The Role of Endothelin System in Renal Structure and Function during the Postnatal Development of the Rat Kidney.

Renal development in rodents, unlike in humans, continues during early postnatal period. We aimed to evaluate whether the pharmacological inhibition of Endothelin system during this period affects renal development, both at structural and functional level in male and female rats. Newborn rats were treated orally from postnatal day 1 to 20 with vehicle or bosentan (Actelion, 20 mg/kg/day), a dual endothelin receptor antagonist (ERA). The animals were divided in 4 groups: control males, control females, ERA males and ERA females. At day 21, we evaluated renal function, determined the glomerular number by a maceration method and by morphometric analysis and evaluated possible structural renal alterations by three methods: 〈alpha〉-Smooth muscle actin (α-SMA) immunohistochemistry, Masson's trichrome and Sirius red staining. The pharmacological inhibition of Endothelin system with a dual ERA during the early postnatal period of the rat did not leads to renal damage in the kidneys of male and female rats. However, ERA administration decreased the number of glomeruli, the juxtamedullary filtration surface area and the glomerular filtration rate and increased the proteinuria. These effects could predispose to hypertension or renal diseases in the adulthood. On the other hand, these effects were more pronounced in male rats, suggesting that there are sex differences that could be greater later in life. These results provide evidence that Endothelin has an important role in rat renal postnatal development. However these results do not imply that the same could happen in humans, since human renal development is complete at birth

Sirius red staining of the renal cortex.

<p>Representative images of Sirius red staining of the renal cortex; CA: cortical area; JA: juxtamedullary area. A = control males (CA); B = ERA-treated males (CA); C = control females (CA); D = ERA-treated females (CA); E = control males (JA); F = ERA-treated males (JA); G = control females (JA); H = ERA-treated females (JA). ERA: Endothelin receptor antagonist. Total magnification: 400x.</p

Morphometric parameters in juxtamedullary and cortical renal tissue.

<p>Morphometric parameters in juxtamedullary and cortical renal tissue.</p

Renal functional parameters.

<p>Renal functional parameters.</p

Renal histochemical evaluation by Sirius red and Masson´s thrichrome.

<p>Renal histochemical evaluation by Sirius red and Masson´s thrichrome.</p

Number of total glomeruli.

<p>Number of total glomeruli obtained by the maceration method. ERA: Endothelin receptor antagonist. Values are mean ± SEM (n = 10). Data were analyzed using two way ANOVA followed by Bonferroni posttest. Two-way ANOVA showed a statistically significant interaction (p < 0.05) between the effects of ERA treatment and sex. *p<0.05 vs control males.</p

Masson´s trichrome staining of the renal cortex.

<p>Representative images of Masson´s trichrome staining of the renal cortex; CA: cortical area; JA: juxtamedullary area. A = control males (CA); B = ERA-treated males (CA); C = control females (CA); D = ERA-treated females (CA); E = control males (JA); F = ERA-treated males (JA); G = control females (JA); H = ERA-treated females (JA). ERA: Endothelin receptor antagonist. Total magnification: 400x.</p

Histological structure of the renal cortex.

<p>Representative images of Hematoxylin-eosin staining of the renal cortex. A = control males; B = ERA-treated males; C = control females; D = ERA-treated females. ERA: Endothelin receptor antagonist. Total magnification: 400x.</p

A: Proteinuria (mg/24 h/100g). B: Creatinine Clearance (ml/min/100g).

<p>ERA: Endothelin receptor antagonist. Values are mean ± SEM. Data were analyzed using two way ANOVA followed by Bonferroni posttest.</p

Renal response to L-arginine in diabetic rats. A possible link between nitric oxide system and aquaporin-2.

The aim of this study was to evaluate whether L-Arginine (L-Arg) supplementation modifies nitric oxide (NO) system and consequently aquaporin-2 (AQP2) expression in the renal outer medulla of streptozotocin-diabetic rats at an early time point after induction of diabetes. Male Wistar rats were divided in four groups: Control, Diabetic, Diabetic treated with L-Arginine and Control treated with L-Arginine. Nitric oxide synthase (NOS) activity was estimated by [14C] L-citrulline production in homogenates of the renal outer medulla and by NADPH-diaphorase staining in renal outer medullary tubules. Western blot was used to detect the expression of AQP2 and NOS types I and III; real time PCR was used to quantify AQP2 mRNA. The expression of both NOS isoforms, NOS I and NOS III, was decreased in the renal outer medulla of diabetic rats and L-Arg failed to prevent these decreases. However, L-Arg improved NO production, NADPH-diaphorase activity in collecting ducts and other tubular structures, and NOS activity in renal homogenates from diabetic rats. AQP2 protein and mRNA were decreased in the renal outer medulla of diabetic rats and L-Arg administration prevented these decreases. These results suggest that the decreased NOS activity in collecting ducts of the renal outer medulla may cause, at least in part, the decreased expression of AQP2 in this model of diabetes and constitute additional evidence supporting a role for NO in contributing to renal water reabsorption through the modulation of AQP2 expression in this pathological condition. However, we cannot discard that another pathway different from NOS also exists that links L-Arg to AQP2 expression