Effect of insulin and metformin on methylation and glycolipid metabolism of peroxisome proliferator-activated receptor γcoactivator-1A of rat offspring with gestational diabetes mellitus

Objective:To discuss the effect of insulin and metformin on amethylation and glycolipid metabolism of peroxisome proliferator-activated receptor γ coactivator-1A(PPARGC1A) of rat offspring with gestational diabetes mellitus(GDM).Methods:A total of 45 pregnant rats received the intraperitoneal injection of streptozotocin to establish the pregnant rat model of GDM.A total of 21 pregnant rats with GDM were randomly divided into three groups,with 7ruts in each group,namely the insulin group,metformin group and control group.Rats in the insulin group received the abdominal subcutaneous injection of 1 mL/kg recombinant insulin glargine at 18:00 every day.Rats in the metformin group received the intragastric infusion of metformin hydrochloride at 18:00 every day,with the first dose of 300 mg/kg.The doses of two groups were adjusted every 3 d to maintain the blood glucose level at 2.65-7.62 mmol/L.Rats in the control group received the intragastric infusion of 1 mL normal saline at 18:00 every day.After the natural delivery of pregnant rats.10 offspring rats were randomly selected from each group.At birth,4 wk and 8 wk after the birth of offspring rats,the weight of offspring rats was measured.The blood glucose level of offspring rats was measured at 4wk and 8 wk,while the level of serum insulin,triglyceride and leptin was measured at 8 wk.Results:The weight of offspring rats at birth in the insulin group and metformin group was significantly lower than the one in the control group(P<0.05),and there was no significant difference at 4 wk and 8 wk among three groups(P>0.05).The fasting blood glucose and random blood glucose in the insulin group and metformin group at 4 wk and 8 wk were all significantly lower than ones in the control group(P<0.05);there was no significant difference between the insulin group and metformin group(P>0.05).The expression of PPARGC1 A mRNA in the insulin group and metformin group was significantly higher and the methylation level of PPARGC1 A was significantly lower than the one in the control group(P<0.05),but there was no significant difference between the insulin group and metformin group(P>0.05).Insulin and leptin at 8 wk in the insulin group and metformin group were significantly higher,while triglyceride was significantly lower than the one in the control group(P<0.05);triglyceride level of rats in the insulin group was significantly higher than the one in the metformin group(P<0.05).There was no significant difference in insulin and leptin level of offspring rats between the insulin group and metformin group(P>0.05).Conclusions:GDM can induce the methylation of PPARGC1 A of offspring rats to reduce the expression of PPARGC1 A mRNA and then cause the disorder of glycolipid metabolism when the offspring rats grow up;the insulin or metformin in the treatment of pregnant rats with GDM can reduce the methylation level of PPARGC1 A and thus improve the abnormal glycolipid metabolism of offspring rats.

Role of Neuropeptide Y and Peroxisome Proliferator-activated Receptor γ Coactivator-1α in Stress Cardiomyopathy

Death following situations of intense emotional stress has been linked to the cardiac pathology described as stress cardiomyopathy, whose pathomechanism is still not clear. In this study, we sought to determine, via an animal model, whether the transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α) and the amino peptide neuropeptide Y (NPY) play a role in the pathogenesis of this cardiac entity. Male Sprague-Dawley rats in the experimental group were subjected to immobilization in a plexy glass box for 1 h, which was followed by low voltage elec-tric foot shock for about 1h at 10s intervals in a cage fitted with metallic rods. After 25 days the rats were sacrificed and sections of their hearts were processed. Hematoxylin-eosin staining of cardiac tissues revealed the characteristic cardiac lesions of stress cardiomyopathy such as contraction band necrosis, inflammatory cell infiltration and fibrosis. The semi-quantitative RT-PCR analysis for PGC-1α mRNA expression showed significant overexpression of PGC1-α in the stress-subjected rats (P<0.05). Fluorescence immunohistochemistry revealed a higher production of NPY in the stress-subjected rats as compared to the control rats (P=0.0027). Thus, we are led to conclude that following periods of intense stress, an increased expression of PGC1-α in the heart and an overflow of NPY may lead to stress car-diomyopathy and even death in susceptible victims. Moreover, these markers can be used to identify stress cardiomyopathy as the cause of sudden death in specific cases.

Down-regulation of peroxisome proliferator-activated receptor γ coactivator-1α expression in fatty acid-induced pancreatic betacell apoptosis involves nuclear factor-κB pathway

Background Pancreatic beta-cell apoptosis induced by lipotoxicity, to a large extent, contributes to the progression of type 2 diabetes. To investigate the mechanism of free fatty acid induced apoptosis, we aimed to study the effects of palmitic acid （PA） on the apoptosis and peroxisome proliferator-activated receptor y coactivator-1α （PGC-1α） expression in βTC3 cells as well as the possible role of nuclear factor-KB （NF-KB） in this process. Methods Hoechst 33258 was used to detect βTC3 cell apoptosis, which was induced by PA stimulation for 12 hours. PGC-1α expression was analyzed by reverse transcription polymerase chain reaction, IκB kinase β （IKKβ）, IκBα NF-KB-inducing kinase （NIK） and ReI-B expressions were analyzed by Western blotting. MGβ2 was employed to block the endogenous IκBαdegradation before PA administration, and then its effect on PA-inducing cell apoptosis and PGC-1α mRNA expression was analyzed. Results Significant increased cell apoptosis was found at the concentration of 0.5 mmol/L and 1.0 mmol/L PA administration. PA （0.5 mmol/L） could extensively reduced the expression of PGC-1α mRNA. After exposing βTC3 cells to 0.5 mmol/L PA for different time periods （0, 4, 6, 8, 10 and 12 hours）, IKKβ protein expression increased while IκBα NIK and ReI-B protein expression declined in a time-dependent manner. Pretreatment with MGβ2 to inhibit the degradation of IκBα partially prevented the down-regulation of PGC-1α mRNA expression after 12-hour PA treatment in accordance with the decrease of PA induced apoptosis. Conclusions NF-KB canonical pathway was activated in PA-mediated βTC3 cell apoptosis, whereas non-canonical pathway was inhibited. Reduced PGC-1α expression by PA in βTC3 cells could involve the activation of canonical NF-KB pathway, so as to deteriorate the PA induced apoptosis.

Small interfering RNA targeting PGC-1α inhibits VEGF expression and tube formation in human retinal vascular endothelial cells

AIMTo determine whether small interfering RNA (siRNA) of PGC-1α could inhibit vascular endothelial growth factor (VEGF) expression and tube formation in human retinal vascular endothelial cells (hRVECs).METHODShRVECs transfected with peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) siRNA were incubated for 24h and then placed into a normoxic (20%, O2) or hypoxic (1%, O2) environment for another 16h. PGC-1α mRNA and protein levels were detected by real-time PCR and Western blot. VEGF mRNA and protein levels were detected by real-time PCR and ELISA. Cell proliferation was evaluated by BrdU incorporation assay. Forty-eight hours after siRNA transfection, hRVECs were planted into Matrigel-coated plates and cultured under normoxic (20%, O2) or hypoxic (1%, O2) conditions for another 48h. The tube formation of hRVECs was observed under an optical microscope and quantified by counting the number of branch points and calculating the total tube length.RESULTSPGC-1α mRNA and protein levels were significantly reduced by PGC-1α siRNA, and VEGF mRNA and protein levels also decreased significantly. The percentage of BrdU-labeled cells in siPGC-1α groups were significantly decreased compared with control siRNA groups under normoxia and hypoxia in cell proliferation assay. In the tube formation assay, PGC-1α siRNA treated cells formed significantly fewer tubes.CONCLUSIONBlocking PGC-1α expression can inhibit VEGF expression in hRVECs and inhibit their ability to form tubes under both normoxic and hypoxic conditions.

Down-regulated expressions of PPAR_γ and its coactivator PGC-1 are related to gastric carcinogenesis and Lauren's classification in gastric carcinoma

Objective： To explore the relationship between peroxisome proliferator activated receptor-gamma （PPARγ） and peroxisome proliferator-activated receptor-gamma coactivator-1 （PGC-1） expression in gastric carcinoma （GC）, and analyze their correlations with clinicopathological features and clinical outcomes of patients. Methods：The two-step immunohistochemical method was used to detect the expression of PPARγ and PGC-1 in 179 cases of GC, and 108 cases of matched normal gastric mucosa. Besides, 16 cases of fresh GC specimens and corresponding normal gastric mucosa were detected for PGC-1 expression with Western blotting. Results： The positive rates of PPART and PGC-1 expression were significantly lower in GC （54.75%, 49.16%） than in normal gastric mucosa （70.37%, 71.30%）, respectively （P〈0.05）. The decreased expression of PGC-1 in GC was confirmed ha our Western blot analysis （P=0.004）. PPAR7 and PGC-1 expressions were related to Lauren＇s types ofGC （P〈0.05）. Positive correlation was found between PPART and PGC-1 expression in GC （rk=0.422, P〈0.001）. The survival time of PPART negative and positive patients was 36.6±3.0 vs. 38.5_＋2.7 months, and no statistical difference was found between the 5-year survival rates of two groups （34.4% vs. 44.1%, P=0.522, log-rank test）; the survival time of PGC-1 negative and positive patients was 36.2±2.8 vs. 39.9±2.9 months, while no statistical difference was found between the 5-year survival rates of the two groups （32.0% vs. 48.2%, P=0.462, log-rank test） Conclusions＇. Decreased expression of PPARγand PGC-1 in GC was related to the Lauren＇s classification. Their expressions in GC were positively correlated, indicating that their fimctions in gastric carcinogenesis may be closely related.

Effectiveness of sub-maximal intermittent exercise on muscle glycogen depletion, PGC-1<i>α</i>and PDK-4 gene expression

Several metabolic gene expressions are regulated in concert with muscle glycogen status. We hypothesized that intermittent exercise performed at high but sub-maximal intensities with long recovery periods would induce a low glycogen state that would stimul- ate peroxisome proliferator-activated receptor-γ coa- ctivator-1α (PGC1-α) and pyruvate dehydrogenase kinase-4 (PDK-4) gene expression in muscle. Nine young human subjects performed two intermittent exercise sessions. One session consisted of 60 s cycling bouts at VO2max (IE100%), and the other session consisted of 75 s cycling bouts at 80% VO2max (IE80%). Twelve bouts of exercise were completed in both sessions with a 4 min rest between each bout. Muscle specimens were obtained at pre-exercise and immediately, 1.5 h and 3 h post-exercise. Muscle gly- cogen was significantly decreased after both sessions (IE100%, 94.1 ± 5.8 to 38.7 ± 5.5 mmol/kg w.w.;IE80%, 94.6 ± 9.1 to 53.3 ± 4.8 mmol/kg w.w.;both P α and PDK- 4 mRNA expression were significantly increased after exercise in both IE100% and IE80% (PGC-1α: ~3.7 and ~2.9-fold, respectively;PDK-4: ~11.1 and ~3.5-fold, respectively;all P 100% than in IE80% (P a and PDK-4 mRNA expression, suggesting that increasing exercise intensity contributes to muscle glycogen depletion and PDK-4 mRNA expression in human skeletal muscle.

Danqi Tablet(丹七片)Regulates Energy Metabolism in Ischemic Heart Rat Model through AMPK/SIRT1-PGC-1α Pathway

Objective To investigate the cardioprotective effect of Danqi Tablet(DQT,丹七片)on ischemic heart model rats and the regulative effect on energy metabolism through peroxisome proliferator-activated receptor-γcoactivator-1α(PGC-1α).Methods Rat ischemic heart model was induced by ligation of left anterior descending coronary artery.Totally 40 Sprague-Dawley rats were randomly divided into sham group,model group,DQT group(1.5 mg/kg daily)and trimetazidine(TMZ)group(6.3 mg/kg daily)according to a random number table,10 rats in each group.Twenty-eight days after continuous administration,cardiac function was assessed by echocardiography and the structures of myocardial cells were observed by hematoxylin-eosin staining.The level of adenosine triphosphate(ATP)in myocardial cells was measured by ATP assay kit.Expressions level of key transcriptional regulators,including PGC-1α,Sirtuin 1(SIRT1),AMP-activated protein kinase(AMPK),and downstream targets of PGC-1α,such as mitofusin 1(MFN1),mitofusin 2(MFN2)and superoxide dismutase 2(SOD2)were measured by Western blot.Expression level of PGC-1αwas examined by immunohistochemical staining.Results The rat ischemic heart model was successfully induced and the heart function in model group was compromised.Compared with the model group,DQT exerted cardioprotective effects,up-regulated the ATP production in myocardial cells and inhibited the infiltration of inflammatory cells in the margin area of infarction of the myocardial tissues(P<0.01).The expressions of PGC-1α,SIRT1 and AMPK were increased in the DQT group(all P<0.05).Furthermore,the downstream targets,including MFN1,MFN2 and SOD2 were up-regulated(P<0.05 or P<0.01).Compared with the TMZ group,the expression levels of PGC-1α,MFN1 and SOD2 were increased by DQT treatment(P<0.05 or P<0.01).Conclusion DQT regulated energy metabolism in rats with ischemic heart model through AMPK/SIRT1-PGC-1αpathway.PGC-1αmight serve as a promising target in the treatment of ischemic heart disease.

Effect of the Shensong Yangxin Capsule on Energy Metabolism in Angiotensin II-lnduced Cardiac Hypertrophy

Background：Shensong Yangxin Capsule （SSYX）,traditional Chinese medicine,has been used to treat arrhythmias,angina,cardiac remodeling,cardiac fibrosis,and so on,but its effect on cardiac energy metabolism is still not clear.The objective of this study was to investigate the effects of SSYX on myocardium energy metabolism in angiotensin （Ang） Ⅱ-induced cardiac hypertrophy.Methods：We used 2 μl （10-6 mol/L） AngⅡ to treat neonatal rat cardiomyocytes （NRCMs） for 48 h.Myocardial α-ac tinin staining showed that the myocardial cell volume increased.Expression of the cardiac hypertrophic marker-brain natriuretic peptide （BNP） messenger RNA （mRNA） also increased by real-time polymerase chain reaction （PCR）.Therefore,it can be assumed that the model of hypertrophic cardiomyocytes was successfully constructed.Then,NRCMs were treated with 1 μl of different concentrations of SSYX （0.25,0.5,and 1.0 μg/ml） for another 24 h.To explore the time-depend effect of SSYX on energy metabolism,0.5 μg/ml SSYX was added into cells for 0,6,12,24,and 48 h.Mitochondria was assessed by MitoTracker staining and confocal microscopy.mRNA and protein expression of mitochondrial biogenesis-related genes-Peroxisome proliferator-activated receptor-γ coactivator-1 α （PGC-1 α）,energy balance key factor -adenosine monophosphate-activated protein kinase （AMPK）,fatty acids oxidation factor-camitine palmitoyltransferase-1 （CPT-1）,and glucose oxidation factor-glucose transporter-4 （GLUT-4） were measured by PCR and Western blotting analysis.Results：With the increase in the concentration of SSYX （from 0.25 to 1.0 μg/ml）,an increased mitochondrial density in Angll-induced cardiomyocytes was found compared to that of those treated with Angll only （0.25 μg/ml,18.3300 ± 0.8895 vs.24.4900 ± 0.9041,t =10.240,P 〈 0.0001;0.5 μg/ml,18.3300 ± 0.8895 vs.25.9800 ± 0.8187,t =12.710,P 〈 0.0001;and 1.0 μg/ml,18.3300 ± 0.8895 vs.24.2900 ± 1.3120,t =9.902,P 〈 0.0001;n =5 per dosage group）.SSYX also increased the mRNA and protein expression ofPGC-1α （0.25 μg/ml,0.8892 ± 0.0848 vs.1.0970 ± 0.0994,t =4.319,P =0.0013;0.5 μg/ml,0.8892 ± 0.0848 vs.1.2330 ± 0.0564,t =7.150,P 〈 0.0001;and 1.0 μg/ml,0.8892 ± 0.0848 vs.1.1640 ± 0.0755,t =5.720,P 〈 0.0001;n =5 per dosage group）,AMPK （0.25 μg/ml,0.8872 ± 0.0779 vs.1.1500 ± 0.0507,t =7.239,P 〈 0.0001;0.5 μg/ml,0.8872 ± 0.0779 vs.1.2280 ± 0.0623,t =9.379,P 〈 0.0001;and 1.0 μg/ml,0.8872 ± 0.0779 vs.1.3020 ± 0.0450,t =11.400,P 〈 0.0001;n =5 per dosage group）,CPT-1 （1.0 μg/ml,0.7348 ± 0.0594 vs.0.9880 ± 0.0851,t =4.994,P =0.0007,n =5）,and GLUT-4 （0.5 μg/ml,1.5640 ± 0.0599 vs.1.7720 ± 0.0660,t =3.783,P =0.0117;1.0 μg/ml,1.5640 ± 0.0599 vs.2.0490 ± 0.1280,t =8.808,P 〈 0.0001;n =5 per dosage group）.The effect became more obvious with the increasing concentration of SSYX.When 0.5 μg/ml SSYX was added into cells for 0,6,12,24,and 48 h,the expression of AMPK （6 h,14.6100 ± 0.6205 vs.16.5200 ± 0.7450,t =3.456,P =0.0250;12 h,14.6100 ± 0.6205 vs.18.3200 ± 0.9965,t =6.720,P 〈 0.0001;24 h,14.6100 ± 0.6205 vs.21.8800 ± 0.8208,t =13.160,P 〈 0.0001;and 48 h,14.6100 ± 0.6205 vs.23.7400 ± 1.0970,t =16.530,P 〈 0.0001;n =5 per dosage group）,PGC-1α （12 h,11.4700 ± 0.7252 vs.16.9000 ± 1.0150,t =7.910,P 〈 0.0001;24 h,11.4700 ± 0.7252 vs.20.8800 ± 1.2340,t =13.710,P 〈 0.0001;and 48 h,11.4700 ± 0.7252 vs.22.0300 ± 1.4180,t =15.390;n =5 per dosage group）,CPT-1 （24 h,15.1600 ± 1.0960 vs.18.5800 ± 0.9049,t =6.048,P 〈 0.0001,n =5）,and GL UT-4 （6 h,10.2100 ± 0.9485 vs.12.9700 ± 0.8221,t =4.763,P =0.0012;12 h,10.2100± 0.9485 vs.16.9100± 0.8481,t=1 1.590,P〈 0.0001;24 h,10.2100±0.9485 vs.19.0900± 0.9797,t=15.360,P〈 0.0001;and 48 h,10.2100 ± 0.9485 vs.14.1900 ± 0.9611,t =6.877,P 〈 0.0001;n =5 per dosage group） mRNA and protein increased gradually with the prolongation of drug action time.Conclusions：SSYX could increase myocardial energy metabolism in AngⅡ-induced cardiac hypertrophy.Therefore,SSYX might be considered to be an alternative therapeutic remedy for myocardial hypertrophy.

Sirtuins Function as the Modulators in Aging-related Diseases in Common or Respectively

INTRODUCTIONAccording to the demographics, the world population over 60 years will double from 605 million to 2 billion people between 2000 and 2050. Aging is a complex process in which the organism and its ability to respond to external stresses become progressive decline.