Hepatoprotective and cardioprotective effects of empagliflozin in spontaneously hypertensive rats fed a high-fat diet.

A combination of liver and heart dysfunction worsens the prognosis of human survival. The aim of this study was to investigate whether empagliflozin (a sodium-glucose transporter-2 inhibitor) has beneficial effects not only on cardiac and renal function but also on hepatic function. Adult (6-month-old) male spontaneously hypertensive rats (SHR) were fed a high-fat diet (60% fat) for four months to induce hepatic steatosis and mild heart failure. For the last two months, the rats were treated with empagliflozin (empa, 10 mg.kg-1.day-1 in the drinking water). Renal function and oral glucose tolerance test were analyzed in control (n=8), high-fat diet (SHR+HF, n=10), and empagliflozin-treated (SHR+HF+empa, n=9) SHR throughout the study. Metabolic parameters and echocardiography were evaluated at the end of the experiment. High-fat diet feeding increased body weight and visceral adiposity, liver triglyceride and cholesterol concentrations, and worsened glucose tolerance. Although the high-fat diet did not affect renal function, it significantly worsened cardiac function in a subset of SHR rats. Empagliflozin reduced body weight gain but not visceral fat deposition. It also improved glucose sensitivity and several metabolic parameters (plasma insulin, uric acid, and HDL cholesterol). In the liver, empagliflozin reduced ectopic lipid accumulation, lipoperoxidation, inflammation and pro-inflammatory HETEs, while increasing anti-inflammatory EETs. In addition, empagliflozin improved cardiac function (systolic, diastolic and pumping) independent of blood pressure. The results of our study suggest that hepatoprotection plays a decisive role in the beneficial effects of empagliflozin in preventing the progression of cardiac dysfunction induced by high-fat diet feeding.

Renal sympathetic denervation improves pressure-natriuresis relationship in cardiorenal syndrome: insight from studies with Ren-2 transgenic hypertensive rats with volume overload induced using aorto-caval fistula.

The aim was to evaluate the effects of renal denervation (RDN) on autoregulation of renal hemodynamics and the pressure-natriuresis relationship in Ren-2 transgenic rats (TGR) with aorto-caval fistula (ACF)-induced heart failure (HF). RDN was performed one week after creation of ACF or sham-operation. Animals were prepared for evaluation of autoregulatory capacity of renal blood flow (RBF) and glomerular filtration rate (GFR), and of the pressure-natriuresis characteristics after stepwise changes in renal arterial pressure (RAP) induced by aortic clamping. Their basal values of blood pressure and renal function were significantly lower than with innervated sham-operated TGR (p < 0.05 in all cases): mean arterial pressure (MAP) (115 ± 2 vs. 160 ± 3 mmHg), RBF (6.91 ± 0.33 vs. 10.87 ± 0.38 ml.min-1.g-1), urine flow (UF) (11.3 ± 1.79 vs. 43.17 ± 3.24 µl.min-1.g-1) and absolute sodium excretion (UNaV) (1.08 ± 0.27 vs, 6.38 ± 0.76 µmol.min-1.g-1). After denervation ACF TGR showed improved autoregulation of RBF: at lowest RAP level (80 mmHg) the value was higher than in innervated ACF TGR (6.92 ± 0.26 vs. 4.54 ± 0.22 ml.min-1.g-1, p < 0.05). Also, the pressure-natriuresis relationship was markedly improved after RDN: at the RAP of 80 mmHg UF equaled 4.31 ± 0.99 vs. 0.26 ± 0.09 µl.min-1.g-1 recorded in innervated ACF TGR, UNaV was 0.31 ± 0.05 vs. 0.04 ± 0.01 µmol min-1.g-1 (p < 0.05 in all cases). In conclusion, in our model of hypertensive rat with ACF-induced HF, RDN improved autoregulatory capacity of RBF and the pressure-natriuresis relationship when measured at the stage of HF decompensation.

Renal denervation improves cardiac function independently of afterload and restores myocardial norepinephrine levels in a rodent heart failure model.

Renal nerves play a critical role in cardiorenal interactions. Renal denervation (RDN) improved survival in some experimental heart failure (HF) models. It is not known whether these favorable effects are indirect, explainable by a decrease in vascular afterload, or diminished neurohumoral response in the kidneys, or whether RDN procedure per se has direct myocardial effects in the failing heart. To elucidate mechanisms how RDN affects failing heart, we studied load-independent indexes of ventricular function, gene markers of myocardial remodeling, and cardiac sympathetic signaling in HF, induced by chronic volume overload (aorto-caval fistula, ACF) of Ren2 transgenic rats. Volume overload by ACF led to left ventricular (LV) hypertrophy and dysfunction, myocardial remodeling (upregulated Nppa, MYH 7/6 genes), increased renal and circulating norepinephrine (NE), reduced myocardial NE content, increased monoaminoxidase A (MAO-A), ROS production and decreased tyrosine hydroxylase (+) nerve staining. RDN in HF animals decreased congestion in the lungs and the liver, improved load-independent cardiac function (Ees, PRSW, Ees/Ea ratio), without affecting arterial elastance or LV pressure, reduced adverse myocardial remodeling (Myh 7/6, collagen I/III ratio), decreased myocardial MAO-A and inhibited renal neprilysin activity. RDN increased myocardial expression of acetylcholinesterase (Ache) and muscarinic receptors (Chrm2), decreased circulating and renal NE, but increased myocardial NE content, restoring so autonomic control of the heart. These changes likely explain improvements in survival after RDN in this model. The results suggest that RDN has remote, load-independent and favorable intrinsic myocardial effects in the failing heart. RDN therefore could be a useful therapeutic strategy in HF.

The treatment with sGC stimulator improves survival of hypertensive rats in response to volume-overload induced by aorto-caval fistula.

Heart failure (HF) has been declared as global pandemic and current therapies are still ineffective, especially in patients that develop concurrent cardio-renal syndrome. Considerable attention has been focused on the nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine monophosphate (cGMP) pathway. In the current study, we aimed to investigate the effectiveness of sGC stimulator (BAY41-8543) with the same mode of action as vericiguat, for the treatment of heart failure (HF) with cardio-renal syndrome. As a model, we chose heterozygous Ren-2 transgenic rats (TGR), with high-output heart failure, induced by aorto-caval fistula (ACF). The rats were subjected into three experimental protocols to evaluate short-term effects of the treatment, impact on blood pressure, and finally the long-term survival lasting 210 days. As control groups, we used hypertensive sham TGR and normotensive sham HanSD rats. We have shown that the sGC stimulator effectively increased the survival of rats with HF in comparison to untreated animals. After 60 days of sGC stimulator treatment, the survival was still 50% compared to 8% in the untreated rats. One-week treatment with sGC stimulator increased the excretion of cGMP in ACF TGR (109 ± 28 nnmol/12 h), but the ACE inhibitor decreased it (-63 ± 21 nnmol/12 h). Moreover, sGC stimulator caused a decrease in SBP, but this effect was only temporary (day 0: 117 ± 3; day 2: 108 ± 1; day 14: 124 ± 2 mmHg). These results support the concept that sGC stimulators might represent a valuable class of drugs to battle heart failure especially with cardio-renal syndrome, but further studies are necessary.

Endothelin type A receptor blockade increases renoprotection in congestive heart failure combined with chronic kidney disease: Studies in 5/6 nephrectomized rats with aorto-caval fistula.

BACKGROUND: Association of congestive heart failure (CHF) and chronic kidney disease (CKD) worsens the patient's prognosis and results in poor survival rate. The aim of this study was to examine if addition of endothelin type A (ETA) receptor antagonist to the angiotensin-converting enzyme inhibitor (ACEi) will bring additional beneficial effects in experimental rats. METHODS: CKD was induced by 5/6 renal mass reduction (5/6 NX) and CHF was elicited by volume overload achieved by creation of aorto-caval fistula (ACF). The follow-up was 24 weeks after the first intervention (5/6 NX). The treatment regimens were initiated 6 weeks after 5/6 NX and 2 weeks after ACF creation. RESULTS: The final survival in untreated group was 15%. The treatment with ETA receptor antagonist alone or ACEi alone and the combined treatment improved the survival rate to 64%, 71% and 75%, respectively, however, the difference between the combination and either single treatment regimen was not significant. The combined treatment exerted best renoprotection, causing additional reduction in albuminuria and reducing renal glomerular and tubulointerstitial injury as compared with ACE inhibition alone. CONCLUSIONS: Our results show that treatment with ETA receptor antagonist attenuates the CKD- and CHF-related mortality, and addition of ETA receptor antagonist to the standard blockade of RAS by ACEi exhibits additional renoprotective actions.

Endothelin type A receptor blockade attenuates aorto-caval fistula-induced heart failure in rats with angiotensin II-dependent hypertension.

OBJECTIVE: Evaluation of the effect of endothelin type A (ET A ) receptor blockade on the course of volume-overload heart failure in rats with angiotensin II-dependent hypertension. METHODS: Ren-2 renin transgenic rats (TGR) were used as a model of hypertension. Heart failure was induced by creating an aorto-caval fistula (ACF). Selective ET A receptor blockade was achieved by atrasentan. For comparison, other rat groups received trandolapril, an angiotensin-converting enzyme inhibitor (ACEi). Animals first underwent ACF creation and 2 weeks later the treatment with atrasentan or trandolapril, alone or combined, was applied; the follow-up period was 20 weeks. RESULTS: Eighteen days after creating ACF, untreated TGR began to die, and none was alive by day 79. Both atrasentan and trandolapril treatment improved the survival rate, ultimately to 56% (18 of 31 animals) and 69% (22 of 32 animals), respectively. Combined ACEi and ET A receptor blockade improved the final survival rate to 52% (17 of 33 animals). The effects of the three treatment regimens on the survival rate did not significantly differ. All three treatment regimens suppressed the development of cardiac hypertrophy and lung congestion, decreased left ventricle (LV) end-diastolic volume and LV end-diastolic pressure, and improved LV systolic contractility in ACF TGR as compared with their untreated counterparts. CONCLUSION: The treatment with ET A receptor antagonist delays the onset of decompensation of volume-overload heart failure and improves the survival rate in hypertensive TGR with ACF-induced heart failure. However, the addition of ET A receptor blockade did not enhance the beneficial effects beyond those obtained with standard treatment with ACEi alone.

Empagliflozin Is Not Renoprotective in Non-Diabetic Rat Models of Chronic Kidney Disease.

Gliflozins (sodium-glucose transporter-2 inhibitors) exhibited renoprotective effects not only in diabetic but also in non-diabetic patients with chronic kidney disease (CKD). Controversial results were reported in experimental non-diabetic models of CKD. Therefore, we examined empagliflozin effects in three CKD models, namely, in fawn-hooded hypertensive (FHH) rats, uninephrectomized salt-loaded (UNX + HS) rats, and in rats with Goldblatt hypertension (two-kidney, one-clip 2K1C) that were either untreated or treated with empagliflozin (10 mg/kg/day) for eight weeks. Plethysmography blood pressure (BP) was recorded weekly, and renal parameters (proteinuria, plasma urea, creatinine clearance, and sodium excretion) were analyzed three times during the experiment. At the end of the study, blood pressure was also measured directly. Markers of oxidative stress (TBARS) and inflammation (MCP-1) were analyzed in kidney and plasma, respectively. Body weight and visceral adiposity were reduced by empagliflozin in FHH rats, without a significant effect on BP. Experimentally induced CKD (UNX + HS and 2K1C) was associated with a substantial increase in BP and relative heart and kidney weights. Empagliflozin influenced neither visceral adiposity nor BP in these two models. Although empagliflozin increased sodium excretion, suggesting effective SGLT-2 inhibition, it did not affect diuresis in any experimental model. Unexpectedly, empagliflozin did not provide renoprotection because proteinuria, plasma urea, and plasma creatinine were not lowered by empagliflozin treatment in all three CKD models. In line with these results, empagliflozin treatment did not decrease TBARS or MCP-1 levels in either model. In conclusion, empagliflozin did not provide the expected beneficial effects on kidney function in experimental models of CKD.

Antihypertensive and metabolic effects of empagliflozin in Ren-2 transgenic rats, an experimental non-diabetic model of hypertension.

The new antidiabetic drugs, gliflozins, inhibit sodium-glucose transporter-2 in renal proximal tubules promoting glucose and sodium excretion. This leads not only to a significant improvement of glucose control but also to the reduction of blood pressure and body weight in both diabetic patients and experimental models. We examined whether these beneficial effects can also be achieved in a non-diabetic hypertensive model, namely in Ren-2 transgenic rats (TGR). Adult 6-month-old hypertensive TGR and their normotensive controls (Hannover Sprague-Dawley rats), were either untreated or treated with empagliflozin (10 mg/kg/day) for two months. Telemetric blood pressure monitoring, renal parameters as well as cardiac function via echocardiography were analyzed during the experiment. At the end of the study, the contribution of major vasoactive systems to blood pressure maintenance was studied. Metabolic parameters and markers of oxidative stress and inflammation were also analyzed. Empagliflozin had no effect on plasma glucose level but partially reduced blood pressure in TGR. Although food consumption was substantially higher in empagliflozin-treated TGR compared to the untreated animals, their body weight and the amount of epididymal and perirenal fat was decreased. Empagliflozin had no effect on proteinuria, but it decreased plasma urea, attenuated renal oxidative stress and temporarily increased urinary urea excretion. Several metabolic (hepatic triglycerides, non-esterified fatty acids, insulin) and inflammatory (TNF-α, leptin) parameters were also improved by empagliflozin treatment. By contrast, echocardiography did not reveal any effect of empagliflozin on cardiac function. In conclusion, empagliflozin exerted beneficial antihypertensive, anti-inflammatory and metabolic effects also in a non-diabetic hypertensive model.

Effects of Epoxyeicosatrienoic Acid-Enhancing Therapy on the Course of Congestive Heart Failure in Angiotensin II-Dependent Rat Hypertension: From mRNA Analysis towards Functional In Vivo Evaluation.

This study evaluates the effects of chronic treatment with EET-A, an orally active epoxyeicosatrienoic acid (EETs) analog, on the course of aorto-caval fistula (ACF)-induced heart failure (HF) in Ren-2 transgenic rats (TGR), a model characterized by hypertension and augmented activity of the renin-angiotensin system (RAS). The results were compared with standard pharmacological blockade of the RAS using angiotensin-converting enzyme inhibitor (ACEi). The rationale for employing EET-A as a new treatment approach is based on our findings that apart from increased RAS activity, untreated ACF TGR also shows kidney and left ventricle (LV) tissue deficiency of EETs. Untreated ACF TGR began to die 17 days after creating ACF and were all dead by day 84. The treatment with EET-A alone or ACEi alone improved the survival rate: in 156 days after ACF creation, it was 45.5% and 59.4%, respectively. The combined treatment with EET-A and ACEi appeared to improve the final survival to 71%; however, the difference from either single treatment regimen did not reach significance. Nevertheless, our findings support the notion that targeting the cytochrome P-450-dependent epoxygenase pathway of arachidonic acid metabolism should be considered for the treatment of HF.

Renal Sympathetic Denervation Attenuates Congestive Heart Failure in Angiotensin II-Dependent Hypertension: Studies with Ren-2 Transgenic Hypertensive Rats with Aortocaval Fistula.

OBJECTIVE: We examined if renal denervation (RDN) attenuates the progression of aortocaval fistula (ACF)-induced heart failure or improves renal hemodynamics in Ren-2 transgenic rats (TGR), a model of angiotensin II (ANG II)-dependent hypertension. METHODS: Bilateral RDN was performed 1 week after creation of ACF. The animals studied were ACF TGR and sham-operated controls, and both groups were subjected to RDN or sham denervation. In separate groups, renal artery blood flow (RBF) responses were determined to intrarenal ANG II (2 and 8 ng), norepinephrine (NE) (20 and 40 ng) and acetylcholine (Ach) (10 and 40 ng) 3 weeks after ACF creation. RESULTS: In nondenervated ACF TGR, the final survival rate was 10 versus 50% in RDN rats. RBF was significantly lower in ACF TGR than in sham-operated TGR (6.2 ± 0.3 vs. 9.7 ± 0.5 mL min-1 g-1, p < 0.05), the levels unaffected by RDN. Both doses of ANG II decreased RBF more in ACF TGR than in sham-operated TGR (-19 ± 3 vs. -9 ± 2% and -47 ± 3 vs. -22 ± 2%, p < 0.05 in both cases). RDN did not alter RBF responses to the lower dose, but increased it to the higher dose of ANG II in sham-operated as well as in ACF TGR. NE comparably decreased RBF in ACF TGR and sham-operated TGR, and RDN increased RBF responsiveness. Intrarenal Ach increased RBF significantly more in ACF TGR than in sham-operated TGR (29 ± 3 vs. 17 ± 3%, p < 0.05), the changes unaffected by RDN. ACF creation induced marked bilateral cardiac hypertrophy and lung congestion, both attenuated by RDN. In sham-operated but not in ACF TGR, RDN significantly decreased mean arterial pressure. CONCLUSION: The results show that RDN significantly improved survival rate in ACF TGR; however, this beneficial effect was not associated with improvement of reduced RBF or with attenuation of exaggerated renal vascular responsiveness to ANG II.

AT1 receptor blocker, but not an ACE inhibitor, prevents kidneys from hypoperfusion during congestive heart failure in normotensive and hypertensive rats.

To provide novel insights into the pathogenesis of heart failure-induced renal dysfunction, we compared the effects of ACE inhibitor (ACEi) and AT1 receptor blocker (ARB) on systemic and kidney hemodynamics during heart failure in normotensive HanSD and hypertensive transgenic (TGR) rats. High-output heart failure was induced by creating an aorto-caval fistula (ACF). After five weeks, rats were either left untreated or treatment with ACEi or ARB was started for 15 weeks. Subsequently, echocardiographic, renal hemodynamic and biochemical measurements were assessed. Untreated ACF rats with ACF displayed significantly reduced renal blood flow (RBF) (HanSD: 8.9 ± 1.0 vs. 4.7 ± 1.6; TGR: 10.2 ± 1.9 vs. 5.9 ± 1.2 ml/min, both P < .001), ACEi had no major RBF effect, whereas ARB completely restored RBF (HanSD: 5.6 ± 1.1 vs. 9.0 ± 1.5; TGR: 7.0 ± 1.2 vs. 10.9 ± 1.9 ml/min, both P < .001). RBF reduction in untreated and ACEi-treated rats was accompanied by renal hypoxia as measured by renal lactate dehydrogenase activity, which was ameliorated with ARB treatment (HanSD: 40 ± 4 vs. 42 ± 3 vs. 29 ± 5; TGR: 88 ± 4 vs. 76 ± 4 vs. 58 ± 4 milliunits/mL, all P < .01). Unlike improvement seen in ARB-treated rats, ACE inhibition didn't affect urinary nitrates compared to untreated ACF TGR rats (50 ± 14 vs. 22 ± 13 vs. 30 ± 13 µmol/mmol Cr, both P < .05). ARB was more effective than ACEi in reducing elevated renal oxidative stress following ACF placement. A marker of ACEi efficacy, the angiotensin I/angiotensin II ratio, was more than ten times lower in renal tissue than in plasma. Our study shows that ARB treatment, in contrast to ACEi administration, prevents renal hypoperfusion and hypoxia in ACF rats with concomitant improvement in NO bioavailability and oxidative stress reduction. The inability of ACE inhibition to improve renal hypoperfusion in ACF rats may result from incomplete intrarenal RAS suppression in the face of depleted compensatory mechanisms.

Deleterious Effects of Hyperactivity of the Renin-Angiotensin System and Hypertension on the Course of Chemotherapy-Induced Heart Failure after Doxorubicin Administration: A Study in Ren-2 Transgenic Rat.

Doxorubicin's (DOX) cardiotoxicity contributes to the development of chemotherapy-induced heart failure (HF) and new treatment strategies are in high demand. The aim of the present study was to characterize a DOX-induced model of HF in Ren-2 transgenic rats (TGR), those characterized by hypertension and hyperactivity of the renin-angiotensin-aldosterone system, and to compare the results with normotensive transgene-negative, Hannover Sprague-Dawley (HanSD) rats. DOX was administered for two weeks in a cumulative dose of 15 mg/kg. In HanSD rats DOX administration resulted in the development of an early phase of HF with the dominant symptom of bilateral cardiac atrophy demonstrable two weeks after the last DOX injection. In TGR, DOX caused substantial impairment of systolic function already at the end of the treatment, with further progression observed throughout the experiment. Additionally, two weeks after the termination of DOX treatment, TGR exhibited signs of HF characteristic for the transition stage between the compensated and decompensated phases of HF. In conclusion, we suggest that DOX-induced HF in TGR is a suitable model to study the pathophysiological aspects of chemotherapy-induced HF and to evaluate novel therapeutic strategies to combat this form of HF, which are urgently needed.

Combined treatment with epoxyeicosatrienoic acid analog and 20-hydroxyeicosatetraenoic acid antagonist provides substantial hypotensive effect in spontaneously hypertensive rats.

OBJECTIVES: The global morbidity and mortality related to hypertension and associated disorders increases continuously and novel therapeutic strategies are still in high demand. Increasing evidence suggests the important role in blood pressure regulation of cytochrome P-450-dependent metabolites of arachidonic acid. Epoxyeicosatrienoic acids (EETs) induce vasodilation and natriuresis, and have renoprotective and anti-inflammatory properties. 20-HETE is an arachidonic acid metabolite with both prohypertensive and antihypertensive activities. To explore the pathophysiological role of arachidonic acid metabolites in more detail, we examined the antihypertensive efficiency of EET-A, a stable analog of 14,15-EET, and of AAA, a novel antagonist of the 20-HETE receptors. METHODS: Male spontaneously hypertensive rats (SHR) were treated for 5 weeks with EET-A, AAA or the combination; age-matched untreated SHR and normotensive Wistar-Kyoto rats served as controls. EET-A and AAA were administered in drinking water at 10 mg/kg/day each. SBP was measured by telemetry and urine, blood, and tissue samples were collected for relevant analyses. RESULTS: EET-A or AAA given alone had no significant effect on SHR blood pressure. In contrast, combined treatment with AAA and EET-A was significantly antihypertensive, causing a decrease in SBP from 180 ±â€Š3 to 160 ±â€Š5 mmHg (P < 0.05). Additionally, the combined treatment attenuated cardiac hypertrophy, decreased kidney ANG II level, increased natriuresis, and increased the excretion of nitric oxide metabolites. CONCLUSION: Considering the beneficial impact of the combined treatment with EET-A and AAA on SHR blood pressure and cardiovascular and renal function, we suggest that the treatment is a promising therapeutic strategy for human hypertension.

Addition of Endothelin A-Receptor Blockade Spoils the Beneficial Effect of Combined Renin-Angiotensin and Soluble Epoxide Hydrolase Inhibition: Studies on the Course of Chronic Kidney Disease in 5/6 Nephrectomized Ren-2 Transgenic Hypertensive Rats.

INTRODUCTION: Previous studies in Ren-2 transgenic hypertensive rats (TGR) after 5/6 renal ablation (5/6 NX) have shown that besides pharmacological blockade of the renin-angiotensin system (RAS) also increasing kidney tissue epoxyeicosatrienoic acids (EET) levels by blocking soluble epoxide hydrolase (sEH), an enzyme responsible for degradation of EETs, and endothelin type A (ETA) receptor blockade retards chronic kidney disease (CKD) progression. This prompted us to evaluate if this progression will be alleviated by the addition of sEH inhibitor and ETA receptor antagonist to the standard complex blockade of RAS (angiotensin-converting enzyme inhibitor plus angiotensin II type 1 receptor blocker) in rats with established CKD. METHODS: The treatment regimens were initiated 6 weeks after 5/6 NX in TGR, and the follow-up period was 60 weeks. RESULTS: The addition of sEH inhibition to RAS blockade improved survival rate, further reduced albuminuria and renal glomerular and kidney tubulointerstitial injury, and attenuated the decline in creatinine clearance - all this as compared with 5/6 NX TGR treated with RAS blockade alone. Addition of ETA receptor antagonist to the combined RAS and sEH blockade not only offered no additional renoprotection but, surprisingly, also abolished the beneficial effects of adding sEH inhibitor to the RAS blockade. CONCLUSION: These data indicate that pharmacological strategies that combine the blockade of RAS and sEH could be a novel tool to combat the progression of CKD. Any attempts to further extend this therapeutic regimen should be made with extreme caution.

Renoprotection Provided by Additional Diuretic Treatment in Partially Nephrectomized Ren-2 Transgenic Rats Subjected to the Combined RAS and ETA Blockade.

OBJECTIVE: Our previous study in heterozygous Ren-2 transgenic rats (TGR) demonstrated that long-term treatment with endothelin receptor A (ETA) blocker atrasentan added to the renin-angiotensin system (RAS) blockade had renoprotective effects in a model of chronic kidney disease (CKD) induced by partial nephrectomy. Since ETA blockade is known to cause edema, we were interested whether diuretic treatment added to this therapy would be beneficial. DESIGN AND METHODS: Partial nephrectomy (NX) was performed at the age of 3 months in TGR rats which were subjected to: (i) RAS blockade alone (angiotensin receptor blocker losartan and angiotensin converting enzyme inhibitor trandolapril), (ii) combined RAS (losartan and trandolapril) and ETA receptor blockade (atrasentan), or (iii) diuretic (hydrochlorothiazide) added to the combined RAS + ETA blockade for 50 weeks following NX. RESULTS: At the end of the study systolic blood pressure and cardiac hypertrophy were similarly decreased in all treated groups. Survival was significantly improved by ETA receptor blockade added to RAS blockade with no further effects of diuretic treatment. However, additional diuretic treatment combined with RAS + ETA blockade decreased body weight and had beneficial renoprotective effects - reductions of both kidney weight and kidney damage markers. Proteinuria gradually increased in rats treated with RAS blockade alone, while it was substantially lowered by additional ETA blockade. In rats treated with additional diuretic, proteinuria was progressively reduced throughout the experiment. CONCLUSION: A diuretic added to the combined RAS and ETA blockade has late renoprotective effects in CKD induced by partial nephrectomy in Ren-2 transgenic rats. The diuretic improved: renal function (evaluated as proteinuria and creatinine clearance), renal morphology (kidney mass, glomerular volume), and histological markers of kidney damage (glomerulosclerosis index, tubulointerstitial injury).

Epoxyeicosatrienoic Acid-Based Therapy Attenuates the Progression of Postischemic Heart Failure in Normotensive Sprague-Dawley but Not in Hypertensive Ren-2 Transgenic Rats.

Epoxyeicosatrienoic acids (EETs) and their analogs have been identified as potent antihypertensive compounds with cardio- and renoprotective actions. Here, we examined the effect of EET-A, an orally active EET analog, and c-AUCB, an inhibitor of the EETs degrading enzyme soluble epoxide hydrolase, on the progression of post-myocardial infarction (MI) heart failure (HF) in normotensive Hannover Sprague-Dawley (HanSD) and in heterozygous Ren-2 transgenic rats (TGR) with angiotensin II-dependent hypertension. Adult male rats (12 weeks old) were subjected to 60-min left anterior descending (LAD) coronary artery occlusion or sham (non-MI) operation. Animals were treated with EET-A and c-AUCB (10 and 1 mg/kg/day, respectively) in drinking water, given alone or combined for 5 weeks starting 24 h after MI induction. Left ventricle (LV) function and geometry were assessed by echocardiography before MI and during the progression of HF. At the end of the study, LV function was determined by catheterization and tissue samples were collected. Ischemic mortality due to the incidence of sustained ventricular fibrillation was significantly higher in TGR than in HanSD rats (35.4 and 17.7%, respectively). MI-induced HF markedly increased LV end-diastolic pressure (Ped) and reduced fractional shortening (FS) and the peak rate of pressure development [+(dP/dt)max] in untreated HanSD compared to sham (non-MI) group [Ped: 30.5 ± 3.3 vs. 9.7 ± 1.3 mmHg; FS: 11.1 ± 1.0 vs. 40.8 ± 0.5%; +(dP/dt)max: 3890 ± 291 vs. 5947 ± 309 mmHg/s]. EET-A and c-AUCB, given alone, tended to improve LV function parameters in HanSD rats. Their combination amplified the cardioprotective effect of single therapy and reached significant differences compared to untreated HanSD controls [Ped: 19.4 ± 2.2 mmHg; FS: 14.9 ± 1.0%; +(dP/dt)max: 5278 ± 255 mmHg/s]. In TGR, MI resulted in the impairment of LV function like HanSD rats. All treatments reduced the increased level of albuminuria in TGR compared to untreated MI group, but neither single nor combined EET-based therapy improved LV function. Our results indicate that EET-based therapy attenuates the progression of post-MI HF in HanSD, but not in TGR, even though they exhibited renoprotective action in TGR hypertensive rats.

The exaggerated salt-sensitive response in hypertensive transgenic rats (TGR mRen-2) fostered by a normotensive female.

Suboptimal conditions during prenatal and early postnatal development can increase risk of hypertension later in life. We studied consequences of a changed perinatal environment by initiating the cross-fostering of homozygous Ren-2 transgenic rat (TGR) offspring to normotensive, transgene-negative control mothers, and vice versa. We hypothesized that cross-fostering to a normotensive female can attenuate the development of malignant hypertension in TGR offspring (TGRx) and change their salt-sensitive response. Blood pressure (BP) was monitored by the telemetry system under normal salt intake, and BP responses to increased salt intake in the phase of established hypertension. Under normal salt conditions, BP was not markedly different in cross-fostered animals compared with controls. However, BP responses to 2% salt intake led to a stronger BP response in TGRx during the active phase when compared with the control TGR group. The TGRx also exhibited increased albuminuria, lower sodium excretion, and creatinine clearance under higher salt intake compared with control salt intake. Higher salt intake resulted in a significant increase of aldosterone concentrations only in the TGRx group; moreover, TGRx rats exhibited more pronounced renal injury compared with controls. In conclusion, our data indicate that cross-fostering in TGR not only did not attenuate the development of hypertension but, on the contrary, led to the deterioration of BP regulation, particularly due to exaggerated salt sensitivity and sodium retention in TGRx. Results underline the important role of the mother during lactation in postnatal development of the offspring, since these changes reflected different ion content in milk of a particular strain of rats.

Two pharmacological epoxyeicosatrienoic acid-enhancing therapies are effectively antihypertensive and reduce the severity of ischemic arrhythmias in rats with angiotensin II-dependent hypertension.

OBJECTIVE: We examined the effects of treatment with soluble epoxide hydrolase inhibitor (sEHi) and epoxyeicosatrienoic acids (EETs) analogue (EET-A), given alone or combined, on blood pressure (BP) and ischemia/reperfusion myocardial injury in rats with angiotensin II (ANG II)-dependent hypertension. METHODS: Ren-2 transgenic rats (TGR) were used as a model of ANG II-dependent hypertension and Hannover Sprague-Dawley rats served as controls. Rats were treated for 14 days with sEHi or EET-A and BP was measured by radiotelemetry. Albuminuria, cardiac hypertrophy and concentrations of ANG II and EETs were determined. Separate groups were subjected to acute myocardial ischemia/reperfusion injury and the infarct size and ventricular arrhythmias were determined. RESULTS: Treatment of TGR with sEHi and EET-A, given alone or combined, decreased BP to a similar degree, reduced albuminuria and cardiac hypertrophy to similar extent; only treatment regimens including sEHi increased myocardial and renal tissue concentrations of EETs. sEHi and EET-A, given alone or combined, suppressed kidney ANG II levels in TGR. Remarkably, infarct size did not significantly differ between TGR and Hannover Sprague-Dawley rats, but the incidence of ischemia-induced ventricular fibrillations was higher in TGR. Application of sEHi and EET-A given alone and combined sEHi and EET-A treatment were all equally effective in reducing life-threatening ventricular fibrillation in TGR. CONCLUSION: The findings indicate that chronic treatment with either sEHi or EET-A exerts distinct antihypertensive and antiarrhythmic actions in our ANG II-dependent model of hypertension whereas combined administration of sEHi and EET-A does not provide additive antihypertensive or cardioprotective effects.

Fenofibrate Attenuates Hypertension in Goldblatt Hypertensive Rats: Role of 20-Hydroxyeicosatetraenoic Acid in the Nonclipped Kidney.

BACKGROUND: There is vast evidence that the renin-angiotensin system is not the sole determinant of blood pressure (BP) elevation in human renovascular hypertension or the relevant experimental models. This study tested the hypothesis that kidney deficiency of 20-hydroxyeicosatetraenoic acid (20-HETE), a product of cytochrome P450 (CYP)-dependent ω-hydroxylase pathway of arachidonic acid metabolism, is important in the pathophysiology of the maintenance phase of 2-kidney, 1-clip (2K1C) Goldblatt hypertension. MATERIALS AND METHODS: In 2K1C Goldblatt rats with established hypertension, angiotensin II, angiotensin 1-7, 20-HETE concentrations and gene expression of CYP4A1 enzyme (responsible for 20-HETE formation) of the nonclipped kidney were determined. We examined if 14 days׳ administration of fenofibrate, a lipid-lowering drug, would increase CYP4A1 gene expression and renal 20-HETE formation, and if increased 20-HETE concentrations in the nonclipped kidney would decrease BP (telemetric measurements). RESULTS: CYP4A1 gene expression, 20-HETE and angiotensin 1-7 concentrations were lower and angiotensin II levels were higher in the nonclipped kidney of 2K1C rats than in sham-operated rats. Fenofibrate increased CYP4A1 gene expression and 20-HETE concentration in the nonclipped kidney and significantly decreased BP in 2K1C rats but did not restore it to normotensive range. The treatment did not change BP in sham-operated rats. CONCLUSIONS: Our results suggest that alterations in the RAS and CYP-dependent ω-hydroxylase metabolites of arachidonic acid in the nonclipped kidneys are both important in the pathophysiology of the maintenance phase of 2K1C Goldblatt hypertension. Therefore, fenofibrate treatment effectively attenuated hypertension, probably via stimulation of 20-HETE formation in the nonclipped kidney.

Intrarenal alterations of the angiotensin-converting enzyme type 2/angiotensin 1-7 complex of the renin-angiotensin system do not alter the course of malignant hypertension in Cyp1a1-Ren-2 transgenic rats.

The role of the intrarenal renin-angiotensin system (RAS) in the pathophysiology of malignant hypertension is not fully understood. Accumulating evidence indicates that the recently discovered vasodilator axis of the RAS, angiotensin-converting enzyme (ACE) type 2 (ACE2)/angiotensin 1-7 (ANG 1-7), constitutes an endogenous system counterbalancing the hypertensiogenic axis, ACE/angiotensin II (ANG II)/AT1 receptor. This study aimed to evaluate the role of the intrarenal vasodilator RAS axis in the pathophysiology of ANG II-dependent malignant hypertension in Cyp1a1-Ren-2 transgenic rats. ANG II-dependent malignant hypertension was induced by 13 days' dietary administration of indole-3-carbinol (I3C), a natural xenobiotic that activates the mouse renin gene in Cyp1a1-Ren-2 transgenic rats. It was hypothesized that pharmacologically-induced inhibition of the ACE2/ANG 1-7 complex should aggravate, and activation of this axis should attenuate, the course of ANG II-dependent malignant hypertension. Blood pressure (BP) was monitored by radiotelemetry. ACE2 inhibitor (DX 600, 0.2 µg/day) and ACE2 activator (DIZE, 1 mg/day) were administrated via osmotic minipumps. Even though ACE2 inhibitor significantly decreased and ACE2 activator increased intrarenal ANG 1-7 concentrations, the course of BP, as well as of albuminuria, cardiac hypertrophy and renal glomerular damage, were not altered. It was shown that intrarenal alterations in the ACE2/ANG 1-7 complex did not significantly modify the course of malignant hypertension in I3C-induced Cyp1a1-Ren-2 transgenic rats. Thus, in our experimental setting alterations of this intrarenal vasodilator complex of the RAS do not significantly modify the form of malignant hypertension that clearly depends on the inappropriately increased activity of the ACE/ANG II/AT1 receptor axis.