Marcela Pandolfo

Controversy about COOPERATE ABPM Trial Data

mm Hg, 63% between 120 and 150 mm Hg, and the remaining 19% more than 150 mm Hg. A similar incidence was observed among treatment groups. Interestingly, the high variability of the systolic BP values was more characteristic among patients who were able to effectively restrict their salt intake. As compared with the systolic values, the nighttime values were constant throughout the measurements, partly because the drugs were taken at night or bedtime. Naoyuki Nakao, MD, PhD Division of Nephrology Rokko Island Hospital Koh-Yoh Cho Naka 2-11 Higashinada, Kobe, Japan

Different protective actions of losartan and tempol on the renal inflammatory response to acute sodium overload

The aim of this work was to study the role of local intrarenal angiotensin II (Ang II) and the oxidative stress in the up‐regulation of pro‐inflammatory cytokines expression observed in rats submitted to an acute sodium overload. Sprague–Dawley rats were infused for 2 h with isotonic saline solution (Control group) and with hypertonic saline solution alone (Na group), plus the AT1 receptor antagonist losartan (10 mg kg−1 in bolus) (Na–Los group), or plus the superoxide dismutase mimetic tempol (0.5 mg min−1 kg−1) (Na–Temp group). Mean arterial pressure, glomerular filtration rate, and fractional sodium excretion (FENa) were measured. Ang II, NF‐κB, hypoxia inducible factor‐1α (HIF‐1α), transforming growth factor β1 (TGF‐β1), smooth muscle actin (α‐SMA), endothelial nitric oxide synthase (eNOS), and RANTES renal expression was evaluated by immunohistochemistry. Ang II, NF‐κB, and TGF‐β1 and RANTES early inflammatory markers were overexpressed in Na group, accompanied by enhanced HIF‐1α immunostaining, lower eNOS expression, and unmodified α‐SMA. Losartan and tempol increased FENa in sodium overload group. Although losartan reduced Ang II and NF‐κB staining and increased eNOS expression, it did not restore HIF‐1α expression and did not prevent inflammation. Conversely, tempol increased eNOS and natriuresis, restored HIF‐1α expression, and prevented inflammation. Early inflammatory markers observed in rats with acute sodium overload is associated with the imbalance between HIF‐1α and eNOS expression. While both losartan and tempol increased natriuresis and eNOS expression, only tempol was effective in restoring HIF‐1α expression and down‐regulating TGF‐β1 and RANTES expression. The protective role of tempol, but not of losartan, in the inflammatory response may be associated with its greater antioxidant effects. J. Cell. Physiol. 224:41–48, 2010

Renal Protective Role of Atrial Natriuretic Peptide in Acute Sodium Overload-Induced Inflammatory Response

Background: The present study was performed to explore the effect of exogenous infusions of atrial natriuretic peptide (ANP) on the early inflammatory response during acute sodium overload in normal rats. Methods: Sprague Dawley rats were exposed to acute sodium overload (Na 1.5 M). Nonhypotensive doses of ANP (1 and 5 µg ·kg–1 ·h–1) were infused simultaneously with sodium or after sodium infusion in order to evaluate prevention or reversion of the inflammatory response, respectively. We determined inflammation markers in renal tissue by immunohistochemistry. Results: Creatinine clearance was not reduced in any case. Sodium tubular reabsorption increased after sodium overload (334.3 ± 18.7 vs. control 209.6 ± 27.0 mEq·min–1, p < 0.05) without changes in mean arterial pressure. This increase was prevented (228.9 ± 26.4; p < 0.05) and reversed (231.5 ± 13.9; p < 0.05) by ANP-5 µg ·kg–1 ·h–1. Sodium overload increased the expression of: RANTES (38.4.3 ± 0.8 vs. 2.9 ± 0.6%, p < 0.001), transforming-growth-factor-β1 (35.3 ± 1.0 vs. 5.0 ± 0.7%, p < 0.001), α-smooth muscle actin (15.6 ± 0.7 vs. 3.1 ± 0.3%, p < 0.001), NF-ĸB (9.4 ± 1.3 to 2.2 ± 0.5 cells/mm2, p < 0.001), HIF-1α (38.2 ± 1.7 to 8.4 ± 0.8 cells/mm2, p < 0.001) and angiotensin II (35.9 ± 1.3 to 8.2 ± 0.5%, p < 0.001). ANP-5 µg ·kg–1 ·h–1 prevented and reversed inflammation: RANTES (9.2 ± 0.5 and 6.9 ± 0.7, p < 0.001); transforming growth factor-β1 (13.2 ± 0.7 and 10.2 ± 0.5, p < 0.001) and α-smooth muscle actin (4.1 ± 0.4 and 5.2 ± 0.4, p < 0.001). Both prevention and reversion by ANP were associated with downregulation of NF-ĸB (3.2 ± 0.4 and 2.8 ± 0.5, p < 0.001) and angiotensin II (8.2 ± 0.5 and 9.1 ± 0.7, p < 0.001) and diminished hypoxia evaluated through HIF-1α expression (8.4 ± 0.8 and 8.8 ± 0.7, p < 0.001). Conclusion: Our study provides evidence supporting a protective role of ANP in both prevention and reversion of renal inflammation in rats with acute sodium overload.

High-sodium diet promotes a profibrogenic reaction in normal rat kidneys: effects of Tempol administration

BACKGROUND Studies carried out in vitro have recently shown that salt loading induces an increasing mechanical stretch and a flow-induced superoxide production in the thick ascending limb of Henles loop. In this regard, we hypothesized that the oxidative stress induced by salt overload could stimulate inflammatory and fibrogenic signaling pathways in normal rats. METHODS Sprague Dawley rats were fed with an 8% NaCl high- (HS) or 0.4% NaCl normal-salt (NS) diet for 3 weeks, with or without Tempol (T) administration (1 mM, administered in drinking water). Mean arterial pressure (MAP), glomerular filtration rate (GFR) and urinary sodium excretion (UVNa) were measured. NAD(P)H oxidase p47phox, angiotensin II (Ang II), transforming growth factor ß1 (TGF-ß1), a-smooth muscle actin (a-SMA) and nuclear factor-kappa B (NF-kB) expression were evaluated in renal tissues by immunohistochemistry. RESULTS A high NaCl diet produced a slight but significant increase in MAP and enhanced UVNa and oxidative stress. Administration of a high NaCl diet induced the overexpression of TGF-ß1, a-SMA and NF-?B in cortex and medulla, while Ang II increased in proximal convoluted tubules, and decreased in cortical collecting ducts. Tempol administration prevented these changes and simultaneously normalized MAP accompanied by an enhancement in GFR and UVNa. CONCLUSION The results showed that a high NaCl diet is able to produce a renal profibrotic response also in normal rats, which could be associated with oxidative stress rather than intrarenal Ang II expression.

Angiotensin II Increases Intrarenal Transforming Growth Factor-β1 in Rats Submitted to Sodium Overload Independently of Blood Pressure

Angiotensin II (Ang II) promotes sodium-retention, cell growth and fibrosis in addition to its classical effects on blood pressure and fluid homeostasis. In this study we examined whether low and non-hypertensive doses of exogenous Ang II could enhance the intrarenal expression of transforming growth factor-β1 (TGF-β1) observed in rats submitted to sodium overload. Sprague-Dawley-rats were infused for 2 h with 0.1 and 5 μg kg−1 h−1 Ang II (Ang 0.1 and Ang 5, respectively) together with saline solution at four different concentrations (isotonic and Na 0.5 mol L−1, Na 1.0 mol L−1 and Na 1.5 mol L−1). Renal function and mean arterial blood pressure (BP) were measured. The renal distributions of TGF-β1, α-smooth-muscle-actin (α-SMA) and nuclear factor-κB (NF-κB) were evaluated by immunohistochemistry. While the Ang 0.1 groups were normotensive, the Ang 5 groups developed arterial hypertension progressively, and the highest blood pressure values were observed when rats were simultaneously infused with Na 1.5 mol L−1. Glomerular function was not altered in any group. In cortical tubules, all groups infused with Ang II (0.1 and 5) and hypertonic saline solution (HSS) showed an increase in TGF-β1 immunostaining compared to those infused with HSS alone. In medullary tubules, only the Ang 5–Na 0.5 group showed a significant increase in TGF-β 1 immunostaining compared to the Na 0.5 group. Peritubular positive staining for α-SMA was present in groups receiving Ang alone or Ang-Na, in a sodium concentration–dependent manner. In cortical-tubules, NF-κB immunostaining was significantly increased in the Ang groups in comparison with the control and in Ang–Na 0.5 and Ang–Na 1.0 groups in comparison with the Na 0.5 mol L−1 and Na 1.5 mol L−1 groups, respectively, except in the case of the Ang 0.1–Na 1.5 mol L−1 and Ang 5–Na 1.5 mol L−1 groups. Moreover, Ang II and sodium overload induced additional changes in TGF-β1, α-SMA and NF-κB immunostanding in glomeruli, medullary tubules and renal vessels. In conclusion, the interaction of Ang II with acute-sodium overload exacerbated intrarenal TGF-β1, α-SMA and NF-κB expression, independently from changes in blood pressure levels, in normal rats.

Salt-induced downregulation of renal aquaporins is prevented by losartan.

AIMS The purpose of this study was to investigate the expression of aquaporin-1 (AQP-1) and aquaporin-2 (AQP-2) in the renal tubule of rats fed with a high-salt diet and its modulation by the AT1 receptor blocker losartan. MAIN METHODS The experiments were performed in four groups of rats fed for 3 weeks with the following diets: regular rat chow (NS); high-salt (8% NaCl) chow (HS), NS plus losartan (NS-L) and HS plus losartan (HS-L). Losartan (40 mg x kg(-1)) was administered in the drinking water. Systolic blood pressure (SBP) and renal function were evaluated. The intrarenal levels of angiotensin II (Ang II), TGF-β(1), α-smooth muscle actin (α-SMA), endothelial nitric oxide synthase (eNOS), AQP-1 and AQP-2 were determined by immunohistochemistry. AQP-1 and AQP-2 protein levels were measured by western blot analysis. KEY FINDINGS A high-sodium diet downregulated AQP-1 and AQP-2 expression levels in the proximal tubule and collecting duct, respectively. The high-sodium diet also induced Ang II, TGF-β(1) and α-SMA overexpression and decreased eNOS expression in the renal cortex and medulla. Losartan increased the diuresis and natriuresis, favoring urinary sodium concentration. Additionally, losartan prevented the profibrogenic response, decreasing Ang II, TGF-β(1) and α-SMA levels and normalizing AQP-2 expression in the HS-L group. AQP-1 expression was upregulated by losartan in both the NS-L and HS-L groups. SIGNIFICANCE These results show that increased intrarenal Ang II in rats fed with a high-salt diet downregulates renal AQP-1 and AQP-2 expressions. In addition, although losartan increased diuresis and natriuresis, it prevented the downregulation of aquaporins, favoring urinary sodium concentration.

Atrial Natriuretic Peptide Stimulates Dopamine Tubular Transport by Organic Cation Transporters: A Novel Mechanism to Enhance Renal Sodium Excretion.

The aim of this study was to demonstrate the effects of atrial natriuretic peptide (ANP) on organic cation transporters (OCTs) expression and activity, and its consequences on dopamine urinary levels, Na+, K+-ATPase activity and renal function. Male Sprague Dawley rats were infused with isotonic saline solution during 120 minutes and randomized in nine different groups: control, pargyline plus tolcapone (P+T), ANP, dopamine (DA), D-22, DA+D-22, ANP+D-22, ANP+DA and ANP+DA+D-22. Renal functional parameters were determined and urinary dopamine concentration was quantified by HPLC. Expression of OCTs and D1-receptor in membrane preparations from renal cortex tissues were determined by western blot and Na+, K+-ATPase activity was determined using in vitro enzyme assay. 3H-DA renal uptake was determined in vitro. Compared to P+T group, ANP and dopamine infusion increased diuresis, urinary sodium and dopamine excretion significantly. These effects were more pronounced in ANP+DA group and reversed by OCTs blockade by D-22, demonstrating that OCTs are implied in ANP stimulated-DA uptake and transport in renal tissues. The activity of Na+, K+-ATPase exhibited a similar fashion when it was measured in the same experimental groups. Although OCTs and D1-receptor protein expression were not modified by ANP, OCTs-dependent-dopamine tubular uptake was increased by ANP through activation of NPR-A receptor and protein kinase G as signaling pathway. This effect was reflected by an increase in urinary dopamine excretion, natriuresis, diuresis and decreased Na+, K+-ATPase activity. OCTs represent a novel target that links the activity of ANP and dopamine together in a common mechanism to enhance their natriuretic and diuretic effects.

Serum free light chains and oligoclonal bands in patients with multiple myeloma and autologous stem cell transplantation.

Abstract Background: To establish stringent complete remission (SCR) in patients with multiple myeloma (MM), it is currently recommended to obtain a normal serum free light chains (sFLC) ratio. The appearance of serum oligoclonal bands (OB) after autologous stem cell transplantation (ASCT) is considered a favorable prognostic factor. The objective of this study was to examine sFLC for assessing SCR in patients with MM, and ASCT with OB. We also examined how capillary electrophoresis (CE) compares with agarose gel electrophoresis (Aga) in identifying oligoclonal bands. Methods: Out of 238 patients studied in our institution between April 1992 and December 2008 a serum protein electrophoresis (SPE) was performed by means of CE and sFLC determination on 37 patients with MM in complete remission (CR), ASCT and OB presence were assigned by conventional Aga electrophoresis and IF. Results: Statistically significant differences (SSD) were found when comparing CE vs. Aga, regarding BO visualization in SPE, favoring the latter. In connection with sFLC, the group of patients with an abnormal ratio presented elevated values in the γ-globulin zone of the SPE, whereas the group of patients with a normal ratio of sFLC presented with normal values resulting in SSD between the groups. Conclusions: It is essential to perform immunofixation to certify the presence of OB, especially if CE is used as it is difficult to distinguish them using this method. A normal sFLC was observed in most of the patients with OB and normal values of the SPE γ-globulin zone. The above-mentioned information might demonstrate a limitation of sFLC test in SCR evaluation for patients with MM, ASCT and CR if OB has been detected.

Sodium Load Combined with Low Doses of Exogenous Angiotensin II Upregulate Intrarenal Angiotensin II

Background/Aims: The present study was designed to evaluate the effects of a salt load combined with exogenous low nonhypertensive angiotensin II (Ang II) doses on Ang II intrarenal regulation. Methods: Sprague-Dawley rats were infused with Ang II nonhypertensive doses (0.1 μg·kg–1·h–1 and 5 μg·kg–1·h–1) and saline overload (Na 0.5 M, Na 1.0 M and Na 1.5 M) for 2 h (0.04 ml·min–1). Sodium tubular reabsorption, sodium urinary excretion and mean arterial pressure (MAP) were measured. Ang II was evaluated in the kidneys by immunohistochemistry. Results: Ang II levels in glomeruli and vessels were exacerbated when sodium load and Ang II were given simultaneously, independently of MAP elevation. In tubules, Ang II staining in the presence of sodium overload was greater in the Ang 0.1 groups than in the Ang 5 groups. Compared with the controls, sodium tubular reabsorption rose in the Ang 0.1-Na 0.5 and Ang 0.1-Na 1 groups and sodium urinary excretion decreased in the Ang 5-Na 0.5 and Ang 5-Na 1 groups. MAP increased in the Ang 5-Na 1 and Ang 5-Na 1.5 groups. Conclusion: We conclude that local renal Ang II levels were upregulated when acute sodium overload and nonhypertensive Ang II doses were administered simultaneously in normal rats, independently from blood pressure and glomerular function changes.

Losartan prevents the imbalance between renal dopaminergic and renin angiotensin systems induced by fructose overload. l-Dopa/dopamine index as new potential biomarker of renal dysfunction

BACKGROUND The renin angiotensin system (RAS) and the renal dopaminergic system (RDS) act as autocrine and paracrine systems to regulate renal sodium management and inflammation and their alterations have been associated to hypertension and renal damage. Nearly 30-50% of hypertensive patients have insulin resistance (IR), with a strong correlation between hyperinsulinemia and microalbuminuria. OBJECTIVE The aim of this study was to demonstrate the existence of an imbalance between RAS and RDS associated to IR, hypertension and kidney damage induced by fructose overload (FO), as well as to establish their prevention, by pharmacological inhibition of RAS with losartan. MATERIALS/METHODS Ninety-six male Sprague-Dawley rats were randomly divided into four groups and studied at 4, 8 and 12 weeks: control group (C4, C8 and C12; tap water to drink); fructose-overloaded group (F4, F8 and F12; 10% w/v fructose solution to drink); losartan-treated control (L) group (L4, L8 and L12; losartan 30 mg/kg/day, in drinking water); and fructose-overloaded plus losartan group (F + L4, F + L8 and F + L12, in fructose solution). RESULTS FO induced metabolic and hemodynamic alterations as well as an imbalance between RAS and RDS, characterized by increased renal angiotensin II levels and AT1R overexpression, reduced urinary excretion of dopamine, increased excretion of l-dopa (increased l-dopa/dopamine index) and down-regulation of D1R and tubular dopamine transporters OCT-2, OCT-N1 and total OCTNs. This imbalance was accompanied by an overexpression of renal tubular Na+, K+-ATPase, pro-inflammatory (NF-kB, TNF-α, IL-6) and pro-fibrotic (TGF-β1 and collagen) markers and by renal damage (microalbuminuria and reduced nephrin expression). Losartan prevented the metabolic and hemodynamic alterations induced by FO from week 4. Increased urinary l-dopa/dopamine index and decreased D1R renal expression associated to FO were also prevented by losartan since week 4. The same pattern was observed for renal expression of OCTs/OCTNs, Na+, K+-ATPase, pro-inflammatory and pro-fibrotic markers from week 8. The appearance of microalbuminuria and reduced nephrin expression was prevented by losartan at week 12. CONCLUSION The results of this study provide new insight regarding the mechanisms by which a pro-hypertensive and pro-inflammatory system, such as RAS, downregulates another anti-hypertensive and anti-inflammatory system such as RDS. Additionally, we propose the use of l-dopa/dopamine index as a biochemical marker of renal dysfunction in conditions characterized by sodium retention, IR and/or hypertension, and as a predictor of response to treatment and follow-up of these processes.