Phosphoenolpyruvate Carboxykinase Research Articles

Effects of Vine Water Status on Malate Metabolism and γ-Aminobutyric Acid (GABA) Pathway-Related Amino Acids in Marselan (Vitis vinifera L.) Grape Berries.

Malic acid is the predominant organic acid in grape berries, and its content is affected by abiotic factors such as temperature (fruit zone microclimate) and water (vine water status). The objectives of this study were to explore the potential mechanisms behind the effects of vine water status on the biosynthesis and degradation of berry malic acid and the potential downstream effects on berry metabolism. This study was conducted over two growing seasons in 2021 and 2022, comprising three watering regimes: no water stress (CK), light water stress (LWS), and moderate water stress (MWS). Compared to CK, a significantly higher level of malic acid was found in berries from the MWS treatment when the berry was still hard and green (E-L 33) in both years. However, water stress reduced the malic acid content at the ripe berry harvest (E-L 38) stage. The activities of NAD-malate dehydrogenase (NAD-MDH) and pyruvate kinase (PK) were enhanced by water stress. Except for the E-L 33 stage, the activity of phosphoenolpyruvate carboxylase (PEPC) was reduced by water stress. The highest phosphoenolpyruvate carboxykinase (PEPCK) activity was observed at the berry veraison (E-L 35) stage and coincided with the onset of a decrease in the malate content. Meanwhile, the expression of VvPEPCK was consistent with its enzyme activity. This study showed that water stress changed the content of some free amino acids (GABA, proline, leucine, aspartate, and glutamate), two of which (glutamate and GABA) are primary metabolites of the GABA pathway.

Combination of transcriptomics, metabolomics and physiological traits reveals the effects of polystyrene microplastics on photosynthesis, carbon and nitrogen metabolism in cucumber (Cucumis sativus L.)

Although microplastic pollution has been widely studied, the mechanism by which they influence plant photosynthesis and carbon and nitrogen metabolism remains unclear. We aimed to explore the effects of polystyrene microplastics (PS) on photosynthesis and carbon and nitrogen metabolism in cucumber using 5 μm and 0.1 μm PS particles. The PS treatments significantly reduced the stability of cucumber mesophyll cells and photosynthetic parameters and increased the soluble sugar content in cucumber leaves. The 5 μm PS affected the photosynthetic pathway by changing the expression of enzyme genes required for the synthesis of NADPH and ATP, which decreased the photosynthetic capacity in cucumber leaves. However, 0.1 μm PS altered the genes expression of phosphoenolpyruvate carboxykinase (PEPCK) and phosphoenolpyruvate carboxylase (PEPC), which affected the intercellular CO2 concentration and attenuated the negative effects on photosynthetic efficiency. Additionally, PS reduced the expression levels of nitrate/nitrite transporter (NRT) and nitrate reductase (NR), reducing the nitrogen use efficiency in cucumber leaves and mesophyll cells damage through increased accumulation of reduced glutathione (GSH), γ-glutamylcysteine (γ-GC), and citrulline. This study provides a new scientific basis for exploring the effects of microplastics on plant photosynthesis and carbon and nitrogen metabolism.

Mango (Mangifera indica L.) seed kernel extract suppresses hyperglycemia by modulating pancreatic β cell apoptosis and dysfunction and hepatic glucose metabolism in diabetic rats.

This study investigated the anti-hyperglycemic action of mango seed kernel extract (MKE) and various mechanisms involved in its actions to improve pancreatic β cells and hepatic carbohydrate metabolism in diabetic rats. An intraperitoneal injection of 60mg/kg of streptozotocin (STZ) followed by 30 consecutive days of treatment with MKE (250, 500, and 1000mg/kg body weight) was used to establish a study group of diabetic rats. Using liquid chromatography-electrospray ionization-quadrupole time-of-flight mass spectrometry (LC-ESI-QTOF-MS/MS) for identification, 26 chemical compounds were found in MKE and the high-performance liquid chromatography (HPLC) analysis of the MKE also revealed the existence of mangiferin, gallic acid, and quercetin. The results confirmed that in each diabetes-affected rat, MKE mitigated the heightened levels of fasting blood glucose, diabetic symptoms, glucose intolerance, total cholesterol (TC), and low-density lipoprotein-cholesterol (LDL-C). As demonstrated by a remarkable increment in serum and pancreatic insulin, the diabetic pancreatic β cell function was potentiated by treating with MKE. The effect of MKE on diabetic pancreatic apoptosis clearly reduced the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells, which was related to diminished levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and Bax and an increase in Bcl-xL protein expression. Furthermore, diabetes-induced liver damage was clearly ameliorated along with a notable reduction in serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels and abnormal liver histology. By enhancing anti-oxidant superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities, MKE alleviated diabetes-induced pancreatic and liver oxidative damage, as demonstrated by diminished levels of malondialdehyde. In minimizing the expression levels of glucose 6-phosphatase and phosphoenolpyruvate carboxykinase-1 proteins in the diabetic liver, MKE also enhanced glycogen content and hexokinase activity. Collectively, these findings indicate that by suppressing oxidative and inflammatory processes, MKE exerts a potent anti-hyperglycemic activity in diabetic rats which serve to protect pancreatic β cell apoptosis, enhance their function, and improve hepatic glucose metabolism.

Effects of an Hourly Bolus Postruminal Infusion of Flaxseed Oil or Palm Oil on Circulating Fatty Acid Concentrations and Hepatic Expression of Pyruvate Carboxylase and Phosphoenolpyruvate Carboxykinase in Dairy Cattle.

Palmitic (C16:0), α-linolenic acid (C18:3n-3 cis), and propionate regulate bovine pyruvate carboxylase (PC) and phosphoenolpyruvate carboxykinase (PCK1) expression in vitro. The objective of this experiment was to determine the impact of C16:0, C18:3n-3 cis, propionate, and acetate postruminal infusions on hepatic PC and PCK1 expression. We hypothesized that circulating fatty acids alter hepatic PC and PCK1 in lactating dairy cows. Acetate, propionate, palm oil, and flaxseed oil were supplied postruminally to lactating cows (n = 4) using two 4 × 4 Latin square studies. For Experiment 1, cows were infused on an hourly basis with either a bolus of propionate, acetate, or the combination of propionate and palm oil, or acetate and palm oil, and Experiment 2 was similar, but flaxseed oil replaced palm oil. Flaxseed infusions increased plasma concentration and the molar percent of C18:3n-3 cis and decreased C16:0 but did not affect PC or PCK1 expression. Palm infusions did not affect blood metabolites or the hepatic expression of PC or PCK1. The lack of responses to short-chain fatty acid infusions and changes in circulating long-chain fatty acids in mature cattle are not suitable models to study the effects of α-linolenic acid and propionate on bovine PC and PCK1 expression previously observed in vitro.

SCH772984 ameliorates lipopolysaccharide-induced hypoglycemia in mice through reversing MEK/ERK/Foxo1-mediated gluconeogenesis suppression.

Lipopolysaccharide (LPS) results in a lethal hypoglycemic response. However, the main molecular mechanism involved in LPS-induced glucose metabolism disorder is poorly understood. This study intends to investigate the signaling pathways involved in LPS-induced hypoglycemia and potential efficacy of extracellular signal-regulated kinase (ERK) inhibitor SCH772984. The effects of LPS and SCH772984 on gluconeogenesis, glucose absorption, and glycogenolysis were evaluated by pyruvate tolerance test, oral glucose tolerance test, and glucagon test, respectively. After a single intraperitoneal injection of 0.5mg/kg LPS, the mice's blood glucose levels and gluconeogenesis ability were significantly lower than that of control group. Besides, mRNA and protein expression of glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK) decreased significantly after LPS treatment. LPS induced the phosphorylation of ERK1/2, MEK1/2 (mitogen-activated protein kinase), and Foxo1 while inhibited Foxo1 expression in the nucleus, indicating an important role of the MEK/ERK/Foxo1 signaling in the inhibition of gluconeogenesis by LPS. Furthermore, SCH772984 elevated blood glucose, increased the G6Pase and PEPCK expression, and inhibited pERK1/2 and pFoxo1 expression in LPS-induced mice. In summary, LPS inhibited gluconeogenesis and induced hypoglycemia through the MEK/ERK/Foxo1 signal pathway, and ERK inhibitor could effectively reverse decreased blood glucose in mice with LPS treatment. These findings provide a novel therapeutic target for LPS-induced hypoglycemia.

Wheat germ peptide improves glucose metabolism and insulin resistance in HepG2 hepatocytes via regulating SOCS3/IRS1/Akt pathway

Evidence has demonstrated that oxidative stress plays a crucial role in regulating cellular glucose metabolism. In previous studies, wheat germ peptide (WGP) was found to effectively mitigate oxidative stress induced by high glucose. Based on the information provided, we hypothesized that WGP could exhibit antihyperglycemic and anti–insulin-resistant effects in cells. The insulin-resistant cell model was established by insulin stimulation. The glucose consumption, glycogen content, and the activities of hexokinase and pyruvate kinase following WGP treatment were measured. The protein expression of SOCS3, phosphorylated insulin receptor substrate-1 (p-IRS1), IRS1, phosphorylated protein kinase B (p-Akt), Akt, glucose transporter 2 (GLUT2), phosphorylated GSK 3β, GSK 3β, FOXO1, G6P, and phosphoenolpyruvate carboxykinase were assessed by western blot analysis. Our results demonstrated that WGP treatment increased cellular glucose consumption and glycogen synthesis and enhanced hexokinase and pyruvate kinase activities. Additionally, WGP treatment was observed to cause a significant reduction in the expression of SOCS3, FOXO1, G6P, and phosphoenolpyruvate carboxykinase, as well as in the ratio of p-IRS1/IRS1. Conversely, the expression of GLUT2 and the ratios of p-Akt/Akt and p-GSK3β/GSK3β were upregulated by WGP. These findings suggested that WGP can activate the SOCS3/IRS1/Akt signaling pathway, thus promoting the phosphorylation of GSK-3β and increasing the expression of FOXO1 and GLUT2, which contribute to enhancing glycogen synthesis, inhibiting gluconeogenesis, and promoting glucose transport in insulin-resistant HepG2 cells.

Long noncoding RNA X-inactive specific transcript (lncRNA XIST) inhibits hepatic insulin resistance by competitively binding microRNA-182-5p.

What is highlighted in this study refers to the role and molecular mechanism of long noncoding RNA (lncRNA) X-inactive specific transcript(XIST) in cells with insulin resistance (IR). In this study, LX-2 cells were applied to establish IR model in vitro. The expressions of lncRNA XIST, phosphoenolpyruvate carboxykinase(PEPCK,) and glucose-6-phosphatase (G6Pase) were quantified by quantitativereverse transcription polymerase chain reaction. The 2-deoxy-d-glucose-6-phosphate (2-DG6P) level was detected utilizing 2-deoxy-d-glucose (2-DG) uptake measurement kit. Western blot was adopted to measure the protein expressions of insulin-like growth factor-1 receptor (IGF-1R), G6Pase, PEPCK, and phosphatidylinositol 3-kinase(PI3K)/Akt pathway-related genes. StarBase was used to predict the targeting relationship between lncRNA XIST or IGF-1R with miR-182-5p, the results of which were verified by dual-luciferase reporter, RNA pull-down, and RNA immunoprecipitation assays. Rescue experiments were conducted to investigate the effect of miR-182-5p on IR cells. Next, low-expressed lncRNA XIST and high-expressed miR-182-5p were observed in IR cells. Upregulation of lncRNA XIST increased IGF-1R and 2-DG6P levels, decreased G6Pase and PEPCK expressions, and promoted PI3K/Akt pathway activation in IR cells. LncRNA XIST sponged miR-182-5p which targeted IGF-1R. MiR-182-5p mimic reversed the above effects of lncRNA XIST overexpression on IR cells. In conclusion, lncRNA XIST/miR-182-5p axis alleviates hepatic IR in vitro via IGF-1R/PI3K/Akt signaling pathway, which could be the promising therapeutic target.

CRISPR/Cas9-mediated knockout of the Vanin-1 gene in the Leghorn Male Hepatoma cell line and its effects on lipid metabolism.

Vanin-1 (VNN1) is a pantetheinase that catalyses the hydrolysis of pantetheine to produce pantothenic acid and cysteamine. Our previous studies have shown that the VNN1 is specifically expressed in chicken liver which negatively regulated by microRNA-122. However, the functions of the VNN1 in lipid metabolism in chicken liver haven't been elucidated. First, we detected the VNN1 mRNA expression in 4-week chickens which were fasted 24 hours. Next, knocked out VNN1 via CRISPR/Cas9 system in the chicken Leghorn Male Hepatoma cell line. Detected the lipid deposition via oil red staining and analysis the content of triglycerides (TG), low-density lipoprotein-C (LDL-C), and highdensity lipoprotein-C (HDL-C) after VNN1 knockout in Leghorn Male Hepatoma cell line. Then we captured various differentially expressed genes (DEGs) between VNN1-modified LMH cells and original LMH cells by RNA-seq. Firstly, fasting-induced expression of VNN1. Meanwhile, we successfully used the CRISPR/Cas9 system to achieve targeted mutations of the VNN1 in the chicken LMH cell line. Moreover, the expression level of VNN1 mRNA in LMH-KO-VNN1 cells decreased compared with that in the wild-type LMH cells (p<0.0001). Compared with control, lipid deposition was decreased after knockout VNN1 via oil red staining, meanwhile, the contents of TG and LDL-C were significantly reduced, and the content of HDL-C was increased in LMH-KO-VNN1 cells. Transcriptome sequencing showed that there were 1,335 DEGs between LMH-KO-VNN1 cells and original LMH cells. Of these DEGs, 431 were upregulated, and 904 were downregulated. Gene ontology analyses of all DEGs showed that the lipid metabolism-related pathways, such as fatty acid biosynthesis and long-chain fatty acid biosynthesis, were enriched. KEGG pathway analyses showed that "lipid metabolism pathway", "energy metabolism", and "carbohydrate metabolism" were enriched. A total of 76 DEGs were involved in these pathways, of which 29 genes were upregulated (such as cytochrome P450 family 7 subfamily A member 1, ELOVL fatty acid elongase 2, and apolipoprotein A4) and 47 genes were downregulated (such as phosphoenolpyruvate carboxykinase 1) by VNN1 knockout in the LMH cells. These results suggest that VNN1 plays an important role in coordinating lipid metabolism in the chicken liver.

TW68, cryptochromes stabilizer, regulates fasting blood glucose levels in diabetic ob/ob and high fat-diet-induced obese mice

Cryptochromes (CRYs), transcriptional repressors of the circadian clock in mammals, inhibit cAMP production when glucagon activates G-protein coupled receptors. Therefore, molecules that modulate CRYs have the potential to regulate gluconeogenesis. In this study, we discovered a new molecule called TW68 that interacts with the primary pockets of mammalian CRY1/2, leading to reduced ubiquitination levels and increased stability. In cell-based circadian rhythm assays using U2OS Bmal1-dLuc cells, TW68 extended the period length of the circadian rhythm. Additionally, TW68 decreased the transcriptional levels of two genes, Phosphoenolpyruvate carboxykinase 1 (PCK1) and Glucose-6-phosphatase (G6PC), which play crucial roles in glucose biosynthesis during glucagon-induced gluconeogenesis in HepG2 cells. Oral administration of TW68 in mice showed good tolerance, a good pharmacokinetic profile, and remarkable bioavailability. Finally, when administered to fasting diabetic animals from ob/ob and HFD-fed obese mice, TW68 reduced blood glucose levels by enhancing CRY stabilization and subsequently decreasing the transcriptional levels of Pck1 and G6pc. These findings collectively demonstrate the antidiabetic efficacy of TW68 in vivo, suggesting its therapeutic potential for controlling fasting glucose levels in the treatment of type 2 diabetes mellitus.

12 Amino Acid Nutrition in Dairy Cattle: Beyond Milk Protein

Abstract The transition from gestation to lactation, also known as the transition period, is a critical time for dairy cows. This phase is typically defined as 3 weeks before parturition through 3 weeks after parturition. Peak disease incidence (shortly after parturition) corresponds with the time of greatest negative energy balance (NEB) and negative protein balance (NPB), the peak in blood concentrations of nonesterified fatty acids, and the greatest acceleration of milk yield. Decreased fertility in the face of increasing milk production may be attributable to greater severity of postpartum NEB and NPB resulting from inadequate transition management or increased disease rates. The depth and duration of NEB and NPB are highly related to dry matter intake. Formulating and delivering appropriate diets that limit total energy intake to requirements but also provide proper intakes of all other nutrients [including the indispensable amino acids (IAA) Met and Lys] before calving can help lessen the extent of NEB and NPB after calving. Supplementation of rumen-protected methionine (RPM) during the transition period improved uterine immune function through improved glandular morphology, increased neutrophil infiltration after calving, and discovery of neutrophil extracellular trap formation in bovine endometrial tissue. Its supplementation has been reported to reduce the incidence of health complications, better lactation performance, and more favorable biomarker and plasma AA profiles, common in cows with increased liver functionality index. Cows supplemented with rumen-protected lysine (RPL) had decreased liver oxidative stress (SOD1, superoxide dismutase 1) and acute phase response (SAA3, serum amyloid A3) and reduced gene expression of NFKB1 (nuclear factor kappa B1) leading to increased proinflammatory IL1B postpartum, likely indicating improved immune activation when cows consumed RPL, a necessary component of early postpartum health. Our group has reported that a prepartum supply of RPL altered uteroplacental metabolism and glucose transport. This was demonstrated through increased expression of uterine transcripts involved in energy metabolism (GLUT3, glucose transporter 3; PCK1, phosphoenolpyruvate carboxykinase 1) and placental metabolism (FGF2, fibroblast growth factor 2; FGF2R, fibroblast growth factor 2 receptor; and PGF, placental growth factor). We also have demonstrated that RPL prepartum increased the placental protein abundance of FGF2 and LRP1 (low-density lipoprotein receptor-related protein 1), indicating possible enhanced placental metabolic activity, probably linked to trophoblast proliferation and migration processes. Finally, calves from dams fed RPL prepartum had improved overall growth and intake than calves from dams not receiving RPL. In conclusion, providing RPM and RPL during the transition period impacts dairy cows' health, uterine environment, and offspring.

Peculiarities of the relationship between the terminal site of glycolysis and the initial segment of gluconeogenesis in the myocardium and skeletal muscles of animals irradiated at different doses

Existing types of muscles differ not only structurally, but also metabolically (direction of energy exchange, choice of biosubstrates, dependence on the action of mediators, hormones, etc.). A number of studies have been devoted to the study of the impact of ionizing radiation on metabolism in muscle tissue. After irradiation, thickened muscle cells with enlarged nuclei appear in the heart muscle, the number of which does not decrease for 6 months, which indicates a violation of nuclear-cytoplasmic relations in myocardial cells. Phosphoenolpyruvate carboxykinase which ensures the utilization of cytoplasmic oxaloacetate and its transformation into phosphoenolpyruvate, completes the initial stage of gluconeogenesis and can limit the rate of gluconeogenesis from lactate. It should be emphasized that phosphoenolpyruvate carboxykinase is more active in skeletal muscles, where the activity of pyruvate kinase and lactate dehydrogenase is increased. The purpose of the work is to investigate the peculiarities of the relationship between the terminal site of glycolysis and the initial segment of gluconeogenesis in the myocardium and skeletal muscles of animals irradiated at different doses. The authors proved that in animals irradiated at a dose of 0.5 Gy, the activity of pyruvate kinase in the myocardium and skeletal muscle increases compared to intact animals. In the blood, on the contrary, there is a decrease in the activity of this enzyme compared to intact animals. When animals are irradiated at a dose of 1.0 Gy, diametrically opposite changes are observed. When animals are irradiated at a dose of 0.5 Gy, there is a slight decrease in the activity of lactate dehydrogenase in the myocardium and blood against the background of an increase in the activity of this enzyme in skeletal muscle. Irradiation of animals at a dose of 1.0 Gy leads to an acute increase in lactate dehydrogenase activity in the myocardium and skeletal muscle. The data obtained revealed that in animals irradiated at a dose of 0.5 Gy, the activity of phosphoenolpyruvate carboxykinase in skeletal muscle increases, while in cardiac muscle, on the contrary, its activity decreases as well as in blood. With an increase in the radiation dose to 1.0 Gy, diametrically opposite changes in the activity of phosphoenolpyruvate carboxykinase are observed. The authors concluded that outlined changes in the waу of intramuscular glycolysis and gluconeogenesis reactions show the pathophysiologcal mechanisms of biochemical supply restructuring as the result of ionizing irradiation influence. From the fundamental point of view these results show the direction of pathophysiologically oriented pharmacological correction of radiation-provoked muscle disturbances.

How Antioxidants, Osmoregulation, Genes and Metabolites Regulate the Late Seeding Tolerance of Rapeseeds (Brassica napus L.) during Wintering.

Rapeseed seeding dates are largely delayed under the rice-rape rotation system, but how rapeseeds adapt to the delayed environment remains unclear. Here, five seeding dates (20 October, 30 October, 10 November, 20 November and 30 November, T1 to T5) were set and the dynamic differences between two late-seeding-tolerant (LST) and two late-seeding-sensitive (LSS) rapeseed cultivars were investigated in a field experiment. The growth was significantly repressed and the foldchange (LST/LSS) of yield increased from 1.50-T1 to 2.64-T5 with the delay in seeding. Both LST cultivars showed higher plant coverage than the LSS cultivars according to visible/hyperspectral imaging and the vegetation index acquired from an unmanned aerial vehicle. Fluorescence imaging, DAB and NBT staining showed that the LSS cultivars suffered more stress damage than the LST cultivars. Antioxidant enzymes (SOD, POD, CAT, APX) and osmoregulation substances (proline, soluble sugar, soluble protein) were decreased with the delay in seeding, while the LST cultivar levels were higher than those of the LSS cultivars. A comparative analysis of transcriptomes and metabolomes showed that 55 pathways involving 123 differentially expressed genes (DEGs) and 107 differentially accumulated metabolites (DAMs) participated in late seeding tolerance regulation, while 39 pathways involving 60 DEGs and 68 DAMs were related to sensitivity. Levanbiose, α-isopropylmalate, s-ribosyl-L-homocysteine, lauroyl-CoA and argino-succinate were differentially accumulated in both cultivars, while genes including isocitrate dehydrogenase, pyruvate kinase, phosphoenolpyruvate carboxykinase and newgene_7532 were also largely regulated. This study revealed the dynamic regulation mechanisms of rapeseeds on late seeding conditions, which showed considerable potential for the genetic improvement of rapeseed.

Aerobic exercise-induced decrease of chemerin improved glucose and lipid metabolism and fatty liver of diabetes mice through key metabolism enzymes and proteins

Our previous studies have implicated an important role of adipokine chemerin in exercise-induced improvements of glycolipid metabolism and fatty liver in diabetes rat, but the underlying mechanisms remain unknown. This study first used an exogenous chemerin supplement to clarify the roles of decreased chemerin in exercised diabetes mice and possible mechanisms of glucose and lipid metabolism key enzymes and proteins [such as adipose triglyceride lipase (ATGL), lipoprotein lipase (LPL), phosphoenolpyruvate carboxykinase (PEPCK), and glucose transporter 4 (GLUT4)]. In addition, two kinds of adipose-specific chemerin knockout mice were generated to demonstrate the regulation of chemerin on glucose and lipid metabolism enzymes and proteins. We found that in diabetes mice, exercise-induced improvements of glucose and lipid metabolism and fatty liver, and exercise-induced increases of ATGL, LPL, and GLUT4 in liver, gastrocnemius and fat were reversed by exogenous chemerin. Furthermore, in chemerin knockdown mice, chemerin(−/−)∙adiponectin mice had lower body fat mass, improved blood glucose and lipid, and no fatty liver; while chemerin(−/−)∙fabp4 mice had hyperlipemia and unchanged body fat mass. Peroxisome proliferator-activated receptor γ (PPARγ), ATGL, LPL, GLUT4 and PEPCK in the liver and gastrocnemius had improve changes in chemerin(−/−)·adiponectin mice while deteriorated alterations in chemerin(−/−)·fabp4 mice, although PPARγ, ATGL, LPL, and GLUT4 increased in the fat of two kinds of chemerin(−/−) mice. ConclusionsDecreased chemerin exerts an important role in exercise-induced improvements of glucose and lipid metabolism and fatty liver in diabetes mice, which was likely to be through PPARγ mediating elevations of ATGL, LPL and GLUT4 in peripheral metabolic organs.

Enhanced glucose homeostasis via Clostridium symbiosum-mediated glucagon-like peptide 1 inhibition of hepatic gluconeogenesis in mid-intestinal bypass surgery.

The small intestine is known to play a crucial role in the development and remission of diabetes mellitus (DM). However, the exact mechanism by which mid-small intestinal bypass improves glucose metabolism in diabetic rats is not fully understood. To elucidate the mechanisms by which mid-small intestinal bypass improves glucose metabolism. Streptozotocin (STZ) was used to induce DM in Sprague-Dawley (SD) rats at a dose of 60 mg/kg. The rats were then randomly divided into two groups: The mid-small intestine bypass (MSIB) group and the sham group (underwent switch laparotomy). Following a 6-wk recovery period post-surgery, the rats underwent various assessments, including metabolic parameter testing, analysis of liver glycogen levels, measurement of key gluconeogenic enzyme activity, characterization of the gut microbiota composition, evaluation of hormone levels, determination of bile acid concentrations, and assessment of the expression of the intestinal receptors Takeda G protein-coupled receptor 5 and farnesoid X receptor. The MSIB group of rats demonstrated improved glucose metabolism and lipid metabolism, along with increased hepatic glycogen content. Furthermore, there was a decrease in the expression of the key gluconeogenic enzymes phosphoenolpyruvate carboxykinase 1 and glucose-6-phosphatase. Importantly, the MSIB group exhibited a substantial increase in the abundances of intestinal Lactobacillus, Clostridium symbiosum, Ruminococcus gnavus, and Bilophila. Moreover, higher levels of secondary bile acids, such as intestinal lithocholic acid, were observed in this group. Remarkably, the changes in the gut microbiota showed a significant correlation with the expression of key gluconeogenic enzymes and glucagon-like peptide 1 (GLP-1) at 6 wk postoperatively, highlighting their potential role in glucose regulation. These findings highlight the beneficial effects of mid-small intestine bypass on glucose metabolism and the associated modulation of the gut microbiota. The findings of this study demonstrate that the introduction of postoperative intestinal Clostridium symbiosum in the mid-small intestine contributes to the enhancement of glucose metabolism in nonobese diabetic rats. This improvement is attributed to the increased inhibition of hepatic gluconeogenesis mediated by GLP-1, resulting in a favorable modulation of glucose homeostasis.

Sex differences in FASN protein concentrations in urinary exosomes related to serum triglycerides levels in healthy adults

BackgroundDysregulation of lipid metabolism is the most prominent metabolic alteration observed in obesity, cancer, and cardiovascular diseases. The present study aimed to explore the sex differences associated with lipid metabolism in urinary exosome proteins, and evaluate the correlation of urinary exosome proteins with serum lipid biomarkers.MethodsThe key enzymes regulating lipid metabolism in healthy adults were screened using urinary exosome data. Urinary exosomes were isolated from 120 healthy subjects and the expression of urinary proteins was assessed by Western blotting and ELISA. The correlation between urinary protein concentrations and the levels of serum lipid biomarkers was analyzed using correlation analysis.ResultsThree urinary exosome proteins, namely fatty acid synthase (FASN), phosphoenolpyruvate carboxykinase (PCK1), and ATP-citrate synthase (ACLY) were identified, and only FASN showed sex differences. Sex differences were also observed in the serum triglyceride (TG) levels. Healthy males had higher FASN levels than females, and a moderate positive correlation was found between FASN concentrations and serum TG levels in healthy males (r = 0.479, P < 0.05). FASN concentrations in different age groups were positively correlated with the level of serum TG (18 ~ 30 years, r = 0.502; 31 ~ 44 years, r = 0.587; 45 ~ 59 years, r = 0.654; all P < 0.05). In addition, FASN concentrations was positively related to the increase in serum TG levels (range:1.0 ~ 1.7 mmol/L; r = 0.574, P < 0.05).ConclusionsSex differences were observed in urinary exosome FASN protein levels in healthy adults. FASN protein levels positively correlated with increased serum TG levels. FASN may serve as a novel biomarker to evaluate fatty acid synthesis in the human body.

Breast Cancer Molecular Subtypes Differentially Express Gluconeogenic Rate-Limiting Enzymes-Obesity as a Crucial Player.

Breast cancer is a heterogeneous entity, where different molecular subtypes (MS) exhibit distinct prognostic and therapeutic responses. A series of 62 breast cancer samples stratified by MS was obtained from the tumor biobank of IPO-Porto. The expression of glycolysis and gluconeogenesis-regulating enzymes was investigated by immunohistochemistry. Data analysis included stratification according to MS, body mass index (BMI), and BMI with MS (mBMI). We observed significant differences in pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PCK), and fructose-1,6-bisphosphatase (FBP) tumor cell expression when stratified by MS and mBMI. The expression of these enzymes was also statistically dependent on hormonal receptors and HER2 status and correlated with pathological stage and histological grade. Obesity tended to attenuate these differences, particularly in PC expression, although these were not affected by adipocyte deposition or inflammatory infiltration at the tumor microenvironment. Nonetheless, PCK and FBP expression was also modified by the presence of obesity-associated disorders like diabetes, hypertension, and dyslipidemia. Taken together, these findings identify metabolic fingerprints for breast cancer as distinct histological types, which are affected by the presence of obesity and obesity-associated conditions. Despite the biological role of the differential expression of enzymes remaining unknown, the current study highlights the need to identify the expression of gluconeogenic-regulating enzymes as a tool for personalized medicine.

Effect of Prunetin on Streptozotocin-Induced Diabetic Nephropathy in Rats - a Biochemical and Molecular Approach.

In the modern era, chronic kidney failure due to diabetes has spread across the globe. Prunetin (PRU), a component of herbal medicines, has a broad variety of pharmacological activities; these may help to slow the onset of diabetic kidney disease. The anti-nephropathic effects of PRU have not yet been reported. The present study explored the potential nephroprotective actions of PRU in diabetic rats. For 28 days, nephropathic rats were given oral doses of PRU (20, 40, and 80 mg/kg). Body weight, blood urea, creatinine, total protein, lipid profile, liver marker enzymes, carbohydrate metabolic enzymes, C-reactive protein, antioxidants, lipid peroxidative indicators, and the expression of insulin receptor substrate 1 (IRS-1) and glucose transporter 2 (GLUT-2) mRNA genes were all examined. Histological examinations of the kidneys, liver, and pancreas were also performed. The oral treatment of PRU drastically lowered the blood glucose, HbA1c, blood urea, creatinine, serum glutamic-oxaloacetic transaminase, serum glutamic pyruvic transaminase, alkaline phosphatase, lipid profile, and hexokinase. Meanwhile, the levels of fructose 1,6-bisphosphatase, glucose-6-phosphatase, and phosphoenol pyruvate carboxykinase were all elevated, but glucose-6-phosphate dehydrogenase dropped significantly. Inflammatory marker antioxidants and lipid peroxidative markers were also less persistent due to this administration. PRU upregulated the IRS-1 and GLUT-2 gene expression in the nephropathic group. The possible renoprotective properties of PRU were validated by histopathology of the liver, kidney, and pancreatic tissues. It is therefore proposed that PRU (80 mg/kg) has considerable renoprotective benefits in diabetic nephropathy in rats.

SAT039 Chronic Glucocorticoid Exposure Induced A S1PR2-Rorc Axis To Enhance Hepatic Gluconeogenesis

Abstract Disclosure: R.A. Lee: None. M. Chang: None. A. Tsay: None. Y. Lee: None. D. Li: None. N. Yiv: None. S. Tian: None. J. Wang: None. Glucocorticoids are steroid hormones whose circulating levels are elevated in response to stress to maintain metabolic homeostasis. Glucocorticoids are also used to treat inflammatory and autoimmune diseases due to their potent anti-inflammatory and immunomodulatory activities. However, chronic glucocorticoid treatment causes various metabolic disorders, such as hyperglycemia and insulin resistance. Glucocorticoids convey their functions through an intracellular glucocorticoid receptor (GR) which is a transcriptional regulator. While GR stimulates hepatic gluconeogenic gene transcription, additional mechanisms activated specifically by chronic glucocorticoid exposure can further enhance hyperglycemia. We found that chronic glucocorticoid exposure elevated hepatic production of sphingosine-1-phosphate (S1P) and hepatic knockdown of sphingosine-1-phosphate receptor 2 (S1PR2) attenuated chronic glucocorticoid-induced glucose intolerance and hepatic gluconeogenesis while having no effect on insulin signaling. Hepatic S1PR2 knockdown reduced GR occupancy on glucocorticoid response elements (GREs) of gluconeogenic genes, such as phosphoenolpyruvate carboxykinase (Pck1) and the catalytic subunit of glucose 6 phosphatase (G6pc), thus reducing their expressions. Interestingly, RAR-related orphan receptor C (Rorc) expression was reduced in the liver of glucocorticoid treated hepatic S1PR2 knockdown mice. We found that Rorc was also recruited to the promoter of Pck1 upon glucocorticoid treatment and reducing Rorc activity decreased the GR occupancy at the GRE of Pck1. In contrast, overexpressing Rorc in the liver of hepatic S1PR2 knockdown mice restored glucocorticoid-induced glucose intolerance, GR recruitment to GREs of gluconeogenic genes, and gluconeogenic gene expression. Overall, we propose that chronic glucocorticoid exposure induces a novel S1PR2-Rorc axis to promote the transcription of gluconeogenic genes to enhance hyperglycemia. Presentation: Saturday, June 17, 2023

SAT034 Exploring The Role Of Zfp664 In Hepatic Glucose Regulation

Abstract Disclosure: A. Tsay: None. J. Wang: None. Genome wide association studies (GWAS) identify the association between variants at genomic loci and common diseases. These variants consist of single nucleotide polymorphisms (SNPs), which can be localized within coding or non-coding regions. The former could affect the functions of proteins encoded by these genes. The latter could modulate the expression of genes contained or localized nearby these SNPs. Several SNPs associated with metabolic disease such as diabetes, obesity, and dyslipidemia are found in and nearby the human ZNF664 gene, which encodes a zinc finger protein that is predicted to be a transcriptional regulator. The mouse homolog of ZNF664 is Zfp664, whose amino acid sequence is identical to ZNF664. To learn whether Zfp664 contains a transcriptional activation or repression function, the expression plasmid harboring the cDNA of full length or deletion mutants of mouse Zfp664 fused to yeast GAL4 DNA binding domain (DBD) was transfected into HEK293T cells along with a luciferase reporter plasmid containing 5 copies of the GAL4 binding sites. We saw decreased luciferase activity in cells overexpressing the GAL4 DBD and full length Zfp664 fusion protein. Thus, Zfp664 acts as a transcriptional repressor. The deletion analysis identified that amino acid 7 to 95 of Zfp664 was required for its transcriptional repressive activity. To test the metabolic function of hepatic Zfp664, 8 weeks old male C57Bl/6J mice were infected with adeno-associated virus (AAV) expressing either scramble small hairpin RNA (shRNA, as a control) or shRNA targeting Zfp664 and then fed a normal chow diet or western diet (42% kcal from fat, 0.2% total cholesterol) for 12 weeks. We found that mice infected with AAV expressing Zfp664 shRNA had a lower expression of Zfp664 in the liver but not in epididymal white adipose tissue and gastrocnemius muscle. Under chow diet feeding, not only did hepatic Zfp664 knockdown not affect glucose tolerance and insulin sensitivity, but it also did not affect fasting glucose, insulin, triglyceride, and total cholesterol levels. In contrast, in mice fed a western diet, hepatic Zfp664 knockdown worsened glucose tolerance and elevated fasting insulin levels. Moreover, the expression of hepatic gluconeogenic genes, such as phosphoenolpyruvate carboxykinase (Pck1) and the catalytic subunit of glucose-6-phosphatase (G6pc), was increased. The fasting plasma triglyceride and total cholesterol levels were not affected by hepatic Zfp664 knockdown in western diet fed mice. Overall, these results discover the novel role of Zfp664 in the regulation of hepatic glucose homeostasis. Presentation: Saturday, June 17, 2023

In vitro, in vivo, and in silico evaluation of the glucocorticoid receptor antagonist activity of 3,6-dibromocarbazole.

3,6-Dibromocarbazole is a novel environmental contaminant which is currently detected in several environmental media worldwide. This work aims to investigate the anti-glucocorticoid potency and endocrine disrupting effects of 3,6-dibromocarbazole. In vitro experiments indicated that 3,6-dibromocarbazole possessed glucocorticoid receptor (GR) antagonistic activity and inhibited dexamethasone-induced GR nuclear translocation. 3,6-Dibromocarbazole reduced the expression levels of glucocorticoid responsive genes including glucose-6-phosphatase (G6Pase), phosphoenolpyruvate carboxykinase (PEPCK), fatty acid synthase (FAS), and tyrosine aminotransferase (TAT), and further disrupted the protein expression of two key enzymes PEPCK and FAS in gluconeogenesis. In vivo experiments showed that 3,6-dibromocarbazole induced abnormal development of zebrafish embryos and disrupted the major neurohormones involved in activation of hypothalamic-pituitary-adrenocortical (HPA) axis in zebrafish larvae. The results of molecular docking and molecular dynamics simulation contributed to explain the antagonistic effect of 3,6-dibromocarbazole. Taken together, this work identified 3,6-dibromocarbazole as a GR antagonist, which might exert endocrine disrupting effects by interfering the pathway of gluconeogenesis.