.2 mM) for 0.5, 1, 2, 3, 5, 7, 9, or 12 h. Treatment with NEFA elevated abundance of phosphorylated IRE1α and PERK, and cleavage of ATF6, along with the ER stress-associated genes XBP1, ATF4, and DNAJC3, resulting in both linear and quadratic effects. Furthermore, ER Tracker red staining and transmission electron microscopy results indicated that ER was dilated and degranulated in response to NEFA treatment, suggesting that ER stress was induced by NEFA treatment in bovine hepatocytes. Second, to assess the effect of ER stress on NEFA-induced insulin resistance, hepatocytes were treated with different concentrations of NEFA (0, 0.6, 1.2, or 2.4 mM) for 5 h with or without tauroursodeoxycholic acid (TUDCA, a canonical inhibitor of ER stress). Here, NEFA induced insulin resistance by increasing the abundance of insulin receptor substrate-1 (IRS1) phosphorylation at the inhibitory residue Ser 307 (S307) and decreasing the abundance of phosphorylated protein kinase B (AKT) and glycogen synthase kinase-3β (GSK3β) in a dose-dependent manner. This was accompanied by upregulation of an abundance of gluconeogenic genes [phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6-Pase)]. These detrimental effects of NEFA on insulin signaling could be reversed with TUDCA treatment, indicating a mechanistic link between ER stress and NEFA-induced insulin resistance. In a third experiment, pGPU6/GFP/Neo vectors containing short hairpin RNA targeting IRE1α were used to silence IRE1α transcription, and GSK2656157 (PERK phosphorylation inhibitor) and 4-(2-aminoethyl) benzenesulfonyl fluoride (AEBSF; an inhibitor of ATF6) were used to block PERK and ATF6 branches, respectively. Notably, the silencing of the IRE1α branch improved NEFA-induced insulin resistance by decreasing phosphorylation of IRS1 (S307) and increasing phosphorylation of AKT and GSK3β, and reducing PEPCK and G6-Pase mRNA abundance, which was likely dependent on IRE1α kinase activity. Similarly, blockage of the PERK branch increased phosphorylation of AKT and GSK3β, and reduced PEPCK and G6-Pase mRNA abundance, but had no effect on phosphorylation of IRS1 (S307). However, results showed that inhibition of the ATF6 branch had no effects on phosphorylation of IRS1, AKT, and GSK3β, and instead found increasing PEPCK and G6-Pase mRNA abundance. Taken together, data in the present study found that impeding IRE1α and PERK signaling might aid in relieving hepatic insulin resistance. However, the more detailed mechanisms of how IRE1α and PERK signaling contribute to hepatic insulin resistance in dairy cows remain to be determined.

Salinity levels affect the lysine nutrient requirements and nutrient metabolism of juvenile genetically improved farmed tilapia (Oreochromis niloticus).

This 62-d research aimed to evaluate the effects of dietary lysine levels (DLL) and salinity on growth performance and nutrition metabolism of genetically improved farmed tilapia (GIFT) juveniles (Oreochromis niloticus). Six diets with lysine supplementation (1·34, 1·70, 2·03, 2·41, 2·72 and 3·04 % of DM) were formulated under different cultured salinities in a two-factorial design. The results indicated that supplemental lysine improved the specific growth rate (SGR) and weight gain (WG) and decreased the feed conversion ratio (FCR). Meanwhile, the fish had higher SGR and WG and lower FCR at 8 ‰ salinity. Except for moisture, the whole-body protein, lipid and ash content of GIFT were increased by 8 ‰ salinity, which showed that DLL (1·34 %) increased the whole-body fat content and DLL (2·41 %) increased whole-body protein content. Appropriate DLL up-regulated mRNA levels of protein metabolism-related genes such as target of rapamycin, 4EBP-1 and S6 kinase 1. However, 0 ‰ salinity reduced these protein metabolism-related genes mRNA levels, while proper DLL could improve glycolysis and gluconeogenesis mRNA levels but decrease lipogenesis-related genes mRNA levels in liver. 0 ‰ salinity improved GLUT2, glucokinase and G6 Pase mRNA levels; however, sterol regulatory element-binding protein 1 and fatty acid synthase mRNA levels were higher at 8 ‰ salinity. Moreover, 8 ‰ salinity also increased plasma total protein and cholesterol levels and decreased glucose levels. These results indicated that the recommended range of lysine requirement under different salinity was 2·03-2·20 % (0 ‰) and 2·20-2·41 % (8 ‰) and 8 ‰ salinity resulted in higher lysine requirements due to changes in the related nutrient metabolism, which might provide useful information for designing more effective feed formulations for GIFT cultured in different salinity environment.

Sodium-glucose co-transporter 1 (SGLT1) differentially regulates gluconeogenesis and GLP-1 receptor (GLP-1R) expression in different diabetic rats: a preliminary validation of the hypothesis of "SGLT1 bridge" as an indication for "surgical diabetes".

BackgroundSodium-glucose co-transporter 1 (SGLT1) may play a synergistic role in gluconeogenesis (GNG) and glucagon-like peptide-1 (GLP-1) expression. We proposed the hypothesis of a “SGLT1 bridge” as an indication for “surgical diabetes” that was preliminary validated in the present study.MethodsWe selected nonobese diabetic Goto-Kakizaki (GK) rats and Zuker diabetic fat (ZDF) rats to represent advanced and early diabetes, respectively. Based on glucose gavage with or without SGLT1 inhibitor phlorizin, the rats were divided into 4 groups: Gk-Glu, GK-P, ZDF-Glu, and ZDF-P. The expressions of SGLT1, GLP-1 receptor (GLP-1R), glucose-6 phosphatase (G6Pase), and phosphoenolpyruvate carboxykinase-1 (Pck1) were determined by immunohistochemistry (IHC) or quantitative reverse transcription polymerase chain reaction (RT-qPCR), and the effects of phlorizin were analyzed.ResultsGlucose tolerance was worse in GK rats and the homeostasis model assessment-insulin resistance (HOMA-IR) was higher in ZDF rats, indicating different pathophysiological conditions between the different diabetic rats. GK rats showed higher activity of duodenal SGLT1 (P=0.022) and jejunal SGLT1 mRNA expression (P=0.000) and lower SGLT1 mRNA expression in the liver (P=0.000) and pancreas (P=0.000). Phlorizin effectively inhibited the activity of duodenal SGLT1 in both GK rats (P=0.000) and ZDF rats (P=0.000). In ZDF rats, the expression of GLP-1R mRNA was downregulated in the jejunum (P=0.001) and upregulated in the pancreas (P=0.021) by phlorizin, but there were no regulatory effects on GLP-1R mRNA in the jejunum and pancreas of GK rats. As for the regulatory effects on GNG, phlorizin upregulated Pck1 mRNA in the duodenum (P=0.000) and the jejunum (P=0.038), whereas it downregulated hepatic G6Pase mRNA in ZDF rats (P=0.005) and Pck1 mRNA expression in GK rats (P=0.001), suggesting that SGLT1 inhibitor may have upregulated intestinal GNG in ZDF rats and downregulated hepatic GNG in both ZDF and GK rats.ConclusionsSGLT1 showed synergistic regulatory effects on the entero-insular axis (EIA) and the gut-brain-liver axis (GBLA), preliminarily validating the hypothesis of a “SGLT1 bridge”. The distinct expression of SGLT1 and its differentially regulatory effects on diabetic rats with different pathophysiological conditions may provide probable potential indications involved in the “Surgical Diabetes” that is supposed as the inclusion for diabetic surgery.

A study to explore the effects of low dietary protein levels on the growth performance and nutritional metabolism of grass carp (Ctenopharyngodon idella) fry

An 8-week feeding trial was conducted to investigate the effects of low dietary protein levels (DPLs) on the growth performance and nutritional metabolism of grass carp (Ctenopharyngodon idella) fry. The results showed that the average final weight (AFW), weight gain rate (WGR) and specific growth rate (SGR) reached the maximum values at 43.30% DPL, and then became constant (P < 0.05). The feed conversion ratio (FCR) showed an opposite trend (P < 0.05). Low DPLs (18.20% and 24.30%) suppressed the survival rate (SR) (P < 0.05). The protein efficiency ratio (PER) reached the maximum value at 36.40% DPL (P < 0.05). The moisture content was lowered by at 36.40% DPL (P < 0.05). The crude protein content was improved by 43.30% DPL (P < 0.05). Low DPLs (18.20% and 24.30%) significantly increased the fat composition compared to other DPLs (P < 0.05). 43.30% DPL significantly promoted the rapamycin target protein (TOR) and ribosomal protein S6 kinase-polypeptide 1 (S6K1) mRNA levels, but inhibited the 4E binding protein 1 (4EBP1) and adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK) mRNA levels (P < 0.05). Low DPLs (18.20% and 24.30%) increased fat synthesis-related genes, the mRNA levels of lipolysis-related genes were significantly higher at 43.30% DPL (P < 0.05). The glucokinase (GK) mRNA levels at 18.20%, 24.30% and 30.60% DPLs were higher (P < 0.05). The glucose 6 phosphatase (G6Pase) mRNA level was higher at 43.30% DPL (P < 0.05). Hepatopancreas transcriptome results showed that compared with the low DPL (18.20%), the appropriate DPL (43.30%) significantly affected the amino acid metabolism and lipid metabolism. This study therefore, demonstrated that low DPLs adversely affected the growth and nutrient metabolism in grass carp fry.

A novel amino acid signaling process governs glucose-6-phosphatase transcription

SummaryEmerging evidence has shown that amino acids act as metabolic regulatory signals. Here, we showed that glucose-6-phosphatase (G6Pase) mRNA levels in cultured hepatocyte models were downregulated in an amino-acid-depleted medium. Inversely, stimulation with amino acids increased G6Pase mRNA levels, demonstrating that G6Pase mRNA level is directly controlled by amino acids in a reversible manner. Promoter assay revealed that these amino-acid-mediated changes in G6Pase mRNA levels were attributable to transcriptional regulation, independent of canonical hormone signaling pathways. Metabolomic analysis revealed that amino acid starvation induces a defect in the urea cycle, decreasing ornithine, a major intermediate, and supplementation of ornithine in an amino-acid-depleted medium fully rescued G6Pase mRNA transcription, similar to the effects of amino acid stimulation. This pathway was also independent of established mammalian target of rapamycin complex 1 pathway. Collectively, we present a hypothetical concept of “metabolic regulatory amino acid signal,” possibly mediated by ornithine.

The gluconeogenic glucose-6-phosphatase gene is expressed during oxygen-limited conditions in the white shrimp Penaeus (Litopenaeus) vannamei: Molecular cloning, membrane protein modeling and transcript modulation in gills and hepatopancreas.

The white shrimp Penaeus (Litopenaeus) vannamei is the most economically important crustacean species cultivated in the Western Hemisphere. This crustacean shifts its metabolism to survive under extreme environmental conditions such as hypoxia, although for a limited time. Glucose-6-phosphatase (G6Pase) is a key enzyme contributing to maintain blood glucose homeostasis through gluconeogenesis and glycogenolysis. To our knowledge, there are no current detailed studies about cDNA or gene sequences of G6Pase from any crustacean reported. Herein we report the shrimp P. (L.) vannamei cDNA and gene sequences. The gene contains seven exons interrupted by six introns. The deduced amino acid sequence has 35% identity to other homolog proteins, with the catalytic amino acids conserved and phylogenetically close to the corresponding invertebrate homologs. Protein molecular modeling predicted eight transmembrane helices with the catalytic site oriented towards the lumen of the endoplasmic reticulum. G6Pase expression under normoxic conditions was evaluated in hepatopancreas, gills, and muscle and the highest transcript abundance was detected in hepatopancreas. In response to different times of hypoxia, G6Pase mRNA expression did not change in hepatopancreas and became undetectable in muscle; however, in gills, its expression increased after 3h and 24h of oxygen limitation, indicating its essential role to maintain glycemic control in these conditions.

Effect of dietary Clostridium autoethanogenum protein on growth, body composition, plasma parameters and hepatic genes expression related to growth and AMPK/TOR/PI3K signaling pathway of the genetically improved farmed tilapia (GIFT: Oreochromis niloticus) juveniles

This study investigated the effect of dietary Clostridium autoethanogenum (C. autoethanogenum) protein on growth, whole-body composition, plasma parameters, and hepatic genes expression related to growth and AMPK/TOR/PI3K signaling pathway of the genetically improved farmed tilapia (GIFT:Oreochromis niloticus) juveniles. The fish (initial weight of 0.7 ± 0.05 g) were fed with diets containing C. autoethanogenum protein at 0 g/kg (Control, diet 1), 50 g/kg (Diet 2), 100 g/kg (Diet 3), 150 g/kg (Diet 4) and 200 g/kg (Diet 5) to partially replace mainly soybean meal in triplicates. After 8 weeks feeding trial, the growth was significantly (P < 0.05) improved by dietary C. autoethanogenum protein compared with the control. Whole-body crude lipid increased in the fish group fed over 50 g/kg C. autoethanogenum protein (P < 0.05). Plasma triglycerides (TG) was significantly (P < 0.05) elevated by dietary C. autoethanogenum protein at 200 g/kg. C. autoethanogenum protein supplementation significantly (P < 0.05) up-regulated the mRNA expression of the growth-related insulin-like growth factor-1 (IGF-1). 50 g/kg and 100 g/kg dietary C. autoethanogenum protein improved the phosphorylation and protein levels of the eukaryotic initiation factor 4E binding protein 1 (4E-BP1) and ribosomal protein S6 Kinase 1 (S6K1), and the phosphorylation of the adenosine monophosphate-activated protein kinase (AMPK) compared with the control diet. The phosphorylation level of the phosphoinositide 3-kinase (PI3K) was lowered by 200 g/kg autoethanogenum protein compared with the control. The pyruvate kinase (PK) mRNA level was significantly (P < 0.05) up regulated by 100 g/kg C. autoethanogenum protein; while glucose 6-phosphatase (G6Pase) mRNA expression was down-regulated by 150 g/kg and 200 g/kg C. autoethanogenum protein compared with the control (P < 0.05). The mRNA expression of fatty acid transport protein 5 (FATP5) was significantly down-regulated by 100–200 g/kg; peroxisome proliferator-activated receptor alpha (PPARα) was significantly down-regulated by 50 g/kg; while fatty acid synthase (FAS) was significantly up-regulated by 150 g/kg and 200 g/kg C. autoethanogenum protein compared with the control (P < 0.05). The results further showed that GIFT juveniles fed with dietary C. autoethanogenum protein could regulate whole-body energy homeostasis through the AMPK signaling pathway. Therefore, C. autoethanogenum protein could successfully be used as a novel and sustainable protein in juvenile GIFT diets.

Puerarin suppresses the hepatic gluconeogenesis via activation of PI3K/Akt signaling pathway in diabetic rats and HepG2 cells

Pueraria, a Chinese herbal medicine, plays an important role in many classic prescriptions for the treatment of diabetes. Puerarin is the main component of pueraria. The current in vivo and in vitro research mainly focus on exploring the potential mechanism of puerarin in inhibiting hepatic gluconeogenesis. The type 2 diabetic rats were established by a combination of small dosage of streptozotocin (STZ) injection with high-fat diet. After the administration of puerarin 4 weeks, the parameters of the glucose and lipid metabolism were determined. HepG2 cells were treated by palmitic acid (PA) to induce the insulin resistance in vitro model. After the treatment of puerarin, the glucose consumption and cell viability were examined. Then, the protein expression of PI3K, Akt, pAkt, pFOXO1, FOXO1, PEPCK and G6pase in liver tissue and HepG2 cells were evaluated by western blot. RT-PCR was used to measure the content of PEPCK, G6pase mRNA in liver tissue. The results showed that puerarin administration significantly decrease the level of FBG, HbA1C and triglycerides in diabetic rats. Mechanistic research showed that puerarin activating PI3K/Akt is puerarin-mediated beneficial effects and can be reversed by inhibitor of PI3K or Akt. In conclusion, puerarin inhibits hepatic gluconeogenesis by activating PI3K/Akt signaling pathway.

Effects of dietary lysine levels on growth performance, whole body composition and gene expression related to glycometabolism and lipid metabolism in grass carp, Ctenopharyngodon idellus fry

8 weeks rearing trial was designed to estimate the dietary lysine requirement, and evaluate the effects of dietary lysine on glycometabolism and lipid metabolism of grass carp fry. Six practical diets were prepared to feed 720 fish (initial weight 0.36 ± 0.00 g) with graded lysine levels (1.44%, 1.79%, 1.97%, 2.44%, 2.56% and 2.87%, dry diet). The results displayed that the final body weight (FBW), weight gain rate (WGR) and specific growth rate (SGR) were significantly improved by 2.44% dietary lysine, the same as feed intake (FI) and feed conversion ratio (FCR), compared with those of the control group (1.44% dietary lysine). Whole-body lipid contents diminished with raising dietary lysine levels from 1.44% to 2.44% and thereafter the content increased. Regarding glycometabolism, compared to the control diet, the 2.44% dietary lysine level significantly improved glycolysis by increasing glucokinase (GK) and pyruvate kinase (PK) mRNA levels, meanwhile the 2.87% dietary lysine level promoted gluconeogenesis by significantly increasing phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) mRNA levels. Glycogen synthase (GS) mRNA levels were independent of dietary treatment. For the lipolysis pathway, the 2.44% dietary lysine level caused the upregulation of hormone sensitive lipase (HSL), adipose triglyceride lipase (ATGL) and carnitine palmitoyl transferase-1 (CPT1) mRNA levels in the PPAR signaling pathway. Like the fatty acid synthase (FAS), acetyl-CoA carboxylase-1 (ACC1) and stearyl-CoA desaturase (SCD), the relative mRNA expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) related to lipid synthesis in the PPAR signaling pathway, was not remarkably influenced by dietary lysine levels. Furthermore, sterol-regulatory element binding protein-1 (SREBP1) mRNA levels in the intestine were not clearly affected among all the treatments. According to the broken-line regression analysis, based on the SGR and FCR, the optimum requirements of grass carp fry for dietary lysine were 2.39% and 2.13% of diet (6.34% and 5.65% of dietary protein), respectively. Overall, appropriate dietary lysine levels (2.44%) could improve the growth performance, reduce the whole-body lipid, promote glycolysis and lipolysis.

Quercetin and Isoquercitrin Inhibiting Hepatic Gluconeogenesis Through LKB1-AMPKα Pathway.

To observe the impact of quercetin and isoquercitrin on gluconeogenesis in hepatocytes. Mouse primary hepatocytes were cultured with lactic acid and pyruvic acid. After treatment with quercetin and isoquercitrin for 24 hours, the glucose concentration in the culture supernatant was determined. RT-PCR was used to detect the mRNAs of PEPCK, G6Pase, LKB1, and AMPKα. Protein levels of LKB1, AMPKα, and Thr172 phosphorylation were evaluated by Western blot. The glucose concentration in the gluconeogenesis group (GN) was significantly higher than in the control group (C), but the glucose concentrations in the high level quercetin(group 80Q) and high level isoquercitrin (group 80I) were significantly lower than in the group GN, P<0.01. In the group 80Q, and group 80I, the mRNA levels of PEPCK and LKB1were significantly lower than in the group GN (P<0.01), and the G6Pase mRNA were significantly lower than in the group GN (P<0.05). The protein levels of LKB1 and the phosphorylation of AMPKα Thr172 in the group 80Q, group 40I, and group 80I were higher than in the group GN. The effects of quercetin and isoquercitrin on LKB1 and AMPKα were similar to those of metformin. Quercetin and isoquercitrin inhibit gluconeogenesis in hepatocytes, which may be related to the LKB1 upregulation and phosphorylation of AMPKα.

Acute Exercise Intervention Combined with Metformin’s Influences on Glucose Homeostasis in T2D Mice

PURPOSE: The aim of our investigation was to determine the effects of different ways of acute exercise intervention combined with high dose of metformin on glucose homeostasis and its relative molecular mechanisms in type 2 diabetic mice. METHODS: 4-week high fat diet (HFD) and one-time Streptozocin (100mg/kg) intraperitoneal injection were used for building T2D mice. 24 mice were divided into normal control (NC), normal acute resistance training (NCR) and normal acute endurance training (NCE) group, all n=8, fed in normal chow. Finally 48 mice were developing T2D and divided into diabetic control (DC), diabetic acute resistance training (DCR), diabetic acute endurance training (DCE), high dose of metformin (200mg/kg) control (HMC), metformin combined with acute resistance training (HMR) and metformin combined with acute endurance continuous training (HME) group, all n=8. RESULTS: The two ways also enhanced blood glucose and lipid metabolism in T2D mice. Compared to HMC group, hepatic G6Pase mRNA expression in HMR and HME group was significantly escalated and hepatic FBP1 mRNA expression of both groups were significantly declined. Compared to HMC group, hepatic GLUT2 and Gck mRNA expression in HMR and HME group showed opposite trends, one was down and the other was up. Compared to HMC group, hepatic PEPCK mRNA expression in HMR group mice was notably raised and hepatic AMPKα2, PGC-1α and CREB mRNA expression in HMR and HME group mice were notably increased and only hepatic AMPKα1 mRNA expression in HMR group was significantly increased. CONCLUSIONS: Acute resistance training (ART) and acute endurance training (AET) combined with metformin can effectively improve glucose homeostasis in T2D mice. And the two ways can improve blood glucose and lipid metabolism in T2D mice. ART combined with metformin was better to improve glucose homeostasis and inhibit hepatic gluconeogenesis relative mRNA expression in T2D mice probably via the signaling pathway of AMPKα-PGC-1α-CREB.

SAT-183 Activating Transcription Factor 4 (ATF4) As A Possible Mediator Of Gluconeogenic And Lipogenic Actions Of Fat Mass And Obesity Associated (FTO) In Liver

Hepatic metabolism is critical for the maintenance of whole body glucose and lipid metabolism. Excessive hepatic glucose production and de novo lipogenesis contributes to fasting hyperglycemia in diabetes and hepatic steatosis, respectively. Common genetic variants of the fat mass and obesity associated (FTO) gene are strongly associated with obesity and type 2 diabetes. However, the role of FTO in the regulation of hepatic metabolism is not well understood. We have demonstrated that increased FTO level leads to an increase in gluconeogenic glucose-6-phosphatase (G6Pase) mRNA expression in hepatic cells. Conversely, reduced FTO level or inhibition of FTO activity caused a reduction of G6Pase mRNA expression. The transcription factor activating transcription factor 4 (ATF4) is a positive regulator of the expression of gluconeogenic and lipogenic genes in hepatocytes. These findings led to the hypothesis that increased hepatic FTO expression or activity leads to an increase expression of not only gluconeogenic genes but also lipogenic genes by activating ATF4, while reduced hepatic FTO expression or activity produces opposite effects. To address this hypothesis, we examined the effect of altered FTO expression on ATF4 and lipogenic stearoyl-CoA desaturase (SCD1) mRNA levels in alpha mouse liver 12 (AML12) immortalized hepatic cell line. We also treated AML12 cells with a small molecule FTO inhibitor rhein and measured the expression level of these genes. FTO overexpression resulted in increased ATF4 and SCD1 mRNA levels. FTO knockdown resulted in a reduction of SCD1 mRNA levels. Similarly, rhein treatment (1 μM) for 24 h reduced levels of ATF4 and SCD1 mRNA without causing cytotoxicity. These data support the hypothesis that FTO acts to shift hepatic metabolism toward anabolism, leading to increased gluconeogenesis and de novo lipogenesis and ATF4 is a possible mediator of this action. Our data also suggest that reduced activity of hepatic signaling pathways involving FTO and ATF4 may be effective in reversing hyperglycemic and hepatic steatosis partly by reducing gluconeogenesis and de novo lipogenesis in the liver. Sources of Research Support: Supported by University Research Grants Program (URGP) from University of Manitoba.

PO-062 Acute exercise intervention combined with metformin’s influences on glucose homeostasis in T2D mice

PO-062 Acute exercise intervention combined with metformin’s influences on glucose homeostasis in T2D mice

Fc Gamma Receptor IIb Expressed in Hepatocytes Promotes Lipid Accumulation and Gluconeogenesis

Non-alcoholic fatty liver disease (NAFLD) is characterized by ectopic lipid accumulation in the liver, usually combined with hepatic insulin resistance. Fc-gamma receptor-IIb (FcγRIIb) and its ligand are reported to be associated with obesity and type 2 diabetes mellitus (T2DM). As knowledge about FcγRIIb in the literature is mostly generated from studies on skeletal muscle tissue, the expression and function of FcγRIIb in the liver and hepatocytes are largely unknown. In this study, we identified the expression of FcγRIIb in primary cultured mouse hepatocytes: FcγRIIb was upregulated in response to oleic acid (OA) in a dose dependent manner. FcγRIIb knockdown using shRNA suppressed the lipid and triglyceride accumulation, and mRNA expression of ACC1, FASn, CD36, MTTP, and ApoB in OA-treated HepG2 cells. FcγRIIb deficiency mice fed with high fat diet (HFD) had significantly lower liver weight and liver to body weight ratio, as well as less triglyceride accumulation in the livers. In glycometabolism, FcγRIIb hindered insulin-induced phosphorylation of AKT and FOXO1, and in turn upregulated G6Pase and PEPCK mRNA expression, suggesting that FcγRIIb promotes gluconeogenesis by suppressing the AKT/FOXO1/G6Pase/PEPCK pathway in hepatocytes. This study reveals a novel role for FcγRIIb in regulating lipid metabolism and glycometabolism, and provides a new therapeutic target to improve NAFLD.

Nesfatin-1 Regulates Feeding, Glucosensing and Lipid Metabolism in Rainbow Trout

Nesfatin-1 is an 82 amino acid peptide that has been involved in a wide variety of physiological functions in both mammals and fish. This study aimed to elucidate the role of nesfatin-1 on rainbow trout food intake, and its putative effects on glucose and fatty acid sensing systems. Intracerebroventricular administration of 25 ng/g nesfatin-1 resulted in a significant inhibition of appetite, likely mediated by the activation of central POMC and CART. Nesfatin-1 stimulated the glucosensing machinery (changes in sglt1, g6pase, gsase, and gnat3 mRNA expression) in the hindbrain and hypothalamus. Central fatty acid sensing mechanisms were unaltered by nesfatin-1, but this peptide altered the expression of mRNAs encoding factors regulating lipid metabolism (fat/cd36, acly, mcd, fas, lpl, pparα, and pparγ), suggesting that nesfatin-1 promotes lipid accumulation in neurons. In the liver, intracerebroventricular nesfatin-1 treatment resulted in decreased capacity for glucose use and lipogenesis, and increased the potential of fatty acid oxidation. Altogether, the present results demonstrate that nesfatin-1 is involved in the homeostatic regulation of food intake and metabolism in fish.