Angiotensin AT₂ receptor stimulation inhibits early renal inflammation in renovascular hypertension

Abstract

Angiotensin II type 2 receptor (AT₂R) counteracts most effects of angiotensin II type 1 receptor (AT(1)R). We hypothesized that direct AT₂R stimulation reduces renal production of the inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and transforming growth factor-β1 (TGF-β1) and enhances the production of nitric oxide (NO) and cyclic guanosine 3',5'-monophosphate (cGMP) in the clipped kidney of 2-kidney, 1-clip (2K1C) hypertension rat model. We used Sprague-Dawley rats to evaluate changes in renal interstitial fluid recovery levels of TNF-α, IL-6, NO, and cGMP; renal expression of AT₁R, AT₂R, TGF-β1, TNF-α, and IL-6 in sham and 2K1C rats treated for 4 days with vehicle, AT₂R agonist compound 21 (C21), or AT₂R antagonist PD123319 (PD), alone and combined (n=6, each group). Systolic blood pressure increased significantly in 2K1C and was not influenced by any treatment. Clipped kidneys showed significant increases in renal expression of AT₁R, AT₂R, TNF-α, IL-6, TGF-β1 and decreases in NO and cGMP levels. These factors were not influenced by PD treatment. In contrast, C21 caused significant decrease in renal TNF-α, IL-6, TGF-β1 and an increase in NO and cGMP levels. Combined C21 and PD treatment partially reversed the observed C21 effects. Compared to sham, there were no significant changes in TNF-α, IL-6, TGF-β1, NO, or cGMP in the nonclipped kidneys of 2K1C animals. We conclude that direct AT₂R stimulation reduces early renal inflammatory responses and improves production of NO and cGMP in renovascular hypertension independent of blood pressure reduction.

Figure 1

Renal angiotensin II type 1 (AT₁R) and type 2 (AT₂R) receptor expressions in right (R) and left (L) kidneys of sham and non-clipped (open bars) and clipped (solid bars) kidneys of 2-kidney, 1-clip (2K1C) hypertension rats treated with vehicle, compound 21 (C21), PD123319 (PD), or C21 plus PD. A and C, mRNA levels of AT₁R and AT₂R, respectively. B and D, Western blot analyzes of AT₁R and AT₂R. Top, Representative blots. Bottom, Quantitative results normalized to β-actin. n=5, each group. Data are mean±SEM. *P<0.05 vs sham and corresponding nonclipped kidneys. †P<0.05 vs 2K1C vehicle-treated. ‡P<0.05 vs 2K1C treated with PD.

Figure 2

Renal mRNA expression and renal interstitial fluid (RIF) levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in right (R) and left (L) kidneys of sham and nonclipped (open bars) and clipped (solid bars) kidneys of 2-kidney, 1-clip (2K1C) hypertension rats treated with vehicle, compound 21 (C21), PD123319 (PD), or C21 plus PD. A and C, mRNA levels of TNF-α and IL-6, respectively (n=5). B and D, RIF levels of TNF-α and IL-6 (n=6). Data are mean±SEM. *P<0.05 vs sham and corresponding nonclipped kidneys. †P<0.05 vs 2K1C vehicle-treated. ‡P<0.05 vs 2K1C treated with PD.

Figure 3

Renal expression of transforming growth factor-β1 (TGF-β1) in right (R) and left (L) kidneys of sham and nonclipped (open bars) and clipped (solid bars) kidneys of 2-kidney, 1-clip (2K1C) hypertension rats treated with vehicle, compound 21 (C21), PD123319 (PD), or C21 plus PD. A, mRNA levels of TGF-β1. B, Western blot analyzes of TGF-β1. Top, Representative blots. Bottom, Quantitative results normalized to β-actin. n=5, each group. Data are mean±SEM. *P<0.05 vs sham and corresponding nonclipped kidneys. †P<0.05 vs 2K1C vehicle-treated. ‡P<0.05 vs 2K1C treated with PD.

Figure 4

Renal interstitial fluid (RIF) levels of nitric oxide (NO; A) and cGMP (B) in right (R) and left (L) kidneys of sham and non-clipped (open bars) and clipped (solid bars) kidneys of 2-kidney, 1-clip (2K1C) hypertension rats treated with vehicle, compound 21 (C21), PD123319 (PD), or C21 plus PD. n=6, each group. Data are mean±SEM. *P<0.05 vs sham and corresponding nonclipped kidneys. †P<0.05 vs 2K1C vehicle-treated. ‡P<0.05 vs 2K1C treated with PD.

Figure 5

Hematoxylin and eosin of renal cortex (upper panel) and medulla (lower panel) of sham (A) and 2-kidney, 1-clip hypertension rats treated with vehicle (B), compound 21 (C21; C), PD123319 (PD; D), or combined C21 and PD (E). Inflammatory cell (blue) infiltration was observed in renal cortex and medulla of the vehicle-treated clipped kidney group (B) that was not influenced by PD treatment (D). Cellular infiltration was reduced significantly in rats treated with C21 (C) and slightly decreased in the combined C21 and PD treatment group (×200 magnification).