Pressure-independent effects of angiotensin II on hypertensive myocardial fibrosis

Angiotensin II (Ang II) is implicated in the proinflammatory process in various disease situations. Thus, we sought to determine the role of Ang II in early inflammation-induced fibrosis of pressure-overloaded (PO) hearts. PO was induced by suprarenal aortic constriction (AC) at day 0 in male Wistar rats, and they were orally administered 0.1 mg/kg per day candesartan every day from day -7. This was the maximum dose of candesartan that did not change arterial pressure in hypertensive rats with AC (AC rats). In AC rats, cardiac angiotensin-converting enzyme (ACE) activity was transiently enhanced after day 1 and peaked at day 3, declining to lower levels by day 14, whereas serum ACE activity was not changed. In AC rats, PO induced early fibroinflammatory changes (monocyte chemoattractant factor [MCP]-1 and transforming growth factor [TGF]-beta expression, perivascular macrophage accumulation, and fibroblast proliferation), and thereafter, left ventricular hypertrophy developed, featuring myocyte hypertrophy, intramyocardial arterial wall thickening, and perivascular and interstitial fibroses. Candesartan suppressed the induction of MCP-1 and TGF-beta and reduced macrophage accumulation and fibroblast proliferation in PO hearts. Candesartan significantly prevented perivascular and interstitial fibrosis. However, candesartan did not affect myocyte hypertrophy and arterial wall thickening. In conclusion, a subdepressor dose of candesartan prevented the MCP-1-mediated inflammatory process and reactive myocardial fibrosis in PO hearts. Ang II might play a key role in reactive fibrosis in hypertensive hearts, independent of arterial pressure changes.     

Diastolic dysfunction in hypertensive hearts: roles of perivascular inflammation and reactive myocardial fibrosis.

There is increasing evidence that myocardial fibrosis plays a role in the pathogenesis of diastolic dysfunction in hypertensive heart disease. However, it has been difficult to explore the mechanisms of isolated diastolic dysfunction in hypertensive hearts because of the lack of adequate animal models. Recently, we demonstrated that Wistar rats with a suprarenal aortic constriction (AC) can be used as a model of cardiac hypertrophy associated with preserved systolic, but impaired diastolic function without overt congestive heart failure. In this model, acute pressure elevation induces reactive myocardial fibrosis (perivascular fibrosis followed by intermuscular interstitial fibrosis) and myocyte/left ventricular (LV) hypertrophy. Perivascular macrophage infiltration, which is mediated by monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule-1, exerts a key role in myocardial fibrosis, but not in myocyte/LV hypertrophy. Transforming growth factor (TGF)-beta is crucial for reactive fibrosis in AC rats. MCP-1 function blocking not only inhibits macrophage infiltration and TGF-beta induction but also prevents reactive fibrosis and diastolic dysfunction, without affecting blood pressure, myocyte/LV hypertrophy, or systolic function. Accordingly, a substantial role of inflammation is indicated in myocardial fibrosis and diastolic dysfunction in hypertensive hearts. Currently, the precise mechanisms whereby acute pressure elevation triggers inflammation remain unknown, but it is likely that activation of the tissue angiotensin system is involved in the induction of the inflammatory process.     

抗氧化剂对心肌肥厚单核细胞趋化蛋白-1的影响

目的观察压力负荷大鼠心肌中单核细胞趋化蛋白-1(MCP-1)的表达变化,并探讨活性氧(ROS)与MCP-1的关系。方法应用腹主动脉缩窄法(AC)建立大鼠压力负荷模型,42只SD大鼠随机分为7组,分别为:假手术组;AC手术组;抗氧化剂N-乙酰半胱氨酸(NAC)2、4周组。NAC组为:AC+NAC(160mg/kg.d腹腔注射2周或4周)。各组均为6只SD大鼠。在观察期后颈动脉插管测大鼠血压及计算左心室质量分数(LVW/BW),采用分光光度计检测大鼠心肌ROS水平,用酶联免疫吸附测定(ELISA)法和逆转录聚合酶链式反应(RT-PCR)分别检测心肌组织MCP-1蛋白和mRNA的表达。结果(1)AC术后1~4周大鼠平均动脉压(MBP)明显高于假手术组(P<0.05)。(2)术后2~4周大鼠LVW/BW与假手术组大鼠比较,差异有统计学意义(P<0.05);术后1周组大鼠LVW/BW与假手术组比较,无显著性差异(P>0.05)。(3)术后1周心肌组织ROS表达量为(30.4±1.5)U/mgpro,2周为(42.3±1.3)U/mgpro,3、4周分别为(34.9±1.6)和(32.3±1.4)U/mgpro,各组与假手术组(17.3±1.8)U/mgpro比较,差异有非常显著意义(P<0.01)。NAC2周组ROS为(11.8±0.7)U/mgpro,明显低于AC2周组,差异有非常显著意义(P<0.01);NAC4周组ROS为(14.0±0.7)U/mgpro,与AC4周组比较,差异有非常显著意义(P<0.01)。(4)术后1~4周心肌组织MCP-1蛋白表达量分别为(67.0±4.5)、(84.8±5.9)、(43.1±3.1)、(36.2±3.2)pg/mgpro,各组与假手术组(12.7±1.3)pg/mgpro比较,差异有非常显著意义(P<0.01)。NAC2周组MCP-1表达量为(33.7±2.4)pg/mgpro,与AC2周组比较,差异有非常显著意义(P<0.01);NAC4周组为(19.4±2.8),低于AC4周组,差异有非常显著意义(P<0.01)。(5)术后1~4周MCP-1mRNA水平分别为(0.83±0.08)、(0.57±0.07)、(0.45±0.06)、(0.3±0.07),各组与假手术组(0.18±0.05)比较,差异有非常显著意义(P<0.01)。NAC2周组MCP-1 mRNA水平为(0.26±0.04),与AC2周组比较,差异有非常显著意义(P<0.01);NAC4周组为(0.21±0.04),与AC4周组比较,差异有非常显著意义(P<0.01)。结论在压力负荷下,心肌组织MCP-1表达增加,参与心室肥厚的发生发展,NAC阻断后MCP-1表达显著下调,表明其机制可能与ROS的诱导作用有关。     

Hypertensive myocardial fibrosis and diastolic dysfunction: another model of inflammation?

Excessive myocardial fibrosis deteriorates diastolic function in hypertensive hearts. Involvement of macrophages is suggested in fibrotic process in various diseased situations. We sought to examine the role of macrophages in myocardial remodeling and cardiac dysfunction in pressure-overloaded hearts. In Wistar rats with suprarenal aortic constriction, pressure overload induced perivascular macrophage accumulation and fibroblast proliferation with a peak at day 3, decreasing to lower levels by day 28. Myocyte chemoattractant protein (MCP)-1 mRNA was upregulated after day 1, peaking at day 3 and returning to insignificant levels by day 28, whereas transforming growth factor (TGF)-beta induction was observed after day 3, with a peak at day 7, and remained relatively elevated at day 28. After day 7, concentric left ventricular (LV) hypertrophy developed, associated with reactive fibrosis and myocyte hypertrophy. At day 28, echocardiography showed normal LV fractional shortening but decreased ratio of early to late filling wave of transmitral Doppler velocity, and hemodynamic studies revealed elevated LV end-diastolic pressure, suggesting normal systolic but impaired diastolic function. Chronic treatment with an anti-MCP-1 monoclonal neutralizing antibody inhibited not only macrophage accumulation but also fibroblast proliferation and TGF-beta induction. Furthermore, the neutralizing antibody attenuated myocardial fibrosis, but not myocyte hypertrophy, and ameliorated diastolic dysfunction without affecting blood pressure and systolic function. In conclusion, roles of MCP-1-mediated macrophage accumulation are suggested in myocardial fibrosis in pressure-overloaded hearts through TGF-beta-mediated process. Inhibition of inflammation may be a new strategy to prevent myocardial fibrosis and resultant diastolic dysfunction in hypertensive hearts.     

Tranilast attenuates myocardial fibrosis in association with suppression of monocyte/macrophage infiltration in DOCA/salt hypertensive rats

OBJECTIVE: In order to study the association between myocardial fibrosis and inflammatory cell infiltration in the hypertensive heart, we investigated whether N(3,4-dimethoxycinnamoyl) anthranilic acid (tranilast), an anti-inflammatory drug, would suppress myocardial fibrosis via inhibition of inflammatory cell infiltration in deoxycorticosterone-acetate (DOCA) hypertensive rats. METHODS: Sprague-Dawley rats treated with DOCA combined with the addition of 1% NaCl and 0.2% KCl in the drinking water after left nephrectomy were given tranilast (100 mg/kg per day, n = 15) or vehicle (n = 15) for up to 4 weeks. Systolic blood pressure (SBP), amount of myocardial interstitial fibrosis, perivascular fibrosis and type I and III collagen, and mRNA expression of procollagen I (PI) and procollagen III (PIII), transforming growth factor (TGF)-beta1, type-1 plasminogen activator inhibitor (PAI-1), monocyte chemoattractant protein (MCP)-1 and interleukin (IL)-6 were determined. RESULTS: SBP was increased significantly 2 weeks after treatment with DOCA and salt. Myocardial interstitial fibrosis, perivascular fibrosis and collagen accumulation increased significantly 4 weeks after the treatment. Two weeks after the treatment with DOCA and salt, mRNA expression of PI and PIII, TGF-beta1, PAI-1, MCP-1 and IL-6 increased significantly. Although the SBP was similar in animals treated with tranilast or vehicle, monocyte/macrophage infiltration was suppressed, mRNA expression of TGF-beta1, PAI-1, MCP-1, IL-6, PI and PIII was attenuated, and myocardial fibrosis and collagen accumulation were suppressed in hypertensive animals receiving tranilast. CONCLUSION: Myocardial fibrosis seen in DOCA/salt hypertensive rats might be associated with the inflammation/wound healing response. Tranilast suppresses both infiltration of monocytes/macrophages and myocardial fibrosis.     

Inducible nitric oxide synthase deficiency protects the heart from systolic overload-induced ventricular hypertrophy and congestive heart failure

Inducible nitric oxide synthase (iNOS) protein is expressed in cardiac myocytes of patients and experimental animals with congestive heart failure (CHF). Here we show that iNOS expression plays a role in pressure overload-induced myocardial chamber dilation and hypertrophy. In wild-type mice, chronic transverse aortic constriction (TAC) resulted in myocardial iNOS expression, cardiac hypertrophy, ventricular dilation and dysfunction, and fibrosis, whereas iNOS-deficient mice displayed much less hypertrophy, dilation, fibrosis, and dysfunction. Consistent with these findings, TAC resulted in marked increases of myocardial atrial natriuretic peptide 4-hydroxy-2-nonenal (a marker of lipid peroxidation) and nitrotyrosine (a marker for peroxynitrite) in wild-type mice but not in iNOS-deficient mice. In response to TAC, myocardial endothelial NO synthase and iNOS was expressed as both monomer and dimer in wild-type mice, and this was associated with increased reactive oxygen species production, suggesting that iNOS monomer was a source for the increased oxidative stress. Moreover, systolic overload-induced Akt, mammalian target of rapamycin, and ribosomal protein S6 activation was significantly attenuated in iNOS-deficient mice. Furthermore, selective iNOS inhibition with 1400W (6 mg/kg per hour) significantly attenuated TAC induced myocardial hypertrophy and pulmonary congestion. These data implicate iNOS in the maladaptative response to systolic overload and suggest that selective iNOS inhibition or attenuation of iNOS monomer content might be effective for treatment of systolic overload-induced cardiac dysfunction.     

Salt excess causes left ventricular diastolic dysfunction in rats with metabolic disorder

Metabolic syndrome is a highly predisposing condition for cardiovascular disease and could be a cause of excess salt-induced organ damage. Recently, several investigators have demonstrated that salt loading causes left ventricular diastolic dysfunction associated with increased oxidative stress and mineralocorticoid receptor activation. We, therefore, investigated whether excess salt induces cardiac diastolic dysfunction in metabolic syndrome via increased oxidative stress and upregulation of mineralocorticoid receptor signals. Thirteen-week-old spontaneously hypertensive rats and SHR/NDmcr-cps, the genetic model of metabolic syndrome, were fed a normal salt (0.5% NaCl) or high-salt (8% NaCl) diet for 4 weeks. In SHR/NDmcr-cps, salt loading induced severe hypertension, abnormal left ventricular relaxation, and perivascular fibrosis. Salt-loaded SHR/NDmcr-cps also exhibited overproduction of reactive oxygen species and upregulation of mineralocorticoid receptor-dependent gene expression, such as Na(+)/H(+) exchanger-1 and serum- and glucocorticoid-inducible kinase-1 in the cardiac tissue. However, in spontaneously hypertensive rats, salt loading did not cause these cardiac abnormalities despite a similar increase in blood pressure. An antioxidant, tempol, prevented salt-induced diastolic dysfunction, perivascular fibrosis, and upregulation of mineralocorticoid receptor signals in SHR/NDmcr-cps. Moreover, a selective mineralocorticoid receptor antagonist, eplerenone, prevented not only diastolic dysfunction but also overproduction of reactive oxygen species in salt-loaded SHR/NDmcr-cps. These results suggest that metabolic syndrome is a predisposed condition for salt-induced left ventricular diastolic dysfunction, possibly via increased oxidative stress and enhanced mineralocorticoid receptor signals.     

The antioxidant edaravone attenuates pressure overload-induced left ventricular hypertrophy

The free radical scavenger 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone) is used to treat patients with ischemic brain damage. We and others reported previously that in vitro and in vivo reactive oxygen species (ROS) act as second messengers to develop cardiac hypertrophy. In this study, we used an in vivo murine model of pressure overload-induced cardiac hypertrophy to examine the effects of edaravone on left ventricular hypertrophy. The animals were subjected to the transverse thoracic aorta constriction, and edaravone (10 mg/kg) was infused intraperitoneally twice daily. Seven days after the operation, we observed a significant increase in ROS production in hearts, which was eliminated by the treatment with edaravone. Pressure-overloaded hearts showed a significant increase in left ventricular weight/body weight ratio and the expression level of atrial natriuretic factor mRNA, which were attenuated by edaravone. It also reduced perivascular and intermuscular fibrosis and inhibited pressure overload-induced activation of apoptosis signal-regulating kinase 1 (ASK1) and its downstream kinases of c-Jun N-terminal protein kinase and p38 mitogen-activated protein kinase. Edaravone attenuated the hypertrophic response even when the treatment was started after the onset of cardiac hypertrophic response. These findings indicate that edaravone significantly attenuates pressure overload-induced cardiac hypertrophy mediated through its antioxidative function and subsequent inhibition of ASK1 signaling pathway.     

Pioglitazone, a thiazolidinedione derivative, attenuates left ventricular hypertrophy and fibrosis in salt-sensitive hypertension.

Thiazolidinediones, which stimulate peroxisome proliferator-activated receptor gamma, have been shown to prevent cardiovascular injury. However, little is known about their effects on salt-sensitive hypertension. We thus investigated whether or not pioglitazone affects left ventricular (LV) hypertrophy in Dahl salt-sensitive rats, then compared its effects to those of an angiotensin II receptor blocker, candesartan. Rats were used at 16 weeks of age after they had been fed either a low-salt (0.3%; DSL) or high-salt (8%; DSH) diet for 10 weeks; some of the DSH rats were treated with pioglitazone (10 mg/kg/day) or candesartan (4 mg/kg/day). Both drugs decreased the elevated blood pressure in DSH rats, although it was still higher than in DSL rats. Both drugs decreased plasma insulin levels, but neither affected plasma glucose levels. The thiobarbituric acid reactive substance level in the LV was decreased by both drugs. LV hypertrophy evaluated by echocardiography in DSH rats was nearly normalized by both drugs, whereas only candesartan decreased LV diameter. In histological analysis, both drugs ameliorated LV fibrosis and myocardial cell hypertrophy. Both drugs decreased elevated gene expression levels of transforming growth factor-beta1 and collagen type I, although the pioglitazone action was slightly modest. The metalloproteinase activity was increased in DSH rats, but both drugs decreased this level. Taken together, these findings indicate that pioglitazone reduced LV hypertrophy and fibrosis in salt-sensitive hypertension. Improvement in blood pressure, insulin level, and oxidative stress may be associated with this beneficial action of pioglitazone.     

AT1 receptor blocker added to ACE inhibitor provides benefits at advanced stage of hypertensive diastolic heart failure

Diastolic heart failure (DHF) has become a social burden; however, evidences leading to its therapeutic strategy are lacking. This study investigated effects of addition of angiotensin II type 1 receptor blocker (ARB) to angiotensin-converting enzyme inhibitor (ACEI) at advanced stage of DHF in hypertensive rats. Dahl salt-sensitive rats fed 8% NaCl diet from age 7 weeks served as DHF model, and those fed a normal chow served as control. The DHF model rats were arbitrarily assigned to 3 treatment regimens at age 17 weeks: ACEI (temocapril 0.4 mg/kg per day), combination of ACEI (temocapril 0.2 mg/kg per day) with ARB (olmesartan 0.3 mg/kg per day), or placebo. At age 17 weeks, this model represents progressive ventricular hypertrophy and fibrosis, relaxation abnormality, and myocardial stiffening. Data were collected at age 20 weeks. As compared with the monotherapy with ACEI, the addition of ARB induced more prominent suppression of ventricular hypertrophy and fibrosis, leading to suppression of myocardial stiffening, improvement of relaxation, and inhibition of hemodynamic deterioration. Such benefits were associated with greater decreases in reactive oxygen species (ROS) generation, macrophage infiltration, and gene expression of transforming growth factor (TGF)-beta(1) and interleukin (IL)-1beta, but not with changes in gene expression of monocyte chemoattractant protein (MCP)-1 and tumor necrosis factor (TNF)-alpha. Thus, ARB added to ACEI provides more benefits as compared with ACEI alone in DHF when initiated at an advanced stage. The additive effects are likely provided through more prominent suppression of ROS generation and inflammatory changes without effects on expression of MCP-1 and TNF-alpha.     

Therapeutic effects of angiotensin II type 1 receptor blocker at an advanced stage of hypertensive diastolic heart failure

OBJECTIVE: Angiotensin II type 1 receptor blocker (ARB) is increasingly prescribed for the treatment of systolic heart failure with a growing body of clinical evidence. The roles of ARB, however, remain to be clarified in the treatment of diastolic heart failure (DHF), particularly at its advanced stage. This experimental study investigated the effects of ARB administered at an advanced stage of hypertensive DHF. METHODS: Dahl salt-sensitive rats fed an 8% NaCl diet from age 7 weeks represent overt DHF at age 20 weeks, as noted in previous studies (hypertensive DHF model). The DHF model rats were randomly divided into two groups at age 17 weeks when left ventricular diastolic dysfunction, hypertrophy, fibrosis, macrophage infiltration and reactive oxygen species generation were already augmented; six rats treated for 3 weeks with a subdepressor dose of ARB (olmesartan 0.6 mg/kg per day), and six untreated rats. RESULTS: The 3-week administration of ARB significantly decreased the left ventricular end-diastolic pressure in association with attenuation of left ventricular hypertrophy, fibrosis and diastolic dysfunction. Macrophage infiltration was attenuated with decreased gene expression of transforming growth factor-beta1 and monocyte chemoattractant protein-1 in the left ventricular myocardium of the ARB-treated rats. The production of reactive oxygen species also decreased with NADPH oxidase activity. CONCLUSIONS: ARB provides beneficial effects in hypertensive DHF independent of its antihypertensive effects even if initiated at an advanced stage. The beneficial effects are at least partly attributed to the attenuation of inflammatory changes and oxidative stress through the suppression of cytokine and chemokine production and of NADPH oxidase activity.     

Cardioprotective Action of Perindopril versus Candesartan in Renovascular Hypertensive Rats

We investigated the effects of an angiotensin-converting enzyme inhibitor and an angiotensin II type 1 receptor blocker on cardiac hypertrophy in rats with renovascular hypertension. Renovascular hypertensive (Goldblatt) rats were surgically prepared from Wistar rats. Four weeks later, the rats showed a significant increase in blood pressure. At high doses, both the perindopril (1 mg/kg/day) and the candesartan (2 mg/kg/day) decreased the systolic pressure in these rats to the level of control Wistar rats. At low doses (perindopril 0.1 mg/kg/day and candesartan 0.1 mg/kg/day), these drugs lowered blood pressure to 85% of that in hypertensive rats. Echocardiographic and morphological studies revealed severe cardiac hypertrophy and fibrosis in untreated Goldblatt rats. High-dose treatment with both drugs suppressed the progression of hypertrophy and fibrosis. Also, low-dose perindopril prevented cardiac hypertrophy and fibrosis. In contrast, at the same levels of blood-pressure reduction, low-dose candesartan did not prevent cardiac fibrosis nor the upregulation of cardiac collagen types I and III mRNA observed in untreated Goldblatt rats. Atrial natriuretic peptide mRNA was up-regulated in untreated Goldblatt rats. These changes were significantly decreased by both doses of perindopril or the high dose of candesartan. Serum levels of angiotensin II and aldosterone were significantly higher in untreated Goldblatt rats. Both doses of perindopril inhibited activation of the renin-angiotensin system, whereas candesartan had weaker effects. In particular, serum aldosterone was 347 ± 20 pg/ml in low-dose perindopril versus 1796 ± 324 pg/ml in low-dose candesartan. These results suggest that there were no differences between the cardioprotective actions of perindopril and candesartan at high dosages. On the other hand, low-dose treatment with perindopril was more effective in preventing cardiac fibrosis than was low-dose treatment with candesartan, despite similar changes in blood pressure. It is possible that changes in aldosterone secretion are related to this difference.     

坎地沙坦对肝纤维化大鼠Ang-(1-7)的影响

目的研究血管紧张素Ⅱ1型受体拮抗剂坎地沙坦抗大鼠肝纤维化的疗效及对血管紧张素1-7[Ang-(1-7)]的影响。方法制备四氯化碳诱导大鼠肝纤维化模型,同时应用坎地沙坦灌胃,共8周。肝组织进行常规HE与Masson三色染色,并测定门脉压及血清肝功能,血浆中Ang-(1-7)应用酶联免疫方法检测。结果与模型组大鼠比较,坎地沙坦可改善肝纤维化程度及降低门脉压,血浆Ang-(1-7)水平增加。结论坎地沙坦抗肝纤维化可能与升高Ang-(1-7)有关。     

Increased expression of gp91phox homologues of NAD(P)H oxidase in the aortic media during chronic hypertension: involvement of the renin-angiotensin system.

Although vascular cells express multiple members of the Nox family of nicotinamide adenine dinucleotide phosphate (NAD(P)H) oxidase, including gp91phox, Nox1, and Nox4, the reasons for the different expressions and specific roles of these members in vascular injury in chronic hypertension have remained unclear. Thus, we quantified the mRNA expressions of these NAD(P)H oxidase components by real-time polymerase chain reaction and evaluated superoxide production and morphological changes in the aortas of 32-week-old stroke-prone spontaneously hypertensive rats (SHRSP) and age-matched Wistar Kyoto rats (WKY). The aortic media of SHRSP had an approximately 2.5-fold greater level of Nox4 mRNA and an approximately 10-fold greater level of Nox1 mRNA than WKY. The mRNA expressions of gp91phox and p22phox in SHRSP and WKY were comparable. SHRSP were treated from 24 weeks of age for 8 weeks with either high or low doses of candesartan (4 mg/kg/day or 0.2 mg/kg/day), or a combination of hydralazine (30 mg/kg/day) and hydrochlorothiazide (4.5 mg/kg/day). The high-dose candesartan or the hydralazine plus hydrochlorothiazide decreased the blood pressure of SHRSP to that of WKY, whereas the low-dose candesartan exerted no significant antihypertensive action. Media thickening and fibrosis, as well as the increased production of superoxide in SHRSP, were nearly normalized with high-dose candesartan and partially corrected with low-dose candesartan or hydralazine plus hydrochlorothiazide. These changes by antihypertensive treatment paralleled the decrease in mRNA expression of Nox4 and Nox1. These results suggest that blood pressure and angiotensin II type 1 receptor activation are involved in the up-regulation of Nox1 and Nox4 expression, which could contribute to vascular injury during chronic hypertension.     

Differential effects of AT1 receptor and Ca2+ channel blockade on atherosclerosis, inflammatory gene expression, and production of reactive oxygen species

Angiotensin II receptor blockade has been shown to inhibit atherosclerosis in several different animal models. We sought to determine if this effect was the result of blood pressure reduction per se or a result of the anti-inflammatory effects of receptor blockade. ApoE-deficient mice were fed a high fat diet and treated with either an angiotensin II receptor antagonist, candesartan (0.5 mg/kg/day, s.c.) or a calcium channel blocker, amlodipine (7.5 mg/kg/day, mixed with food). Atherosclerotic lesion area, aortic inflammatory gene expression as well as aortic H2O2 and superoxide production were assayed. We found that candesartan but not amlodipine treatment dramatically attenuated the development of atherosclerosis despite a similar reduction in blood pressure. Similarly, candesartan treatment inhibited aortic expression of inflammatory genes and production of reactive oxygen species, effects not seen with amlodipine. These data demonstrate that angiotensin II receptor blockade inhibits atherosclerosis by reducing vascular oxidative stress and inflammatory gene production independent of blood pressure reduction.     

Beneficial effects of pioglitazone on hypertensive cardiovascular injury are enhanced by combination with candesartan

The effect of pioglitazone, a peroxisome proliferator-activated receptor gamma agonist, on hypertensive cardiovascular injury is unknown. We examined the effect of pioglitazone on hypertensive cardiovascular injury and the significance of combination of pioglitazone with angiotensin type 1 receptor blocker. Stroke-prone spontaneously hypertensive rats (SHRSP) were orally given pioglitazone, candesartan, or combined pioglitazone and candesartan for 4 weeks to compare their effects on cardiovascular injury. Pioglitazone, without lowering blood pressure, significantly suppressed cardiac inflammation and fibrosis and reduced vascular endothelial dysfunction, and these beneficial effects were associated with the reduction of superoxide by inhibition of cardiovascular NADPH oxidase. Thus, pioglitazone protects against hypertensive cardiovascular injury, by inhibiting reactive oxygen species (ROS). Combination of pioglitazone and candesartan suppressed cardiac hypertrophy, inflammation, and interstitial fibrosis of SHRSP to a greater extent than either monotherapy, and reduced vascular endothelial dysfunction of SHRSP more than either monotherapy. Furthermore, more beneficial effects of their combination on cardiovascular injury were associated with more reduction of NADPH oxidase-mediated cardiovascular ROS. To elucidate the underlying molecular mechanism, we examined cardiovascular NADPH oxidase subunits. Pioglitazone monotherapy significantly attenuated cardiovascular p22(phox) and Rac1 in SHRSP, whereas pioglitazone combined with candesartan more attenuated p22(phox) and significantly reduced Nox1. Thus, additive suppression of cardiovascular NADPH oxidase by the combination was attributed to its additive attenuation of p22(phox) and Nox1 protein levels. In conclusion, we showed that pioglitazone protected against hypertensive cardiovascular damage, and the combination of pioglitazone and candesartan exerted more beneficial effects on hypertensive cardiovascular injury by more suppressing ROS.     

Role of xanthine oxidoreductase in the reversal of diastolic heart failure by candesartan in the salt-sensitive hypertensive rat

The role of angiotensin II and reactive oxygen species in the exacerbation of diastolic heart failure is unknown. We examined the therapeutic effect of angiotensin blockade on hypertensive diastolic heart failure, focusing on the role of xanthine oxidoreductase and reduced nicotinamide-adenine dinucleotide phosphate oxidase, major enzymes producing reactive oxygen species. Dahl salt-sensitive hypertensive rats (DS rats) with established diastolic heart failure were given vehicle, candesartan (an angiotensin II receptor subtype 1 receptor blocker), oxypurinol (a xanthine oxidoreductase inhibitor), apocynin (a reduced nicotinamide-adenine dinucleotide phosphate oxidase inhibitor), or hydralazine (a vasodilator), and their therapeutic effects on diastolic heart failure were compared. Candesartan treatment of DS rats with established diastolic heart failure reversed cardiac remodeling, improved cardiac relaxation abnormality, and prolonged survival, being accompanied by the attenuation of the increase in cardiac superoxide, reduced nicotinamide-adenine dinucleotide phosphate oxidase, and xanthine oxidoreductase activities. Thus, the beneficial effect of candesartan in DS rats appears to be mediated by the inhibition of cardiac reactive oxygen species. Cardiac xanthine oxidoreductase inhibition with oxypurinol significantly reduced cardiac superoxide, prevented the progression of cardiac remodeling, and delayed the mortality in DS rats. Apocynin, which significantly inhibited cardiac reduced nicotinamide-adenine dinucleotide phosphate oxidase activity, prevented the exacerbation of diastolic heart failure more than hydralazine. However, compared with candesartan or oxypurinol, apocynin did not improve cardiac reactive oxygen species, remodeling, and function in DS rats. In conclusion, candesartan slowed the exacerbation of hypertensive diastolic heart failure in DS rats by causing reverse cardiac remodeling. Cardiac xanthine oxidoreductase contributed to these beneficial effects of candesartan.     

肾血管性高血压大鼠心肌纤维化发展过程的实验研究

采用二肾一夹肾血管性高血压（２Ｋ１Ｃ－ＲＨＴ）大鼠为模型，观察在反应性和修复性纤维化阶段心肌胶原形态和生化的改变。方法建立２Ｋ１Ｃ－ＲＨＴ模型，将大鼠随机分为４组：术后４，１２周高血压组和对照组。苦味酸天狼星红染色观察胶原形态，图像分析系统测量胶原容积分数（ＣＶＦ）和血管周围胶原面积（ＰＶＣＡ），羟脯氨酸法测定心肌胶原浓度（Ｃｏｌ）。结果在反应性纤维化阶段，心肌胶原从血管周围向邻近间质短距离延伸，同时ＣＶＦ、ＰＶＣＡ和Ｃｏｌ显著增高；在修复性纤维化阶段，胶原在间质中长距离延伸，出现取代缺失心肌细胞的疤痕灶，同时ＣＶＦ、ＰＶＣＡ和Ｃｏｌ进一步升高。结论提示２Ｋ１Ｃ－ＲＨＴ大鼠心肌纤维化呈进行性发展。     

Angiotensin II receptor activation in youth triggers persistent insulin resistance and hypertension--a legacy effect?

Although the involvement of angiotensin II (Ang II) in insulin resistance and hypertension has been established, the temporal relationships between Ang II receptor activation and changes in insulin sensitivity and blood pressure are not clear. To better understand this issue, we infused rats with Ang II (200?ng?kg(-1)?min(-1)) or vehicle for 4 weeks and assessed the residual effects after the discontinuation of the infusion on blood pressure, insulin sensitivity and tissue parameters of inflammation. Four weeks after the discontinuation of the Ang II infusion, the blood pressure was higher by 12.8?mm?Hg, and insulin sensitivity as determined by a euglycemic hyperinsulinemic glucose clamp was reduced (glucose infusion rate: 11.1±0.7 vs. 17.6±0.5?mg?kg(-1)?min(-1)) in the Ang II-treated group compared with controls. The persistent hypertension and insulin resistance were associated with greater than two-fold increases in macrophage chemoattractant protein-1, tumor necrosis factor-α and thiobarbituric acid-reactive substrates in the soleus muscle. Furthermore, total and activated forms of Rac-1, a regulatory subunit of the NADPH oxidase complex, were increased by 144±14% and 277±82%, respectively, in the skeletal muscle of Ang II-treated rats. These residual effects after Ang II infusion were all attenuated by the co-administration of tempol, a free radical scavenger, or candesartan with Ang II. The effects of candesartan were not mimicked by hydralazine at an equidepressant dose. These findings suggest that Ang II receptor activation in youth triggers the upregulation of inflammatory cytokines and the production of reactive oxygen species, thereby inducing later insulin resistance and hypertension.