PEPCK Expression Research Articles

Hovenia dulcis (Guaizao) polysaccharide ameliorates hyperglycemia through multiple signaling pathways in rats with type 2 diabetes mellitus

Type 2 diabetes mellitus (T2DM) poses a significant threat to human health, with its incidence and mortality rates increasing annually. This study investigated the hypoglycemic effects and underlying mechanisms of pure Hovenia dulcis (Guaizao) polysaccharide (HDPs-2A) in rats subjected to a high-fat and high-sugar diet combined with streptozotocin-induced T2DM. Oral administration of HDPs-2A resulted in significant increases in body weight and liver glycogen levels compared to untreated controls. Moreover, a reduction in fasting blood glucose levels, alleviation of hyperinsulinemia, enhanced glucose tolerance, and improved insulin resistance were observed in the HDPs-2A-treated group. HDPs-2A also effectively reversed diabetes-induced dyslipidemia, as evidenced by decreased total cholesterol and triglyceride levels, alongside increased high-density lipoprotein cholesterol levels. Histopathological analyses confirmed that HDPs-2A partially repaired liver tissue damage by mitigating oxidative stress responses in the liver. Additionally, treatment with HDPs-2A significantly elevated short-chain fatty acid levels in T2DM rats. Real-time quantitative PCR and Western blot analyses indicated that HDPs-2A significantly enhanced the expression of InsR, IRS2, PI3K, Akt, and GLUT4, suggesting that HDPs-2A regulates insulin resistance and glycometabolism through the activation of the PI3K/Akt signaling pathway. Furthermore, HDPs-2A appeared to modulate the expression of GS, GSK-3β, and FoxO1 to improve glucose metabolism and reduce insulin resistance. It also improved glucose metabolism by activating the AMPK pathway and modulating G6Pase and PEPCK expression. This study provides novel insights into the antidiabetic effects of HDPs, positioning them as promising nutritional agents for the management of T2DM.

Research on Function of Ribosomal Protein S6 Kinases, 1α and β, Based on Molecular Cloning and siRNA-Based Interference in Juvenile Blunt Snout Bream (Megalobrama amblycephala).

The aim of this study was to investigate the effects of S6K1α and β on the expression of glycolysis- and gluconeogenesis-related genes in juvenile blunt snout bream. Two isoforms, α and β, of ribosomal protein S6 kinase 1 in blunt snout bream were cloned and characterized, and their expression patterns were examined in vivo. The sequence analysis showed that s6k1α and s6k1β contain open reading frames of 2217 and 1497 bp, encoding 738 and 498 amino acids, respectively. Both S6K1α and S6K1β consist of an S_TKc domain and an extended S_TK_X domain. s6k1α and s6k1β were abundantly expressed in the heart and gonads. siRNAs were designed, and the experiment showed that α-siRNA inhibited s6k1α and s6k1β expression, but β-siRNA exclusively inhibited s6k1α expression (p < 0.05). α-siRNA upregulated the expression levels of gk and pk, while β-siRNA upregulated pepck and g6p expression (p < 0.05). The expression of g6pdh was found to be downregulated, but the gs mRNA level was overexpressed after treatment with α-siRNA and β-siRNA (p < 0.05). In the present experiment, S6K1α was more intimately involved in the regulation of gluconeogenesis when only S6K1α was inhibited, whereas the inhibition of both S6K1α and S6K1β collectively co-regulated glycolysis.

Protopanaxadiol synergizes with glucocorticoids to enhance the therapeutic effect in adriamycin-induced nephrotic syndrome

To date, glucocorticoids remain the mainstay of treatment of nephrotic syndrome (NS). However, serious side effects and development of drug-resistance following long-term use limit the application of glucocorticoids. Protopanaxadiol (PPD) possesses activity of dissociating transactivation from transrepression by glucocorticoid receptor (GR), which may serve as a potential selective GR modulator. However, steroid-like effects of PPD in vivo are unclear and not defined. How to translate PPD into clinical practice remains to be explored. The current study explored the renoprotection and potential mechanism of PPD and its combination with steroid hormones using adriamycin-induced NS rats. Adriamycin was given intravenously to rats to induce nephropathy. The determination of proteinuria, biochemical changes and inflammatory cytokines were performed, and pathological changes were examined by histopathological examination. Immunostaining and PCR were used to analyze the expression of interesting proteins and genes. The results showed that PPD, alone and in combination with prednisone, efficiently alleviate the symptoms of NS, attenuate nephropathy, improve adriamycin-induced podocyte injury by reducing desmin and increasing synaptopodin expression. In addition, the combined treatment reduced the expression of NF-κB protein and mRNA, as well as cytokine levels, and yet increased the expression of GR protein and mRNA. PPD modulated the transactivation of GR, manifested as repressing TAT, PEPCK and ANGPTL4 mRNA expressions mediated by GR. Meanwhile, PPD inhibited elevation of blood glucose and immune organ atrophy induced by prednisone. In summary, PPD increases the therapeutic effect of prednisone in NS while effectively prevents or decreases the appearance of side effects of glucocorticoids.

Insulin Signaling Pathway Mediates FoxO-Pepck Axis Regulation of Glucose Homeostasis in Drosophila suzukii.

The agricultural pest Drosophila suzukii exhibits a strong preference for feeding on fresh fruits, demonstrating high adaptability to sugary environments. Meanwhile, high sugar levels stimulate insulin secretion, thereby regulating the steady state of sugar metabolism. Understanding the mechanisms related to sugar metabolism in D. suzukii is crucial due to its adaptation to these specific environmental conditions. The insulin signaling pathway is an evolutionarily conserved phosphorylation cascade with significant roles in development and metabolism. We observed that the activation of the insulin signaling pathway inhibited FoxO activity and downregulated the expression of Pepck, thereby activating glycolysis and reducing glucose levels. By contrast, inhibiting insulin signaling increased the FoxO activity and upregulated the expression of Pepck, which activated gluconeogenesis and led to increased glucose levels. Our findings demonstrated the crucial role of the insulin signaling pathway in mediating glucose metabolism through the FoxO-Pepck axis, which supports the ecological adaptation of D. suzukii to high-sugar niches, thereby providing insights into its metabolic control and suggesting potential strategies for pest management. Elucidating these molecular processes is important for understanding metabolic regulation and ecological specialization in D. suzukii.

Changes of glycolysis and gluconeogenesis key enzymes in the muscle of the shrimp Penaeus vannamei in response to hypoxia and reoxygenation

Although the shrimp Penaeus vannamei can tolerate oxygen-limited conditions, the organisms may suffer detrimental effects in several processes, including growth, reproduction, immune responses, and others. During hypoxia, the shrimp makes metabolic adjustments in energy use and glucose homeostasis. Glucose is hydrolyzed during glycolysis and synthesized from non-glycosidic metabolites in gluconeogenesis. Although these pathways share many enzymes, the processes are not exactly the reverse of each other. We studied the first and last irreversible steps of glycolysis and gluconeogenesis catalyzed by hexokinase/glucose-6-phosphatase (HK/G6Pase) and pyruvate kinase/pyruvate carboxylase/phosphoenolpyruvate carboxykinase (PK/PC/PEPCK), respectively, in muscle under hypoxia and hypoxia followed by reoxygenation to gain insights about the coordinated metabolic responses that maintain glucose balance during oxygen stress. For the first group, HK gene expression was induced in hypoxia while G6Pase was reduced in hypoxia and reoxygenation, whereas for enzymes activity, HK was maintained and G6Pase was only detected in reoxygenation. For the second group, gene expression increased in PK in hypoxia, in PC in reoxygenation and in both cases, the enzymatic activities were maintained. Mitochondrial PEPCK expression increased during hypoxia and cytosolic PEPCK decreased, while total PEPCK activity was maintained. Intracellular lactate decreased during hypoxia while glucose levels were maintained. Altogether the results show that in limited oxygen, the expression of glycolytic enzymes-genes are induced, the gluconeogenic counterparts are active, although with more varied responses to oxygen limited conditions, and intracellular glucose levels are maintained.

Protopanaxadiol derivative: A plant origin of novel selective glucocorticoid receptor modulator with anti-inflammatory effect

Constant efforts have been made to move towards maintaining the positive anti-inflammatory functions of glucocorticoids (GCs) while minimizing side effects. The anti-inflammatory effect of GCs is mainly attributed to the inhibition of major inflammatory pathways such as NF-κB through GR transrepression, while its side effects are mainly mediated by transactivation. Here, we investigated the selective glucocorticoid receptor modulator (SGRM)-like properties of a plant-derived compound. In this study, glucocorticoid receptor (GR)-mediated alleviation of inflammation by SP-8 was investigated by a combination of in vitro, in silico, and in vivo approaches. Molecular docking and cellular thermal shift assay suggested that SP-8 bound stably to the active site of GR via hydrogen bonding and hydrophobic interactions. SP-8 activated GR, induced GR nuclear translocation, and inhibited NF-κB pathway activation. Furthermore, SP-8 did not up-regulate the gene and protein expression of PEPCK and TAT in HepG2 cells, and it did not induce fat deposition like GC and has little effect on bone metabolism. Interestingly, SP-8 upregulated GR protein expression and did not cause GR phosphorylation at Ser211 in RAW264.7 cells. This work proved that SP-8 dissociated characteristics of transrepression and transactivation can be separated. In addition, the in vitro and in vivo anti-inflammatory effects of SP-8 were confirmed in LPS-induced RAW 264.7 cells and in a mouse model of DSS-induced ulcerative colitis, respectively. In conclusion, SP-8 might serve as a potential SGRM and might hold great potential for therapeutic use in inflammatory diseases.

Effects of undescribed iridoids in Patrinia punctiflora on insulin resistance in HepG2 cells

Patrinia punctiflora is a medical and edible Chinese herb with high nutritional and medicinal value. The continuing study of its chemical constituents led to the isolation of six iridoids, which were previously unreported compounds, patriscabioins PU (1–6). Their structures were characterized and confirmed with NMR (1D & 2D), HRMS, IR and UV. Among them, compound 5 was screened to evaluate its insulin resistance activity on an IR-HepG-2 cell model. Compound 5 had no cytotoxicity compared with the control group and could promote glucose uptake in IR-HepG-2 cells. Through further mechanism studies, the undescribed compound 5 could increase the expression levels of PI-3 K, p-AKT, GLUT4 and p-GSK3β proteins. Moreover, the expression of PEPCK and G6Pase proteins, which are key gluconeogenic enzymes, was also inhibited. Thus, compound 5 promotes the transfer of GLUT4 to the plasma membrane by activating the PI-3 K/AKT signaling pathway, at the same time, promotes glycogen synthesis and inhibits the onset of gluconeogenesis, which in turn ameliorates insulin resistance.

Tyrosol-loaded Nano-niosomes Attenuate Diabetic Injury by TargetingGlucose Metabolism, Inflammation, and Glucose Transfer.

The increasing prevalence of type 2 diabetes, has become a global concern, making it imperative to control. Chemical drugs commonly recommended for diabetes treatment cause many complications and drug resistance over time. The polyphenol tyrosol has many health benefits, including anti-diabetes properties. Tyrosol's efficacy can be significantly increased when it is used as a niosome in the treatment of diabetes. In this study, Tyrosol and nano-Tyrosol are examined for their effects on genes implicated in type 2 diabetes in streptozotocin-treated rats. Niosome nanoparticles containing 300 mg surfactant (span60: tween60) and 10 mg cholesterol were hydrated in thin films with equal molar ratios. After 72 hours, nano-niosomal formulas were assessed for their physicochemical properties. MTT assays were conducted on HFF cells to assess the cellular toxicity of the nano niosome contacting optimal Tyrosol. Finally, the expression of PEPCK, GCK, TNF-ɑ, IL6, GLUT2 and GLUT9 was measured by real-time PCR. Physiochemical properties of the SEM images of niosomes loaded with Tyrosol revealed the nanoparticles had a vehicular structure. In this study, there were two stages of release: initial release (8 hours) and sustainable release (72 hours). Meanwhile, free-form drugs were considerably more toxic than niosomal drugs in terms of their cellular toxicity. An in vivo comparison of oral Tyrosol gavage with nano-Tyrosol showed a significant increase in GCK (P < 0.001), GLUT2 (P < 0.001), and GLUT9 (P < 0.001). Furthermore, nano-Tyrosol decreased the expression of TNF-ɑ (P < 0.05), PEPCK (P < 0.001), and IL-6 (P < 0.05) which had been increased by diabetes mellitus. The results confirmed nano-Tyrosol's anti-diabetes and anti-inflammatory effects. These findings suggest that nano-Tyrosol has potential applications in diabetes treatment and associated inflammation. Further research is needed to better understand the mechanism of action.

GHSR signalling in perinatal phases is involved in liver metabolism at puberty.

Ghrelin has effects that range from the maturation of the central nervous system to the regulation of energy balance. The production of ghrelin increases significantly during the first weeks of life. Studies have addressed the metabolic effects of liver-expressed antimicrobial peptide 2 (LEAP2) in inhibiting the effects evoked by ghrelin, mainly in glucose homeostasis, insulin resistance, and lipid metabolism. Despite the known roles of ghrelin in the postnatal development, little is known about the long-term metabolic influences of modulation with the endogenous expressed growth hormone secretagogue receptor (GHSR) inverse agonist LEAP2. This study aimed to evaluate the contribution of GHSR signalling during perinatal phases, to neurodevelopment and energy metabolism in young animals, under inverse antagonism by LEAP2[1-14]. For this, two experimental models were used: (i) LEAP2[1-14] injections in female rats during the pregnancy. (ii) Postnatal modulation of GHSR with LEAP2[1-14] or MK677. Perinatal GHSR modulation by LEAP2[1-14] impacts glucose homeostasis in a sex and phase-dependent manner, despite no effects on body weight gain or food intake. Interestingly, liver PEPCK expression was remarkably impacted by LEAP2 injections. The observed results suggests that perinatal LEAP2 exposure can modulate liver metabolism and systemic glucose homeostasis. In addition, these results, although not expressive, may just be the beginning of the metabolic imbalance that will occur in adulthood.

DOES THE COMBINATION OF ROSMARINUS OFFICINALIS AND CENTELLA ASIATICA NANOEMULSIONS AFFECT BDNF EXPRESSION IN THE GESTATIONAL DIABETES MELLITUS MODEL IN ZEBRAFISH LARVAE

Background: Pancreatic β-cells compensate for the increased insulin demand due to increasing glucose levels during pregnancy or in gestational diabetes mellitus (GDM). The enzyme that regulates the rate of gluconeogenesis production is PEPCK. Circulatory and metabolic disturbances in the brain caused by hyperglycemia could affect brain-derived neurotrophic factor (BDNF) expression as an agent for neurodevelopment. Objective: The purpose of this work is to ascertain the impact of combining nanoemulsions of Rosmarinus officinalis and Centella asiatica on the expression of BDNF and PEPCK. Methods: This study was a true experimental laboratory using zebrafish larvae. The larvae were exposed to 3% glucose and a combination of nanoemulsion extracts. The study sample of zebrafish larvae at 72 hpf (hour post fertilization) was divided into control, glucose, treatment 1, 2, and 3, with the combination of Rosmarinus officinalis and Centella asiatica nanoemulsions at 2.5 μg/ml, 5 μg/ml, and 10 μg/ml, respectively. PEPCK and BDNF expression were measured by the real-time PCR method. Results: The expression of PEPCK increased significantly in the glucose group (3.05 ± 0.27). The combination of Rosmarinus officinalis and Centella asiatica nanoemulsions significantly decreased PEPCK (p-value 0,001 &lt; 0.05). The BDNF level did not significantly differ (p-value 0.253 &gt;0.05) among groups, but the highest level (1.06 ± 0.60) was gained in the T2 group, which received an embryonic medium (EM), 3% glucose, and the combination of Rosmarinus officinalis and Centella asiatica nanoemulsions at 5μg/ml. Conclusion: The combination of Rosmarinus officinalis and Centella asiatica nanoemulsions significantly reduces PEPCK expression but does not affect BDNF expression.

Network Pharmacology Analysis, Molecular Docking Integrated Experimental Verification Reveal the Mechanism of Gynostemma pentaphyllum in the Treatment of Type II Diabetes by Regulating the IRS1/PI3K/Akt Signaling Pathway.

Gynostemma pentaphyllum (Thunb.) Makino (GP), a plant with homology of medicine and food, as a traditional Chinese medicine, possesses promising biological activities in the prevention and treatment of type 2 diabetes mellitus (T2DM). However, the material basis and the mechanism of action of GP in the treatment of T2DM have not been fully elucidated. This study aimed to clarify the active components, potential targets and signaling pathways of GP in treating T2DM. The chemical ingredients of GP were collected by combining UPLC-HRMS analysis and literature research. Network pharmacology revealed that GP had 32 components and 326 potential targets in treating T2DM. The results showed that GP affected T2DM by mediating the insulin resistance signaling pathway, PI3K/Akt signaling pathway and FoxO1 signaling pathway, which had a close relationship with T2DM. Molecular docking results showed that STAT3, PIK3CA, AKT1, EGFR, VEGFA and INSR had high affinity with the active compounds of GP. In vitro, GP extracts obviously increased the glucose uptake and glucose consumption in IR-HepG2 cells. GP extracts increased the levels of PI3K, p-AKT, p-GSK3β and p-FoxO1 and decreased the expression of p-IRS1, p-GS, PEPCK and G6Pase, which indicated that GP could promote glycogen synthesis and inhibit gluconeogenesis by regulating the IRS1/PI3K/Akt signaling pathway. The results demonstrated that GP could improve insulin resistance by promoting glucose uptake and glycogen synthesis and inhibiting gluconeogenesis through regulating the IRS1/PI3K/Akt signaling pathway, which might be a potential alternative therapy for T2DM.

Spirulina versus metformin for controlling some insulin signaling pathway genes in induced polycystic ovary syndrome rat model

Polycystic ovary syndrome (PCOS) is a prevalent endocrinologic and gynecologic disorder that affects women of reproductive age; besides, insulin resistance (IR) occurs in 50–70 % of PCOS cases. Metformin (Met) is commonly prescribed for IR management; however, it does not affect IR with some gastrointestinal symptoms. Spirulina platensis (SP) is a blue-green alga that may increase insulin sensitivity. Therefore, our study aims to evaluate SP as an alternative treatment to Met for improving glucose homeostasis by assessing the expression of 11 crucial genes involved in the insulin signaling pathway. After induction of the PCOS model using dehydroepiandrosterone (DHEA) (60 mg/kg bwt) for 30 consecutive days, rats were allocated into six groups. Relative liver weight, glutamic pyruvic transaminase (GPT) serum levels, glutamic-oxaloacetic transaminase (GOT), and insulin were determined. Furthermore, the gene expression of Ins1, Irs1, Pik3ca, Prkcz, Foxo1, Srebf1, Ppargc1a, Pklr, Gk, G6pc, and Pepck in the rat's liver tissue was determined using qRT-PCR. Treatment of the PCOS control group with Met or SP revealed a decrease in all these parameters compared with the PCOS model. Additionally, we found a statistically significant difference in the expression of both the Gk and Prkcz genes. To summarize our study results, SP or Met supplementation to PCOS rats had almost the same effect on assessed relative liver weight, GOT, GPT, and insulin levels compared with PCOS control rats. If further studies confirm and detect more impact of SP on IR in PCOS, SP could be used instead of Met since the latter causes many side effects.

237 Disrupted insulin signaling due to an acute immune response in swine

Abstract Insulin is an anabolic hormone involved in glucose uptake and synthesis of fats, proteins, and glycogen. Altered insulin signaling can occur due to metabolic state, pathogen exposure, and autoimmune disorders. Lipopolysaccharide (LPS), an endotoxin secreted produced by gram-negative bacteria, can induce a severe, systemic immune response. In pigs, chronic LPS exposure has induced insulin resistance. However, the effects of acute exposure to LPS on insulin signaling and resistance have not been elucidated. Crossbred nursery pigs [n = 58, initial body weight (BW) = 12.1 kg ± 0.23 kg] were randomly assigned into two experimental subsets, where 10 animals were designated for repeated blood sampling and 48 animals underwent timed euthanasia for tissue collection. For repeated blood sampling, jugular catheters were placed under anesthesia and the animals were given 48 h to recover before being randomly assigned to treatments. These treatments were an intraperitoneal injection of either 15µg/kg BW LPS or an equal volume of 0.9% Saline (n = 5 pigs/treatment). Blood was collected for 48 h followed by euthanasia. The other 48 pigs were randomly assigned to the same treatments, and then randomly assigned to euthanasia timepoints of 0, 3, 6, 12, 24 or 48 h post injection for tissue collection. Among the catheterized pigs, 2-h post LPS challenge there was a decrease in serum insulin concentration (P &amp;lt; 0.05) and mild hyperglycemia (P &amp;lt; 0.05) post challenge. At 12- and 24-h post LPS challenge there was a decrease in insulin (P &amp;lt; 0.05) and a trend hyperglycemia was also apparent at 24 h (P = 0.063). In liver tissue, insulin related genes exhibited alterations post LPS injection. Phosphoinositide 3-kinase (pi3k) and insulin receptor substrate 1 (irs1) expression were decreased at 3 and 6 h (p&amp;lt; 0.05) and protein kinase b (akt) expression were decreased at 3 h (P &amp;lt; 0.05) post challenge. The expression of insulin inhibitor growth factor receptor bound protein 10 (grb10) was decreased at 6-, 12-, and 24-h after challenge (P &amp;lt; 0.05). Fructose-1-6-bisphosphatase (f16bp) expression was reduced at 3-, 6-, and 12-h (P &amp;lt; 0.05) in the liver while phosphoenolpyruvate carboxykinase (pepck) expression was decreased at 3- and 6-h (P &amp;lt; 0.01) post challenge. Other metabolic genes were decreased at 3-, 6-, and 12-h post LPS challenge, including fatty acid synthase (fas; P &amp;lt; 0.05), carbohydrate response element binding protein (chrebp; P &amp;lt; 0.05), and acetyl CoA carboxylase (acc; P &amp;lt; 0.05). Protein abundance of Akt 3 h after LPS injection was decreased in the liver (P &amp;lt; 0.05). In skeletal muscle 6 h post LPS injection, insulin receptor (insr), insulin-like growth factor receptor (igfr) and fas were all increased (P &amp;lt; 0.05) These results indicate disrupted insulin signaling at various levels after an acute immune response.

Disrupted Insulin Signaling Due to an Acute Immune Response in Swine

Insulin is an anabolic hormone involved in glucose uptake and synthesis of fats, proteins, and glycogen. Altered insulin signaling can occur due to metabolic state, pathogen exposure, and autoimmune disorders. Lipopolysaccharide (LPS), an endotoxin secreted produced by gram-negative bacteria, can induce a severe, systemic immune response. In pigs, chronic LPS exposure has induced insulin resistance. However, the effects of acute exposure to LPS on insulin signaling and resistance has not been elucidated. Crossbred nursery pigs (58, 12.1 kg ± 0.23kg BW) were randomly assigned into two experimental subsets, where 10 animals were designated for repeated blood sampling and 48 animals underwent timed euthanasia for tissue collection. For repeated blood sampling, jugular catheters were placed under anesthesia and the animals were given 48 hours to recover before being randomly assigned to treatments. These treatments were an intraperitoneal injection of either 15μg/kg BW LPS or an equal volume of 0.9% Saline (n=5 pigs/treatment). Blood was collected for 48 hours followed by euthanasia. The other 48 pigs were randomly assigned to the same treatments, and then randomly assigned to euthanasia timepoints of 0, 3, 6, 12, 24 or 48 hours post injection for tissue collection. Among the catheterized pigs, two hours post LPS challenge there was a decrease in serum insulin concentration (p&lt;0.05) and mild hyperglycemia (p&lt;0.05) post challenge. At 12 and 24 hours post LPS challenge there was a decrease in insulin (p&lt;0.05) and a trend hyperglycemia was also apparent at 24 hours (p=0.063). In liver tissue, insulin related genes exhibited alterations post LPS injection. Phosphoinositide 3-kinase ( pi3k) and insulin receptor substrate 1 ( irs1) expression were decreased at 3 and 6 hours (p&lt;0.05) and protein kinase b ( akt) expression were decreased at 3 hours (p&lt;0.05) post challenge. The expression of insulin inhibitor growth factor receptor bound protein 10 ( grb10) was decreased at 6, 12, and 24 hours after challenge (p&lt;0.05). Fructose-1-6-bisphosphatase ( f16bp) expression was reduced at 3, 6, and 12 hours (p&lt;0.05) in the liver while phosphoenolpyruvate carboxykinase ( pepck) expression was decreased at 3 and 6 hours (p&lt;0.01) post challenge. Other metabolic genes were decreased at 3, 6, and 12 hours post LPS challenge, including fatty acid synthase ( fas) (p&lt;0.05), carbohydrate response element binding protein ( chrebp) (p&lt;0.05), and acetyl CoA carboxylase ( acc)(p&lt;0.05). Protein abundance of Akt 3 hours after LPS injection was decreased in the liver (p&lt;0.05). In skeletal muscle 6 hours post LPS injection, insulin receptor ( insr), insulin-like growth factor receptor ( igfr) and fas were all increased (p&lt;0.05) These results indicate disrupted insulin signaling at various levels after an acute immune response. Research reported in this abstract was supported by Eunice Kennedy Shriver National Institute of Child Health &amp; Human Development (NICHD) of the National Institutes of Health under award number R01HD092304A. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sex Differences in Ammoniagenesis in Obese, Diabetic, ZSF1 Rats

Diabetic ketoacidosis develops when insulin is insuffcient to clear blood sugar into cells for energy, then the liver breaks down fat for fuel with production of ketoacids. Lower pH activates proximal tubule PEPCK expression to replenish bicarbonate and excrete acid via NHE3 as ammonia, known as, “ammoniagenesis.” Baseline female (F) rodents exhibit greater ammoniagenesis and less NHE3 activity than males (M). We examined ammoniagenesis in male (M) and female (F) ZSF1 rats, a model of obesity and diabetic kidney disease. ZSF1 rats are born lean (L) or obese (O). OM display hyperglycemia and glucosuria at 10 wk, not yet evident in OF at 18 wk. We tested three hypotheses in ZSF1 rats (n=4-6/group): 1) ammoniagenesis rate is greater in OM and OF than LM and LF, 2) LF exhibit a higher rate of ammoniagenesis than OF, and 3) the blood glucose lowering SGLT2 inhibitor empagliflozin (SGLT2i) reduces ammoniagenesis in OM. Ammoniagenesis was measured as ammonia/24 hr/tibia length because OM and OF weigh 50% more than LM and LF. Abundance of PEPCK and NHE3 was assessed by semiquantitative immunoblots. Titratable acidity (TA) was measured as classically defined. In OM, ammoniagenesis was 6X greater (P&lt; 0.0001), and TA elevated (P=0.0003) versus LM. Ammoniagenesis was not different in OF vs LF (P=0.99). Ammoniagenesis was only 2X greater in OM vs OF (P=0.001) indicative of higher baseline ammoniagenesis in F vs M. PEPCK abundance was 2.8X greater in both OM and OF vs LM and LF (P&lt; 0.001), and NHE3 abundance was lower by 35 and 20%, respectively, in OM and OF vs LM and LF (P&lt; 0.003). Oral SGLT2i treatment (10 mg/d, n=5-6/group, 6 wks starting at 20 wk old) vs vehicle lowered plasma glucose 35%, doubled urine glucose excretion, but did not significantly alter ammoniagenesis, PEPCK or NHE3 abundance (but tended to reduce urine TA). These findings demonstrate that at 16 weeks, OM exhibit highly elevated ammoniagenesis and TA along with hyperglycemia. In contrast, OF do not exhibit hyperglycemia nor elevated ammoniagenesis, illustrating a resistance to diabetic ketoacidosis at 16 wk, despite obesity, higher TA (P= 0.003), higher PEPCK and lower NHE3. In 20 wk old OM ZSF1, SGLT2i treatment does not reverse ammoniagenesis despite lowering plasma glucose for 6 weeks. R01 DK083785, R25DK113659, NIH Step Up U24 DK115255,. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Dysregulation of phosphoenolpyruvate carboxykinase in cancers: A comprehensive analysis

BackgroundPhosphoenolpyruvate carboxykinase (PEPCK) plays a crucial role in gluconeogenesis, glycolysis, and the tricarboxylic acid cycle by converting oxaloacetate into phosphoenolpyruvate. Two distinct isoforms of PEPCK, specifically cytosolic PCK1 and mitochondrial PCK2, have been identified. Nevertheless, the comprehensive understanding of their dysregulation in pan-cancer and their potential mechanism contributing to signaling transduction pathways remains elusive. MethodsWe conducted comprehensive analyses of PEPCK gene expression across 33 diverse cancer types using data from The Cancer Genome Atlas (TCGA). Multiple public databases such as HPA, TIMER 2.0, GEPIA2, cBioPortal, UALCAN, CancerSEA, and String were used to investigate protein levels, prognostic significance, clinical associations, genetic mutations, immune cell infiltration, single-cell sequencing, and functional enrichment analysis in patients with pan-cancer. PEPCK expression was analyzed about different clinical and genetic factors of patients using data from TCGA, GEO, and CGGA databases. Furthermore, the role of PCK2 in Glioma was examined using both in vitro and in vivo experiments. ResultsThe analysis we conducted revealed that the expression of PEPCK is involved in both clinical outcomes and immune cell infiltration. Initially, we verified the high expression of PCK2 in GBM cells and its role in metabolic reprogramming and proliferation in GBM. ConclusionOur study showed a correlation between PEPCK (PCK1 and PCK2) expression with clinical prognosis, gene mutation, and immune infiltrates. These findings identified two possible predictive biomarkers across different cancer types, as well as a comprehensive analysis of PCK2 expression in various tumors, with a focus on GBM.

Tyrosine-phosphorylated DNER sensitizes insulin signaling in hepatic gluconeogenesis by inducing proteasomal degradation of TRB3

ObjectiveHepatic insulin resistance, which leads to increased hepatic gluconeogenesis, is a major contributor to fasting hyperglycemia in type 2 diabetes mellitus (T2DM). However, the mechanism of impaired insulin-dependent suppression of hepatic gluconeogenesis remains elusive. Delta/Notch-like epidermal growth factor (EGF)-related receptor (DNER), firstly described as a neuron-specific Notch ligand, has been recently identified as a susceptibility gene for T2DM through genome-wide association studies. We herein investigated whether DNER regulates hepatic gluconeogenesis and whether this is mediated by enhanced insulin signaling. MethodsThe association between DNER, tribbles homolog 3 (TRB3) and Akt signaling was evaluated in C57BL/6J, ob/ob and db/db mice by western blot analysis. DNER loss-of-function and gain-of-function in hepatic gluconeogenesis were analyzed by western blot analysis, quantitative real-time PCR, glucose uptake and output assay in AML-12 cells and partially validated in primary mouse hepatocytes. Hepatic DNER knockdown mice were generated by tail vein injection of adenovirus to confirm the effects of DNER in vivo. The interaction between DNER and TRB3 was investigated by rescue experiments, cycloheximide chase analysis, co-immunoprecipitation and immunofluorescence. The potential insulin-stimulated phosphorylation sites of DNER were determined by co-immunoprecipitation, LC-MS/MS analysis and site-specific mutagenesis. ResultsHere we show that DNER enhanced hepatic insulin signaling in gluconeogenesis by inhibiting TRB3, an endogenous Akt inhibitor, through the ubiquitin-proteasome degradation pathway. In AML-12 hepatocytes, insulin-stimulated activation of Akt and suppression of gluconeogenesis are attenuated by DNER knockdown, but potentiated by DNER over-expression. In C57BL/6J mice, hepatic DNER knockdown is accompanied by impaired glucose and pyruvate tolerance. Furthermore, the in vitro effects of DNER knockdown or over-expression on both Akt activity and hepatic gluconeogenesis can be rescued by TRB3 knockdown or over-expression, respectively. In response to insulin stimulation, DNER interacted directly with insulin receptor and was phosphorylated at Tyr677. This site-specific phosphorylation is essential for DNER to upregulate Akt activity and then downregulate G6Pase and PEPCK expression, by interacting with TRB3 directly and inducing TRB3 proteasome-dependent degradation. ConclusionsTaken together, the crosstalk between insulin-Akt and DNER-TRB3 pathways represents a previously unrecognized mechanism by which insulin regulates hepatic gluconeogenesis.

Dihydromyricetin ameliorates hepatic steatosis and insulin resistance via AMPK/PGC-1α and PPARα-mediated autophagy pathway

BackgroundDihydromyricetin (DHM), a flavonoid compound of natural origin, has been identified in high concentrations in ampelopsis grossedentata and has a broad spectrum of biological and pharmacological functions, particularly in regulating glucose and lipid metabolism. The objective of this research was to examine how DHM affected nonalcoholic fatty liver disease (NAFLD) and its underlying mechanisms involved in the progression of NAFLD in a rat model subjected to a high-fat diet (HFD). Additionally, the study examines the underlying mechanisms in a cellular model of steatohepatitis using palmitic acid (PA)-treated HepG2 cells, with a focus on the potential correlation between autophagy and hepatic insulin resistance (IR) in the progress of NAFLD.MethodsSD rats were exposed to a HFD for a period of eight weeks, followed by a treatment with DHM (at doses of 50, 100, and 200 mg·kg−1·d−1) for additional six weeks. The HepG2 cells received a 0.5 mM PA treatment for 24 h, either alone or in conjunction with DHM (10 µM). The histopathological alterations were assessed by the use of Hematoxylin–eosin (H&E) staining. The quantification of glycogen content and lipid buildup in the liver was conducted by the use of PAS and Oil Red O staining techniques. Serum lipid and liver enzyme levels were also measured. Autophagic vesicle and autolysosome morphology was studied using electron microscopy. RT-qPCR and/or western blotting techniques were used to measure IR- and autophagy-related factors levels.ResultsThe administration of DHM demonstrated efficacy in ameliorating hepatic steatosis, as seen in both in vivo and in vitro experimental models. Moreover, DHM administration significantly increased GLUT2 expression, decreased G6Pase and PEPCK expression, and improved IR in the hepatic tissue of rats fed a HFD and in cells exhibiting steatosis. DHM treatment elevated Beclin 1, ATG 5, and LC3-II levels in hepatic steatosis models, correlating with autolysosome formation. The expression of AMPK levels and its downstream target PGC-1α, and PPARα were decreased in HFD-fed rats and PA-treated hepatocytes, which were reversed through DHM treatment. AMPK/ PGC-1α and PPARα knockdown reduced the impact of DHM on hepatic autophagy, IR and accumulation of hepatic lipid.ConclusionsOur findings revealed that AMPK/ PGC-1α, PPARα-dependent autophagy pathways in the pathophysiology of IR and hepatic steatosis has been shown, suggesting that DHM might potentially serve as a promising treatment option for addressing this disease.Graphical

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.

Resveratrol attenuates high glucose-induced inflammation and improves glucose metabolism in HepG2 cells

Diabetes mellitus (DM) is characterized by impaired glucose and insulin metabolism, resulting in chronic hyperglycemia. Hyperglycemia-induced inflammation is linked to the onset and progression of diabetes. Resveratrol (RES), a polyphenol phytoalexin, is studied in diabetes therapeutics research. This study evaluates the effect of RES on inflammation and glucose metabolism in HepG2 cells exposed to high glucose. Inflammation and glucose metabolism-related genes were investigated using qPCR. Further, inflammatory genes were analyzed by applying ELISA and Bioplex assays. High glucose significantly increases IKK-α, IKB-α, and NF-kB expression compared to controls. Increased NF-kB expression was followed by increased expression of pro-inflammatory cytokines, such as TNF-α, IL-6, IL-β, and COX2. RES treatment significantly reduced the expression of NF-kB, IKK-α, and IKB-α, as well as pro-inflammatory cytokines. High glucose levels reduced the expression of TGFβ1, while treatment with RES increased the expression of TGFβ1. As glucose levels increased, PEPCK expression was reduced, and GCK expression was increased in HepG2 cells treated with RES. Further, HepG2 cells cultured with high glucose showed significant increases in KLF7 and HIF1A but decreased SIRT1. Moreover, RES significantly increased SIRT1 expression and reduced KLF7 and HIF1A expression levels. Our results indicated that RES could attenuate high glucose-induced inflammation and enhance glucose metabolism in HepG2 cells.