Urotensin II in the development and progression of chronic kidney disease following ⅚ nephrectomy in the rat

Figures

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  Figure 1. Pharmacokinetic profile of SB‐611812. Concentration of SB‐611812 in rat blood, after administration p.o. at 5 mg kg−1. The concentration peaked at 1643 nm 6 h after dosing. Data are shown as means ± SD from n = 4 rats

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  Figure 2. Collagen IV α1 (Col4a1; a), laminin‐β1 (Lamb1; b) and fibronectin 1 (Fn1; c) mRNA expression normalized to Ywhaz in the renal cortex of rats after subtotal nephrectomy (SNx), as chronic kidney disease progresses. Glomerular basement membrane marker expression is shown at 8 weeks postsurgery [early (E)] and at end‐stage renal failure [late (L)]. Data are shown as a scatter plot with median; statistical comparisons were by two‐way ANOVA of log10‐transformed data followed by Tukey's post hoc test; *** P < 0.001 SNx‐L versus sham‐L

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  Figure 3. Immunolocalization of urotensin system proteins in the renal cortex as chronic kidney disease progresses. Urotensin II (UII; a–d) and urotensin receptor (UT; e–h) proteins in the cortex of sham and SNx rats during the early and late stages of renal disease. Urotensin II immunoreactivity was generally diffuse and localized to the cytoplasm of cells in the proximal tubules (P), with some staining also seen in the distal tubules (D) in the sham groups (a and b). Similar staining was seen in the early SNx group (c). In the late SNx group (d), staining was seen in the tubular epithelia (arrows), with further diffuse staining within the glomeruli and in areas of peritubular hypercellularity (red star). Diffuse UT immunoreactivity was observed in the sham groups (e and f) and in the early SNx group (g). Staining in the late SNx cohort (h) was particularly abundant in areas of peritubular hypercellularity (arrowheads). (i–p) Representative negative controls (primary antibody omitted) across each group. All images are representative and were captured at ×40 magnification with scale bars representing 50 μm (d, f, n), ×20 magnification with scale bars representing 100 μm (a–c, e, h; i–m, p)…

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  Figure 4. Immunolocalization of urotensin system proteins in the renal medulla as chronic kidney disease progresses. Urotensin II (a–d) and UT (e–h) proteins in the medulla of sham and SNx rats during the early and late stages of renal disease. Typically, UII immunoreactivity was localized within the cytoplasm of collecting duct epithelial cells of sham‐E (a) and sham‐L rats (b). In the SNx‐E (c) and SNx‐L (d) groups, the staining was localized in a similar manner, with additional faint staining visible in the surrounding interstitium. Expression of UT was localized within the collecting ducts (arrows). Expression of UT in SNx‐E (g) was comparable with that in sham‐E animals (e). In the SNx‐L group (h), the staining was localized in a pattern similar to that of the time‐matched shams (f) but with a pronounced apical distribution, and additional faint staining was visible in the surrounding cells. (i–p) Representative negative controls (primary antibody omitted) across each group. All images are representative and were captured at ×40 magnification with scale bars representing 50 μm (e–h, n, o), ×20 magnification with scale bars representing 100 μm (a–d, i–m) or ×10 magnification with scale…

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  Figure 5. Systolic blood pressure (SBP; a) and urinary albumin:creatinine ratio (uACR; b) during subchronic treatment of SNx and sham rats with the UT antagonist SB‐611812 (30 mg kg−1 day−1) or vehicle. Data are shown as means ± SD; in panel (b), where downward error bars have negative values these have been omitted because data are plotted on a logarithmic scale. Statistical comparisons were by three‐way ANOVA with repeated measures; * P < 0.05, *** P < 0.001 SNx‐V versus sham‐V; ## P < 0.01, ### P < 0.001 SNx‐A versus sham‐A

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  Figure 6. Collagen IV α1 (Col4a1; a), laminin‐β1 (Lamb1; b) and fibronectin 1 (Fn1; c) mRNA expression normalized to B2m in the renal cortex of SNx and sham rats after subchronic treatment with the UT antagonist SB‐611812 (30 mg kg−1 day−1) or vehicle. Compared with the SNx‐V group, 13 weeks of UT antagonist treatment in SNx rats had no effect on Col4a1 (P = 0.49), Lamb1 (P > 0.99) or Fn1 (P = 0.83) mRNA expression. Data are shown as a scatter plot with median; statistical comparisons were by two‐way ANOVA of log10‐transformed data followed by Tukey's post hoc test

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  Figure 7. Immunolocalization of urotensin system proteins in the renal cortex of SNx and sham rats after subchronic treatment with the UT antagonist SB‐611812 (30 mg kg−1 day−1) or vehicle. Urotensin II (a–d) and UT (e–h) proteins in the cortex of sham and SNx rats receiving the UT antagonist or vehicle. Urotensin II immunoreactivity was diffuse in both sham (a, b) and SNx (c, d) groups, with the most prominent staining localized to the proximal tubules (P) (compare with distal tubules (D) and glomeruli (G)). There was limited immunostaining for UT in cortical sections from both sham groups (e, f). In SNx rats receiving vehicle (g) or UT antagonist (h), staining was observed primarily within areas of hypercellular expansion (arrowheads) rather than within intact cortical structures; some additional staining was seen in the parietal layer (arrows). (i–p) Representative negative controls (primary antibody omitted) across each group. All images are representative and were captured at ×20 magnification with scale bars representing 100 μm

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  Figure 8. Immunolocalization of urotensin system proteins in the renal medulla of SNx and sham rats after subchronic treatment with the UT antagonist SB‐611812 (30 mg kg−1 day−1) or vehicle. Urotensin II (a–d) and UT (e–h) proteins in the medulla of sham and SNx rats receiving the UT antagonist or vehicle. In the medulla of sham rats receiving vehicle (sham‐V; a) or UT antagonist (sham‐A; b), immunohistochemical staining for urotensin II was localized primarily within the collecting duct epithelium (arrows, panels a–d). This pattern was also seen in SNx rats receiving vehicle (SNx‐V; c) or UT antagonist (SNx‐A; d). Expression of the UT receptor was consistent across sham‐V (e), sham‐A (f), SNx‐V (g) and SNx‐A (h) groups. Staining was most obvious in the collecting ducts (arrowheads, all panels); however, staining was also seen in the thin limbs of the loop of Henle (arrows in panels e and h). (i–p) Representative negative controls (primary antibody omitted) across each group. All images are representative and were captured at ×20 magnification with scale bars representing 100 μm