Disparate effects of eplerenone, amlodipine and telmisartan on podocyte injury in aldosterone-infused rats

Abstract

Background: Several studies in patients with primary aldosteronism (PA) have suggested that aldosterone (ALD) is directly contributing to albuminuria. However, there are limited data pertaining to the direct role of ALD in in vivo models in regard to the induction of renal injury and the involved mechanisms. In the present study, we established a high-dose ALD-infused rat model to evaluate urinary albumin excretion rate (UAER) and podocyte damage. Moreover, we studied the effect of eplerenone (EPL), telmisartan (TEL) and amlodipine (AML) on ALD-induced renal structural and functional changes.

Methods: Immunohistochemical and real-time PCR analyses, and TUNEL assays were performed to evaluate nephrin expression and podocyte injury.

Results: ALD-receiving rats (ARR) showed a progressive increase in BP, UAER and proteinuria when compared with control rats (CR). Conversely, BP was significantly reduced in ALD + EPL (A/ERR)-, ALD + AML (A/ARR)- and ALD + TEL (A/TRR)-treated rats. However, UAER and proteinuria were decreased only in A/ERR and A/TRR, but not in A/ARR. Only EPL administration provided protection against ALD-induced podocyte apoptosis. Renal tissue of ARR revealed enhanced expression of nephrin protein and mRNA. This effect of ALD was inhibited by EPL, but not by TEL or AML. Conclusions. ALD induces direct glomerular injury independent of its haemodynamic effects; this effect of ALD is, at least in part, mediated through activation of the mineralocorticoid receptor.

Fig. 1

The temporal changes of systolic blood pressure, albuminuria and proteinuria in the different groups at 0, 7, 14, 21 and 28 days. A. Systolic blood pressure measured by tail-cuff plethysmography. B, C. Urinary albumin and proteinuria in the different groups, respectively. All graphs represent mean and SEM. CR, vehicle-treated; ARR, aldosterone-infused; A/ERR, A/ARR and A/TRR eplerenone-, amlodipine- and telmisartan-treated; UAER, urinary albumin excretion rate. a, P < 0.01 vs. CR; b, P < 0.01 vs. ARR; c, P < 0.05 vs. A/ERR at the same time point; d, P < 0.05 vs. 0 day; e, P < 0.05 vs. 14 days.

Fig. 2

Histological findings of glomeruli from different group rats. A–E. Representative photomicrographs of glomerular sections from vehicle-treated controls (CR), aldosterone-infused rats (ARR), and eplerenone (A/ERR)-, amlodipine (A/ARR)- and telmisartan (A/TRR)-treated animals, respectively. F. Result of the semi-quantitative analysis. G. Distribution of glomeruli with different glomerular sclerosis score. Means and SD are given with arbitrary units. a, P < 0.01 vs. CR; b, P < 0.05 vs. ARR; c, P < 0.05 vs. A/ERR (PAS stain, original magnification × 400).

Fig. 3

Representative micrographs of ultrastructural changes in glomeruli under transmission electron microscopy. The foot process in control rats appeared normal (A), while aldosterone infusion caused the lost of arrangement of foot processes and segmental fusion of foot process (black arrow, B), and enriched euchromatin along with shrinkaged nuclear membrane (white arrow) of podocyte was detected in aldosterone-infused rats (C). D–F display the changes of foot process after using eplerenone, amlodipine and telmisartan, respectively (A, B, D, E, and F × 10 000; C × 5000).

Fig. 4

TUNEL assay of glomerular cell apoptosis. A–E display the representative micrographs of TUNEL-stained glomeruli from vehicle-treated controls (CR), aldosterone-infused rats (ARR), and eplerenone (A/ERR)-, amlodipine (A/ARR)- and telmisartan (A/TRR)-treated animals, respectively. Black arrows indicate apoptotic podocytes in aldosterone-infused rats. F. Comparison of apoptosis ratio of podocyte on a glomerular cross section. Values represent means ± SD. a, P < 0.01 vs. CR; b, P < 0.01 vs. ARR; c, P < 0.01 vs. A/ERR (original magnification × 400).

Fig. 5

Immunostaining for nephrin expression in glomeruli from different groups. A–E indicate the representative photomicrographs of nephrin-stained glomeruli from vehicle-treated controls (CR), aldosterone-infused rats (ARR), and eplerenone (A/ERR)-, amlodipine (A/ARR)- and telmisartan (A/TRR)-treated animals, respectively. F. Analysis of the intensity of nephrin staining with mean integrated optical density (IOD) expressed as means ± SD (arbitrary units). a, P < 0.01 vs. CR; b, P < 0.01 vs. ARR (original magnification × 400).

Fig. 6

The mRNA expression of nephrin analysed by real-time PCR. Amplification of β-actin served as a control, and the levels were compared with CR kidneys taken as 100% (n = 6). a, P < 0.01 vs. CR; b, P < 0.01 vs. ARR.