Renal Rats Research Articles

Castration Has Antihypertensive and Organoprotective Effects in Male but Not in Female Heterozygous Ren-2 Rats

Gender differences in the regulation of blood pressure (BP) and the contributions of the renin-angiotensin system remain controversial. We assessed the effect of castration on BP and organ damage, as well as angiotensin II (ANG II), estradiol and testosterone levels in heterozygous Ren-2 transgenic (TGR) rats and transgene-negative Hannover Sprague-Dawley control rats. Male TGR had severe hypertension throughout the experiment, while the BP of female TGR declined after 5 months to normotensive levels. Ovariectomy had no effect on BP, cardiac hypertrophy or proteinuria in female Ren-2 TGR. On the contrary, BP and cardiac hypertrophy were significantly reduced in castrated male TGR as compared to sham-operated TGR. Moreover, proteinuria in these animals was normalized to the levels of control rats. ANG II levels did not differ between male and female TGR, and no effect of castration on plasma and tissue ANG II levels was found either in male or female TGR at the end of the experiment. In conclusion, the contribution of the renin-angiotensin system to the gender difference in BP homeostasis seems to be negligible in aging heterozygous Ren-2 transgenic rats.

Comparative analysis of telmisartan and olmesartan on cardiac function in the transgenic (mRen2)27 rat

Telmisartan, an angiotensin receptor blocker, may have unique benefits as it possesses partial peroxisome proliferator-activated receptor (PPAR)-γ agonist activity in addition to antihypertensive effects. In this study, we test whether treatment with telmisartan ameliorates cardiovascular abnormalities to a greater extent than olmesartan, which has little PPAR-γ activity. The hypertensive rodent model of tissue renin-angiotensin system activation, transgenic (mRen2)27 (Ren2) rats and their littermate Sprague-Dawley controls were used. Rats were treated with telmisartan (2 mg · kg(-1) · day(-1)), olmesartan (2.5 mg · kg(-1) · day(-1)), or vehicle via drinking water for 3 wk; these doses achieved similar blood pressure control, as measured by telemetry. Ren2 rats displayed impaired diastolic and systolic function using left ventricular (LV) pressure-volume (P-V) analysis. Load-independent diastolic indexes, including the time constant of isovolumic relaxation and the slope of the end-diastolic P-V relationship, as well as systolic indexes, including preload recruitable stroke work, the dP/dt(max)-end-diastolic volume (EDV) relationship, and the P-V area-EDV relationship, were elevated in Ren2 rats compared with Sprague-Dawley controls (P < 0.05). The Ren2 myocardium exhibited parallel increases in the oxidant markers NADPH oxidase and 3-nitrotyrosine. The increase in the prohypertrophic protein Jak2 in Ren2 rats was associated with cardiac structural abnormalities using light microscopic and ultrastructural analysis, which included interstitial fibrosis, cardiomyocyte and LV hypertrophy, and mitochondrial derangements. Both angiotensin receptor blockers attenuate these abnormalities to a similar extent. Our data suggest that the beneficial effect of telmisartan and olmesartan on cardiac structure and function may be predominantly pressor-related or angiotensin type 1 receptor dependent in this model of renin-angiotensin system activation.

Candesartan Attenuates Diabetic Retinal Vascular Pathology by Restoring Glyoxalase-I Function

OBJECTIVEAdvanced glycation end products (AGEs) and the renin-angiotensin system (RAS) are both implicated in the development of diabetic retinopathy. How these pathways interact to promote retinal vasculopathy is not fully understood. Glyoxalase-I (GLO-I) is an enzyme critical for the detoxification of AGEs and retinal vascular cell survival. We hypothesized that, in retina, angiotensin II (Ang II) downregulates GLO-I, which leads to an increase in methylglyoxal-AGE formation. The angiotensin type 1 receptor blocker, candesartan, rectifies this imbalance and protects against retinal vasculopathy.RESEARCH DESIGN AND METHODSCultured bovine retinal endothelial cells (BREC) and bovine retinal pericytes (BRP) were incubated with Ang II (100 nmol/l) or Ang II+candesartan (1 μmol/l). Transgenic Ren-2 rats that overexpress the RAS were randomized to be nondiabetic, diabetic, or diabetic+candesartan (5 mg/kg/day) and studied over 20 weeks. Comparisons were made with diabetic Sprague-Dawley rats.RESULTSIn BREC and BRP, Ang II induced apoptosis and reduced GLO-I activity and mRNA, with a concomitant increase in nitric oxide (NO•), the latter being a known negative regulator of GLO-I in BRP. In BREC and BRP, candesartan restored GLO-I and reduced NO•. Similar events occurred in vivo, with the elevated RAS of the diabetic Ren-2 rat, but not the diabetic Sprague-Dawley rat, reducing retinal GLO-I. In diabetic Ren-2 rats, candesartan reduced retinal acellular capillaries, inflammation, and inducible nitric oxide synthase and NO•, and restored GLO-I.CONCLUSIONSWe have identified a novel mechanism by which candesartan improves diabetic retinopathy through the restoration of GLO-I.

Reactive Oxygen Species, NADPH Oxidases, and Hypertension

Reactive oxygen species (ROS) produced in the neuronal, renal, and vascular systems not only influence cardiovascular physiology but are also strongly implicated in pathological signaling leading to hypertension. Different sources of ROS have been identified, ranging from xanthine-xanthine oxidase and mitochondria to NADPH oxidase (Nox) enzymes. Of 7 Nox family members, Nox1, Nox2, and Nox4 (and Nox5 in humans) influence the cardiovascular system. Their activation processes and cell and tissue distribution vary widely, adding complexity to understanding their functional roles. Whether these systems act collectively or independently in disease conditions is unclear, but recently feed forward mechanisms have been established between ROS sources. Studies published in Hypertension over the last few years are the focus of this review, and they provide a framework with which to consider the roles of Nox enzymes in neuronal, renal, and vascular hypertensive mechanisms, as well as cardiac remodeling, and their relationships with other ROS-generating systems. ### Neuronal ROS in Hypertension Redox signaling in the central nervous system is well recognized in neuronal control of blood pressure (BP), as well as in response to angiotensin II (Ang II) and aldosterone, which are linked to ROS-dependent hypertension. Recently, new roles for ROS have been described in the hypothalamus and brain stem, nucleus tractus solitarius (NTS), subfornical organ (SFO), rostral ventrolateral medulla, and area postrema (Figure 1). Figure 1. Neuronal NADPH oxidase–dependent ROS involved in central regulation of hypertension. NADPH oxidase homologues, mainly Nox2 and Nox4, are found in different regions of the neuronal system and are reported to have a role in the neuropathogenesis of hypertension by enhancing the sympathetic nerve activity. Nox-induced ROS initiate a forward loop in cross-activation of different receptors and between Nox and mitochondrial ROS. OVLT indicates organum vasculosum of the lamina terminalis; PVN, paraventricular nucleus; PP, posterior pituitary; AP, area postrema; RVLM, rostral ventrolateral medulla. Several …

Tranilast attenuates the up-regulation of thioredoxin-interacting protein and oxidative stress in an experimental model of diabetic nephropathy

Diabetic nephropathy is the leading cause of kidney failure in the developed world. Tranilast has been reported to not only act as an anti-inflammatory and anti-fibrotic compound, but it also exerts anti-oxidative stress effects in diabetic nephropathy. Thioredoxin-interacting protein (Txnip) is the endogenous inhibitor of the anti-oxidant thioredoxin and is highly up-regulated in diabetic nephropathy, leading to oxidative stress and fibrosis. In this study, we aimed to investigate whether tranilast exerts its anti-oxidant properties through the inhibition of Txnip. Heterozygous Ren-2 rats were rendered diabetic with streptozotocin. Another group of rats were injected with citrate buffer alone and treated as non-diabetic controls. After 6 weeks of diabetes, diabetic rats were divided into two groups: one group gavaged with tranilast at 200 mg/kg/day and another group with vehicle. Diabetic rats had a significant increase in albuminuria, tubulointerstitial fibrosis, peritubular collagen IV accumulation, reactive oxygen species (ROS) and macrophage infiltration (all P < 0.05). These changes were associated with an increase in Txnip mRNA and protein expression in the tubules and glomeruli of diabetic kidney. Treatment with tranilast for 4 weeks significantly attenuated Txnip up-regulation in diabetic rats and this was associated with a reduction in ROS, fibrosis and macrophage infiltration (all P < 0.05). This is the first study to demonstrate that tranilast not only has anti-inflammatory and anti-fibrotic effects as previously reported but also attenuates the up-regulation of Txnip and oxidative stress in diabetic nephropathy.

THE EFFECT OF REPEATED ANTISENSE THERAPY ON VARIOUS VASOACTIVE SYSTEMS IN YOUNG REN-2 TRANSGENIC RATS: PP.29.140

Objective: We demonstrated that single injection of antisense oligodeoxynucleotides (AS) against AT1 receptor had transient antihypertensive effect in TGR. We were interested whether repeated AS therapy will have durable effect on blood pressure (BP) and on the contribution of principle vasoactive systems to BP regulation in this model of ANG II-dependent hypertension. Methods: From the age of 30 days heterozygous Ren-2 rats were injected in 10-day intervals with AS until the age of 60 days. On days 40, 50 and 70 of age, basal BP and acute responses to consecutive blockade of renin-angiotensin (10 mg/kg captopril), sympathetic (5 mg/kg pentolinium) and nitric oxide (NO) (30 mg/kg L-NAME) systems were determined in conscious rats. Results: AS therapy decreased BP and reduced cardiac hypertrophy. Both these effects disappeared after the third AS injection. Captopril-induced BP decrease was higher on day 70 in untreated than in AS-treated TGR, suggesting higher contribution of RAS in established hypertension. On the contrary, AS therapy decreased the contribution of sympathetic system on day 40 (during hypertension development). BP decrease was double after SNS than after RAS blockade. AS-treated animals had higher BP increase after NO blockade on day 40. The impact of NO on BP control increased with age, suggesting its role as a compensatory mechanism against augmented activity of pressor systems. The dependency of principle vasoactive systems on the number of renin gene copies was evaluated in three groups of animals: HanSD (-/-),TGR heterozygous for renin gene (+/−) and homozygous TGR (+/+). The contribution of angiotensin-dependent and nifedipine-sensitive vasoconstriction increased with the number of renin gene copies, while heterozygous and homozygous TGR had reduced NO vasodilatory component. Conclusions: Repeated AS therapy of Ren-2 TGR has transient antihypertensive effect, which correlates with cardiac hypertrophy. Augmented sympathetic activity together with reduced NO activity contribute to hypertension in initial phases, while increased RAS influence prevails in established hypertension.

AT1 Receptor Blockade Prevents the Increase in Blood Pressure and the Augmentation of Intrarenal ANG II Levels in Hypertensive Cyp1a1-Ren2 Transgenic Rats Fed With a High-Salt Diet

IntroductionThis study was performed to determine the effects of high-salt diet on the magnitude of the increases in systolic blood pressure (SBP) and kidney tissue angiotensin (ANG) II levels that occur after induction of ANG II-dependent malignant hypertension in Cyp1a1-Ren2 transgenic rats with inducible expression of the mouse Ren2 renin gene [strain name: TGR(Cyp1a1Ren2)]. MethodsCyp1a1- Ren2 rats (n=6) were fed a normal diet containing 0.3% indole-3- carbinol (I3C) for 10days to induce ANG II-dependent malignant hypertension. ResultsRats induced with I3C exhibited increases in SBP and elevations of ANG II levels in kidney cortex and medulla. In a second group of rats (n=6), high-salt intake alone did not alter basal SBP; however, subsequent dietary administration of 0.3% I3C during continued high-salt intake elicited a substantially greater increase in SBP than observed in rats fed a normal salt diet. ANG II levels in kidney cortex and medulla of rats induced with I3C and fed a high-salt diet were elevated similarly to those in rats induced with I3C alone. Chronic administration of the AT1 receptor antagonist, losartan (100mg/L in drinking water, n=6), markedly attenuated the I3C-induced increase in SBP and prevented the augmentation of ANG II levels in kidney cortex and medulla in rats induced with I3C and maintained on a high-salt diet. ConclusionsActivation of AT1 receptors contributes to the augmented blood pressure and elevated kidney tissue ANG II levels that occur in Cyp1a1-Ren2 transgenic rats with malignant hypertension maintained on a high-salt diet.

Stimulation of the renin angiotensin system (RAS) in an early stage of life causes sustained hypertension in rats

Transient administration of inhibitors of the renin angiotensin system (RAS) during the prehypertensive period of spontaneously hypertensive rats leads to a long‐lasting lowering of blood pressure (BP) and a reduction of end organ damage. We tested if a transient stimulation of the RAS at young age would exert the opposite effect and permanently increases BP.Studies were performed in Cyp1a1 Ren2 transgenic rats. In this model the production of mouse renin is enhanced when 0.3% Indole‐3 carbinol (I3C) is added to the diet. After withdrawal of I3C the production of mouse renin stops almost immediately. Cyp1a1 Ren2 rats received I3C‐treatment between 4 to 8 weeks of age (WOA) or between 30–34 WOA. Non‐treated rats served as control. BP follow‐up was performed via tailcuff and intra‐arterial BP was determined at 4, 8, 12 and 20 WOA or 34 and 38 WOA. Data are presented as mean ± SEM.At 12 WOA, i.e. 4 weeks after I3C withdrawal, the I3C‐treated rats show a statistically significant increase in BP compared with control rats (156±18 versus 129±6 mmHg). This increase in BP is still present at 20 WOA, i.e. 12 weeks after I3C withdrawal (141±7 versus 125±6mmHg). In the adult rats (30–34 weeks), BP returned immediately to control values after stopping renin activation.This study shows that, at least in rats, sustained hypertension can be induced by a transient stimulation of the RAS in an early stage of life.

Nebivolol Attenuates Maladaptive Proximal Tubule Remodeling in Transgenic Rats

Background/Aims: The impact of nebivolol therapy on the renal proximal tubular cell (PTC) structure and function was investigated in a transgenic (TG) rodent model of hypertension and the cardiometabolic syndrome. The TG Ren2 rat develops nephropathy with proteinuria, increased renal angiotensin II levels and oxidative stress, and PTC remodeling. Nebivolol, a β₁-antagonist, has recently been shown to reduce albuminuria, in part, through reductions in renal oxidative stress. Accordingly, we hypothesized that nebivolol therapy would attenuate PTC damage and tubulointerstitial fibrosis. Methods: Young Ren2 (R2-N) and SD (SD-N) rats were treated with nebivolol (10 mg/kg/day) or vehicle (R2-C; SD-C) for 3 weeks. PTC structure and function were tested using transmission electron microscopy and functional measurements. Results: Nebivolol treatment decreased urinary N-acetyl-β-D-glucosaminidase, tubulointerstitial ultrastructural remodeling and fibrosis, NADPH oxidase activity, 3-nitrotyrosine levels, and increased megalin and lysosomal-associated membrane protein-2 immunostaining in PTCs. Ultrastructural abnormalities that were improved with therapy included altered canalicular structure, reduced endosomes/lysosomes and PTC vacuoles, basement membrane thickening, and mitochondrial remodeling/fragmentation. Conclusion: These observations support the notion that nebivolol may improve PTC reabsorption of albumin and other glomerular filtered small molecular weight proteins in association with the attenuation of oxidative stress, tubulointerstitial injury and fibrosis in this rat model of metabolic kidney disease.

Pericytopathy: Oxidative Stress and Impaired Cellular Longevity in the Pancreas and Skeletal Muscle in Metabolic Syndrome and Type 2 Diabetes

The pericyte's role has been extensively studied in retinal tissues of diabetic retinopathy; however, little is known regarding its role in such tissues as the pancreas and skeletal muscle. This supportive microvascular mural cell plays an important and novel role in cellular and extracellular matrix remodeling in the pancreas and skeletal muscle of young rodent models representing the metabolic syndrome and type 2 diabetes mellitus (T2DM). Transmission electron microscopy can be used to evaluate these tissues from young rodent models of insulin resistance and T2DM, including the transgenic Ren2 rat, db/db obese insulin resistantߞT2DM mouse, and human islet amyloid polypeptide (HIP) rat model of T2DM. With this method, the earliest pancreatic remodeling change was widening of the islet exocrine interface and pericyte hypercellularity, followed by pericyte differentiation into islet and pancreatic stellate cells with early fibrosis involving the islet exocrine interface and interlobular interstitium. In skeletal muscle there was a unique endothelial capillary connectivity via elongated longitudinal pericyte processes in addition to pericyte to pericyte and pericyte to myocyte cellcell connections allowing for paracrine communication. Initial pericyte activation due to moderate oxidative stress signaling may be followed by hyperplasia, migration and differentiation into adult mesenchymal cells. Continued robust oxidative stress may induce pericyte apoptosis and impaired cellular longevity. Circulating antipericyte autoantibodies have recently been characterized, and may provide a screening method to detect those patients who are developing pericyte loss and are at greater risk for the development of complications of T2DM due to pericytopathy and rarefaction. Once detected, these patients may be offered more aggressive treatment strategies such as early pharmacotherapy in addition to lifestyle changes targeted to maintaining pericyte integrity. In conclusion, we have provided a review of current knowledge regarding the pericyte and novel ultrastructural findings regarding its role in metabolic syndrome and T2DM.

Despite Similar Reduction of Blood Pressure and Renal ANG II and ET-1 Levels Aliskiren but not Losartan Normalizes Albuminuria in Hypertensive Ren-2 Rats

The relationship between angiotensin II (ANG II) and endothelin-1 (ET-1) is known to be complex; both peptides can initiate and potentiate the gene expression of each other. This pilot study investigated the effects of the AT(1) receptor blocker losartan or the direct renin inhibitor aliskiren on mean arterial pressure (MAP) and albuminuria and the renal ANG II and ET-1 levels. 3-month-old male Ren-2 transgenic rats (TGR) were treated either with losartan (5 mg kg(-1) day(-1)) or aliskiren (10 mg kg(-1) day(-1)) for 10 weeks. At the end of the experiment, rats were decapitated and cortical and papillary parts of kidneys were separated. Plasma and tissue ANG II levels were measured by RIA and tissue ET-1 concentrations by ELISA. In all four groups of animals ET-1 levels were lowest in renal cortex and more than 100-fold higher in the papilla. Cortical and papillary ET-1 concentrations in untreated TGR significantly exceeded those of control HanSD rats and were significantly depressed by both drugs. In both strains, papillary ANG II concentrations were moderately but significantly higher than cortical ANG II, TGR exhibited higher ANG II levels both in cortex and papilla as compared to control HanSD rats. Aliskiren and losartan at the doses used depressed similarly the levels of ANG II in cortex and papilla and reduced ET-1 significantly in the renal cortex and papilla below control levels in HanSD rats. Albuminuria, which was more than twice as high in TGR as in HanSD rats, was normalized with aliskiren and reduced by 28% with losartan, although MAP was reduced to a similar degree by both drugs. Despite similar reductions of MAP and renal ET-1 and ANG II levels aliskiren appears to be more effective than losartan, at the doses used, in reducing albuminuria in heterozygous hypertensive Ren-2 rats.

Contribution of oxidative stress to pulmonary arterial hypertension

Recent data implicate oxidative stress as a mediator of pulmonary hypertension (PH) and of the associated pathological changes to the pulmonary vasculature and right ventricle (RV). Increases in reactive oxygen species (ROS), altered redox state, and elevated oxidant stress have been demonstrated in the lungs and RV of several animal models of PH, including chronic hypoxia, monocrotaline toxicity, caveolin-1 knock-out mouse, and the transgenic Ren2 rat which overexpresses the mouse renin gene. Generation of ROS in these models is derived mostly from the activities of the nicotinamide adenine dinucleotide phosphate oxidases, xanthine oxidase, and uncoupled endothelial nitric oxide synthase. As disease progresses circulating monocytes and bone marrow-derived monocytic progenitor cells are attracted to and accumulate in the pulmonary vasculature. Once established, these inflammatory cells generate ROS and secrete mitogenic and fibrogenic cytokines that induce cell proliferation and fibrosis in the vascular wall resulting in progressive vascular remodeling. Deficiencies in antioxidant enzymes also contribute to pulmonary hypertensive states. Current therapies were developed to improve endothelial function, reduce pulmonary artery pressure, and slow the progression of vascular remodeling in the pulmonary vasculature by targeting deficiencies in either NO (PDE-type 5 inhibition) or PGI(2) (prostacyclin analogs), or excessive synthesis of ET-1 (ET receptor blockers) with the intent to improve patient clinical status and survival. New therapies may slow disease progression to some extent, but long term management has not been achieved and mortality is still high. Although little is known concerning the effects of current pulmonary arterial hypertension treatments on RV structure and function, interest in this area is increasing. Development of therapeutic strategies that simultaneously target pathology in the pulmonary vasculature and RV may be beneficial in reducing mortality associated with RV failure.

Comparative effect of direct renin inhibition and AT1R blockade on glomerular filtration barrier injury in the transgenic Ren2 rat

Renin-angiotensin system (RAS) activation contributes to kidney injury through oxidative stress. Renin is the rate-limiting step in angiotensin (ANG II) generation. Recent work suggests renin inhibition improves proteinuria comparable to ANG type 1 receptor (AT1R) blockade (ARB). Thereby, we investigated the relative impact of treatment with a renin inhibitor vs. an ARB on renal oxidative stress and associated glomerular structural and functional changes in the transgenic Ren2 rat, which manifests hypertension, albuminuria, and increased tissue RAS activity. Young Ren2 and age-matched Sprague-Dawley (SD) controls (age 6-9 wk) were treated with a renin inhibitor (aliskiren), an ARB (irbesartan), or vehicle for 21 days. Ren2 rats exhibited increases in systolic pressure (SBP), albuminuria, and renal 3-nitrotyrosine content as well as ultrastructural podocyte foot-process effacement and diminution of the podocyte-specific protein nephrin. Structural and functional alterations were accompanied by increased renal cortical ANG II, AT1R, as well as NADPH oxidase subunit (Nox2) expression compared with SD controls. Abnormalities were attenuated to a similar extent with both aliskiren and irbesartan treatment. Despite the fact the dose of irbesartan used caused a greater reduction in SBP than aliskerin treatment (P < 0.05), the effects on proteinuria, nephrin, and oxidative stress were similar between the two treatments. Our results highlight both the importance of pressor-related reductions on podocyte integrity and albuminuria as well as RAS-mediated oxidant stress largely comparable between ARB and renin inhibition treatment.

Rosuvastatin ameliorates the development of pulmonary arterial hypertension in the transgenic (mRen2)27 rat

We have recently reported that transgenic (mRen2)27 rats (Ren2 rats) exhibit pulmonary arterial hypertension (PAH), which is, in part, mediated by oxidative stress. Since 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors (statins) exhibit beneficial vascular effects independent of cholesterol synthesis, we hypothesized that rosuvastatin (RSV) treatment ameliorates PAH and pulmonary vascular remodeling in Ren2 rats, in part, by reducing oxidative stress. Six-week-old male Ren2 and Sprague-Dawley rats received RSV (10 mg x kg(-1) x day(-)1 ip) or vehicle for 3 wk. After treatment, right ventricular systolic pressure (RVSP) and mean arterial pressure (MAP) were measured. To evaluate treatment effects on pulmonary arteriole remodeling, morphometric analyses were performed to quantitate medial thickening and cell proliferation, whereas whole lung samples were used to quantitate the levels of 3-nitrotyrosine, superoxide, stable nitric oxide (NO) metabolites [nitrates and nitrites (NO(x))], and expression of NO synthase isoforms. In the Ren2 rat, RVSP is normal at 5 wk of age, PAH develops between 5 and 7 wk of age, and the elevated pressure is maintained with little variation through 13 wk. At 8 wk of age, left ventricular function and blood gases were normal in the Ren2 rat. Ren2 rats exhibited elevations in medial hypertrophy due to smooth muscle cell proliferation, 3-nitrotyrosine, NO(x), NADPH oxidase activity, and endothelial NO synthase expression compared with Sprague-Dawley rats. RSV significantly blunted the increase in RVSP but did not reduce MAP in the Ren2 rat; additionally, RSV significantly attenuated the elevated parameters examined in the Ren2 rat. These data suggest that statins may be a clinically viable adjunct treatment of PAH through reducing peroxynitrite formation.

Diabetic renal failure and associated pathology are ameliorated by Perindopril treatment in transgenic Ren-2 rats

Diabetic renal failure and associated pathology are ameliorated by Perindopril treatment in transgenic Ren-2 rats

T1609 Exercise Attenuates Hepatic Oxidative Stress and Prevents Fatty Liver Disease in Hypertensive Transgenic Ren2 Rats

T1609 Exercise Attenuates Hepatic Oxidative Stress and Prevents Fatty Liver Disease in Hypertensive Transgenic Ren2 Rats

Rosuvastatin Attenuates Pulmonary Arterial Hypertension in the Transgenic (mREN2)27 (Ren2) Rat

Activation of the intrapulmonary renin‐angiotensin system in male transgenic Ren2 rats is associated with increases in lung oxidative stress, medial thickening of pulmonary arterioles, and pulmonary hypertension (PH). Statins are potent antioxidants and are vasculoprotective. We hypothesized that rosuvastatin (RSV) ameliorates PH and pulmonary vascular remodeling in Ren2 rats, in part, by reducing oxidative stress. Ren2 and Sprague‐Dawley (SD) rats were injected daily with RSV (10 mg/kg) for 3 wk, after which we measured right ventricular systolic pressure (RVSP) and mean arterial pressure (MAP). To evaluate remodeling, we measured the thickness of pulmonary arterioles. To evaluate oxidative stress in the lung we measured 3‐nitrotyrosine, NADPH oxidase activity, NOX2 and Rac1 expression, and ROS. Ren2 rats had elevations in MAP, RVSP, thickness of pulmonary arterioles, NADPH oxidase activity, Nox2 and Rac1, ROS, and 3‐nitrotyrosine compared to SD rats. RSV blunted the increase in RVSP, but not MAP, in the Ren2. RSV attenuated elevations in surface area of small pulmonary arteries, reduced NADPH oxidase activity and ROS, and 3‐nitrotyrosine, but did not blunt elevated levels of Nox2 and Rac1 protein expression in the Ren2. In summary, RSV reduces oxidative stress, pulmonary vascular remodeling and PH in male Ren2 rats. NIH R01‐HL‐63904 and VA Merit (JRS), and MU Research Board (VD).

Direct renin inhibition with Aliskiren lowers blood pressure and improves renal dysfunction in Ren2 rats

Renin has gained notoriety over recent times regarding its involvement and/or facilitation of pro‐inflammatory processes. Investigations into the mechanism of action of renin have revealed that this enzyme can not only activate the RAAS but can also act through the renin receptor directly, to affect kidney function. The transgenic Ren2 rat harbors stable copies of the mouse renin gene and has been well characterized to have hypertension, endothelial dysfunction and insulin resistance. Mouse renin transgene is expressed at high levels in the kidneys and is likely responsible for the pathophysiology that is so characteristic of this animal. We sought to reverse the pathology with a mouse specific direct renin inhibitor, Aliskiren (50mg/kg). We observed that Aliskiren lowered blood pressure significantly as expected (N=8, p&lt;0.05). In addition, kidney oxidative stress, perivascular fibrosis and most importantly albuminuria were lowered in Ren2 transgenic animals (N=10, p&lt;0.05). Interestingly, Aliskiren‐treated Ren2 rats demonstrated improved insulin signaling as measured with increase in total IRS‐1, decrease in phosphorylated Akt (thr308), and improved GLUT4 translocation to the plasma membrane fraction (N=5, p&lt;0.05). Our data suggests that, Aliskiren treatment improves kidney function likely via inhibition of overactivity of RAAS and improved insulin signaling.

Oxidative Stress-Mediated Mitochondrial Dysfunction Contributes to Angiotensin II-Induced Nonalcoholic Fatty Liver Disease in Transgenic Ren2 Rats

Emerging evidence indicates that impaired mitochondrial fatty acid beta-oxidation plays a key role in liver steatosis. We have recently demonstrated that increased angiotensin (ANG) II causes progressive hepatic steatosis associated with oxidative stress; however, the underlying mechanisms remain unclear. We hypothesized that ANG II causes hepatic mitochondrial oxidative damage and impairs mitochondrial beta-oxidation, thereby leading to hepatic steatosis. We used the Ren2 rat with elevated endogenous ANG II levels to evaluate mitochondrial ultrastructural changes, gene expression levels, and beta-oxidation. Compared with Sprague-Dawley littermates, Ren2 livers exhibited mitochondrial damage and reduced beta-oxidation, as evidenced by ultrastructural abnormalities, decrease of mitochondrial content, percentage of palmitate oxidation to CO(2), enzymatic activities (beta-HAD and citrate synthase), and the expression levels of cytochrome c, cytochrome c oxidase subunit 1, and mitochondrial transcription factor A. These abnormalities were improved with either ANG II receptor blocker valsartan or superoxide dismutase/catalase mimetic tempol treatment. Both valsartan and tempol substantially attenuated mitochondrial lipid peroxidation in Ren2 livers. Interestingly, there was no difference in the expression of key enzymes (ACC1 and FAS) for fatty acid syntheses and their transcription factors (SREBP-1c and ChREBP) between Sprague-Dawley, untreated Ren2, and valsartan- or tempol-treated Ren2 rats. These results document that ANG II induces mitochondrial oxidative damage and impairs mitochondrial beta-oxidation, contributing to liver steatosis.

Mineralcorticoid Receptor (MR) Antagonism Attenuates Glomerular Filtration Barrier Remodeling in the Transgenic Ren2 Rat

Renin‐angiotensin‐aldosterone system (RAAS) activation is associated with glomerular filtration barrier/podocyte injury &amp; accompanying proteinuria. To explore potential MR mediated renal effects, Ren2 transgenic rats, which manifest increased tissue RAAS activity with elevated serum aldosterone levels, were used. Six week old Ren2 and Sprague‐Dawley rats were treated with a low dose of the MR antagonist spironolactone (SP)(0.24 mg/day) or vehicle for 21 days. Ren2 rats exhibited albuminuria, loss of podocyte‐specific proteins and podocyte foot‐process effacement on ultrastructural analysis, along with reduced expression of mediators of insulin metabolic signaling. Ren2 kidneys also manifested increased NADPH oxidase/ROS and redox‐sensitive Rho kinase activation in conjunction with enhanced renal tissue levels of MR, angiotensin type 1 receptor, angiotensin II and angiotensin‐(1‐12). SP treatment reduced albuminuria and tissue RAAS expression and improved podocyte structural and protein integrity along with improvements in insulin metabolic signaling. MR blockade directly provides protection in this model of renal RAAS activation, in part, through reductions in redox‐sensitive ROK impairment of insulin metabolic signaling and glomerular remodeling.Supported by NIH R01 HL73101‐01A1 (JRS) &amp; HL‐51952 (CMF), VA Merit (JRS), VA VISN 15 (AWC), &amp; Missouri Kidney Program (AWC).