The purpose of this study was to investigate the change of aortic semicarbazide-sensitive amine oxidase (SSAO) activity in diabetic rats and examine the effect of 2-bromoethylamine (2-BEA) on SSAO activity and vascular endothelium in diabetic rats. SSAO was prepared from rat aorta. For assessment of the inhibitory effect, the enzymes were preincubated in the presence of different concentrations of 2-BEA before the addition of benzylamine in vitro. Type 1 diabetic rat model was induced by a single intraperitoneal injection of streptozotocin (STZ). Sprague Dawley (SD) rats were randomly divided into normal control group (NC), diabetic model group (DM), 2-BEA 5 mg/kg group, 2-BEA 20 mg/kg group (n = 10 in each group). 2-BEA was administered daily via intraperitoneal injection for 8 weeks. At the end of 8 weeks, blood sample was collected from the abdominal aorta. Plasma nitric oxide (NO) was determined by nitrate reductase method. Plasma endothelin-1 (ET-1) was determined by radioimmunoassay. Aorta SSAO was determined by high performance liquid chromatography. The aorta was prepared to observe morphological changes and ultramicroscopic structures. The results were as follows: Compared with NC group, aortic SSAO activity and the plasma ET-1 were significantly increased (P < 0.01), and plasma NO was significantly decreased (P < 0.01) in DM group. 2-BEA decreased plasma ET-1 and elevated plasma NO by inhibiting aortic SSAO activity in diabetic rats (P < 0.01), and 2-BEA 20 mg/kg group was more significant than 2-BEA 5 mg/kg group (P < 0.05). Endothelial injury of 2-BEA group rats was less serious than DM group. These results suggest that 2-BEA protect aortic endothelium by inhibiting aortic SSAO activity.
Amine Oxidase (Copper-Containing) ; metabolism ; Animals ; Aorta, Abdominal ; enzymology ; Diabetes Mellitus, Experimental ; enzymology ; Endothelin-1 ; blood ; Endothelium, Vascular ; drug effects ; Ethylamines ; pharmacology ; Protective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley

Previous evidence supports the important role that oxidative stress (OxS) plays in metabolic syndrome (MetS)-related manifestations. We determined the relationship between the number of MetS components and the degree of OxS in MetS patients. In this comparative cross-sectional study from the LIPGENE cohort, a total of 91 MetS patients (43 men and 48 women; aged between 45 and 68 years) were divided into four groups based on the number of MetS components: subjects with 2, 3, 4 and 5 MetS components (n=20, 31, 28 and 12, respectively). We measured ischemic reactive hyperemia (IRH), plasma levels of soluble vascular cell adhesion molecule-1 (sVCAM-1), total nitrite, lipid peroxidation products (LPO), hydrogen peroxide (H2O2), superoxide dismutase (SOD) and glutathione peroxidase (GPx) plasma activities. sVCAM-1, H2O2 and LPO levels were lower in subjects with 2 or 3 MetS components than subjects with 4 or 5 MetS components. IRH and total nitrite levels were higher in subjects with 2 or 3 MetS components than subjects with 4 or 5 MetS components. SOD and GPx activities were lower in subjects with 2 MetS components than subjects with 4 or 5 MetS components. Waist circumference, weight, age, homeostatic model assessment-beta, triglycerides (TGs), high-density lipoprotein and sVCAM-1 levels were significantly correlated with SOD activity. MetS subjects with more MetS components may have a higher OxS level. Furthermore, association between SOD activity and MetS components may indicate that this variable could be the most relevant OxS biomarker in patients suffering from MetS and could be used as a predictive tool to determine the degree of the underlying OxS in MetS.
Aged ; Anthropometry ; Antioxidants/metabolism ; Biological Markers/metabolism ; Blood Pressure ; Endothelium, Vascular/pathology/physiopathology ; Female ; Glutathione Peroxidase/blood ; Humans ; Hydrogen Peroxide/metabolism ; Hyperemia/blood/physiopathology ; Male ; Metabolic Syndrome X/blood/enzymology/*pathology/physiopathology ; Middle Aged ; Nitrites/blood ; *Oxidative Stress ; Regression Analysis ; Superoxide Dismutase/blood ; Vascular Cell Adhesion Molecule-1/metabolism

The incidence of cardiovascular disease is predicted to increase as the population ages. There is accumulating evidence that arginase upregulation is associated with impaired endothelial function. Here, we demonstrate that arginase II (ArgII) is upregulated in aortic vessels of aged mice and contributes to decreased nitric oxide (NO) generation and increased reactive oxygen species (ROS) production via endothelial nitric oxide synthase (eNOS) uncoupling. Inhibiting ArgII with small interfering RNA technique restored eNOS coupling to that observed in young mice and increased NO generation and decreased ROS production. Furthermore, enhanced vasoconstrictor responses to U46619 and attenuated vasorelaxation responses to acetylcholine in aged vasculature were markedly improved following siRNA treatment against ArgII. These results might be associated with increased L-arginine bioavailability. Collectively, these results suggest that ArgII may be a valuable target in age-dependent vascular diseases.
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid/pharmacology ; Aging ; Animals ; Aorta/enzymology/physiopathology ; Arginase/genetics/*metabolism ; Endothelium, Vascular/*enzymology/physiopathology ; Enzyme Induction ; Gene Knockdown Techniques ; Mice ; Mice, Inbred C57BL ; Nitric Oxide/metabolism ; Nitric Oxide Synthase Type III/*metabolism ; RNA, Small Interfering/genetics ; Reactive Oxygen Species/metabolism ; Up-Regulation ; Vasoconstriction/drug effects

BACKGROUNDIncreasing evidences indicate that an activated renin-angiotensin system (RAS) causes an imbalance between the vasoconstrictive and vasodilator mechanisms involving the pulmonary circulation leading to the development of pulmonary arterial hypertension (PAH). Angiotensin-converting enzyme 2 (ACE2), a primary component of the vasoprotective axis in RAS, is recently identified that it has regulatory actions in lung pathophysiology, but the mechanism in these processes is uncertain yet.

METHODSSevere PAH was induced by monocrotaline injection one week following pneumonectomy in rats. The activation of ACE2 by continuous injection of resorcinolnaphthalein was studied by real time-polymerase chain reaction (RT-PCR), Western blotting and fluorogenic peptide assay. Endothelial functions were evaluated by the response to acetylcholine and cytokines were measured by RT-PCR seven days after monocrotaline injection. The PAH-related hemodynamics and pathological changes were examined at day 21 when severe PAH was completely established.

RESULTSResorcinolnaphthalein caused significant activation of ACE2 in both normal and diseased rats in 7 days after treatment. The pulmonary arterial pressure (PAP) started to increase at least 7 days after monocrotaline injection, and the rats developed severe PAH in 21 days with high PAP, right ventricular hypertrophy and neointimal formation. Treatment with resorcinolnaphthalein prevented these features. Resorcinolnaphthalein caused an improved endothelia-dependent vasorelaxation and decrease in proinflammatory cytokines (tumor necrosis factor (TNF)-α, monocyte chemoattractant protein-1 (MCP-1), interleukin (IL)-6) and increase in anti-inflammatory cytokine IL-10 in the early stage of the pathogenesis.

CONCLUSIONSThese results demonstrated that activation of ACE2 by continuous injection of resorcinolnaphthalein prevented the development of PAH through improving early endothelial dysfunction and mediating the level of proinflammatory and anti-inflammatory cytokines.

Animals ; Cytokines ; biosynthesis ; Endothelium, Vascular ; physiology ; Enzyme Activation ; drug effects ; Familial Primary Pulmonary Hypertension ; Hypertension, Pulmonary ; enzymology ; prevention & control ; Inflammation ; prevention & control ; Male ; Peptidyl-Dipeptidase A ; physiology ; Rats ; Rats, Sprague-Dawley ; Resorcinols ; pharmacology

Hypertension is associated with endothelial dysfunction and increased cardiovascular risk. Caveolin-1 regulates nitric oxide (NO) signaling by modulating endothelial nitric oxide synthase (eNOS). The purpose of this study was to examine whether HMG-CoA reductase inhibitor improves impaired endothelial function of the aorta in spontaneous hypertensive rat (SHR) and to determine the underlying mechanisms involved. Eight-week-old male SHR were assigned to either a control group (CON, n=11) or a rosuvastatin group (ROS, n=12), rosuvastatin (10 mg/kg/day) administered for eight weeks. Abdominal aortic rings were prepared and responses to acetylcholine (10-9-10-4 M) were determined in vitro. To evaluate the potential role of NO and caveolin-1, we examined the plasma activity of NOx, eNOS, phosphorylated-eNOS and expression of caveolin-1. The relaxation in response to acetylcholine was significantly enhanced in ROS compared to CON. Expression of eNOS RNA was unchanged, whereas NOx level and phosphorylated-eNOS at serine-1177 was increased accompanied with depressed level of caveolin-1 in ROS. We conclude that 3-Hydroxy-3-methylglutaryl Coenzyme-A (HMG-CoA) reductase inhibitor can improve impaired endothelial dysfunction in SHR, and its underlying mechanisms are associated with increased NO production. Furthermore, HMG-CoA reductase inhibitor can activate the eNOS by phosphorylation related to decreased caveolin-1 abundance. These results imply the therapeutic strategies for the high blood pressure-associated endothelial dysfunction through modifying caveolin status.
Acetylcholine/metabolism ; Animals ; Aorta/metabolism/physiopathology ; Blood Pressure/drug effects ; Caveolin 1/*metabolism ; Down-Regulation ; Drug Administration Schedule ; Endothelium, Vascular/*drug effects/physiopathology ; Fluorobenzenes/administration & dosage/*pharmacology ; Hydroxymethylglutaryl-CoA Reductase Inhibitors/administration & dosage/*pharmacology ; Hypertension/enzymology/metabolism/*physiopathology ; Male ; Nitric Oxide/blood ; Nitric Oxide Synthase Type III/*metabolism ; Phosphorylation ; Pyrimidines/administration & dosage/*pharmacology ; Rats ; Rats, Inbred SHR ; Sulfonamides/administration & dosage/*pharmacology ; Vasodilation/drug effects

OBJECTIVETo explore the molecular biological mechanism for tanshinone II A reversing left ventricular hypertrophy, it would be studying the effect of tashinone on the endothelial nitric oxide synthase (eNOS) and protein kinase C (PKC) in the hypertrophic cadiocyte of rats suffered abdominal aorta constriction.

METHODSD rats were operated with abdominal aorta constriction and 8 rats were done with sham surgery. After 4 weeks, all rats were divided into 4 groups: myocardial hypertrophy group, low dose tanshinone II A group (10 mg x kg(-1) x d(-1)), high dose tanshinone II A group (20 mg x kg(-1) x d(-1)) and valsartan group (10 mg x kg(-1) d(-1) intragastric administration). 8 weeks later, the rats were used to measure the left ventricular mass index (LVMI) with the tissue of left ventricle and myocardial fiber dimension (MFD) by pathological section and HE stain, to detect the nitric oxide content by nitrate reductase, to detect the genic expression of eNOS by RT-PCR and to detect the activity of protein kinase C (PKC) by Western blotting.

RESULT1) The blood pressure in group myocardial hypertrophy [(186 +/- 13) mmHg] and tansginone II A [low and high dose (188 +/- 11,187 +/- 14) mmHg] was obviously higher than that in group sham surgery and valsartan group [vs (117 +/- 8, 136 +/- 15) mmHg, P < 0.01]. But there was no difference between group myocardial hypertrophy and group tanshinone II A (low and high dose). 2) The LVMI and MFD were obviously higher in group tanshinone II A low and high dose) and group valsartan than those in group sham surgery (P < 0.05), and lower than those in group myocardial hypertrophy (P < 0.01). 3) The NO level was obviously higher in group tanshinone II A (low and high dose) and group valsartan than that in group myocardial hypertrophy (12.78 +/- 1.66, 11.95 +/- 1.39, 12.26 +/- 2.08 vs 5.83 +/- 1.06) micromol x L(-1), (P < 0.01 ), and lower than that in group sham surgery (vs 19.35 +/- 1.47) micromol x L(-1), (P < 0.05). 4) The expressive level of eNOS mRNA and protein in myocardial hypertrophy group was less than that in other groups (P < 0.01). And valsartan group was less than tanshinone II A groups and sham surgery group (P < 0.05), but there were no difference among the two tanshinone II A groups and sham surgery group. 5) The level of PKC protein in group myocardial hypertrophy was obviously higher than that in all the other groups (1.291 +/- 0.117 vs 0.563 +/- 0.094, 0.605 +/- 0.051, 0.519 +/- 0.062, 0.827 +/- 0.086, P < 0.01), and the level in group valsartan was higher than that in group sham operation and group tanshinone II A (low and high dose).

CONCLUSIONNO/NOS system in local myocardium has close relationship with the pathological process for myocardial hypertrophy. Tanshinone II A can produce the pharmacological action to reverse myocardial hypertrophy by inhibiting the activity of PKC and promoting the genic expression of eNOS in local myocardium and the production of endogenous NO.

Animals ; Aorta, Abdominal ; pathology ; Benzofurans ; pharmacology ; Blood Pressure ; drug effects ; Cardiomyopathy, Hypertrophic ; complications ; enzymology ; physiopathology ; Constriction, Pathologic ; complications ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Endothelium, Vascular ; drug effects ; enzymology ; Female ; Gene Expression Regulation, Enzymologic ; drug effects ; Heart Ventricles ; drug effects ; metabolism ; pathology ; physiopathology ; Male ; Myocytes, Cardiac ; drug effects ; enzymology ; pathology ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; genetics ; metabolism ; Protein Kinase C ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats

AIMTo observe the effects of inulicin on the neointimal hyperplasia and expression of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2) after balloon injury in rat aorta.

METHODSThe intimal hyperplasia model was replicated by balloon injury. The histological changes in vascular wall were observed by HE staining. MMP-2 activation was tested by gelatin zymogram analysis; MMP-2 and TIMP-2 expression was detected by Western blot and immunohistochemistry analysis.

RESULTSNeointimal hyperplasia induced by balloon injury was significantly inhibited by inulicin. The proteolytic activity and the expression of MMP-2 and ratio of MMP-2 and TIMP-2 were also returned to control level by inulicin after balloon injury.

CONCLUSIONInulicin inhibits hyperplasia of neointima by modulating the balance of MMP-2 and TIMP-2.

Animals ; Endothelium, Vascular ; drug effects ; enzymology ; metabolism ; Hyperplasia ; prevention & control ; Male ; Matrix Metalloproteinase 2 ; metabolism ; Neointima ; prevention & control ; Rats ; Rats, Sprague-Dawley ; Sesquiterpenes ; pharmacology ; Tissue Inhibitor of Metalloproteinase-2 ; metabolism

OBJECTIVETo observe the effects of Xiongshao Capsule (XSC) on blood vessel collagenase gene expression in experimental rabbits with arterial restenosis, and to probe its mechanisms for preventing restenosis.

METHODSRestenosis rabbit model was established by injuring endothelium of abdominal aorta by balloon dilation and feeding with high fatty diet for 6 weeks. Eighty rabbits were randomly allocated into 8 groups, Group A, normal rabbit for control; Group B, rabbit with simple injured arterial endothelium; Group C, model rabbits at different times after modeling (3 days for Group C1, 2 weeks for Group C2, and 6 weeks for Group C3); Group D, model rabbit treated with Probucol for 6 weeks; Group E and F, model rabbit treated with small and large dose of XSC respectively. The effect of XSC on collagenase gene expression during the course of restenosis was observed adopting RT-PCR method and computer image analyzer, and its mechanisms in preventing RS were probed by combined analyzing the change of collagen and patho-morphological examination.

RESULTSCompensatory dilation of lumens appeared at the end of the 2nd week; while 6 weeks after modeling, the diameters of lumens obviously diminished with an apparently increased proliferation index. The cell proliferation inhibiting effect in Group D and F was significant. The total amount of collagen increased and reached the peak at the 2nd week but without conspicuous accumulation on intima, which increased gradually and reached its peak at the 6th week. In Group D-F, especially in Group F, the amount of collagen in vascular wall (intima, media and externa) was lesser than that in Groups C. MMP-1 mRNA showed weak expression in Group A and Group C1-C3; significant difference only existed in comparing Group F with C3 (P < 0.05).

CONCLUSIONXSC could markedly increase the MMP-1 mRNA expression in injured portion of vessels, suggesting that its action in preventing RS might be related with the up-regulating of MMP-1 mRNA expression, increasing collagen degradation and reducing collagen deposition in vascular wall.

Animals ; Aorta, Abdominal ; drug effects ; metabolism ; pathology ; Aortic Valve Stenosis ; enzymology ; etiology ; prevention & control ; Capsules ; Catheterization ; adverse effects ; Drugs, Chinese Herbal ; pharmacology ; Endothelium, Vascular ; drug effects ; metabolism ; pathology ; Female ; Gene Expression Regulation, Enzymologic ; drug effects ; Male ; Matrix Metalloproteinase 1 ; genetics ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Rabbits ; Random Allocation ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate the influence of endothelial dysfunction induced by inoculated dendritic cells (DCs) loaded heat shock protein 60 (HSP60) in apolipoprotein (Apo) E-null mice, and the effect of Puerarin on it.

METHODSHSP60 DC (DChsp) acquired after prepared bone marrow-derived DCs of ApoE-null mice and treated with HSP60. In vitro, the function of DCs and the effect of Puerarin were detected. While in vivo, ApoE-null mice fed with high-cholesterol forage were divided into two groups and intravenous inoculated with DCh-sp or normal saline via vein twice respectively. The mice in the two groups were subdivided into the Puerarin group and non-treated group, and they were injected intraperitoneally with Puerarin and normal saline at the beginning of inoculation and the following 3 weeks, respectively. In addition, C57BL/6 mice without inoculation were taken as the normal control group. Two weeks after the last time inoculated, the response of T lymphocytes to HSP60 and endothelial-dependent diastolic function of aortic ring were detected.

RESULTSHSP60 could promote DCs expressed CD86 and stimulate T lymphocytes proliferation in vitro, while Puerarin had significantly inhibitory effect. After inoculated, DChsp activated inflammatory response in vivo and aggravated endothelium-dependent dilation in mice. Puerarin could significantly inhibit inflammatory reaction caused by DChsp and improve endothelium dilation.

CONCLUSIONHsp60 could activate DCs in vitro and in vivo, Puerarin could significantly inhibit specific immunity induced by HSP60 and improve vascular endothelium-dependent dilation.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Apolipoproteins E ; genetics ; B7-2 Antigen ; immunology ; Cell Proliferation ; drug effects ; Chaperonin 60 ; metabolism ; Dendritic Cells ; drug effects ; enzymology ; immunology ; Endothelium, Vascular ; drug effects ; physiology ; Immunity ; drug effects ; Inflammation ; chemically induced ; Isoflavones ; pharmacology ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Protective Agents ; pharmacology ; T-Lymphocytes ; drug effects ; Vasodilator Agents ; pharmacology

Fluid shear stress plays a critical role in vascular health and disease. While protein kinase A (PKA) has been implicated in shear-stimulated signaling events in endothelial cells, it remains unclear whether and how PKA is stimulated in response to shear stress. This issue was addressed in the present study by monitoring the phosphorylation of endogenous substrates of PKA. Shear stress stimulated the phosphorylation of cAMP responsive element binding protein (CREB) in a PKA-dependent manner. Western blot analysis using the antibody reactive against the consensus motif of PKA substrates detected two proteins, P135 and P50, whose phosphorylation was increased by shear stress. The phosphorylation of P135 was blocked by a PKA inhibitor, H89, but not by a phosphoinositide 3-kinase inhibitor, wortmannin. Expression of a constitutively active PKA subunit stimulated P135 phosphorylation, supporting the potential of P135 as a PKA substrate. P135 was identified as endothelial nitric oxide synthase (eNOS) by immunoprecipitation study. PKA appeared to mediate shear stress-stimulated eNOS activation. Shear stress stimulated intracellular translocation of PKA activity from 'soluble' to 'particulate' fractions without involving cellular cAMP increase. Taken together, this study suggests that shear stress stimulates PKA-dependent phosphorylation of target proteins including eNOS, probably by enhancing intracellular site-specific interactions between protein kinase and substrates.
Animals ; Aorta, Thoracic/cytology ; Blotting, Western ; Cattle ; Cell Culture Techniques ; Cell Extracts ; Cells, Cultured ; Comparative Study ; Cyclic AMP-Dependent Protein Kinases/analysis/*metabolism ; Endothelium, Vascular/cytology/*enzymology/*metabolism ; Nitric Oxide Synthase Type III/analysis/*metabolism ; Phosphorylation ; Precipitin Tests ; Research Support, Non-U.S. Gov't ; Stress, Mechanical ; Time Factors