Angiotensin II increases urokinase-type plasminogen activator expression and induces aneurysm in the abdominal aorta of apolipoprotein E-deficient mice

Urokinase-type plasminogen activator (uPA) is increased in human abdominal aortic aneurysm (AAA). Chronic infusion of angiotensin II (Ang II) results in AAA in apolipoprotein E-deficient mice. We tested the hypothesis that Ang II infusion results in an elevation of uPA expression contributing to aneurysm formation. Ang II or vehicle was infused by osmotic pumps into apoE-KO mice. All mice treated with Ang II developed a localized expansion of the suprarenal aorta (75% increase in outer diameter), accompanied by an elevation of blood pressure (22 mmHg), compared to the vehicle-treated group. Histological examination of the dilated aortic segment revealed similarities to human AAA including focal elastin fragmentation, macrophage infiltration, and intravascular hemorrhage. Ang II treatment resulted in a 13-fold increase in the expression of uPA mRNA in the AAA segment in contrast to a twofold increase in the atherosclerotic aortic arch. Increased uPA protein was detected in the abdominal aorta as early as 10 days after Ang II infusion before significant aorta expansion. Thus, Ang II infusion results in macrophage infiltration, increased uPA activity, and aneurysm formation in the abdominal aorta of apoE-KO mice. These data are consistent with a causal role for uPA in the pathogenesis of AAA.     

Reduced cardiac functional reserve in apolipoprotein E knockout mice

Several years ago, the authors reported that aortic flow velocity under resting conditions was significantly higher in apolipoprotein E knockout (apoE-KO) mice than in age-matched C57Black/6J wildtype (WT) controls. The goal of this study was to examine whether the cardiac functional reserve is impacted in response to a pharmacological stress agent in apoE-KO mice. Cardiac function was measured noninvasively by the Doppler ultrasound method at baseline and at 1 min, 5 min, 10 min, and 20 min after intraperitoneal injection of dobutamine at the doses of 1 microg/g, 3 microg/g, or 10 microg/g in 16-month-old male apoE-KO (n = 9) and WT (n = 10) mice under light anesthesia with 1.5% isoflurane via inhalation. The baseline peak and mean aortic flow velocities were 39% to 48% higher, and left ventricular contractility measured by peak acceleration rate of aortic flow velocity was 24% higher in apoE-KO compared with WT mice (P < 0.01). Dobutamine stress dose-dependently increased cardiac function, which, however, was significantly smaller with a right shift of the dose-response curve in apoE-KO mice compared with WT controls. The hypotensive response to dobutamine was not significantly different between the 2 groups. Thus, despite an elevated resting aortic flow velocity and left ventricular contractility, cardiac functional reserve in response to dobutamine stress was significantly reduced in apoE-KO mice, which could be the consequence of coronary atherosclerosis and endothelial dysfunction that limits blood supply to the heart.     

Involvement of the nitric oxide system in the anti-atherosclerotic potential of lacidipine in the ApoE-deficient mouse: a morphological, functional, and electrochemical study

The present study investigated the anti-atherosclerotic activity of lacidipine, a calcium antagonist with antioxidant properties in apoE-deficient mice. These mice show widespread vascular lesions which closely resemble the inflammatory-fibrous plaques seen in humans in atherosclerosis. Mice were fed a Western-type diet (WTD), and treated for 8 weeks with either vehicle or lacidipine at 3 or 10 mg/kg/day. In parallel with histological studies of atherosclerotic lesions in the aorta, functional studies on vascular acetylcholine (ACh) reactivity and analysis of voltammetric levels of nitric oxide (NO) were performed. Recent work has suggested that dihydropyridines (DHPs) modulate vascular relaxation via an increase in the release of NO. Lacidipine treatment had no effect on the plasma lipid profile. However, a significant (p < 0.01) dose-related reduction of 36.4% and 43.3% of the aortic lesion area in respect to methocel-treated mice was observed. Moreover, the aortic ring from control apoE-deficient mice fed a WTD for 8 weeks showed a lower relaxation in response to ACh in comparison to wild-type C57BL/6J mice; on the contrary, lacidipine-treated apoE-deficient mice lacidipine-treated displayed a response similar to that of wildtype C57BL/6J mice. Voltammetric analyses demonstrated a significant decrease of NO release in apoE-deficient mice, while lacidipine-treated mice showed enhanced activity of the NO system. We conclude that lacidipine reduced the extent of atherosclerotic area in hypercholesterolemic apoE-deficient mice, and this reduction may be associated with the capacity of the drug to maintain endothelial NO levels at concentrations useful to protect against vascular damage.     

Nitric oxide counteracts angiotensin II induced contraction in efferent arterioles in mice

AIM: Efferent arterioles (Ef) are one of the final control elements in glomerular haemodynamics. The influence of nitric oxide (NO) on Ef remains ambiguous. METHODS: To test the hypothesis that endothelial NO plays an important role in this context, afferent arterioles (Af) and Ef of wild-type mice (WT), and Ef of mice lacking the endothelial NO synthetase [eNOS(-/-)] were perfused. Perfusion was performed in Ef via Af (orthograde) as well as from the distal end of Ef (retrograde), which provides an estimate for the importance of substances derived from the glomerulus. Angiotensin II (Ang II) was added in doses ranging from 10(-12) to 10(-6) mol L(-1) to the bath solution. RESULTS: Ang II reduced the luminal diameter of Af to 68 +/- 7 and in Ef to 55 +/- 8% during orthograde, and to 35 +/- 6% during retrograde perfusion (10(-6) mol L(-1) Ang II) in WT. Pre-treatment with N(G)-Nitro-L-arginine-methylester (l-NAME) (10(-4) mol L(-1)) increased the Ang II sensitivity in retrograde (17 +/- 9%) and orthograde perfused Ef (19 +/- 9%). The Ang II sensitivity was enhanced in eNOS(-/-) mice compared with WT, too. Already at a dose of Ang II 10(-9) mol L(-1), luminal diameters diminished to 8 +/- 7 and 7 +/- 4%. CONCLUSION: The increased Ang II sensitivity during L-NAME pre-treatment and in eNOS(-/-) mice indicates a strong counteraction of endothelial derived NO on Ang II induced contraction in Ef. Moreover, Ef are similarly sensitive to Ang II during either retrograde or orthograde perfusion in the absence of NO effects, suggesting that NO mediates, at least in part, the action of potential vasodilatory substances from the glomerulus.     

Sodium nitrite therapy rescues ischemia-induced neovascularization and blood flow recovery in hypertension

Arterial hypertension is a major risk factor that can lead to complication of peripheral vascular disease due, in part, to endothelial dysfunction. Because sodium nitrite (SN) can be converted to nitric oxide (NO), which counteracts endothelial dysfunction, we explored the effect of nitrite on neovascularization following hind limb ischemia in different models of hypertension (HT). Chronic delivery of angiotensin II (Ang II, 400?ng/kg/min) or N(omega)-nitro-l-arginine-methyl-ester (L-NAME, 0.1?g/L) was used for a 2-week period to induce hypertension. Mice were subjected to femoral artery ligation-induced ischemia in the hind limb followed by treatment with SN (50?mg/L) for 2?weeks. SN significantly reduced systolic arterial blood pressure in mice receiving Ang II and L-NAME but had no effect in sham animals. After 2?weeks, blood flow and microangiography showed 60?%?±?1.0 recovery in sham compared with 40?%?±?1.3 in HT mice. Importantly, sham and HT mice treated with SN showed a 100?% blood flow recovery associated with normalization in capillary density. The inhibition of xanthine-oxido-reductase (allopurinol) or VEGFR (SU-5416) prevented the neovascularization in HT mice treated with SN. Cyclic GMP (cGMP) content in the hind limb was significantly increased in mice treated with SN compared with non-treated mice. Nitrite/nitrate content was only increased in the sham group treated with SN. Immunoprecipitation and Western blot analysis revealed an increase in eNOS/Akt/VEGFR phosphorylation in skeletal muscle from mice treated with SN compared with non-treated mice. Our findings indicate that SN therapy rescues the neovascularization and blood flow recovery in the ischemic hind limb of sham and HT mice likely through the Akt/NO/cGMP and VEGFR pathways.     

Elevated endothelial nitric oxide bioactivity and resistance to angiotensin-dependent hypertension in 12/15-lipoxygenase knockout mice

12/15-Lipoxygenase (12/15-LOX) plays a pathogenic role in atherosclerosis. To characterize whether 12/15-LOX also contributes to endothelial dysfunction and hypertension, regulation of vessel tone and angiotensin II (ang II) responses were characterized in mice deficient in 12/15-LOX. There was a twofold increase in the magnitude of l-nitroarginine-methyl ester-inhibitable, acetylcholine-dependent relaxation or phenylephrine-dependent constriction in aortic rings isolated from 12/15-LOX(-/-) mice. Plasma NO metabolites and aortic endothelial NO synthase (eNOS) expression were also elevated twofold. Angiotensin II failed to vasoconstrict 12/15-LOX(-/-) aortic rings in the absence of L-nitroarginine-methyl ester, and ang II impaired acetylcholine-induced relaxation in wild-type, but not 12/15-LOX(-/-) rings. In vivo, 12/15-LOX(-/-) mice had similar basal systolic blood pressure measurements to wild type, however, blood pressure elevations in response to ang II infusion (1.1 mg/kg/day) were significantly attenuated (maximal pressure, 143.4 +/- 4 mmHg versus 122.1 +/- 5.3 mmHg for wild type and 12/15-LOX(-/-), respectively). In contrast, vascular hypertrophic responses to ang II, and ang II type 1 receptor (AT1-R) expression were similar in both strains. This study shows that 12/15-LOX(-/-) mice have increased NO biosynthesis and impaired ang II-dependent vascular responses in vitro and in vivo, suggesting that 12/15-LOX signaling contributes to impaired NO bioactivity in vascular disease in vivo.     

Evaluation of the effect of flavangenol on serum lipid peroxide levels and development of atherosclerosis in spontaneously hyperlipidemic B6.KOR-Apoeshl mice

Antioxidative flavonoids are used to reduce the risk of cardiovascular diseases in humans. However, the precise mechanism for the anti-atherosclerotic actions of flavonoids remains to be elucidated. In the present study, to assess the mechanism for the action of antioxidative flavonoids on atherosclerosis, we investigated the effect of flavangenol, one of the most potent antioxidants currently known, on spontaneously hyperlipidemic B6.KOR-Apoeshl mice. Flavangenol was orally administered to B6.KOR-Apoeshl mice ad libitum (6 mg flavangenol/mouse/day). After 6 months, serum levels of lipids (total cholesterol, triglyceride, HDL-cholesterol and LDL-cholesterol) and lipid peroxide were measured, and histopathological changes (lipid accumulation and inflammatory cell infiltration) in the aortic root were evaluated. Serum levels of total cholesterol and LDL-cholesterol were markedly increased, and HDL-cholesterol levels were decreased in B6.KOR-Apoeshl mice compared to C57BL/6 mice used as a control (p<0.001). Among these serum lipids, only HDL-cholesterol levels were significantly increased by flavangenol administration (p<0.05). Moreover, Oil Red O staining (lipid accumulation) was significantly increased in B6.KOR-Apoeshl mice compared to C57BL/6 mice (p<0.001). Notably, flavangenol administration significantly suppressed the increase in Oil Red O staining (p<0.01). Similarly, inflammatory cell infiltration into the intima was significantly increased in B6.KOR-Apoeshl mice compared to C57BL/6 mice (p<0.01), and flavangenol administration significantly suppressed the inflammatory cell infiltration (p<0.01). Importantly, flavangenol administration significantly reduced the increase of serum lipid peroxide levels in B6.KOR-Apoeshl mice (p<0.05). Together, these observations indicate that flavangenol, one of the most potent antioxidants, exerts its anti-atherosclerotic action on spontaneously hyperlipidemic and atherosclerotic B6.KOR-Apoeshl mice, possibly by increasing HDL-cholesterol levels and reducing lipid peroxide levels, thereby suppressing the lipid accumulation (formation of atherosclerotic lesions) and inflammatory cell infiltration (chronic inflammation) in the intima of the aortic root.     

Facilitation of renal autoregulation by angiotensin II is mediated through modulation of nitric oxide

AIMS: This study was designed to investigate the influence of angiotensin II (Ang II) and nitric oxide (NO) on autoregulation of renal perfusion. METHODS: Autoregulation was investigated in isolated perfused kidneys (IPRK) from Sprague-Dawley rats during stepped increases in perfusion pressure. RESULTS: Ang II (75-200 pM) produced dose-dependent enhancement of autoregulation whereas phenylephrine produced no enhancement and impaired autoregulation of GFR. Enhancement by Ang II was inhibited by the AT1 antagonist, Losartan, and the superoxide scavenger, Tempol. Under control conditions nitric oxide synthase (NOS) inhibition by 10 microm N-omega-nitro-L-arginine methyl ester (L-NAME) facilitated autoregulation in the presence of non-specific cyclooxygenase (COX) inhibition by 10 microm indomethacin. Both COX and combined NOS/COX inhibition reduced the autoregulatory threshold concentration of Ang II. Facilitation by 100 pm Ang II was inhibited by 100 microm frusemide. Methacholine (50 nm) antagonised Ang II-facilitated autoregulation in the presence and absence of NOS/COX inhibition. Infusion of the NO donor, 1 microm sodium nitroprusside, inhibited L-NAME enhancement of autoregulation under control conditions and during Ang II infusion. CONCLUSIONS: The results suggest than an excess of NO impairs autoregulation under control conditions in the IPRK and that endogenous and exogenous NO, vasodilatory prostaglandins and endothelium-derived hyperpolarizing factor (EDHF) activity antagonise Ang II-facilitated autoregulation. Ang II also produced a counterregulatory vasodilatory response that included prostaglandin and NO release. We suggest that Ang II facilitates autoregulation by a tubuloglomerular feedback-dependent mechanism through AT1 receptor-mediated depletion of nitric oxide, probably by stimulating generation of superoxide.     

Group V Secretory Phospholipase A(2) Enhances the Progression of Angiotensin II-Induced Abdominal Aortic Aneurysms but Confers Protection against Angiotensin II-Induced Cardiac Fibrosis in ApoE-Deficient Mice

Abdominal aortic aneurysms (AAAs) and heart failure are complex life-threatening diseases whose etiology is not completely understood. In this study, we investigated whether deficiency of group V secretory phospholipase A(2) (GV sPLA(2)) protects from experimental AAA. The impact of GV sPLA(2) deficiency on angiotensin (Ang) II-induced cardiac fibrosis was also investigated. Apolipoprotein E (apoE)(-/-) mice and apoE(-/-) mice lacking GV sPLA(2) (GV DKO) were infused with 1000 ng/kg per minute Ang II for up to 28 days. Increases in systolic blood pressure, plasma aldosterone level, and urinary and heart prostanoids were similar in apoE(-/-) and GV DKO mice after Ang II infusion. The incidence of aortic rupture in Ang II-infused GV DKO mice (10%) was significantly reduced compared with apoE(-/-) mice (29.4%). Although the incidence of AAA in GV DKO mice (81.3%) and apoE(-/-) mice (100%) was similar, the mean percentage increase in maximal luminal diameter of abdominal aortas was significantly smaller in GV DKO mice (68.5% ± 7.7%) compared with apoE(-/-) mice (92.6% ± 8.3%). Deficiency of GV sPLA(2) resulted in increased Ang II-induced cardiac fibrosis that was most pronounced in perivascular regions. Perivascular collagen, visualized by picrosirius red staining, was associated with increased TUNEL staining and increased immunopositivity for macrophages and myofibroblasts and nicotinamide adenine dinucleotide phosphate oxidase (NOX)-2 and NOX-4, respectively. Our findings indicate that GV sPLA(2) modulates pathological responses to Ang II, with different outcomes for AAA and cardiac fibrosis.     

Angiotensin II angiogenic effect in vivo involves vascular endothelial growth factor- and inflammation-related pathways

Although accumulating lines of evidence indicate the proangiogenic role of angiotensin II (Ang II), little is known about the molecular mechanisms associated with such an effect. This study aimed to identify molecular events involved in Ang II-induced angiogenesis in the Matrigel model in mice. C57Bl/6 female mice received a subcutaneous injection of either Matrigel or Matrigel with Ang II (10(-7) M) alone, with Ang II and an AT1 receptor antagonist (candesartan, 10(-6) M), or with Ang II and AT2 receptor antagonist (PD123319, 10(-6) M). After 14 days, angiogenesis was assessed in the Matrigel-plug by histological evaluation and cellular counting. Ang II increased by 1.9-fold the number of cells within the Matrigel (p < 0.01 versus control). Immunohistological analysis revealed the presence of macrophages, endothelial and smooth muscle cells, and the development of vascular-like structure. Such an angiogenic effect was associated with an increase in vascular endothelial growth factor (VEGF) (1.5-fold, p < 0.01), endothelial nitric oxide (eNOS) (1.7-fold, p < 0.01), and cyclooxygenase-2 (1.4-fold, p < 0.05) protein levels measured by Western blotting. Conversely, Ang II treatment did not affect MMP-9 and MMP-2 activity, assessed by zymography. Blockade of AT1 receptor completely prevented the Ang II-induced angiogenesis and protein regulations, whereas that of AT2 was ineffective. Administration of VEGF neutralizing antibody (2.5 microg ip twice a week) and cyclooxygenase-2 selective inhibitor (nimesulide, 30 mg/L) also hampered Ang II proangiogenic effect. In addition, Ang II-induced cell ingrowth was impaired by treatment with nitric oxide synthase inhibitor (L-NAME, 10 mg/kg/day) and in eNOS-deficient mice. Therefore, in an in vivo model, Ang II induced angiogenesis through AT1 receptor, which involved activation of VEGF/eNOS-related pathway and of the inflammatory process.     

Deterioration of atherosclerosis in mice lacking angiotensin II type 1A receptor in bone marrow-derived cells

The renin-angiotensin system (RAS) modulates end-organ damages, resulting in cardiovascular and kidney diseases. Experiments both in vitro and in vivo demonstrate that the angiotensin II (Ang II) type 1 (AT1) receptor pathway also exerts pro-inflammatory and pro-atherogenic effects on bone marrow-derived cells (BMDCs). Here, we investigated how AT1 receptor expression by BMDCs contributes to atherosclerosis and kidney injury in vivo by transplanting BM into RAS-activated transgenic mice. There was no difference in the extent of kidney damage between mice receiving BM transplants from mutant mice lacking the angiotensin II type 1a receptor (AT1a) gene and mice receiving transplants from wild-type (WT) mice. However, mice receiving transplants from AT1a 'knockout' (KO) mice displayed accelerated lethality and atherosclerotic lesions. These results indicated that the effects of AT1a receptor on BMDCs are organ dependent. Microarray expression profiling of macrophages from AT1a-KO mice revealed significant changes in the mRNA levels for a number of genes implicated in atherosclerosis. In accordance with the in vivo atherosclerosis results, AT1a-KO macrophages exhibited greater uptake of modified lipoproteins relative to macrophages from WT mice. We propose that the expression of AT1a receptor by BMDCs limits atherosclerosis in vivo.     

Central nitric oxide blocks vasopressin, oxytocin and atrial natriuretic peptide release and antidiuretic and natriuretic responses induced by central angiotensin II in conscious rats

The presence of nitric oxide synthase (NOS), the enzyme that catalyses the formation of nitric oxide (NO), in the circumventricular organs and magnocellular neurones suggests an important role of NO in the modulation of vasopressin (AVP) and oxytocin (OT) release. Intracerebroventricular (I.C.V.) injection of angiotensin II (Ang II) stimulates the release of AVP, OT and atrial natriuretic peptide (ANP), with the resultant antidiuretic and natriuretic effects. This study investigated the interaction between nitrergic and angiotensinergic pathways on the release of AVP, OT and ANP and on urinary volume and sodium excretion in water-loaded rats. Unanaesthetized, freely moving, male Wistar rats received two water loads followed by an injection into the lateral ventricle of an inhibitor of NOS (L-NAME), a NO donor [3-morpholinylsydnoneimine chloride (SIN-1) or S-nitroso-N-acetyl penicillamine (SNAP)] or vehicle (isotonic saline) and, 20 min after, they received a second I.C.V. injection of Ang II or vehicle. Injections of L-NAME or Ang II produced an increase in plasma levels of AVP, OT and ANP, a reduction in urinary volume and an increase in sodium excretion. Pretreatment with L-NAME enhanced the Ang II-induced increase in AVP, OT and ANP release, as well as the antidiuresis and natriuresis. Injection of SIN-1 or SNAP did not modify hormonal plasma levels and urinary parameters. In contrast SNAP blocked the AVP, OT and ANP release, as well as antidiuretic and natriuretic responses induced by ANG-II. Thus, the central nitrergic system can act to inhibit AVP, OT and ANP secretion and the antidiuretic and natriuretic effects in response to Ang II.     

Angiogenic actions of angiopoietin-1 require endothelium-derived nitric oxide

Angiopoietin1 (Ang1) is a novel angiogenic factor with important actions on endothelial cell (EC) differentiation and vascular maturation. Ang1 has been shown to prevent EC apoptosis through activation of PI3-kinase/Akt, a pathway that is also known to activate endothelium nitric oxide synthase (eNOS). Therefore, we hypothesized that the angiogenic effects of Ang1 would also be dependent on the PI3-kinase/Akt pathway, possibly mediated by increased eNOS activity and NO release. Treatment of human umbilical vein endothelial cells with recombinant Ang1* (300 ng/ml) for 15 minutes resulted in PI3-kinase-dependent Akt phosphorylation, comparable to that observed with vascular endothelial growth factor (VEGF) (50 ng/ml), and increased NO production in a PI3-kinase/Akt-dependent manner. Capillary-like tube formation induced by Ang1* in fibrin matrix at 24 hours (differentiation index, DI: 13.74 +/- 0.76 versus control 1.71 +/- 0.31) was abolished in the presence of the selective PI3-kinase inhibitor, LY294002 (50 micro mol/L) (DI: 0.31 +/- 0.31, P < 0.01) or the NOS inhibitor, L-NAME (3 mmol/L) (DI: 4.10 +/- 0.59, P < 0.01). In subcutaneous Matrigel implants in vivo, addition of recombinant Ang1* or wild-type Ang1 from conditioned media of COS-1 cells transfected with a pFLAG Ang1 expression vector, induced significant neovascularization to a degree similar to VEGF. Finally, angiogenesis in vivo in response to both Ang1 and VEGF was significantly reduced in eNOS-deficient compared with wild-type mice. In summary, our results demonstrate for the first time that endothelial-derived NO is required for Ang1-induced angiogenesis, and that the PI3-kinase signaling mediates the activation of eNOS and NO release in response to Ang1.     

Disruption of mindin exacerbates cardiac hypertrophy and fibrosis

Cardiac hypertrophy is a response of the myocardium to increased workload and is characterised by an increase of myocardial mass and an accumulation of extracellular matrix (ECM). As an ECM protein, an integrin ligand, and an angiogenesis inhibitor, all of which are key players in cardiac hypertrophy, mindin is an attractive target for therapeutic intervention to treat or prevent cardiac hypertrophy and heart failure. In this study, we investigated the role of mindin in cardiac hypertrophy using littermate Mindin knockout (Mindin ( -/- )) and wild-type (WT) mice. Cardiac hypertrophy was induced by aortic banding (AB) or angiotensin II (Ang II) infusion in Mindin ( -/- ) and WT mice. The extent of cardiac hypertrophy was quantitated by echocardiography and by pathological and molecular analyses of heart samples. Mindin ( -/- ) mice were more susceptible to cardiac hypertrophy and fibrosis in response to AB or Ang II stimulation than wild type. Cardiac function was also markedly exacerbated during both systole and diastole in Mindin ( -/- ) mice in response to hypertrophic stimuli. Western blot assays further showed that the activation of AKT/glycogen synthase kinase 3β (GSK3β) signalling in response to hypertrophic stimuli was significantly increased in Mindin ( -/- ) mice. Moreover, blocking AKT/GSK3β signalling with a pharmacological AKT inhibitor reversed cardiac abnormalities in Mindin ( -/- ) mice. Our data show that mindin, as an intrinsic cardioprotective factor, prevents maladaptive remodelling and the transition to heart failure by blocking AKT/GSK3β signalling.     

Salt-Sensitive Hypertension Resulting From Nitric Oxide Synthase Inhibition is Associated with Loss of Regulation of Angiotensin II in the Rat

In the Dahl salt-sensitive hypertensive rat, a diet containing L-arginine, the natural substrate for nitric oxide synthase, abrogates the hypertension. We postulated that nitric oxide synthase inhibition might induce a salt-sensitive form of hypertension and that this salt sensitivity might be linked to a loss of the regulatory effect of sodium ingestion on angiotensin II (Ang II) and angiotensinogen. Male Wistar-Kyoto rats were randomised to a diet containing 0.008 %, 2.2 % or 4.4 % sodium chloride and to treatment with the NO synthase inhibitor L-NAME (10 mg kg(-1) day(-1)) in the drinking water, or drinking water alone (Controls) for 4 weeks. Blood pressure was measured by tail cuff plethysmography twice weekly. After 4 weeks, the rats were anaesthetised and truncal blood collected for determination of angiotensinogen, renin, angiotensin I (Ang I), Ang II and aldosterone concentrations as well as angiotensin-converting enzyme (ACE) activity. Systolic blood pressure increased with increasing dietary sodium intake in the L-NAME-treated rats (P < 0.05). Plasma renin and aldosterone concentrations decreased with increasing dietary sodium intake in both Control and L-NAME-treated rats. Ang I and ACE activity were unchanged by increasing dietary sodium intake. In contrast, the plasma concentration of Ang II and angiotensinogen increased with increasing dietary sodium (P < 0.05 and P < 0.005, respectively). Treatment with the Ang II receptor blocker, losartan, reversed the blood pressure increase. We conclude that treatment with L-NAME induces an increase in blood pressure that is at least in part salt sensitive. Further, the salt-sensitive component appears to be Ang II-dependent, as it was associated with increasing plasma Ang II levels and could be reversed by treatment with an Ang II receptor antagonist.     

促红细胞生成素通过PI3-K/Akt信号通路抑制血管紧张素II诱导的新生大鼠心脏成纤维细胞增殖

目的： 探讨促红细胞生成素（EPO）对血管紧张素Ⅱ(Ang Ⅱ)诱导的心脏成纤维细胞（CFs）增殖的影响,以及磷脂酰肌醇-3-激酶/蛋白激酶B（PI3-K/Akt）信号途径和一氧化氮合酶（NOS）的作用。方法: 应用胰酶和胶原酶双酶和差速贴壁法分离培养新生大鼠CFs细胞,应用EPO、Ang Ⅱ、PI3-K抑制剂LY294002、NOS抑制剂L-NAME不同因素干预。细胞计数和MTT法作出CFs的生长曲线,检测CFs的增殖。化学酶法检测CFs培养液中的一氧化氮（NO）浓度以及总NOS和其亚型的活性。Western blotting检测Akt、p-Akt、内皮型一氧化氮合酶（eNOS）和诱生型一氧化氮合酶（iNOS）蛋白的表达。结果: Ang Ⅱ促CFs增殖的作用显著。EPO剂量依赖性的增加CFs培养液中的NO浓度,同时剂量依赖性的抑制Ang Ⅱ诱导的CFs增殖。在给药后第4 d,和单纯的Ang Ⅱ相比,浓度为5×103 U/L、1×104 U/L和2×104 U/L EPO对CFs增殖的抑制率分别达到了24.4％、41.5%和50.5%。EPO显著提高Akt的磷酸化水平,促进eNOS蛋白的表达。应用PI3-K抑制剂LY294002和NOS抑制剂L-NAME均能使培养液中的NO浓度也随之下降,EPO抑制Ang Ⅱ诱导CFs增殖的作用均被阻断。但LY294002同时阻断了eNOS蛋白表达,而L-NAME对eNOS没有影响。结论: EPO可剂量依赖性的抑制Ang Ⅱ诱导的新生大鼠CFs的增殖。其作用机制是通过激活PI3-K/Akt信号途径促使CFs中eNOS表达来促进NO的生成,从而抑制CFs的增殖。     

四逆汤与维生素E抗血管内皮功能氧化损伤及防治家兔实验性动脉粥样硬化的比较研究

目的:观察比较具有清除氧自由基作用的复方中药四逆汤与抗氧化剂维生素E抗血管内皮功能氧化损伤、防治家兔实验性动脉粥样硬化(AS)的作用。方法:运用喂饲高脂饲料造家兔实验性AS模型,随机分组处理,实验结束时,取主动脉及血样品,分析各组主动脉粥样斑块面积、脂代谢及血管内皮氧化损伤指标(SOD活性、MDA含量、NO水平、ET浓度)。结果:四逆汤中、高剂量组和维生素E组的主动脉内膜脂质斑块面积与内皮氧化损伤变化(除维生素E组的SOD活性外)均明显轻于模型组(P＜0.05),四逆汤组的脂代谢紊乱指标也较模型组明显改善(P均＜0.05)。结论:四逆汤抗血管内皮功能氧化损伤、防治AS的综合疗效较维生素E为优。