Dewan S. A. Majid

Involvement of Tumor Necrosis Factor-α in Angiotensin II–Mediated Effects on Salt Appetite, Hypertension, and Cardiac Hypertrophy

Hypertension is considered a low-grade inflammatory condition induced by various proinflammatory cytokines, including tumor necrosis factor (TNF)-α. Recent studies have implicated an involvement of TNF-α in the development of salt-sensitive hypertension induced by angiotensin II (Ang II). To understand further the relationship between TNF-α and Ang II, we examined the responses to Ang II in TNF-α knockout (TNF-α−/−) mice in the present study. A continuous infusion of Ang II (1 μg/kg per minute) for 2 weeks was given to both TNF-α−/− and wild-type (WT) mice with implanted osmotic minipumps. Daily measurement of water intake, salt intake, and urine output were performed using metabolic cages. Blood pressure was monitored continuously with implanted radiotelemetry. Ang II administration for 2 weeks caused increases in salt (0.2±0.07 to 5.6±0.95 mL/d) and water (5.4±0.34 to 11.5±1.2 mL/d) intake and in mean arterial pressure (115±1 to 151±3 mm Hg) in wild-type mice, but these responses were absent in TNF-α−/− mice (0.2±0.04 to 0.3±0.09 mL/d, 5.5±0.2 to 6.1±0.07 mL/d, and 113±2 to 123±3 mm Hg, respectively). Cardiac hypertrophy induced by Ang II was significantly attenuated in TNF-α−/− mice compared with wild-type mice. In a group of TNF-α−/− mice, when replacement therapy was made with recombinant TNF-α, Ang II induced similar responses in salt appetite, mean arterial pressure, and cardiac hypertrophy, as observed in wild-type mice. These results suggest that TNF-α plays a mechanistic role in mediating chronic Ang II–induced effects on salt appetite and blood pressure, as well as on cardiac hypertrophy.

Nitric oxide in the control of renal hemodynamics and excretory function

Experimental evidence has now been amassed to indicate that inhibition of nitric oxide (NO) synthase reduces total or regional renal blood flow by approximately 25 to 30% and markedly increases the renal vascular resistance, demonstrating that basal release of NO helps to maintain the relatively low vascular resistance that is characteristic for the kidney. It has been demonstrated that intraarterial administration of NO synthase inhibitors causes marked reductions in sodium excretion without changes in filtered load and suppressed the arterial pressure-induced natriuretic responses in the kidney. We also demonstrated that a constant rate infusion of a NO donor in dogs pretreated with a NOS inhibitor resulted in increases in sodium excretion but failed to restore the slope of the relation between arterial pressure and sodium excretion, suggesting that an alteration in intrarenal NO production rate during changes in arterial pressure is involved in the mediation of pressure natriuresis. Further experiments in dogs performed in our laboratory have confirmed that there is a direct relationship between changes in arterial pressure and intrarenal NO activity measured using NO-sensitive microelectrodes in the renal tissue. These arterial pressure-induced changes in intrarenal NO activity were seen positively correlated with the changes in urinary excretion rates of sodium. Collectively, these data suggest that acute changes in arterial pressure alter intrarenal NO production, which inhibits tubular sodium reabsorption to manifest the phenomenon of pressure natriuresis.

Chronic NF-κB blockade reduces cytosolic and mitochondrial oxidative stress and attenuates renal injury and hypertension in SHR

Nuclear factor-kappaB (NF-kappaB) plays an important role in hypertensive renal injury; however, its roles in perpetuating mitochondrial oxidative stress and renal dysfunction remain unclear. In this study, we assessed the effects of chronic NF-kappaB blockade with pyrrolidine dithiocarbamate (PDTC) on renal dysfunction and mitochondrial redox status in spontaneously hypertensive rats (SHR). PDTC (150 mg.kg body wt(-1).day(-1)) or vehicle was administered orally to 8-wk-old SHR and their respective controls for 15 wk. Systolic blood pressure (SBP) was measured by tail-cuff plethysmography at the start of and at every third week throughout the study. After 15 wk of treatment, anesthetized rats underwent acute renal experiments to determine renal blood flow and glomerular filtration rate using PAH and inulin clearance techniques, respectively. Following renal experiments, kidneys were excised from killed rats, and cortical mitochondria were isolated for reactive oxygen species (ROS) measurements using electron paramagnetic resonance. Tissue mRNA and protein levels of NF-kappaB and oxidative stress genes were determined using real-time PCR and immunofluorescence or Western blotting, respectively. PDTC treatment partially attenuated the increase in SBP (196.4 +/- 9.76 vs. 151.4 +/- 2.12; P < 0.05) and normalized renal hemodynamic and excretory parameters and ATP production rates in SHR. PDTC treatment also attenuated the higher levels of cytosolic and mitochondrial ROS generation and tissue mRNA and protein expression levels of NF-kappaB and oxidative stress genes in SHR without any comparable responses in control rats. These findings suggest that NF-kappaB activation by ROS induces the cytosolic and mitochondrial oxidative stress and tissue injury that contribute to renal dysfunction observed in SHR.

Mechanisms mediating pressure natriuresis: what we know and what we need to find out.

1. It is well established that pressure natriuresis plays a key role in long‐term blood pressure regulation, but our understanding of the mechanisms underlying this process is incomplete.

Relation Between Renal Interstitial ATP Concentrations and Autoregulation-Mediated Changes in Renal Vascular Resistance

The present study was performed to examine the hypothesis that autoregulation-related changes in renal vascular resistance (RVR) are mediated by extracellular ATP. By use of a microdialysis method, renal interstitial concentrations of ATP and adenosine were measured at different renal arterial pressures (RAPs) within the autoregulatory range in anesthetized dogs (n=12). RAP was reduced in steps from the ambient pressure (131+/-4 mm Hg) to 105+/-3 mm Hg (step 1) and 80+/-2 mm Hg (step 2). Renal blood flow and glomerular filtration rate exhibited efficient autoregulation in response to these changes in RAP. RVR decreased by 22+/-2% in step 1 (P<0.01) and 38+/-3% in step 2 (P<0.01). The control renal interstitial concentration of ATP was 6.51+/-0.71 nmol/L and decreased to 4. 51+/-0.55 nmol/L in step 1 (P<0.01) and 2.77+/-0.47 nmol/L in step 2 (P<0.01). In contrast, the adenosine concentrations (117+/-6 nmol/L) were not altered significantly. Changes in ATP levels were highly correlated with changes in RVR (r=0.88, P<0.0001). Further studies demonstrated that stimulation of the tubuloglomerular feedback (TGF) mechanism by increasing distal volume delivery elicited with acetazolamide also led to increases in renal interstitial ATP concentrations, whereas furosemide, which is known to block TGF responses, reduced renal interstitial fluid ATP concentrations. The data demonstrate a positive relation between renal interstitial fluid ATP concentrations and both autoregulation- and TGF-dependent changes in RVR and thus support the hypothesis that changes in extracellular ATP contribute to the RVR adjustments responsible for the mechanism of renal autoregulation.

Renal responses to intra-arterial administration of nitric oxide donor in dogs.

Inhibition of nitric oxide synthesis by intra-arterial administration of nitro-L-arginine (NLA) leads to attenuation of the slope of the relation between renal arterial pressure (RAP) and sodium excretion without an alteration in renal autoregulatory efficiency. In the present study, we examined whether only the presence of nitric oxide or, alternatively, changes in nitric oxide production during changes in RAP are required for pressure natriuresis to occur. Anesthetized sodium-replete dogs (n = 8) were treated with NLA (50 micrograms.kg-1 x min-1) to inhibit endogenous nitric oxide formation, and S-nitroso-n-acetylpenicillamine (SNAP) was infused intra-arterially at a constant rate (2 micrograms.kg-1 x min-1) to replenish intrarenal nitric oxide levels. Renal responses to reductions in RAP within the autoregulatory range were assessed before and during NLA infusion followed by SNAP+NLA infusion. As reported previously, NLA infusion alone increased renal vascular resistance and decreased renal blood flow, urine flow, sodium excretion, and fractional excretion of sodium, with no change in glomerular filtration rate. Autoregulatory efficiency remained intact, whereas the pressure-induced natriuretic responses were attenuated. During SNAP+NLA infusion, renal blood flow increased from 2.8 +/- 0.3 to 3.5 +/- 0.3 mL.min-1 x g-1 (P < .001), without significant changes in glomerular filtration rate (0.75 +/- 0.07 to 0.81 +/- 0.05 mL.min-1 x g-1); the autoregulatory efficiency of renal blood flow and glomerular filtration rate remained intact. SNAP increased urine flow (4.8 +/- 1.8 to 10.0 +/- 2.5 microL.min-1 x g-1), sodium excretion (0.63 +/- 0.26 to 1.70 +/- 0.37 mumol.min-1 x g-1), and fractional excretion of sodium (0.55 +/- 0.20% to 1.38 +/- 0.27%).(ABSTRACT TRUNCATED AT 250 WORDS)

Renal Interstitial ATP Responses to Changes in Arterial Pressure During Alterations in Tubuloglomerular Feedback Activity

We recently demonstrated a direct relationship between autoregulation-related changes in renal vascular resistance (RVR) and renal interstitial ATP concentrations. To assess the possible role for extracellular ATP in the regulation of tubuloglomerular feedback (TGF)-mediated autoregulatory adjustments in RVR, renal interstitial ATP concentrations were measured with microdialysis probes in anesthetized dogs at different renal arterial pressures (RAPs) within the autoregulatory range during augmented and diminished activity of the TGF mechanism. Stepwise reductions in RAP from ambient pressure (129±3 mm Hg) to 102±2 mm Hg (step 1) and 75±1 mm Hg (step 2) resulted in significant decreases in ATP concentrations from 9.0±0.8 to 6.3±0.6 nmol/L in step 1 and to 4.2±0.5 nmol/L in step 2. Changes in RVR were highly correlated with changes in ATP concentrations (r =0.86, P <0.001, n=12). Acetazolamide (100 &mgr;g · kg−1 · min−1, n=6), which increases solute delivery to the macula densa, thus augmenting TGF activity, significantly decreased renal blood flow (RBF) by −16±2% and glomerular filtration rate (GFR) by −22±4% and increased ATP concentrations from 8.4±0.7 to 15.5±1.4 nmol/L. Although basal RBF and GFR levels were reduced by the acetazolamide infusion, autoregulation efficiency was maintained, and interstitial ATP concentrations were significantly decreased in response to reductions in RAP by −36±4% in step 1 and by −54±2% in step 2. The relationship between changes in RVR and interstitial ATP concentrations was preserved during acetazolamide treatment (r =0.80, P <0.01). Inhibition of the TGF mechanism by furosemide significantly increased RBF by 33±6% and GFR by 13±2% and decreased ATP concentrations from 8.9±1.4 to 5.0±0.8 nmol/L (n=6). Furosemide caused marked impairment of RBF and GFR autoregulatory efficiency (by −14±3% and −11±3% in step 1 and by −26±2% and −18±4% in step 2, respectively). In the furosemide-treated kidneys, interstitial ATP levels remained low and were not altered during reductions in RAP (4.7±0.7 nmol/L in step 1 and 4.7±0.8 nmol/L in step 2), and changes in RVR did not exhibit a correlation with changes in ATP concentrations (r =0.22, P =0.30). These data support the hypothesis that extracellular ATP contributes to autoregulatory adjustments in RVR that are mediated by changes in activity of the TGF mechanism.

Nitric oxide and superoxide interactions in the kidney and their implication in the development of salt-sensitive hypertension.

1 Enhanced superoxide () activity as a result of the inhibition of the superoxide dismutase (SOD) enzyme results in vasoconstrictor and antinatriuretic responses in the canine kidney; these responses were shown to be greatly enhanced during inhibition of nitric oxide synthase (NOS). Glomerular filtration rate remained mostly unchanged during SOD inhibition in the intact nitric oxide (NO) condition, but was markedly reduced during NOS inhibition. These findings indicate that endogenous NO has a major renoprotective effect against by acting as an anti‐oxidant. Nitric oxide synthase inhibition was also shown to enhance endogenous activity. 2 Experiments in our laboratory using dogs, rats and gene knockout mice have shown that renal vasoconstrictor and antinatriuretic responses to acute or chronic angiotensin (Ang) II administration are mediated, in part, by generation. In the absence of NO, enhanced activity largely contributes to AngII‐induced renal tubular sodium reabsorption. Acute or chronic treatment with the scavenger tempol in experimental models of hypertension (induced by chronic low‐dose treatment with AngII and NO inhibitors) causes an improvement in renal haemodynamics and in excretory function, abolishes salt sensitivity and reduces blood pressure. 3 The present mini review also discusses related studies from many other laboratories implicating a role for and its interaction with NO in the development of salt‐sensitive hypertension. 4 Overall, the collective data support the hypothesis that an imbalance between the production of NO and in the kidney primarily determines the condition of oxidative stress that alters renal haemodynamics and excretory function leading to sodium retention and, thus, contributes to the development of salt‐sensitive hypertension.

Renoprotective effects of nitric oxide in angiotensin II-induced hypertension in the rat

Experiments were performed in anesthetized male Sprague-Dawley rats to determine whether increased nitric oxide (NO) activity during the development of hypertension exerts a protective effect on renal cortical blood flow (CBF) and medullary blood flow (MBF). The effects of acute NO synthase inhibition on renal function and on CBF and MBF, measured by laser-Doppler flow probes, were evaluated in control and ANG II-infused hypertensive rats, prepared by the infusion of ANG II at a rate of 65 ng/min via osmotic minipumps implanted subcutaneously for 13 days. In normotensive rats (n = 8), intravenous infusion of N omega-nitro-L-arginine (NLA; 20 micrograms.100 g-1.min-1) decreased CBF by 21 +/- 4% and MBF by 49 +/- 8% and increased blood pressure from 118 +/- 1 to 140 +/- 2 mmHg. In ANG II-infused rats (n = 7), CBF and MBF decreased by 46 +/- 5% and 25 +/- 6%, respectively, during infusion of NLA. Arterial pressure increased from 160 +/- 5 to 197 +/- 7 mmHg, which was a greater absolute increase than in normotensive controls. Basal renal blood flow (RBF), estimated from p-aminohippurate clearance and hematocrit, was similar in both the control (6.0 +/- 0.5 ml.min-1.g-1) and hypertensive (6.0 +/- 0.6 ml.min-1.g-1) rats. However, NLA-induced reductions in RBF averaged 60 +/- 5% in the hypertensive rats, compared with 31 +/- 9% observed in control rats. GFR in control (0.97 +/- 0.03 ml.min-1.g-1) and hypertensive rats (0.78 +/- 0.12 ml.min-1.g-1) decreased to a similar extent during the first 30-min period of NLA infusion. GFR returned toward control levels in control rats; in contrast, GFR remained significantly decreased in the ANG II-infused rats (0.58 +/- 0.11 ml.min-1.g-1). Basal urinary sodium excretion (0.2 +/- 0.08 mueq.min-1.g-1), fractional excretion of sodium (0.3 +/- 0.13%), and urine flow (4.9 +/- 0.39 microliters.min-1.g-1) in hypertensive rats did not increase significantly after NLA treatment as occurred in normotensive controls. These data suggest that a compensatory increase in nitric oxide activity partially counteracts the vasoconstrictor influence of elevated ANG II levels to regulate renal hemodynamics and maintain cortical perfusion in the renal circulation.

Superoxide Contributes to Development of Salt Sensitivity and Hypertension Induced by Nitric Oxide Deficiency

This study was performed to examine the role of superoxide (O2−) in the development of salt sensitivity and hypertension induced by inhibition of nitric oxide (NO) generation. Male Sprague-Dawley rats were fed with diet containing either normal salt (NS) (0.4% NaCl) or high salt (HS) (4% NaCl). These rats were treated with or without an NO synthase inhibitor, nitro-l-arginine methylester (l-NAME) (15 mg/kg/d) and O2− scavenger, tempol (30 mg/kg per day) in the drinking water for 4 weeks. Systolic blood pressure (SBP) was measured by tail-cuff plethysmography and urine collection was performed during the course of experimental periods. At the end of 4 weeks, l-NAME treatment resulted in greater increases in SBP in HS rats (127±2 to 172±3 mm Hg; n=8) than in NS rats (130±2 to 156±2 mm Hg; n=9). Co-administration of tempol with l-NAME markedly attenuated these SBP responses to a similar level in both HS (128±3 to 147±2 mm Hg; n=8) and NS rats (126±2 to 142±3 mm Hg; n=8). Urinary 8-isoprostane excretion (UIsoV) increased in response to l-NAME treatment that was higher in HS (10.6±0.5 to 21.5±0.8 ng/d) than in NS rats (10.8±0.7 to 16.9±0.6 ng/d). Co-treatment with tempol completely abolished these UIsoV responses to l-NAME in both HS and NS rats but did not alter urinary H2O2 excretion rate. The decreases in urinary nitrate/nitrite excretion in response to l-NAME treatment were not altered by co-administration of tempol in both HS and NS rats. These data suggest that enhancement of O2− activity during NO inhibition contributes to the development of salt sensitivity that is associated with NO-deficient hypertension.