Liliya M. Yamaleyeva

Estrogen Receptor GPR30 Reduces Oxidative Stress and Proteinuria in the Salt-Sensitive Female mRen2.Lewis Rat

The current study assessed whether activation of the novel estrogen receptor GPR30 ameliorates salt-dependent renal damage in intact mRen2.Lewis (mRen2) females. Hemizygous mRen2 rats were maintained on either a normal salt (0.5% Na) or high-salt (HS; 4.0% Na) diet for 10 weeks (5 to 15 weeks of age), and HS animals were treated with the GPR30 agonist G-1 or vehicle for 2 weeks. Systolic blood pressure markedly increased with HS diet (149±3 to 219±5 mm Hg; P<0.01), but G-1 did not influence pressure (P=0.42). G-1 and estradiol induced relaxation of preconstricted mesenteric vessels from normal salt mRen2 rats, but both responses were attenuated in the HS group. Despite the lack of an effect on blood pressure, G-1 decreased renal hypertrophy, proteinuria, urinary 8-isoprostane excretion, and tubular 4-hydroxynonenal staining. HS diet significantly increased GPR30 mRNA (1.01±0.04 versus 1.59±0.13; P<0.01) and protein (0.60±0.31 versus 3.99±0.75; P<0.01) in the renal cortex. GPR30 was highly expressed in the brush border of proximal tubules and colocalized with megalin. Finally, megalin expression was reduced by HS diet and restored with G-1. We conclude that GPR30-mediated beneficial effects in salt-sensitive mRen2 females occurred independent of changes in systolic blood pressure. The failure of G-1 to influence pressure may reflect a salt-induced impairment in GPR30-mediated vasorelaxation. The renoprotective actions of GPR30 may involve attenuation of tubular oxidative stress and activation of megalin-mediated protein reabsorption.

Differential effects of sex steroids in young and aged female mRen2.Lewis rats: a model of estrogen and salt-sensitive hypertension.

BACKGROUND Male-female differences in the expression of hypertension and in end-organ damage are evident in both experimental models and human subjects, with males exhibiting a more rapid onset of cardiovascular disease and mortality than do females. The basis for these male-female differences is probably the balance of the complex effects of sex steroids (androgens, estrogen, progesterone) and their metabolites on the multiple regulatory systems that influence blood pressure (BP). A key target of estrogen and other steroids is likely to be the different components of the renin-angiotensin-aldosterone system (RAAS). OBJECTIVE The aim of this study was to review the current experimental evidence on the protective effects of estrogen in hypertensive models. METHODS The search terms estrogen , renin-aangiotensin-aldosterone system, renin receptor, salt-sensitivity, endorgan damage, hypertension, kidney, mRen2. Lewis, and injury markers were used to identify relevant publications in the PubMed database (restricted to the English language) from January 1990 to October 2007. RESULTS In a new congenic model that expresses the mouse renin 2 gene (mRen2. Lewis), estrogen depletion (via ovariectomy [OVX ]) in young rats was found to have a marked stimulatory effect on the progression of increased BP and cardiac dysfunction. Moreover, estrogen depletion exacerbated salt-sensitive hypertension and the extent of salt-induced cardiac and renal injury in young mRen2. Lewis rats, which probably reflected the inability to appropriately regulate various components of the RAAS. However, OVX in aged mRen2. Lewis rats conveyed renal protective effects from a high-salt diet compared with intact hypertensive littermates (64 weeks), and these effects were independent of changes in BP. CONCLUSION These studies in hypertensive mRen2. Lewis rats underscored the influence of ovarian hormones on BP and tissue injury, as well as the plasticity of this response, apparently due to age and salt status.

Differential regulation of circulating and renal ACE2 and ACE in hypertensive mRen2.Lewis rats with early-onset diabetes

We examined the impact of early diabetes on the circulating and kidney renin-angiotensin system (RAS) in male and female mRen2.Lewis (mRen2) hypertensive rats. Diabetes (DB) was induced by streptozotocin (STZ; 65 mg/kg) at 11 wk of age for 4 wk without insulin replacement. Systolic blood pressures were not increased in DB males or females compared with controls (CON). Circulating angiotensin-converting enzyme 2 (ACE2) increased ninefold (P < 0.05) in DB females and threefold (P < 0.05) in DB males, but circulating ACE and ANG II were higher in the DB groups. Serum C-reactive protein was elevated in DB females but not DB males, and the vascular responses to acetylcholine and estradiol were attenuated in the DB females. Proteinuria, albuminuria, and angiotensinogen excretion increased to a similar extent in both DB females and males. Glomerular VEGF expression also increased to a similar extent in both DB groups. Renal inflammation (CD68(+)cells) increased only in DB females although males exhibited greater inflammation that was not different with DB. Cortical ACE2 did not change in DB females but was reduced (30%) in DB males. Renal neprilysin activity (>75%, P < 0.05) was markedly reduced in the DB females to that in the DB and CON males. ACE activity was significantly lower in both female (75%, P < 0.05) and male (50%; P < 0.05) DB groups, while cortical ANG II and Ang-(1-7) levels were unchanged. In conclusion, female mRen2 rats are not protected from vascular damage, renal inflammation, and kidney injury in early STZ-induced diabetes despite a marked increase in circulating ACE2 and significantly reduced ACE within the kidney.

Angiotensin-Converting Enzyme 2 Deficiency Is Associated With Impaired Gestational Weight Gain and Fetal Growth Restriction

Angiotensin-converting enzyme 2 (ACE2) is a key enzyme of the renin-angiotensin system that influences the relative expression of angiotensin II (Ang II) and Ang-(1-7). Although ACE2 expression increases in normal pregnancy, the impact of ACE2 deficiency in pregnancy has not been elucidated. We determined the influence of ACE2 deficiency on circulating and tissue renin-angiotensin system components, fetal and maternal growth characteristics, and maternal hemodynamics (mean blood pressure and cardiac output) at day 18 of gestation. Gestational body weight gain was lower in the ACE2 knockout (KO) versus C57BL/6 (wild-type) mice (30.3±4.7 versus 38.2±1.0 g; P<0.001). Fetal weight (0.94±0.1 versus 1.24±0.01 g; P<0.01) and length (19.6±0.2 versus 22.2±0.2 mm; P<0.001) were less in KO. Mean blood pressure was significantly reduced in C57BL/6 with pregnancy; it was elevated (P<0.05) in the KO virgin and pregnant mice, and this was associated with an increased cardiac output in both C57BL/6 and KO pregnant mice (P<0.05). Plasma Ang-(1-7) was reduced in pregnant KO mice (P<0.05). Placenta Ang II levels were higher in KO mice (52.9±6.0 versus 22.0±3.3 fmol/mg of protein; P<0.001). Renal Ang II levels were greater in KO virgin mice (30.0±1.7 versus 23.7±1.1 fmol/mg of protein; P<0.001). There was no change in the Ang-(1-7) levels in the KO placenta and virgin kidney. These results suggest that ACE2 deficiency and associated elevated placenta Ang II levels impact pregnancy by impairing gestational weight gain and restricting fetal growth.

Uterine artery dysfunction in pregnant ACE2 knockout mice is associated with placental hypoxia and reduced umbilical blood flow velocity

Angiotensin-converting enzyme 2 (ACE2) knockout is associated with reduced fetal weight at late gestation; however, whether uteroplacental vascular and/or hemodynamic disturbances underlie this growth-restricted phenotype is unknown. Uterine artery reactivity and flow velocities, umbilical flow velocities, trophoblast invasion, and placental hypoxia were determined in ACE2 knockout (KO) and C57Bl/6 wild-type (WT) mice at day 14 of gestation. Although systolic blood pressure was higher in pregnant ACE2 KO vs. WT mice (102.3 ± 5.1 vs. 85.1 ± 1.9 mmHg, n = 5-6), the magnitude of difference was similar to that observed in nonpregnant ACE2 KO vs. WT mice. Maternal urinary protein excretion, serum creatinine, and kidney or heart weights were not different in ACE2 KO vs. WT. Fetal weight and pup-to-placental weight ratio were lower in ACE2 KO vs. WT mice. A higher sensitivity to Ang II [pD2 8.64 ± 0.04 vs. 8.5 ± 0.03 (-log EC50)] and greater maximal contraction to phenylephrine (169.0 ± 9.0 vs. 139.0 ± 7.0% KMAX), were associated with lower immunostaining for Ang II receptor 2 and fibrinoid content of the uterine artery in ACE2 KO mice. Uterine artery flow velocities and trophoblast invasion were similar between study groups. In contrast, umbilical artery peak systolic velocities (60.2 ± 4.5 vs. 75.1 ± 4.5 mm/s) and the resistance index measured using VEVO 2100 ultrasound were lower in the ACE2 KO vs. WT mice. Immunostaining for pimonidazole, a marker of hypoxia, and hypoxia-inducible factor-2α were higher in the trophospongium and placental labyrinth of the ACE2 KO vs. WT. In summary, placental hypoxia and uterine artery dysfunction develop before major growth of the fetus occurs and may explain the fetal growth restricted phenotype.

Increased Angiotensin II Contraction of the Uterine Artery at Early Gestation in a Transgenic Model of Hypertensive Pregnancy Is Reduced by Inhibition of Endocannabinoid Hydrolysis

Increased vascular sensitivity to angiotensin II (Ang II) is a marker of a hypertensive human pregnancy. Recent evidence of interactions between the renin–angiotensin system and the endocannabinoid system suggests that anandamide and 2-arachidonoylglycerol may modulate Ang II contraction. We hypothesized that these interactions may contribute to the enhanced vascular responses in hypertensive pregnancy. We studied Ang II contraction in isolated uterine artery (UA) at early gestation in a rat model that mimics many features of preeclampsia, the transgenic human angiotensinogen×human renin (TgA), and control Sprague–Dawley rats. We determined the role of the cannabinoid receptor 1 by blockade with SR171416A, and the contribution of anandamide and 2-arachidonoylglycerol degradation to Ang II contraction by inhibiting their hydrolyzing enzyme fatty acid amide hydrolase (with URB597) or monoacylglycerol lipase (with JZL184), respectively. TgA UA showed increased maximal contraction and sensitivity to Ang II that was inhibited by indomethacin. Fatty acid amide hydrolase blockade decreased Ang IIMAX in Sprague–Dawley UA, and decreased both Ang IIMAX and sensitivity in TgA UA. Monoacylglycerol lipase blockade had no effect on Sprague–Dawley UA and decreased Ang IIMAX and sensitivity in TgA UA. Blockade of the cannabinoid receptor 1 in TgA UA had no effect. Immunolocalization of fatty acid amide hydrolase and monoacylglycerol lipase showed a similar pattern between groups; fatty acid amide hydrolase predominantly localized in endothelium and monoacylglycerol lipase in smooth muscle cells. We demonstrated an increased Ang II contraction in TgA UA before initiation of the hypertensive phenotype. Anandamide and 2-arachidonoylglycerol reduced Ang II contraction in a cannabinoid receptor 1–independent manner. These renin–angiotensin system-endocannabinoid system interactions may contribute to the enhanced vascular reactivity in early stages of hypertensive pregnancy.

Apelin-13 in blood pressure regulation and cardiovascular disease.

Purpose of reviewDespite extensive pharmacological treatment, hypertension and heart failure still pose as high health and economic burden. Thus, novel therapeutic approaches are needed to promote more effective treatment of hypertension and cardiovascular disease. In this review we summarized recent evidence supporting the therapeutic potential of apelin-13, a recently discovered endogenous ligand for the G-protein coupled receptor APJ. Recent findingsSystemic administration of apelin-13 or its posttranslationally modified form, pyroglutamate apelin-13, exert vasodilatory and antihypertensive effects. Yet, central application of apelin increases blood pressure and its systemic effects may be compromised in the presence of endothelial dysfunction. In addition, positive inotropic effects by exogenous apelin in the normal and failing heart, as well as cardioprotective effects after myocardial infarction, strongly suggest its therapeutic potential in preventing and treating heart failure and consequences of myocardial ischemia. However, therapeutic use of apelin is limited primarily by its short half-life and parenteral administration, and significant effort has been directed to the development of novel agonists, delivery methods, and improving the efficacy of agonists at APJ. SummaryThe apelin/APJ axis may represent a new target for the development of novel therapeutic approaches for the treatment of hypertension and cardiovascular disease.

Amelioration of renal injury and oxidative stress by the nNOS inhibitor L-VNIO in the salt-sensitive mRen2.Lewis congenic rat.

Abstract: Salt sensitivity is a key risk factor for cardiovascular disease and renal injury. Alterations in renal nitric oxide may contribute to salt-dependent increases in blood pressure and tissue damage. Therefore, we assessed the expression of nitric oxide synthase (NOS) isoforms in the kidney and the effects of nNOS inhibition on renal injury, inflammation, and oxidative stress in the female mRen2.Lewis rat (mRen), a model of salt-sensitive hypertension. We find that a high-salt diet (4% sodium) significantly reduced endothelial NOS mRNA (2.6-fold) and protein (1.5-fold) but increased nNOS mRNA (2.4-fold) and protein (1.9-fold) in the renal cortex of these animals. Immunostaining for nNOS also seemed higher in macula densa and cortical tubules of the rats fed a high-salt diet. Circulating nitrate and nitrite levels were reduced, including the tissue levels of the NOS cofactor tetrahydrobiopterin. Cortical markers of oxidative stress (4HNE, 8-OH-deoxyguanosine) and fibrosis were increased; however, mRNA levels of the NAD(P)H oxidase components NOX4, p22phox, and p47phox were reduced. Chronic treatment with the nNOS inhibitor N5-(1-Imino-3-butenyl)-L-ornithine did not influence systolic blood pressure after 4 weeks but significantly attenuated albuminuria, renal fibrosis, inflammation, and indices of oxidative stress. We conclude that an increase in nNOS expression in conjunction with reduced levels of cortical tetrahydrobiopterin may stimulate oxidative stress and renal injury in the salt-sensitive female mRen2.Lewis rat.

Photoacoustic Imaging for the Detection of Hypoxia in the Rat Femoral Artery and Skeletal Muscle Microcirculation.

ABSTRACT Photoacoustic (PA) imaging is an emerging technology that combines structural and functional imaging of tissues using laser and ultrasound energy. We evaluated the ability of PA imaging system to measure real-time systemic and microvascular mean oxygen saturation (mSAO2) in a rat model of hypoxic shock. Male Sprague Dawley rats (n = 6) underwent femoral artery catherization and were subjected to acute hypoxia by lowering the fraction of inspired oxygen (FiO2) from 1.0 to 0.21, and then to 0.08. PA measurements of mSaO2 were taken in the femoral artery near the catheter tip using the Vevo 2100 LAZR at each FiO2 and compared to co-oximetry on blood removed from the femoral catheter. Both co-oximetry and PA imaging measured a similar stepwise decline in femoral artery mSaO2 as FiO2 was lowered. We also measured mSaO2 in the feed arteriole of the rat spinotrapezius muscle and adjacent microvessels (n = 6) using PA imaging. A significant decrease in mSaO2 in both the feed arteriole and adjacent microvessels was recorded as FiO2 was decreased from 1.0 to 0.08. Moreover, we detected a rapid return toward baseline mSaO2 in the feed arteriole and microvessels when FiO2 was increased from 0.08 to 1.0. Thus, PA imaging is noninvasive imaging modality that can accurately measure real-time oxygen saturation in the macro and microcirculation during acute hypoxia. This proof-of-concept study is a first step in establishing PA imaging as an investigational tool in critical illness.

Decidualized pseudopregnant rat uterus shows marked reduction in Ang II and Ang-(1-7) levels.

UNLABELLED Previous studies showed that angiotensin (Ang) II and Ang-(1-7) concentrations were reduced in the implantation site at day 7 of pregnancy in Sprague-Dawley rats as compared to the site immediately adjacent to it, which does not have the embryo attached, clearly showing the importance of the blastocyst in the regulation of renin-angiotensin system (RAS). OBJECTIVE The objective of this study was to evaluate the regulation of the RAS in the decidualized uterus in the pseudopregnant rat, a model without the presence of a conceptus. METHODS Ovariectomized, adult female rats were sensitized for the decidual cell reaction with steroid treatments; decidualization was induced by oil-injection of the right horn; the left horn served as a control. The uterine content of Ang I, Ang II, and Ang-(1-7) was examined in the decidualized and non-decidualized uteri. RESULTS Both Ang-(1-7) and Ang II and ACE and ACE2 mRNA were significantly reduced in the decidualized horn as compared to the non-decidualized horn. Immunocytochemical characterization of Ang II, Ang-(1-7), ACE and ACE2 demonstrated that Ang-(1-7), Ang II, and ACE2 polarize to the anti-mesometrial pole with decidualization. CONCLUSION The decidualization process elicits marked reduction in uterine Ang II and Ang-(1-7) content as compared to the non-decidualized horn. The differential immunocytochemical expression of Ang II and Ang-(1-7) with ACE2, but not ACE in the anti-mesometrial pole of the decidualized horn may favor the formation and action of Ang-(1-7) in the anti-mesometrial pole, an area which plays a role in triggering the decidualization process.