SIRT1 deacetylates mitochondrial trifunctional enzyme α subunit to inhibit ubiquitylation and decrease insulin resistance.

Dysregulation of free acid metabolism is a major contributor to the development of insulin resistance and diabetes. Mitochondrial trifunctional enzyme subunit (MTPα) has a critical role in fatty acid ß-oxidation. However, the association between MTPα and insulin resistance is not definitively known. Here, we aimed to determine how MTPα affects insulin resistance. We tested how MTPα affected glucose uptake in insulin-resistant 3T3-L1 adipocytes and white adipose tissue (WAT) of db/db diabetic mice. We also measured how acetylation and ubiquitylation modifications regulated MTPα activation and stability, using quantitative real-time polymerase chain reactions, immunoblotting, and immunoprecipitation. We found that MTPα overexpression promoted glucose uptake via Glut4 translocation to the plasma membrane in 3T3-L1 adipocytes. Moreover, MTPα upregulation decreased glycemia in db/db mice. Deacetylation increased MTPα protein stability and its ability to reduce insulin resistance. The activation of SIRT1, a major deacetylase, prevented MTPα degradation by decreasing its acetylation in adipocytes. Our study demonstrates a new role for MTPα in reducing insulin resistance. Acetylation and ubiquitylation modifications of MTPα were crucial to regulating its function in glucose metabolism.

Efficacy and Safety of Policosanol Plus Fenofibrate Combination Therapy in Elderly Patients with Mixed Dyslipidemia: A Randomized, Controlled Clinical Study.

BACKGROUND: Policosanol is a mixture of long-chain alcohols isolated from sugar cane. This controlled, randomized clinical trial was designed to compare the efficacy and safety of fenofibrate, policosanol and a combination of these 2 in lowering low-density-lipoprotein cholesterol (LDL-C) in elderly patients with mixed dyslipidemia. METHODS: A total of 102 patients aged ≥60years were randomly assigned into 3 groups: patients receiving a 24-week therapy of fenofibrate (200 mg/day), policosanol (20 mg/day) or fenofibrate + policosanol combination. Lipids were evaluated at baseline, after 16 and after 24 weeks of therapy. Brachial-ankle pulse wave velocity (ba-PWV) was performed, and SF-36 questionnaires were used to evaluate the patients' quality of life. The primary endpoint was the percentage reduction in LDL-C. The secondary end points included percentage change in nonhigh density lipoprotein cholesterol (non-HDL-C), total cholesterol (TC), triglyceride, high-density-lipoprotein cholesterol (HDL-C), ba-PWV and SF-36 scores. Safety was assessed by adverse events and laboratory parameters. RESULTS: LDL-C, non-HDL-C and TC were decreased, respectively after treatment with policosanol for 24 weeks (P < 0.01). Treatment with policosanol + fenofibrate resulted in significantly greater reductions in TC, non-HDL-C and LDL-C compared to fenofibrate alone (P < 0.01, respectively). There were significant increases in SF-36 scores in the policosanol and policosanol + fenofibrate groups (P < 0.05), and significant improvements of ba-PWV in the 2 groups (P < 0.01). There were no serious adverse events or significant changes in laboratory variables after any of the treatment regimens. CONCLUSIONS: Policosanol + fenofibrate combination therapy significantly improved lipid parameters, arterial stiffness, and quality of life, with good tolerability.

9-PAHSA promotes browning of white fat via activating G-protein-coupled receptor 120 and inhibiting lipopolysaccharide / NF-kappa B pathway.

Browning of white adipose tissue is a novel mechanism to counteract obesity in view of its thermogenic activity. Activation of G-protein-coupled receptor 120 (GPR120) can promote the browning of white fat. 9-PAHSA, an endogenous mammalian lipid, which is acting as the ligand of GPR120 to enhance glucose uptake and exert anti-inflammatory effect. In the study, we would like to investigate the biological effects of 9-PAHSA on adipocyte browning. Here, we show that 9-PAHSA induces browning of 3T3-L1 adipocytes via enhanced expression of brown fat specific genes. 9-PAHSA-induced browning in white adipocytes of WT mice and ob/ob mice was investigated by determining expression levels of brown adipocyte-specific genes/proteins by quantitative real-time polymerase chain reaction analysis, immunoblot analysis and immunochemical staining. The effects of 9-PAHSA on brown fat markers in 3T3-L1 cells were decreased when GPR120 gene was silenced. To investigate the molecular mechanism of 9-PAHSA on adipocyte browning, lipopolysaccharide (LPS)-induced inflammatory model was conducted. 9-PAHSA treatment abolished LPS-induced NF-kappa B (NF-κB) activation and inflammatory cytokine secretion. But these anti-inflammatory effects of 9-PAHSA were attenuated by GPR120 knockdown. Our finding demonstrated that the browning of adipocyte was induced by 9-PAHSA through activating GPR120 and inhibiting the LPS/NF-κB pathway. This promising result will help to reveal the potential pathogenesis of obesity.

High Glucose Concentration Impairs 5-PAHSA Activity by Inhibiting AMP-Activated Protein Kinase Activation and Promoting Nuclear Factor-Kappa-B-Mediated Inflammation.

Recently, the endogenous fatty acid palmitic acid-5-hydroxystearic acid (5-PAHSA) was found to increase insulin sensitivity and have anti-inflammatory effects in mice with high-fat diet (HFD)-induced diabetes. However, it is unknown if 5-PAHSA affects glucose and lipid metabolism in db/db mice, which are characterized by extreme hyperglycemia. Here, we aim to determine the effect of continued 5-PAHSA administration on glucose and lipid metabolism in db/db mice. We also used 3T3-L1 cells and HepG2 cells to investigate the mechanism behind this effect. HepG2 cells and 3T3-L1 cells were induced to become models of insulin resistance. The models were used to test the effect of 5-PAHSA on insulin signaling. 5-PAHSA was administered orally to db/db mice for 1 month to assess its effects on glucose and lipid metabolism. We also exposed HepG2 cells to high glucose concentrations to investigate the influence on 5-PAHSA's effects on hepatic lipid metabolism and inflammation. 5-PAHSA improved glucose uptake and insulin signaling in HepG2 cells and 3T3-L1 cells. However, after 1 month of treatment, 5-PAHSA did not reduce blood glucose levels, but increased inflammation and promoted fatty liver in db/db mice. In HepG2 cells under normal glucose conditions, 5-PAHSA treatment reduced lipogenesis and increased lipid oxidation. Notably, a high glucose concentration in cell media abolished the positive effects of 5-PAHSA treatment. These changes were associated with: decreased phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC); upregulation of sterol-regulatory element-binding protein 1c (SREBP1c), and fatty acid synthase (FAS); and downregulation of carnitine palmitoyltransferase 1 (CPT1). Besides, the anti-inflammatory effect of 5-PAHSA was also impaired by high glucose conditions. Thus, high glucose concentrations impaired 5-PAHSA action by inhibiting the AMPK signaling pathway and promoting nuclear factor-kappa-B (NF-κB) mediated inflammation.

Adipocyte differentiation is regulated by mitochondrial trifunctional protein α-subunit via sirtuin 1.

Mitochondrial trifunctional protein α-subunit (MTPα) is involved in the fatty acid ß-oxidation (FAO) pathway. Two MTPα activities, 3-hydroxyacyl-CoA dehydrogenase and long-chain hydratase, have been linked with the occurrence and development of obesity and obesity-related disorders. These activities catalyze two steps in the FAO pathway (the second and third reactions). However, the role of MTPα in the pathogenesis of obesity has not been evaluated, and the functional role of MTPα in adipocyte differentiation has not been determined. Here, we analyzed the functional role of MTPα using in vitro and in vivo models of adipogenesis. MTPα expression was upregulated during the differentiation of 3T3-L1 preadipocyte cells into adipocytes. MTPα gene silencing stimulated peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT-enhancer-binding protein alpha(C/EBPα) expression, which promoted adipocyte differentiation. By contrast, MTPα overexpression blocked adipogenesis in 3T3-L1 cells. Further analysis showed that MTPα positively regulated sirtuin 1 (SIRT1). Injection of preadipocytes overexpressing MTPα into athymic mice significantly impaired de novo fat pad formation compared with that of the control, and furthermore MTPα knockdown enhances fat pad formation at a time point earlier than 5-week, such as week-2 and week-3, when the control fat pad is not fully developed. In summary, our data indicate that MTPα is a novel factor that negatively regulates adipocyte differentiation. We propose a pathway in which MTPα inhibits adipogenesis by promoting SIRT1 expression, which represses PPARγ and attenuates adipogenesis.

Freeze-thaw Caenorhabditis elegans freeze-thaw stress response is regulated by the insulin/IGF-1 receptor daf-2.

BACKGROUND: Adaption to cold temperatures, especially those below freezing, is essential for animal survival in cold environments. Freezing is also used for many medical, scientific, and industrial purposes. Natural freezing survival in animals has been extensively studied. However, the underlying mechanisms remain unclear. Previous studies demonstrated that animals survive in extremely cold weather by avoiding freezing or controlling the rate of ice-crystal formation in their bodies, which indicates that freezing survival is a passive thermodynamic process. RESULTS: Here, we showed that genetic programming actively promotes freezing survival in Caenorhabditis elegans. We found that daf-2, an insulin/IGF-1 receptor homologue, and loss-of-function enhanced survival during freeze-thaw stress, which required the transcription factor daf-16/FOXO and age-independent target genes. In particular, the freeze-thaw resistance of daf-2(rf) is highly allele-specific and has no correlation with lifespan, dauer formation, or hypoxia stress resistance. CONCLUSIONS: Our results reveal a new function for daf-2 signaling, and, most importantly, demonstrate that genetic programming contributes to freezing survival.

CCAAT/enhancer-binding protein CEBP-2 controls fat consumption and fatty acid desaturation in Caenorhabditis elegans.

Mammalian CCAAT/enhancer-binding proteins (C/EBPs) are generally known as regulators in adipocyte differentiation. However, more understanding of the role of C/EBPs in lipid and glucose metabolism remains to be discovered. In this study, we verified the effect of CEBP-2, the homolog of CEBPs, on fat storage in Caenorhabditis elegans. Expressions of 85 genes that encode the major enzymes in energy metabolic pathways were then screened in cebp-2-deficient worms using a quantitative real-time polymerase chain reaction (QRT-PCR). Our data implied that loss of function of CEBP-2 displayed a low-fat phenotype in C. elegans owing to increased expression of ech-1.1 and decreased expression of fat-5. Our findings indicated that cebp-2 controls total body fat content by governing fatty acid mitochondrial ß-oxidation and desaturation in C. elegans. These data provide insights into how C/EBPs may affect lipid metabolism in mammals in addition to regulating adipocyte differentiation.

Differential ERK1/2 Signaling and Hypertrophic Response to Endothelin-1 in Cardiomyocytes from SHR and Wistar-Kyoto Rats: A Potential Target for Combination Therapy of Hypertension.

Extracellular signal regulated kinase½ (ERK1/2) signaling is critical to endothelin-1 (ET-1)-induced cardiomyocyte hypertrophy. This study was to investigate ERK1/2 signaling and hypertrophic response to ET-1 stimulation in cardiomyocytes (CMs) from spontaneous hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). Primary neonatal SHR and WKY CMs were exposed to ET-1 for up to 24 hrs. Minimal basal ERK1/2 phosphorylation was present in WKY CMs, while a significant baseline ERK1/2 phosphorylation was observed in SHR CMs. ET-1 induced a time- and dose-dependent increase in ERK1/2 phosphorylation in both SHR and WKY CMs. However, ET-1-induced ERK1/2 activation occurred much earlier with significantly higher peak phosphorylation level, and stayed elevated for longer duration in SHR CMs than that in WKY CMs. ET-1-induced hypertrophic response was more prominent in SHR CMs than that in WKY CMs as reflected by increased cell surface area, intracellular actin density, and protein synthesis. Pre-treatment with ERK1/2 phosphorylation inhibitor PD98059 completely prevented ET-1-induced ERK1/2 phosphorylation and increases in cell surface area and protein synthesis in SHR and WKY CMs. The specific PI3 kinase inhibitor LY294002 blocked ET-1-induced Akt and ERK1/2 phosphorylation, and protein synthesis in CMs. These data indicated that ERK1/2 signaling was differentially enhanced in CMs, and was associated with increased cardiac hypertrophic response to ET-1 in SHR. ET-1-induced ERK1/2 activation and cardiac hypertrophy appeared to be mediated via PI3 kinase/Akt signaling in SHR and WKY. The differential ERK1/2 activation in SHR CMs by ET-1 might represent a potential target for combination therapy of hypertension.

Adventitial gene transfer of catalase attenuates angiotensin II-induced vascular remodeling.

Vascular adventitia and adventitia­derived reactive oxygen species (ROS) contribute to vascular remodeling following vascular injury. A previous ex vivo study in adventitial fibroblasts showed that catalase, one of most important anti­oxide enzymes, was downregulated by angiotensin II (AngII). The aim of the present study was to investigate whether adventitial gene transfer of catalase affects AngII­induced vascular remodeling in vivo. Adenoviruses co­expressing catalase and enhanced green fluorescent protein (eGFP) or expressing eGFP only were applied to the adventitial surface of common carotid arteries of Sprague­Dawley rats. Alzet minipumps administering AngII (0.75 mg/kg/day) were then implanted subcutaneously for 14 days. Systolic blood pressure and biological parameters of vascular remodeling were measured in each group. Adventitial fibroblasts were cultured and p38 mitogen­activated protein kinase (MAPK) phosphorylation was measured using western blot analysis. The results showed that adventitial gene transfer of catalase had no effect on AngII­induced systolic blood pressure elevation. However, catalase adenovirus transfection significantly inhibited AngII­induced media hypertrophy compared with that of the control virus (P<0.05). In addition, catalase transfection significantly attenuated AngII­induced ROS generation, macrophage infiltration, collagen deposition and adventitial α­smooth muscle actin expression. Furthermore, catalase transfection significantly inhibited the AngII­induced increase in p38MAPK phosphorylation. In conclusion, the results of the present study demonstrated that adventitial gene transfer of catalase significantly attenuated AngII­induced vascular remodeling in rats via inhibition of adventitial p38MAPK phosphorylation.

Effects of atorvastatin on bone mineral density (BMD) and bone metabolism in elderly males with osteopenia and mild dyslipidemia: a 1-year randomized trial.

We explored the effects of atorvastatin on BMD and biochemical markers of bone metabolism in a 1-year, prospective, randomized controlled study. 64 male patients with osteopenia and mild dyslipidemia (mean age 80.1±6.6 years) were randomized to a 1-year atorvastatin treatment or control. BMD of hip and lumbar spine was measured with dual-energy X-ray absorptionmetry (DXA). Bone metabolic markers including resorption markers ß-c-terminal telopeptide of type I collagen (CTx), formative markers osteocalcin (OC), 25-hydroxyvitamin D (25(OH)D) were measured with electrochemiluminescence immunoassay (ECLIA). Other bone metabolism markers including intact parathyroid hormone (iPTH) and testosterone were measured with chemiluminescence enzyme immunoassay (CLEIA). Levels of serum lipid and biochemical parameters were measured with automatic biochemical analyzer. All the parameters were recorded at baseline, and at 6 and 12 months, respectively. Compared with the control group, the atorvastatin treatment group showed significant reduction of triglyceride (TG, P<0.01) and low-density lipoprotein cholesterol (LDL-C, P<0.01). At 12 month, total hip BMD in atorvastatin group was significantly higher (P<0.01) compared with the control group, while there were no similar effect on femoral neck or lumbar spine between the two groups (P=0.48 and 0.53 respectively). Meanwhile, CTx significantly reduced in atorvastatin treatment group (P<0.001) compared with baseline. Our findings suggest that in elderly male patients with osteopenia and mild dyslipidemia, therapeutic doses of atorvastatin were associated with positive effects on BMD, probably mediated by suppressed bone resorption.

Use of improved tracheal catheters in patient of tracheostomy tube-induced tracheoesophageal fistula: a case report.

Tracheostomy tube might cause tracheoesophageal fistula (TEF) due to high cuff pressure or direct mechanical trauma. Surgical repair provides the ideal way to deal with TEF but it necessitates the weaning the patient from mechanical ventilation. Here we report a spontaneous closure of TEF by managing it with improved tracheal catheters in a patient who is dependent on mechanical ventilation.

Gamma-glutamyltransferase level and risk of hypertension: a systematic review and meta-analysis.

BACKGROUND: Several prospective observational studies suggest that gamma-glutamyltransferase(GGT) level is positively associated with risk of hypertension. However, these studies draw inconsistent conclusions. Therefore, we conducted a systematic review and meta-analysis to evaluate the exact association between GGT level and subsequent development of hypertension. METHODS: We searched Pubmed, Embase, and Science Citation Index (ISI Web of Science) for prospective cohort studies examining the association between GGT level and hypertension. Then, pooled effect estimates (RRs) for the association between GGT level and hypertension were calculated. RESULTS: A total of 13 prospective cohort studies including 43314 participants and 5280 cases of hypertension were included. The pooled RR of hypertension was 1.94(95%CI: 1.55-2.43; P<0.001) when comparing the risk of hypertension between the highest versus lowest category of GGT levels. Moreover, the risk of hypertension increased by 23% (summary RR: 1.23; 95%CI: 1.13-1.32; P<0.001) per 1 SD logGGT increment. Subgroup analyses showed significant positive associations in each subgroup except in ≧160/95 subgroup (RR: 2.56, 95%CI: 0.87-7.54; P = 0.088) and nondrinkers subgroup (RR: 1.76, 95%CI: 0.88-3.53; P = 0.113). Sensitivity analyses showed no single study significantly affects the pooled RRs. No publication bias was found in our meta-analysis. CONCLUSIONS: GGT level is positively associated with the development of hypertension. Further studies are needed to confirm our findings and elucidate the exact mechanisms between GGT level and the incidence of hypertension.

The cannabinoid WIN55,212-2 protects against oxidized LDL-induced inflammatory response in murine macrophages.

The endocannabinoid system has recently been attracted interest for its anti-inflammatory and anti-oxidative properties. In this study, we investigated the role of the endocannabinoid system in regulating the oxidized low-density lipoprotein (oxLDL)-induced inflammatory response in macrophages. RAW264.7 mouse macrophages and peritoneal macrophages isolated from Sprague-Dawley (SD) rats were exposed to oxLDL with or without the synthetic cannabinoid WIN55,212-2. To assess the inflammatory response, reactive oxygen species (ROS) and tumor necrosis factor alpha (TNF- alpha) levels were determined, and activation of the mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-kappa B signaling pathways were assessed. We observed that: i) oxLDL strongly induced ROS generation and TNF- alpha secretion in murine macrophages; ii) oxLDL-induced TNF- alpha and ROS levels could be lowered considerably by WIN55,212-2 via inhibition of MAPK (ERK1/2) signaling and NF-kappa B activity; and iii) the effects of WIN55212-2 were attenuated by the selective CB2 receptor antagonist AM630. These results demonstrate the involvement of the endocannabinoid system in regulating the oxLDL-induced inflammatory response in macrophages, and indicate that the CB2 receptor may offer a novel pharmaceutical target for treating atherosclerosis.

Differential expression of alpha-enolase in the normal and pathological cardiac growth.

OBJECTIVES: It was found that alpha-enolase was dramatically up-regulated in the hypertrophic hearts of SHR in our previous study. The purposes of this study were to examine the expression pattern of alpha-enolase in pre- and postnatal myocardium of spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats, and to explore the relationship between the overexpression of alpha-enolase and left ventricular hypertrophy. METHODS: HE staining was used for the measurement of cardiac hypertrophy. Immunohistochemical technique was used to evaluate the location of alpha-enolase. The expressions of alpha-enolase in the left cardiac ventricles at different development times were examined by Real-time RT-PCR and Western blot. RESULTS: Cardiac hypertrophy was found in SHR rats at 4 weeks of age and remained up to 24 weeks of age. The signals of alpha-enolase protein were strong and existed extensively in hypertrophic myocardium in SHR, while in the normal myocardium of WKY, the signals were scarcely found and weak. The levels of alpha-enolase mRNA and protein in SHR and WKY hearts during fetal stage and newborn stage were similar, while from 4 weeks of age to 24 weeks of age, accompanied by the cardiac hypertrophy, the levels of alpha-enolase mRNA and protein in left ventricle of SHR were significantly higher than that in WKY. CONCLUSIONS: The expressions of alpha-enolase in the left ventricle of the rats during normal and pathological cardiac development were different. This phenomenon provides the potential clues to understanding pathophysiological mechanisms in cardiac hypertrophy of SHR.

Alterations of mitochondrial enzymes contribute to cardiac hypertrophy before hypertension development in spontaneously hypertensive rats.

Mitochondrial dysfunction is recently thought to be tightly associated with the development of cardiac hypertrophy as well as hypertension. However, the detailed molecular events in mitochondria at early stages of hypertrophic pathogenesis are still unclear. Applying two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) combined with MALDI-TOF/TOF tandem mass spectrometry, here we identified the changed mitochondrial proteins of left ventricular mitochondria in prehypertensive/hypertensive stages of cardiac hypertrophy through comparing spontaneously hypertensive rats (SHR) and the age-matched normotensive Wistar Kyoto (WKY) rats. The results revealed that in the hypertrophic left ventricle of SHR as early as 4 weeks old with normal blood pressure, 33 mitochondrial protein spots presented significant alterations, with 17 down-regulated and 16 up-regulated. Such alterations were much greater than those in 20-week-old SHR with elevated blood pressure. Of the total alterations, the expression of two mitochondrial enzymes, trifunctional enzyme alpha subunit (Hadha) and NADH dehydrogenase 1 alpha subcomplex 10 (Ndufa10), were found to have special expression modification patterns in SHR strain. These data would provide new clues to investigate the potential contribution of mitochondrial dysfunction to the development of cardiac hypertrophy.

Efficacy and safety of benidipine therapy of essential hypertension in elderly Chinese patients.

BACKGROUND: Benidipine (CAS 105979-17-7) is a dihydropyridine calcium channel blocker used in the treatment of hypertension and angina pectoris. OBJECTIVE: To examine the efficacy and safety of therapy with benidipine in elderly hypertensive patients. METHODS: Chinese patients >60 years of age with mild to moderate essential hypertension were enrolled. The patients were prescribed benidipine at the dose of 8 mg once daily for 12 weeks. Detailed laboratory examinations and 24-h ambulatory blood pressure monitoring were performed before and after the treatment. RESULTS: One hundred and sixty-four of the 180 patients enrolled completed the 12-week active treatment phase. Sitting systolic blood pressure (SBP) and diastolic blood pressure (DBP) reductions at the end of treatment were 21.50 +/- 12.83 and 10.60 +/- 8.04 mmHg, respectively; the proportion of patients showing a good treatment response was 95.1% for SBP and 96.9% for DBP. Benidipine significantly reduced the mean 24-h ambulatory blood pressure (p < 0.001 vs. baseline) exhibiting smooth, sustained effects and high trough-to-peak ratios (T/P ratio) (0.87 for SBP and 0.72 for DBP). Moreover, benidIpine significantly reduced the systolic morning blood pressure surge and urinary albumin, and it was well tolerated. No serious adverse events were noted. CONCLUSION: Benidipine was welltolerated and effective in elderly Chinese patients with essential hypertension.

Hyper-phosphorylation of alpha-enolase in hypertrophied left ventricle of spontaneously hypertensive rat.

Cardiac hypertrophy is one of the main target organ damages of essential hypertension and predicts a poor prognosis of the disease. The molecules involved in this event, especially their posttranslational modifications, remain largely unknown to date. With a combination of phosphoprotein column enrichment and two-dimensional gel electrophoresis separation, here we compared the profiling of enriched phosphoproteins from the left ventricle (LV) of spontaneously hypertensive rats (SHR) to that of age-matched Wistar Kyoto rats. As a result, 19 differential proteins were found in the hypertrophied LV of SHR. Among them, we focused on a glycolysis enzyme alpha-enolase, of which the hyper-phosphorylation was shown in the hypertrophied LV but not in non-hypertrophied atrium and right ventricle of SHR. Furthermore, the alpha-enolase hyper-phosphorylation was accompanied by decreased enzymatic activity. The further investigation based on these results would provide new clues to understand the pathological process of cardiac hypertrophy in SHR.

Peroxisome proliferator-activated receptor-gamma agonists attenuate angiotensin II-induced collagen type I expression in adventitial fibroblasts.

1. Angiotensin (Ang) II-mediated oxidative stress may be important in enhanced adventitial fibroblast collagen formation. The aim of the present study was to test whether PPAR-gamma agonists 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ2) and pioglitazone could alter AngII-induced collagen type I formation in vascular adventitial fibroblasts via reactive oxygen species (ROS). 2. Vascular adventitial fibroblasts were isolated from rat thoracic aortas of male Sprague-Dawley rats and treated with different concentrations of AngII for different periods of time. The expression of collagen type I induced by AngII was examined by western blot. Expression of PPAR-gamma mRNA was examined by reverse transcription-polymerase chain reaction (RT-PCR). Intracellular ROS generation was measured by flow cytometry. Activation of transcription factors nuclear factor (NF)-kappaB and activator protein (AP)-1 was assessed by an electrophoretic mobility shift assay. 3. Angiotensin II increased expression of collagen type I in a time- and dose-dependent manner in adventitial fibroblasts. In addition, AngII stimulated intracellular generation of ROS in adventitial fibroblasts. Pretreatment of cells with 15d-PGJ2 and pioglitazone attenuated collagen type I expression and generation of ROS induced by AngII, respectively. Moreover, we observed that N-acetylcysteine inhibited collagen type I expression induced by AngII as did the PPAR-gamma agonists. Angiotensin II treatment activated the redox-sensitive transcription factors NF-kappaB and AP-1, whereas pretreatment with 15d-PGJ2 and pioglitazone reduced the AngII-induced DNA-binding activity of NF-kappaB but not AP-1. 4 Our data demonstrate that the PPAR-gamma agonists 15d-PGJ2 and pioglitazone attenuate AngII-mediated collagen type I expression in adventitial fibroblasts, which may be mediated by the modulation of ROS release and the redox-sensitive transcription factor NF-kappaB.

Differential protein expression in hypertrophic heart with and without hypertension in spontaneously hypertensive rats.

Although cardiac hypertrophy in hypertension has been well recognized, the molecular mechanisms for the development of hypertrophy are still largely unknown. In this study, the protein expression profiles of left ventricular myocardia in spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats at different ages were analyzed using 2-DE in combination with MALDI-TOF/TOF MS/MS. The results showed that 20 proteins were modulated in the hypertrophic myocardium. Out of these modulated proteins, 13 proteins presented significant changes in SHR at an early stage prior to the development of sustained hypertension, while the changes of the other 7 protein expressions occurred only at a late stage in SHR when the blood pressure was significantly elevated, and were largely reversible by treatment with rennin-angiotensin-aldosterone system inhibitors losartan or enalapril. These data demonstrate that the changes in energy metabolism in the hypertrophied heart favor an increase in glycolysis and a decrease in oxidation of fatty acid and glucose, which occur at an early stage in SHR without hypertension. Our results also provide evidence to support the hypothesis that oxidative stress plays an important role in the development of hypertensive cardiac hypertrophy.