Resveratrol improved kidney function and structure in malignantly hypertensive rats by restoration of antioxidant capacity and nitric oxide bioavailability.

BACKGROUND: The main cause of death among patients with malignant hypertension is a kidney failure. The promising field in essential and malignant hypertension therapy could be centered on the amelioration of oxidative stress using antioxidant molecules like resveratrol. Resveratrol is a potent antioxidative agent naturally occurred in many plants that possess health-promoting properties. METHODS: In the present study, we investigated the therapeutic potential of resveratrol, a polyphenol with anti-oxidative activity, in NG-L-Arginine Methyl Ester (L-NAME) treated spontaneously hypertensive rats (SHR) - malignantly hypertensive rats (MHR). RESULTS: Resveratrol significantly improves oxidative damages by modulation of antioxidant enzymes and suppression of prooxidant factors in the kidney tissue of MHR. Enhanced antioxidant defense in the kidney improves renal function and ameliorates the morphological changes in this target organ. Besides, protective properties of resveratrol are followed by the restoration of the nitrogen oxide (NO) pathway. 4) Conclusion: Antioxidant therapy with resveratrol could represent promising therapeutical approach in hypertension, especially malignant, against kidney damage.

RNA sequencing reveals induction of specific renal inflammatory pathways in a rat model of malignant hypertension.

In malignant hypertension, far more severe kidney injury occurs than in the "benign" form of the disease. The role of high blood pressure and the renin-angiotensin-aldosterone system is well recognized, but the pathogenesis of the renal injury of malignant hypertension (MH) remains incompletely understood. Using the rat model of two-kidney, one-clip renovascular hypertension in which some but not all animals develop MH, we performed a transcriptomic analysis of gene expression by RNA sequencing to identify transcriptional changes in the kidney cortex specific for MH. Differential gene expression was assessed in three groups: MH, non-malignant hypertension (NMH), and normotensive, sham-operated controls. To distinguish MH from NMH, we considered two factors: weight loss and typical renovascular lesions. Mean blood pressure measured intraarterially was elevated in MH (220 ± 6.5 mmHg) as well as in NMH (192 ± 6.4 mmHg), compared to controls (119 ± 1.7 mmHg, p < 0.05). Eight hundred eighty-six genes were exclusively regulated in MH only. Principal component analysis revealed a separated clustering of the three groups. The data pointed to an upregulation of many inflammatory mechanisms in MH including pathways which previously attracted relatively little attention in the setting of hypertensive kidney injury: Transcripts from all three complement activation pathways were upregulated in MH compared to NMH but not in NMH compared with controls; immunohistochemistry confirmed complement deposition in MH exclusively. The expression of chemokines attracting neutrophil granulocytes (CXCL6) and infiltration of myeloperoxidase-positive cells were increased only in MH rats. The data suggest that these pathways, especially complement deposition, may contribute to kidney injury under MH. KEY MESSAGES: The most severe hypertension-induced kidney injury occurs in malignant hypertension. In a rat model of malignant hypertension, we assessed transcriptional responses in the kidney exposed to high blood pressure. A broad stimulation of inflammatory mechanisms was observed, but a few specific pathways were activated only in the malignant form of the disease, notably activation of the complement cascades. Complement inhibitors may alleviate the thrombotic microangiopathy of malignant hypertension even in the absence of primary complement abnormalities.

Correlations Between Interleukin-11 Expression and Hypertensive Kidney Injury in a Rat Model of Renovascular Hypertension.

BACKGROUND: Interleukin-11 (IL-11) is a pleiotropic cytokine of the interleukin-6 family. Recent studies revealed its crucial role in the development of cardiovascular fibrosis. In this study we examined IL-11 expression levels in the heart and the kidney exposed to high blood pressure in renovascular hypertensive rats and their correlations to fibrotic markers and kidney injury. METHODS: Two-kidney, one-clip renovascular hypertension (2K1C) was induced in rats. IL-11 expression was measured by real-time polymerase chain reaction in the left ventricle and the right kidney. The correlation of cardiac IL-11 expression with biomarkers of renal fibrosis was assessed. We further investigated IL-11 expression in 2K1C rats grouped into rats with malignant vs. nonmalignant hypertension (distinguishing criteria: weight loss, number of fibrinoid necrosis, and onion skin lesions). RESULTS: Thirty-five days after clipping, mean arterial pressure was significantly increased in 2K1C. Renal IL-11 expression was elevated in 2K1C. In the heart there was only a trend toward higher IL-11 expression in 2K1C. IL-11 in the kidney in 2K1C correlated with the expression of transforming growth factor (TGF)-ß1/2, collagens, fibronectin, osteopontin, as well as tissue inhibitors of metalloprotease 1/2. There were also correlations of IL-11 with tissue collagen expansion, number of activated fibroblasts and serum creatinine, but no correlation with mean arterial pressure. Renal expression of IL-11 was highest in rats with malignant hypertension. CONCLUSIONS: Renal IL-11 expression of renovascular hypertensive rats is markedly increased and correlates with profibrotic markers and loss of function and might therefore serve as a biomarker for the severity of hypertensive nephrosclerosis.

20-Hydroxyeicosatetraenoic acid antagonist attenuates the development of malignant hypertension and reverses it once established: a study in Cyp1a1-Ren-2 transgenic rats.

We hypothesized that vascular actions of 20-hydroxyeicosatetraenoic acid (20-HETE), the product of cytochrome P450 (CYP450)-dependent ω-hydroxylase, potentiate prohypertensive actions of angiotensin II (ANG II) in Cyp1a1-Ren-2 transgenic rats, a model of ANG II-dependent malignant hypertension. Therefore, we evaluated the antihypertensive effectiveness of 20-HETE receptor antagonist (AAA) in this model. Malignant hypertension was induced in Cyp1a1-Ren-2 transgenic rats by activation of the renin gene using indole-3-carbinol (I3C), a natural xenobiotic. Treatment with AAA was started either simultaneously with induction of hypertension or 10 days later, during established hypertension. Systolic blood pressure (SBP) was monitored by radiotelemetry, indices of renal and cardiac injury, and kidney ANG II levels were determined. In I3C-induced hypertensive rats, early AAA treatment reduced SBP elevation (to 161 ± 3 compared with 199 ± 3 mmHg in untreated I3C-induced rats), reduced albuminuria, glomerulosclerosis index, and cardiac hypertrophy (P<0.05 in all cases). Untreated I3C-induced rats showed augmented kidney ANG II (405 ± 14 compared with 52 ± 3 fmol/g in non-induced rats, P<0.05) which was markedly lowered by AAA treatment (72 ± 6 fmol/g). Remarkably, in TGR with established hypertension, AAA also decreased SBP (from 187 ± 4 to 158 ± 4 mmHg, P<0.05) and exhibited organoprotective effects in addition to marked suppression of kidney ANG II levels. In conclusion, 20-HETE antagonist attenuated the development and largely reversed the established ANG II-dependent malignant hypertension, likely via suppression of intrarenal ANG II levels. This suggests that intrarenal ANG II activation by 20-HETE is important in the pathophysiology of this hypertension form.

Comparison of Renal Outcome among Japanese Patients with or without Microangiopathic Hemolysis in Malignant Phase Hypertension: A Single-Center Retrospective Study.

BACKGROUND/AIMS: Although microangiopathic hemolysis (MAH) is a well-known complication of malignant phase hypertension (MPH), only less data on whether MAH in MPH predicts renal outcome exist. Therefore, we evaluated whether MAH was associated with the renal outcome in patients with MPH. METHODS: We conducted a single-center, retrospective, cohort study. Data from 35 patients diagnosed with MPH between October 1998 and January 2015 were analyzed. MPH was defined as the presence of a diastolic blood pressure of ≥120 mm Hg and grades III/IV hypertensive retinopathy according to the Keith-Wagener-Barker classification. MAH was defined as the presence of a low platelet count (<150 × 109/L) together with either an elevated level of lactate dehydrogenase (LDH; >220 U/L), or the presence of schistocytes, or both and the normalization of platelet and LDH level or schistocyte levels after adequate blood pressure control was achieved. The primary outcome was dialysis induction. RESULTS: Fifteen patients had MAH. Those with MAH had significantly severe renal dysfunction at the onset of MPH. The length of follow-up (median, interquartile range) of patients with MAH and those without MAH were 30 (16-94) and 48 (25-115) months, respectively. Dialysis was induced in 9 of 15 patients with MAH and in 6 of 20 patients without MAH. Renal survival in patients with MAH was worse than that in those without, but this was not statistically significant (p = 0.08). By multivariate Cox regression analysis, MAH was not shown to contribute to dialysis induction. CONCLUSION: MAH did not predict renal outcome in MPH.

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.

The endothelium as the common denominator in malignant hypertension and thrombotic microangiopathy.

The endothelium plays a pivotal role in vascular biology. The endothelium is the primary site of injury in thrombotic microangiopathies including malignant hypertension. Endothelial injury in thrombotic microangiopathies is the result of increased shear stress, toxins, and/or dysregulated complement activation. Endothelial injury can lead to microvascular thrombosis resulting in ischemia and organ dysfunction, the clinical hallmarks of thrombotic microangiopathies. Currently, available therapies target the underlying mechanisms that lead to endothelial injury in these conditions. Ongoing investigations aim at identifying drugs that protect the endothelium.

Mesangial cells are responsible for orchestrating the renal podocytes injury in the context of malignant hypertension.

AIM: Two populations of renal cells fully possess functional contractile cell apparatus: mesangial cells and podocytes. Previous studies demonstrated that in the context of malignant hypertension overproduction of Angiotensin-II by the contracting mesangial cells aggravated hypercellularity and apoptosis of adjacent cell populations. The role of podocytes in pathogenesis of malignant hypertension is unclear. We investigated responsiveness of normal vs. hyperglycaemic podocytes to pressure in a model of malignant hypertension. METHODS: Rat renal podocytes and mesangial cells were subjected to high hydrostatic pressure, using an in vitro model of malignant hypertension. Part of them was pre-exposed to hyperglycaemic medium. Alternatively, the cells were cultured in conditioned medium collected from mesangial cells pre-exposed to pressure. RESULTS: Angiotensin-II was significantly increased in normoglycaemic mesangial cells subjected to pressure, triggering enhanced proliferation and apoptosis. No augmented Angiotensin-II, proliferation or apoptosis were evident in pressure-exposed normoglycaemic podocytes. In hyperglycaemic mesangial cells, but not podocytes, basal Angiotensin-II and apoptosis were augmented, along with abrogated proliferation. Challenge with exogenous Angiotensin-II or Angiotensin-II-containing conditioned medium, induced apoptosis both in podocytes and mesangial cells. CONCLUSIONS: 1. Unlike mesangial cells, podocytes do not respond to high pressure or hyperglycaemia per se. Resultantly, neither high pressure nor hyperglycaemia, trigger apoptosis of podocytes in vitro. However, surplus of Angiotensin-II, amply produced in vivo by the adjacent mesangial cells, would seem to be sufficient for initiating apoptosis of both mesangial cells and podocytes. 2. Hyperglycaemia abrogates cell replication. Resultantly, in diabetic patients regeneration of renal tissue damaged by the incidence of malignant hypertension may become compromised or completely lost.

Oxidative capacity and fatigability in run-trained malignant hyperthermia-susceptible mice.

INTRODUCTION: The purpose of this study was to test the hypothesis that malignant hyperthermia model mice (RyR1Y522S/wt) are more vulnerable to exercise-induced muscle injury and fatigability and adapt less to run training. METHODS: After 6 weeks of voluntary wheel running, we measured anterior crural muscle fatigability, muscle injury, and cytochrome oxidase (COX) and citrate synthase (CS). RESULTS: Although RyR1Y522S/wt mice ran without undergoing MH episodes, they ran 42% less distance than wild-type (WT) mice. Muscles from WT mice exhibited increased fatigue resistance and COX content after training. Muscles from RyR1Y522S/wt mice demonstrated no significant change in fatigability or COX and CS after training. However, muscles from RyR1Y522S/wt mice displayed less intrinsic fatigability and greater COX/CS content and muscle damage than WT mice. CONCLUSIONS: RyR1Y522S/wt mice can run without having rhabdomyolysis, and their inability to adapt to training appears to stem from intrinsic enhancement of mitochondrial enzymes and fatigue resistance.

[Analysis of efficacy and safety of administration of moxonidine in patients with arterial hypertension and hypertensive crises].

In the review we present detailed analysis of antihypertensive action of 3-nd generation sympatholytic moxonidine. Due to selective interaction with imidazoline I1-receptors moxonidine diminishes sympathetic activity causing lowering of peripheral vascular resistance. This leads to significant lowering of systolic and diastolic arterial pressure. Efficacy and safety of the drug has been shown both for the management of uncomplicated hypertensive crises and long term treatment of arterial hypertension (AH). Appropriateness of the use of moxonidine in patients with AH combined with diabetes mellitus, metabolic syndrome, chronic obstructive pulmonary disease has been confirmed. Moxonidine is well tolerated; its bioavailability after oral intake reaches 90%. The drug produces neither hypotensive "first dose" nor rebound effects.

Enhancement of renin and prorenin receptor in collecting duct of Cyp1a1-Ren2 rats may contribute to development and progression of malignant hypertension.

To determine whether in the transgenic rat model [TGR(Cyp1a1Ren2)] with inducible ANG II-dependent malignant hypertension changes in the activation of intrarenal renin-angiotensin system may contribute to the pathogenesis of hypertension, we examined the gene expression of angiotensinogen (AGT) in renal cortical tissues and renin and prorenin receptor [(P)RR] in the collecting duct (CD) of the kidneys from Cyp1a1Ren2 rats (n = 6) fed a normal diet containing 0.3% indole-3-carbinol (I3C) for 10 days and noninduced rats maintained on a normal diet (0.6% NaCl diet; n = 6). Rats induced with I3C developed malignant hypertension and exhibited alterations in the expression of renin and (P)RR expressed by the CD cells. In the renal medullary tissues of the Cyp1a1Ren2 transgenic rats with malignant hypertension, renin protein levels in CD cells were associated with maintained renin content and lack of suppression of the endogenous Ren1c gene expression. Furthermore, these tissues exhibited increased levels of (P)RR transcript, as well as of the protein levels of the soluble form of this receptor, the s(P)RR. Intriguingly, although previous findings demonstrated that urinary AGT excretion is augmented in Cyp1a1Ren2 transgenic rats with malignant hypertension, in the present study we did not find changes in the gene expression of AGT in renal cortical tissues of these rats. The data suggest that upregulation of renin and the s(P)RR in the CD, especially in the renal medullary tissues of Cyp1a1Ren2 transgenic rats with malignant hypertension, along with the previously demonstrated increased availability of AGT in the urine of these rats, may constitute a leading mechanism to explain elevated formation of kidney ANG II levels in this model of ANG II-dependent hypertension.

Increased TRPC3 expression in vascular endothelium of patients with malignant hypertension.

An increased expression of transient receptor potential canonical type 3 (TRPC3) cation channels has been proposed as one of the factors contributing to the pathogenesis of hypertension. To test that hypothesis we compared the expression of TRPC3 and TRPC6 as an endogenous control in human vascular endothelium of preglomerular arterioles in kidney biopsies from six patients with malignant hypertension and from four patients with diarrhea-associated hemolytic-uremic syndrome. Patients with malignant hypertension showed significantly higher systolic blood pressure and more prominent expression of TRPC3 in vascular endothelium of preglomerular arterioles compared to patients with hemolytic-uremic syndrome. The expression of TRPC6 was not different between the two groups. The study supports the hypothesis that the increased expression of TRPC3 is associated with malignant hypertension in humans.

Association between thrombotic microangiopathy and reduced ADAMTS13 activity in malignant hypertension.

The thrombotic microangiopathy observed in malignant hypertension is similar to that of thrombotic thrombocytopenic purpura, which is associated with a deficiency of ADAMTS13, a von Willebrand factor (VWF)-cleaving protease that cleaves large prothrombogenic multimers. We hypothesized that ADAMTS13 is deficient in malignant hypertension and that the severity of thrombotic microangiopathy is associated with decreased ADAMTS13 activity. We included 20 patients with malignant and 20 patients with severe hypertension, and 20 matched normotensive individuals served as control subjects. VWF, active VWF, and free hemoglobin were assessed to explore predictors of ADAMTS13 activity. Patients with malignant hypertension had lower ADAMTS13 activity (80%; interquartile range: 53% to 130%) compared with control subjects (99% interquartile range: 82% to 129%; P<0.01) but not compared with patients with severe hypertension (P=0.14). ADAMTS13 activity negatively correlated with lactic dehydrogenase levels after logarithmic transformation (r=-0.65; P<0.001) and was associated with platelet count (r=0.34; P=0.04) and the presence of schistocytes (r=-0.37; P=0.02). Apart from the association with thrombotic microangiopathy, ADAMTS13 was inversely associated with creatinine (r=-0.42; P=0.008). Increasing levels of VWF were associated with a decrease in ADAMTS13 activity (r=-0.34; P=0.03). There was no significant association between ADAMTS13 activity and other parameters, including blood pressure. In conclusion, ADAMTS13 is decreased in malignant hypertension and associated with the severity of thrombotic microangiopathy, likely because of the release of VWF after endothelium stimulation. A severe deficiency could not be demonstrated. More studies are needed to identify the role of ADAMTS13 in the thrombotic microangiopathy and ischemic complications of malignant hypertension.

Renoprotective effects of neuronal NOS-derived nitric oxide and cyclooxygenase-2 metabolites in transgenic rats with inducible malignant hypertension.

The present study was performed to determine the effects of neuronal nitric oxide synthase (nNOS) and cyclooxygenase-2 (COX-2) inhibition on blood pressure and renal hemodynamics in transgenic rats with inducible ANG II-dependent malignant hypertension [strain name: TGR(Cyp1a1Ren2)]. Male Cyp1a1-Ren2 rats (n = 7) were fed a normal diet containing indole-3-carbinol (I3C; 0.3%) for 6-9 days to induce malignant hypertension. Mean arterial pressure (MAP) and renal hemodynamics were assessed in pentobarbital sodium-anesthetized Cyp1a1-Ren2 rats before and during intravenous infusion of the nNOS inhibitor S-methyl-l-thiocitrulline (l-SMTC; 1 mg/h). In hypertensive Cyp1a1-Ren2 rats, l-SMTC increased MAP from 169 +/- 3 to 188 +/- 4 mmHg (P < 0.01), which was a smaller increase than in noninduced rats (124 +/- 9 to 149 +/- 9 mmHg, P < 0.01, n = 5). Additionally, l-SMTC decreased renal plasma flow (RPF) to a similar extent (-34 +/- 13 vs. -35 +/- 12%) in the hypertensive and normotensive rats (4.1 +/- 0.2 to 2.7 +/- 0.5 and 3.1 +/- 0.3 to 2.0 +/- 0.3 ml x min(-1) x g(-1), respectively, P < 0.01) but did not alter glomerular filtration rate (GFR) in either group. In additional experiments, administration of the COX-2 inhibitor, nimesulide (3 mg/kg i.v.), during simultaneous infusion of l-SMTC decreased MAP in both hypertensive and noninduced rats (182 +/- 2 to 170 +/- 3 mmHg and 153 +/- 3 to 140 +/- 3 mmHg, respectively, P < 0.01). Nimesulide also decreased RPF (1.9 +/- 0.2 to 0.8 +/- 0.1 ml x min(-1) x g(-1), P < 0.01) and GFR (0.9 +/- 0.1 to 0.4 +/- 0.1 ml x min(-1) x g(-1), P < 0.01) in hypertensive rats but did not alter RPF or GFR in noninduced rats. The present findings demonstrate that both nNOS-derived NO and COX-2 metabolites exert pronounced renal vasodilator influences in hypertensive Cyp1a1-Ren2 rats. The data also indicate that the renal vasodilator effects of COX-2-derived prostanoids in hypertensive Cyp1a1-Ren2 rats are not dependent on nNOS activity.

Combination therapy with telmisartan and spironolactone alleviates L-NAME exacerbated nephrosclerosis with an increase in PPAR-gamma and decrease in TGF-beta(1).

To investigate whether the receptor blockades of angiotensin II type 1 and aldosterone receptors can prevent renal tissue injury in relation to the renal tissue mRNA levels of peroxisome proliferation-activated receptors-gamma (PPAR-gamma) and transforming growth factor-beta (1) (TGF-beta(1)) in spontaneously hypertensive rats (SHR) given N(G)-nitro-L-arginine methyl ester (L-NAME), which is considered a model of malignant hypertension. This study was performed in 5 groups of 17-week-old male SHR treated for 3 weeks as follows: group 1, control; group 2, L-NAME (50 mg/L in drinking); group 3, L-NAME plus aldosterone antagonist, spironolactone (SPRL, 100 mg/kg/day); group 4, L-NAME plus angiotensin II type 1 receptor blocker, telmisartan (TELM, 3 mg/kg/day); group 5, L-NAME plus combination therapy (COMB) with low-dose TELM (1 mg/kg/day) and SPRL (100 mg/kg/day). Urinary protein excretion and the glomerular injury score were significantly reduced in the SPRL, TELM, and COMB groups as compared with the L-NAME group, while significant blood pressure reduction was observed only in the TELM group. In the TELM and COMB groups, the perivascular cell infiltration and fibrosis area were significantly reduced together with the PPAR-gamma mRNA increase and TGF- beta(1) mRNA decrease. The urinary excretion of nitric oxides was significantly recovered and the wall to lumen ratio of the interlobular artery was significantly reduced only in the COMB group compared with the L-NAME group. Combined administration of 1 mg/kg/day telmisartan and 100 mg/kg/day spironolactone is thought to be effective in alleviating hypertensive renal injuries independently of blood pressure changes. The anti-inflammatory and antifibrotic effects due to PPAR-gamma activation and TGF-beta(1) inhibition may participate in the renoprotection of this combination therapy.

Aldosterone receptor antagonism alleviates proteinuria, but not malignant hypertension, in Cyp1a1-Ren2 transgenic rats.

The contribution of elevated aldosterone to the pathogenesis of malignant, ANG II-dependent hypertension remains uncertain. Therefore, we examined whether chronic mineralocorticoid receptor blockade attenuates the development of malignant hypertension in transgenic rats (TGRs) with inducible expression of the Ren2 gene [TGR(Cyp1a1Ren2)]. Systolic blood pressure (SBP) was measured by radiotelemetry in male TGRs in three groups: 1) control (n = 9), 2) hypertensives (HT; n = 8), and 3) hypertensives + spironolactone (11 mg.kg(-1).day(-1) sc; HTS; n = 8). Malignant hypertension was induced with dietary indole-3-carbinol (0.3%) for 10 days. Metabolic measurements were taken at the beginning of the study and at days 2 and 9. HT exhibited elevated SBP (125 +/- 3 vs. 187 +/- 5 mmHg), plasma renin activity (5 +/- 1 vs. 29 +/- 10 ng ANG I.ml(-1).h(-1)), plasma ANG II (175 +/- 39 vs. 611 +/- 74 fmol/ml), and plasma aldosterone (0.31 +/- 0.04 vs. 5.42 +/- 1.02 nmol/l). Urinary aldosterone excretion increased 5.5-fold by day 2 and an additional 90% by day 9. HT was associated with a 1.8-fold increase in proteinuria by day 9 that was alleviated by treatment with spironolactone (25 +/- 5 vs. 13 +/- 3 mg/day), suggesting that aldosterone contributes to the renal damage observed in malignant hypertension. Urinary Na+ excretion was decreased 76% on day 2, despite a sixfold increase in urinary aldosterone excretion. Decrease in urinary Na+ excretion on day 2 in HT suggests that Na+ reabsorption was increased in response to the increase in aldosterone; however, the lack of a change in SBP between HT and HTS suggests that mechanisms independent of aldosterone stimulation make a greater contribution to the maintenance of elevated arterial pressure in malignant hypertension in Cyp1a1-Ren2 transgenic rats.

Enhanced angiotensin II production by renal mesangium is responsible for apoptosis/proliferation of endothelial and epithelial cells in a model of malignant hypertension.

OBJECTIVE: The systemic renin-angiotensin system (RAS) plays a crucial role in the pathogenesis of malignant hypertension. However, the intrarenal RAS might be at least equally important. We investigated the relationship between intrarenal RAS and mesangial, epithelial and endothelial cell proliferation/apoptosis in a model of malignant hypertension. METHODS: Cultured murine mesangial cells were subjected to 160 mmHg hydrostatic pressure for 1 h. Angiotensin II was assessed by radio-immunoassay (RIA); pro-metalloproteinase-1 (pro-MMP-1) by enzyme-linked immunosorbent assay (ELISA); hydrogen peroxide (H2O2) by photocolorimetric assay, apoptosis by terminal dUTP (2-deoxyuridine 5'-triphosphate) nick-end labelling (TUNEL), p53 by western blot and proliferation by [H]thymidine incorporation, with or without angiotensin II and/or angiotensin II type 1/angiotensin II type 2 (AT-1/AT-2) receptor blockers. Endothelial and epithelial cells were similarly treated, and the same parameters evaluated. Further, untreated cells of both lines were cultured in conditioned medium of mesangial cells exposed to pressure. Their proliferation, apoptosis and angiotensin II production were also assessed. RESULTS: High hydrostatic pressure increased angiotensin II production by mesangial cells, coinciding with augmented apoptosis and proliferation. Co-stimulation with exogenous angiotensin II amplified both effects. Pressure per se evoked no response in endothelial/epithelial cells, while exogenous angiotensin II stimulated proliferation and apoptosis. No augmentation of p53 expression was evident. These effects were abolished by anti-angiotensin-II peptide, saralasine and losartan, but not by PD123319. Incubation of untreated cells in medium of mesangium subjected to pressure, augmented proliferation and apoptosis. No significant changes were noticed in pro-MMP or H2O2. CONCLUSIONS: Mesangium plays a deleterious role in the pathogenesis of malignant hypertension. High hydrostatic pressure stimulates angiotensin II synthesis by mesangial cells. The latter is responsible for hypercellularity and apoptotic death of mesangial, endothelial and epithelial cells. In this model, exaggerated apoptosis and proliferation are mediated via the angiotensin II pathway independently of p53 gene activation.

Genetic clamping of renin gene expression induces hypertension and elevation of intrarenal Ang II levels of graded severity in Cyp1a1-Ren2 transgenic rats.

INTRODUCTION: Transgenic rats with inducible angiotensin II (Ang II)-dependent hypertension (strain name: TGR[Cyp1a1-Ren2]) were generated by inserting the mouse Ren2 renin gene, fused to the cytochrome P450 1a1 (Cyp1a1) promoter, into the genome of the rat. The present study was performed to characterise the changes in plasma and kidney tissue Ang II levels and in renal haemodynamic function in Cyp1a1-Ren2 rats following induction of either slowly developing or malignant hypertension in these transgenic rats. MATERIALS AND METHODS: Arterial blood pressure (BP) and renal haemodynamics and excretory function were measured in pentobarbital sodium-anaesthetised Cyp1a1- Ren2 rats fed a normal diet containing either a low dose (0.15%, w/w for 1415 days) or high dose (0.3%, w/w for 1112 days) of the aryl hydrocarbon indole-3-carbinol (I3C) to induce slowly developing and malignant hypertension, respectively. In parallel experiments, arterial blood samples and kidneys were harvested for measurement of Ang II levels by radioimmunoassay. RESULTS: Dietary I3C increased plasma renin activity (PRA), plasma Ang II levels, and arterial BP in a dose-dependent manner. Induction of different fixed levels of renin gene expression and PRA produced hypertensive phenotypes of varying severity with rats developing either mild or malignant forms of hypertensive disease. Administration of I3C, at a dose of 0.15% (w/w), induced a slowly developing form of hypertension whereas administration of a higher dose (0.3%) induced a more rapidly developing hypertension and the clinical manifestations of malignant hypertension including severe weight loss. Both hypertensive phenotypes were characterised by reduced renal plasma flow, increased filtration fraction, elevated PRA, and increased plasma and intrarenal Ang II levels. These I3C-induced changes in renal haemodynamics, PRA and kidney Ang II levels were more pronounced in Cyp1a1-Ren2 rats with malignant hypertension. Chronic administration of the AT1-receptor antagonist, hypertension, the associated changes in renal haemodynamics, and the augmentation of intrarenal Ang II levels. CONCLUSIONS: Activation of AT1-receptors by Ang II generated as a consequence of induction of the Cyp1a1-Ren2 transgene mediates the increased arterial pressure and the associated reduction of renal haemodynamics and enhancement of intrarenal Ang II levels in hypertensive Cyp1a1-Ren2 transgenic rats.

Long-term administration of rho-kinase inhibitor ameliorates renal damage in malignant hypertensive rats.

We have shown recently that fasudil, a Rho-kinase inhibitor, has renoprotective effects in salt-sensitive hypertensive rats. We hypothesized that activation of Rho-kinase is involved in the pathogenesis of glomerulosclerosis in malignant hypertensive rats. To test this hypothesis, we studied the following 4 groups: control Wistar-Kyoto rats, untreated deoxycorticosterone-acetate salt spontaneously hypertensive rats (DOCA-SHR), low-dose fasudil-treated DOCA-SHR, and high-dose fasudil-treated DOCA-SHR. After 3 weeks of treatment, the effects of fasudil were examined. DOCA-SHR was characterized by increased blood pressure (BP); increased kidney weight; decreased renal function; increased proteinuria; abnormal histological findings; increased monocyte/macrophage infiltration; increased urinary 8-isoprostran levels; increased gene expression of collagen I, collagen III, transforming growth factor-beta, and reduced nicotinamide-adenine dinucleotide phosphate oxidase subunits (p40phox, p47phox, and p67phox); and decreased gene expression of endothelial NO synthase (eNOS) in the renal cortex as compared with Wistar-Kyoto rats. Long-term high-dose fasudil treatment significantly improved renal function and histological findings without changing BP, as compared with untreated DOCA-SHR. Interestingly, long-term fasudil treatment significantly decreased monocyte/macrophage infiltration and urinary 8-isoprostran excretion, in association with decreased mRNA levels of transforming growth factor-beta, collagen I, collagen III, and NADPH oxidase subunits (p40phox, p47phox, and p67phox), and increased mRNA levels of eNOS in the renal cortex. Long-term low-dose fasudil treatment tended to improve these variables slightly but did not affect most of them significantly. Our results suggest that long-term fasudil treatment provides renoprotective effects independent of BP-lowering activity. These renoprotective effects are associated with inhibition of extracellular matrix gene expression, monocyte/macrophage infiltration, oxidative stress, and upregulation of eNOS gene expression.