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Type 2 diabetes (T2D) is often accompanied with an induction of retinaldehyde dehydrogenase 1 (RALDH1 or ALDH1A1) expression and a consequent decrease in hepatic retinaldehyde (Rald) levels. However, the role of hepatic Rald deficiency in T2D progression remains unclear. In this study, we demonstrated that reversing T2D-mediated hepatic Rald deficiency by Rald or citral treatments, or liver-specific Raldh1 silencing substantially lowered fasting glycemia levels, inhibited hepatic glucogenesis, and downregulated phosphoenolpyruvate carboxykinase 1 (PCK1) and glucose-6-phosphatase (G6PC) expression in diabetic db/db mice. Fasting glycemia and Pck1/G6pc mRNA expression levels were strongly negatively correlated with hepatic Rald levels, indicating the involvement of hepatic Rald depletion in T2D deterioration. A similar result that liver-specific Raldh1 silencing improved glucose metabolism was also observed in high-fat diet-fed mice. In primary human hepatocytes and oleic acid-treated HepG2 cells, Rald or Rald + RALDH1 silencing resulted in decreased glucose production and downregulated PCK1/G6PC mRNA and protein expression. Mechanistically, Rald downregulated direct repeat 1-mediated PCK1 and G6PC expression by antagonizing retinoid X receptor α, as confirmed by luciferase reporter assays and molecular docking. These results highlight the link between hepatic Rald deficiency, glucose dyshomeostasis, and the progression of T2D, whilst also suggesting RALDH1 as a potential therapeutic target for T2D.
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OBJECTIVE@#To observe the effect of electroacupuncture (EA) on liver protein kinase B (Akt)/forkhead box transcription factor 1 (FoxO1) signaling pathway in Zucker diabetic fatty (ZDF) rats, and to explore the possible mechanism of EA on improving liver insulin resistance of type 2 diabetes mellitus.@*METHODS@#Twelve male 2-month-old ZDF rats were fed with high-fat diet for 4 weeks to establish diabetes model. After modeling, the rats were randomly divided into a model group and an EA group, with 6 rats in each group. In addition, six male Zucker lean (ZL) rats were used as the blank group. The rats in the EA group were treated with EA at bilateral "Zusanli" (ST 36), "Sanyinjiao" (SP 6), "Weiwanxiashu" (EX-B 3), and "Pishu" (BL 20). The ipsilateral "Zusanli" (ST 36) and "Weiwanxiashu" (EX-B 3) were connected to EA device, continuous wave, frequency of 15 Hz, 20 min each time, once a day, six times a week, for a total of 4 weeks. The fasting blood glucose (FBG) in each group was compared before modeling, before intervention and after intervention; the serum levels of insulin (INS) and C-peptide were measured by radioimmunoassay method, and the insulin resistance index (HOMA-IR) was calculated; HE staining method was used to observe the liver tissue morphology; Western blot method was used to detect the protein expression of Akt, FoxO1 and phosphoenolpyruvate carboxykinase (PEPCK) in the liver.@*RESULTS@#Before intervention, compared with the blank group, FBG was increased in the model group and the EA group (P<0.01); after intervention, compared with the model group, FBG in the EA group was decreased (P<0.01). Compared with the blank group, the serum levels of INS and C-peptide, HOMA-IR, and the protein expression of hepatic FoxO1 and PEPCK were increased (P<0.01), while the protein expression of hepatic Akt was decreased (P<0.01) in the model group. Compared with the model group, the serum levels of INS and C-peptide, HOMA-IR, and the protein expression of hepatic FoxO1 and PEPCK were decreased (P<0.01), while the protein expression of hepatic Akt was increased (P<0.01) in the EA group. In the model group, the hepatocytes were structurally disordered and randomly arranged, with a large number of lipid vacuoles in the cytoplasm. In the EA group, the morphology of hepatocytes tended to be normal and lipid vacuoles were decreased.@*CONCLUSION@#EA could reduce FBG and HOMA-IR in ZDF rats, improve liver insulin resistance, which may be related to regulating Akt/FoxO1 signaling pathway.
Subject(s)
Male , Animals , Rats , Rats, Zucker , Proto-Oncogene Proteins c-akt/genetics , Diabetes Mellitus, Type 2/therapy , Insulin Resistance , C-Peptide , Electroacupuncture , Liver , Signal Transduction , Insulin , LipidsABSTRACT
ObjectiveTo observe the effect of Momordica charantia extract （MCE） on the gluconeogenesis signaling pathway in diabetes rats. MethodMale Zucker Diabetic Fatty （ZDF） rats aged 5-6 weeks were randomly divided into a model group and an MCE group （administered MCE at a dose of 0.40 g·kg-1 by gavage）. Additionally， seven healthy male ZDF （fa/+） rats were assigned to the normal group and received administration once daily for six consecutive weeks. During the experiment， the general condition of the rats was observed， and body weight was recorded. Fasting blood glucose and random blood glucose levels were measured in the 1st， 3rd， and 5th weeks. In the 6th week， an oral glucose tolerance test （OGTT） was conducted， and serum levels of triglycerides （TG）， free fatty acid （FFA）， total cholesterol （TC）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） were measured. Hematoxylin-eosin （HE） staining was performed to examine liver morphology， periodic acid-Schiff （PAS） staining was used to assess hepatic glycogen storage， and Real-time polymerase chain reaction （PCR） was employed to measure the mRNA expression of phosphoenolpyruvate carboxykinase （PEPCK） and glucose-6-phosphatase （G6Pase） in the liver. Western blot analysis was conducted to measure the phosphorylation level of forkhead box protein O1 （FoxO1） and the protein expression of PEPCK and G6Pase in the liver. ResultCompared with the model group， the MCE group showed significant improvements in body weight， fasting blood glucose， random blood glucose， and glucose tolerance （P<0.05， P<0.01） and reduced serum levels of FFA， TC， and TG （P<0.05， P<0.01）. There were no significant differences in ALT and AST between the two groups. In the MCE group， the HE staining revealed more orderly liver cell arrangement and reduced hepatic steatosis and the PAS staining showed increased hepatic glycogen storage. The protein expression of p-FoxO1 in the liver was significantly elevated （P<0.01）， while there was no significant difference in FoxO1 protein expression. The mRNA and protein expression of PEPCK and G6Pase significantly decreased （P<0.05）. ConclusionMCE exhibits glucose-lowering and lipid-lowering effects， improves glucose tolerance， and enhances hepatic glycogen storage. These effects may be attributed to the upregulation of p-FoxO1， leading to the inhibition of PEPCK and G6Pase expression and the regulation of gluconeogenesis-related processes.
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Objective@#To investigate the role of hepatitis C virus nonstructural protein 5A (NS5A) and its domains I, II, and III in regulating gluconeogenesis in mice and the underlying mechanism.@*Methods@#A total of 60 male C57BL/6J mice were randomly divided into six groups. Recombinant lentiviral particles with specific expression of full-length NS5A, NS5A domain I, NS5A domain II, or NS5A domain III were injected via the caudal vein to establish a mouse model, and the group without injection and the group with the injection of the lentiviral particles containing enhanced green fluorescent protein (EGFP) were established as negative control. The effect of full-length NS5A protein and its domains on fasting blood glucose (FBG) and fasting serum insulin (FINS) were measured. Liver tissue was collected to prepare a paraffin section. Immunohistochemistry was used to measure the expression of phosphoenolpyruvate carboxykinase (PEPCK) in hepatocytes, quantitative real-time PCR and/or Western blot were used to measure the expression of NS5A, phosphorylated adenosine monophosphate-activated protein kinase (p-AMPK), sterol regulatory element-binding protein-1 (SREBP-1), and PEPCK.@*Results@#Compared with the group without injection and the group with the injection of the lentiviral particles containing EGFP, the groups with the injection of the lentiviral particles containing full-length NS5A and NS5A domain II had significant increases in FBG and homeostasis model assessment of insulin resistance index (P < 0.01). Immunohistochemistry and quantitative real-time PCR showed a significant increase in the expression of PEPCK, a key enzyme involved in gluconeogenesis. Western blot showed that full-length NS5A protein and NS5A domain II inhibited the level of p-AMPK and increased the levels of SREBP-1 and PEPCK.@*Conclusion@#NS5A protein and NS5A domain II may affect glucose metabolism in hepatocytes in mice by regulating AMPK/SREBP-1/PEPCK, and NS5A domain II may play an important role in insulin resistance in hepatocytes caused by HCV infection.
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Objective To investigate the effect of PGC-1 on hepatic glucose metabolism of type 2 diabetes by observing its changes in the liver of OLETF rats. Methods OLETF rats were observed,even-aged LETO rats were controled. Oral glucose tolerance test, Fasting Insulin, triglyceride and total cholesterol were measured and then the protein level of PGC-1 , phosphoenolpyruvate earboxykinase and uncoupling protein 2 of liver tissue were detected by Western blot respectively in 8,18 and 28 weeks. Results (1)OLETF rats had significantly higher levels than LETO rats, in body weight, OGTT2h blood glucose and TG at 18th, 28th week. The levels of Fasting Insulin of OLETF rats were higher while insulin sensitive index were lower than that of LETO rats at 28th week. (2)Protein expressions in the livers: PGC-1 of OLETF rats was lower than that of LETO rats at 18th and 28th week. PEPCK of OLETF rats was more while UCP2 was lower than that of LETO rats at 28th week. Conclusions OLETF rats showed pathological phenotypes of type 2 diabetes. The changes of PGC-1 , PEPCK and UCP2 in 28-weeks OLETF rat suggested that PGC-1 plays an important role in the liver glycometabolism in type 2 diabetes.
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Objective To investigate the mechanism of liver X receptor(LXR)signal pathway in regulating glucose metabolism by observing the variety of LXR expression and its impacts on regulating the mRNA expression of PEPCK and GCK, the key enzyme of hepatic glucose metabolism in various glucose metabolic status in rats. Methods SD rats were chosen and divided into four groups:the CON group, the induced DM group, the OB group, and the induced OB+DM group. For each group of rats, body weight, blood glucose, serum triglycerides, and cholesterol were measured. Then the rats were sacrificed and the livers were collected and studied. Real-time PGR was used to measure the expressions of LXR mRNA, PEPCK mRNA, and GCK mRNA in the livers. Finally, the Western Blot assay was used to measure the liver LXR protein expression. Results The expression of LXR mRNA was significantly higher in DM,OB, and OB+DM groups than in CON group(P＜0.05).The Western blot results showed that the levels of protein were in accordance with the mRNA expression. Comparing to the CON and the OB groups, the PEPCK mRNA expression of the OB+DM and the DM groups was significantly higher, while the GCK mRNA expression of these two groups was significantly lower(P＜0.05). Comparing to the CON group, the PEPCK mRNA expression of the OB group was significantly lower, while the GCK mRNA expression of OB group was significantly higher(P＜0.05).Conclusions During non-diabetic phase, LXR could act as a protective receptor for glucose metabolism and keep glucose homeostasis by regulating the key enzymes of the hepatic glucose metabolism. While in the diabetic phase, the protective receptor LXR failed to reverse the change of the related enzymes caused by insulin deficiency, and finally the plasma glucose level was raised.
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The functional signifcance of tyrosine 207 of Saccharomyces cerevisiae phosphoenolpyruvate carboxykinase was explored by examining the kinetic properties of the Tyr207Leu mutant. The variant enzyme retained the structural characteristics of the wild-type protein as indicated by circular dichroism, intrinsic fuorescence spectroscopy, and gel-exclusion chromatography. Kinetic analyses of the mutated variant showed a 15-fold increase in Km CO2, a 32fold decrease in Vmax, and a 6-fold decrease in Km for phosphoenolpyruvate. These results suggest that the hydroxyl group of Tyr 207 may polarize CO2 and oxaloacetate, thus facilitating the carboxylation/decarboxylation steps.
Subject(s)
Mutation/genetics , Phosphoenolpyruvate Carboxylase/genetics , Saccharomyces cerevisiae/enzymology , Tyrosine/genetics , Catalysis , Chromatography, Gel , Circular Dichroism , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Fungal , Phosphoenolpyruvate Carboxylase/chemistry , Spectrometry, Fluorescence , Tyrosine/chemistryABSTRACT
Background Retinol binding protein 4(RBP4) is a novel adipokine which has been related with insulin resistance.Objective To test the hypothesis that telmisartan improves glucose and lipid metabolism may be associated with inhibiting serum RBP4 and phosphoenolpyruvate carboxykinase activity in liver.Methods Thirty wistar male rats were received high fat diet to establish metabolic syndrome model and randomly to receive telmisartan [5 mg/(kg?d),n=8] or pioglitazone [20 mg/(kg?d),n=8] for 8 weeks or high-fat diet placebo(n=10).Plasma triglyceride(TG),total cholesterol(TC),high density lipoprotein cholesterol(HDL-C),low density lipoprotein cholesterol(LDL-C),fasting plasma glucose(FPG),fasting plasma insulin,serum RBP4,and phosphoenolpyruvate carboxykinase specific enzyme activity in liver(PEPCK) were determined.Results Telmisartan improved insulin resistance and the disorders of glucose and lipid metabolism in diet-induced insulin resistance rats [HOMA-IR,telmisartan group(3.4?1.2) vs high-fat diet group:(8.3?1.1),P