BACKGROUND: Gluconeogenesis is a critical metabolic pathway for maintaining glucose homeostasis, and its dysregulation can lead to glycometabolic disorders. This study aimed to identify hub biomarkers of these disorders to provide a theoretical foundation for enhancing diagnosis and treatment. METHODS: Gene expression profiles from liver tissues of three well-characterized gluconeogenesis mouse models were analyzed to identify commonly differentially expressed genes (DEGs). Weighted gene co-expression network analysis (WGCNA), machine learning techniques, and diagnostic tests on transcriptome data from publicly available datasets of type 2 diabetes mellitus (T2DM) patients were employed to assess the clinical relevance of these DEGs. Subsequently, we identified hub biomarkers associated with gluconeogenesis-related glycometabolic disorders, investigated potential correlations with immune cell types, and validated expression using quantitative polymerase chain reaction in the mouse models. RESULTS: Only a few common DEGs were observed in gluconeogenesis-related glycometabolic disorders across different contributing factors. However, these DEGs were consistently associated with cytokine regulation and oxidative stress (OS). Enrichment analysis highlighted significant alterations in terms related to cytokines and OS. Importantly, osteomodulin ( OMD ), apolipoprotein A4 ( APOA4 ), and insulin like growth factor binding protein 6 ( IGFBP6 ) were identified with potential clinical significance in T2DM patients. These genes demonstrated robust diagnostic performance in T2DM cohorts and were positively correlated with resting dendritic cells. CONCLUSIONS Gluconeogenesis-related glycometabolic disorders exhibit considerable heterogeneity, yet changes in cytokine regulation and OS are universally present. OMD , APOA4 , and IGFBP6  may serve as hub biomarkers for gluconeogenesis-related glycometabolic disorders.
Machine Learning ; Humans ; Computational Biology/methods* ; Biomarkers/metabolism* ; Diabetes Mellitus, Type 2/genetics* ; Animals ; Mice ; Gluconeogenesis/physiology* ; Gene Expression Profiling ; Transcriptome/genetics* ; Gene Regulatory Networks/genetics* ; Clinical Relevance

This paper aims to investigate the hypoglycemic effect and mechanism of berberine in improving energy metabolism based on the multi-pathway regulation of brain and muscle aromatic hydrocarbon receptor nuclear translocal protein 1(BMAL1): cyclin kaput complex of day-night spontaneous output cyclin kaput(CLOCK). The dexamethasone-induced hepatic insulin resistance(IR) HepG2 cell model was used; 0.5, 1, 5, 10, 20 μmol·L~(-1) berberine were administered at 15, 18, 21, 24, 30, 36 h. The time-dose effect of glucose content in extracellular fluid was detected by glucose oxidase method. The optimal dosage and time of berberine were determined for the follow-up study. Glucose oxidase method and chemiluminescence method were respectively performed to detect hepatic glucose output and relative content of ATP in cells; Ca~(2+), reactive oxygen species(ROS), mitochondrial structure and membrane potential were detected by fluorescent probes. Moreover, ultraviolet colorimetry method was used to detect the liver type of pyruvate kinase(L-PK) and phosphoenol pyruvate carboxykinase(PEPCK). In addition, pyruvate dehydrogenase E1 subunit α1(PDHA1), phosphate fructocrine-liver type(PFKL), forkhead box protein O1(FoxO1), peroxisome proliferator-activated receptor gamma co-activator 1α(PGC1α), glucose-6-phosphatase(G6Pase), glucagon, phosphorylated nuclear factor-red blood cell 2-related factor 2(p-Nrf2)(Ser40), heme oxygenase 1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), fibroblast growth factor 21(FGF21), uncoupled protein(UCP) 1 and UCP2 were detected by Western blot. BMAL1:CLOCK complex was detected by immunofluorescence double-staining method, combined with small molecule inhibitor CLK8. Western blot was used to detect PDHA1, PFKL, FoxO1, PGC1α, G6Pase, glucagon, Nrf2, HO-1, NQO1, FGF21, UCP1 and UCP2 in the CLK8 group. The results showed that berberine downregulated the glucose content in extracellular fluid in IR-HepG2 cells in a time-and dose-dependent manner. Moreover, berberine inhibited hepatic glucose output and reduced intracellular Ca~(2+) and ROS whereas elevated JC-1 membrane potential and improved mitochondrial structure to enhance ATP production. In addition, berberine upregulated the rate-limiting enzymes such as PFKL, L-PK and PDHA1 to promote glycolysis and aerobic oxidation but also downregulated PGC1α, FoxO1, G6Pase, PEPCK and glucagon to inhibit hepatic gluconeogenesis. Berberine not only upregulated p-Nrf2(Ser40), HO-1 and NQO1 to enhance antioxidant capacity but also upregulated FGF21, UCP1 and UCP2 to promote energy metabolism. Moreover, berberine increased BMAL1, CLOCK and nuclear BMAL1:CLOCK complex whereas CLK8 reduced the nuclear BMAL1:CLOCK complex. Finally, CLK8 decreased PDHA1, PFKL, Nrf2, HO-1, NQO1, FGF21, UCP1, UCP2 and increased FoxO1, PGC1α, G6Pase and glucagon compared with the 20 μmol·L~(-1) berberine group. BMAL1:CLOCK complex inhibited gluconeogenesis, promoted glycolysis and glucose aerobic oxidation pathways, improved the reduction status within mitochondria, protected mitochondrial structure and function, increased ATP energy storage and promoted energy consumption in IR-HepG2 cells. These results suggested that berberine mediated BMAL1:CLOCK complex to coordinate the regulation of hepatic IR cells to improve energy metabolism in vitro.
Humans ; Berberine/pharmacology* ; Gluconeogenesis/drug effects* ; Hep G2 Cells ; Glucose/metabolism* ; Liver/drug effects* ; Energy Metabolism/drug effects* ; Hypoglycemic Agents/pharmacology* ; ARNTL Transcription Factors/genetics* ; Glycolysis/drug effects* ; Oxidation-Reduction/drug effects*

As an important enzyme for gluconeogenesis, mitochondrial phosphoenolpyruvate carboxykinase (PCK2) has further complex functions beyond regulation of glucose metabolism. Here, we report that conditional knockout of Pck2 in osteoblasts results in a pathological phenotype manifested as craniofacial malformation, long bone loss, and marrow adipocyte accumulation. Ablation of Pck2 alters the metabolic pathways of developing bone, particularly fatty acid metabolism. However, metformin treatment can mitigate skeletal dysplasia of embryonic and postnatal heterozygous knockout mice, at least partly via the AMPK signaling pathway. Collectively, these data illustrate that PCK2 is pivotal for bone development and metabolic homeostasis, and suggest that regulation of metformin-mediated signaling could provide a novel and practical strategy for treating metabolic skeletal dysfunction.
Mice ; Animals ; Metformin/pharmacology* ; Phosphoenolpyruvate Carboxykinase (ATP)/metabolism* ; Gluconeogenesis/genetics* ; Mice, Knockout

Six transmembrane protein of prostate 2 (STAMP2) plays a key role in linking inflammatory and diet-derived signals to systemic metabolism. STAMP2 is induced by nutrients/feeding as well as by cytokines such as TNFalpha, IL-1beta, and IL-6. Here, we demonstrated that STAMP2 protein physically interacts with and decreases the stability of hepatitis B virus X protein (HBx), thereby counteracting HBx-induced hepatic lipid accumulation and insulin resistance. STAMP2 suppressed the HBx-mediated transcription of lipogenic and adipogenic genes. Furthermore, STAMP2 prevented HBx-induced degradation of IRS1 protein, which mediates hepatic insulin signaling, as well as restored insulin-mediated inhibition of gluconeogenic enzyme expression, which are gluconeogenic genes. We also demonstrated reciprocal expression of HBx and STAMP2 in HBx transgenic mice. These results suggest that hepatic STAMP2 antagonizes HBx-mediated hepatocyte dysfunction, thereby protecting hepatocytes from HBV gene expression.
Animals ; Female ; Gene Expression ; Gluconeogenesis/genetics ; Hep G2 Cells ; Humans ; Insulin/pharmacology/physiology ; Insulin Receptor Substrate Proteins/genetics/metabolism ; Insulin Resistance ; *Lipid Metabolism ; Liver/*metabolism/physiopathology ; Male ; Membrane Proteins/metabolism/*physiology ; Mice ; Mice, Inbred C57BL ; Mice, Inbred CBA ; Mice, Transgenic ; Oxidoreductases/metabolism/*physiology ; Phosphorylation ; Protein Binding ; Protein Processing, Post-Translational ; Proteolysis ; Receptor, Insulin/metabolism ; Trans-Activators/*physiology ; Transcriptional Activation

During fasting periods, hepatic glucose production is enhanced by glucagon to provide fuels for other organs. This process is mediated via cAMP-dependent induction of the CREB regulated transcriptional coactivator (CRTC) 2, a critical transcriptional activator for hepatic gluconeogenesis. We have previously shown that CRTC2 activity is regulated by AMP activated protein kinase (AMPK) family members. Here we show that adiponectin and thiazolidinedione directly regulate AMPK to modulate CRTC2 activity in hepatocytes. Adiponectin or thiazolidinedione lowered glucose production from primary hepatocytes. Treatment of both reagents reduced gluconeogenic gene expression as well as cAMP-mediated induction of CRE reporter, suggesting that these reagents directly affect CREB/CRTC2- dependent transcription. Furthermore, adiponectin or thiazolidinedione mediated repression of CRE activity is largely blunted by co-expression of phosphorylation defective mutant CRTC2, underscoring the importance of serine 171 residue of this factor. Taken together, we propose that adiponectin and thiazolidinedione promote the modulation of AMPK-dependent CRTC2 activity to influence hepatic gluconeogenesis.
Adiponectin/*pharmacology ; Animals ; Cells, Cultured ; *Gene Expression Regulation ; Gluconeogenesis/*drug effects ; Glucose/metabolism ; Hepatocytes/drug effects/*metabolism ; Humans ; Liver/cytology/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Protein Kinases/genetics/metabolism ; Rats ; Rats, Sprague-Dawley ; Thiazolidinediones/*pharmacology ; Transcription Factors/genetics/*metabolism

OBJECTIVETo explore the molecular mechanism of type 2 diabetes in intrauterine growth restricted adult rats through determination of blood glucose and expression of gluconeogenic enzymes in liver.

METHODSMale intrauterine growth restriction (IUGR) offspring induced by maternal protein-malnutrition and normal controls were studied. The body weights of offspring rats were weighted from birth to 12 weeks of age. Fasting plasma glucose and insulin levels were determined by glucose oxidase method and enzyme-linked immunosorbent assay (ELISA) respectively at 1 week, 8 weeks, and 12 weeks. Peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1 alpha), phosphoenolpyruvate carboxykinase (PEPCK), and glucose-6-phosphatase (G6Pase) mRNA and protein levels in liver were measured by real time RT-PCR and Western blot in newborn rats (Week 1) and adult rats (Week 12).

RESULTSBirth weights of IUGR rats were significantly lower than those of controls until 4 weeks later, when IUGR rats caught up to controls. Between 8 and 12 weeks, the growth of IUGR rats surpassed that of controls. No significant differences were observed in blood glucose and insulin levels at newborn rats between the two groups. However, by the end of 8 weeks IUGR rats developed hyperinsulinemia and high insulin resistance index. At the age of 12 weeks, IUGR rats had mild fasting hyperglycemia. In addition, hepatic PGC-1 alpha mRNA and protein levels as well as hepatic mRNA levels of PEPCK and G6Pase at Week 1 and Week 12 in IUGR rats were all significantly higher than those of controls (P<0.05).

CONCLUSIONSAs a result of intrauterine malnutrition, the expression of gluconeogenic genes is exaggerated in offspring. This change stays through adulthood and may contribute to the pathogenesis of type 2 diabetes.

Animals ; Female ; Fetal Growth Retardation ; metabolism ; Gluconeogenesis ; Glucose ; metabolism ; Glucose-6-Phosphatase ; genetics ; Liver ; metabolism ; Male ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; RNA, Messenger ; analysis ; RNA-Binding Proteins ; genetics ; Rats ; Rats, Wistar ; Transcription Factors ; genetics

OBJECTIVEIntrauterine growth retardation (IUGR) is associated with insulin resistance in later life but the mechanism remains unclear. To explore the molecular mechanism of insulin resistance, we determined the expression of gluconeogenic enzymes as well as the expression of transcription factor which promotes gluconeogenesis in the liver of IUGR rats.

METHODSRat model of IUGR was established by maternal proteindouble ended arrowmalnutrition. Hepatic mRNA levels of the key enzymes for gluconeogenesis, PEPCK and G6Pase, and of peroxisome proliferator-activated receptor-gammacoactivator (PGC) -1alpha were measured by RT- PCR in male IUGR pup rats at 3 and 8 weeks of their lives. Hepatic PGC-1alpha protein levels were determined by Western blot.

RESULTSThe average birth weights of the IUGR group (4.97+/-0.83 g) were significantly lower than normal controls (6.54+/-0.52 g) (P<0.01). Until to 4 weeks of age, the weights of the IUGR rats increased to the control level and were higher than normal controls at 8 weeks of age (P<0.05). There were no significant differences in blood glucose and insulin concentrations between the IUGR rats and normal controls at 3 weeks of age. By 8 weeks of age, the IUGR rats showed high insulin concentrations (P<0.01) and high insulin resistance index (P<0.05) compared with the controls. Hepatic PGC-1alpha mRNA and protein levels as well as hepatic mRNA levels of PEPCK and G6Pase in IUGR rats significantly increased at 3 and 8 weeks compared with controls.

CONCLUSIONSAn increased PGC-1alpha expression may contribute to increased mRNA levels of PEPCK and G6Pase, and thus induce the development of insulin resistance in later life in IUGR rats.

Animals ; Female ; Fetal Growth Retardation ; metabolism ; Gluconeogenesis ; Glucose-6-Phosphatase ; genetics ; Insulin Resistance ; Liver ; enzymology ; Male ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; Phosphoenolpyruvate Carboxykinase (GTP) ; genetics ; RNA, Messenger ; analysis ; RNA-Binding Proteins ; genetics ; Rats ; Rats, Wistar ; Transcription Factors ; genetics

OBJECTIVETo observe the effect of calorie restriction on the high fat diet rats mRNA expressions of liver forkhead box O1(FoxO1), phosphoenolpyruvate carboxykinase (PEPCK), glucose-6-phosphatase (G-6-P) and to explore the possible mechanisms.

METHODS24 normal 6-week-old male Wistar rats were randomly divided into three groups: normal chow group (NC, n = 7), high fat diet group (HF, n = 9) and calorie restriction group (CR, n = 8). They were fed for 12 weeks. At the end of the experiment, the rats were sacrificed and their fasting blood glucose (FBG), insulin (INS), triglycerides (TG), total cholesterol (TC) were measured. Their visceral fat (VF) and body weight (BW) were also measured and VF/BW was calculated. Gene expression was investigated by using semi-quantitative RT-PCR methods. Liver histology was studied with HE stained slides.

RESULTSCompared with the NC group, HF group rats developed visceral obesity which was accompanied by higher FBG, plasma INS, TG, and TC. The levels of FoxO1, PEPCK, and G-6-P increased by 18.9%, 33.8%, and 24.6%, respectively (P less than 0.01). Liver steatosis was observed with microscopy. The BW, VF FBG, INS, TG and TC of the CR group rats were lower in comparison to those of the HF group. The levels of FoxO1, PEPCK and G-6-P were lower by 26.6%, 35.0%, 34.3% (P less than 0.01). Meanwhile, liver steatosis was also milder.

CONCLUSIONCalorie restriction can inhibit the expressions of FoxO1, PEPCK and G-6-P, strengthen insulin signal conduction, suppress gluconeogenesis and thus regulate glycometabolism.

Animals ; Caloric Restriction ; Dietary Fats ; Forkhead Transcription Factors ; genetics ; Gene Expression Regulation ; Gluconeogenesis ; genetics ; Glucose-6-Phosphatase ; genetics ; Liver ; metabolism ; Male ; Nerve Tissue Proteins ; genetics ; Phosphoenolpyruvate Carboxykinase (ATP) ; genetics ; Rats ; Rats, Wistar