Increased susceptibility to structural acute kidney injury in a mouse model of presymptomatic cardiomyopathy

Abstract

The early events that signal renal dysfunction in presymptomatic heart failure are unclear. We tested the hypothesis that functional and mechanistic changes occur in the kidney that precede the development of symptomatic heart failure. We employed a transgenic mouse model with cardiomyocyte-specific overexpression of mutant α-B-crystallin that develops slowly progressive cardiomyopathy. Presymptomatic transgenic mice displayed an increase in serum creatinine (1.17 ± 0.34 vs. wild type 0.65 ± 0.16 mg/dl, P < 0.05) and in urinary neutrophil gelatinase-associated lipocalin (NGAL; 278.92 ± 176.24 vs. wild type 49.11 ± 22.79 ng/ml, P < 0.05) but no renal fibrosis. Presymptomatic transgenic mouse kidneys exhibited a twofold upregulation of the Ren1 gene, marked overexpression of renin protein in the tubules, and a worsened response to ischemia-reperfusion injury based on serum creatinine (2.77 ± 0.66 in transgenic mice vs. 2.01 ± 0.58 mg/dl in wild type, P < 0.05), urine NGAL (9,198.79 ± 3,799.52 in transgenic mice vs. 3,252.94 ± 2,420.36 ng/ml in wild type, P < 0.05), tubule dilation score (3.4 ± 0.5 in transgenic mice vs. 2.6 ± 0.5 in wild type, P < 0.05), tubule cast score (3.2 ± 0.4 in transgenic mice vs. 2.5 ± 0.5 in wild type, P < 0.05), and TdT-mediated dUTP nick-end labeling (TUNEL)-positive nuclei (10.1 ± 2.1 in the transgenic group vs. 5.7 ± 1.6 per 100 cells counted in wild type, P < 0.01). Our findings indicate functional renal impairment, urinary biomarker elevations, and induction of renin gene and protein expression in the kidney that occur in early presymptomatic heart failure, which increase the susceptibility to subsequent acute kidney injury.

Keywords: acute kidney injury; cardiorenal syndrome; ischemia-reperfusion injury; neutrophil gelatinase-associated lipocalin; renin.

Fig. 1.

Renal functional and urinary biomarker changes in presymptomatic cardiomyopathy (transgenic). Serum creatinine (A) and urine neutrophil gelatinase-associated lipocalin (NGAL; B) at 10, 15, and 24 wk of life; n = 20 for each of the 2 groups (transgenic and wild type) at each time point. Bars represent means, and the error bars represent SD. *P < 0.05.

Fig. 2.

Renal structural and gene expression changes in presymptomatic cardiomyopathy (transgenic). A, top: staining with hematoxylin and eosin (H&E), showing mild tubule dilation and glomerular congestion in transgenic kidneys. A, bottom: TdT-mediated dUTP nick-end labeling (TUNEL)-positive nuclei. Each picture is representative of n = 10. B: heatmap comparing gene expression profiles of transgenic (presymptomatic cardiomyopathy) kidneys (n = 3), compared with wild-type kidneys (n = 3), showing 6 genes with fold-change >1.5. Red color indicates upregulated genes, and blue color illustrates downregulated genes. Transgenic kidneys displayed upregulation of Ren1, Nphs2, and Clic3 and downregulation of the Arg2, Serpin, and Defb42.

Fig. 3.

Renal renin protein expression changes in presymptomatic cardiomyopathy (transgenic). A: Western blot showing expression of renin protein in whole kidney lysates from wild-type (n = 3) and transgenic (n = 2) animals. B: immunohistochemistry showing renin expression largely restricted to the juxta-glomerular apparatus in wild-type kidneys (red arrowhead, n = 5). C: immunohistochemistry showing renin expression in the juxta-glomerular apparatus (red arrowhead) as well as diffusely in tubule epithelial cells in transgenic animals (n = 5).

Fig. 4.

Increased susceptibility to acute ischemia-reperfusion injury (IRI) based on renal functional measurements and urinary biomarker changes in presymptomatic cardiomyopathy (transgenic). Serum creatinine (A) and urine NGAL (B) at 24 wk of life pre-IRI and 24 h post-IRI. N = 20 for each of the 2 groups (transgenic and wild type) at each time point. Bars represent means, and the error bars represent SD. *P < 0.05.

Fig. 5.

Increased susceptibility to acute IRI based on structural changes in presymptomatic cardiomyopathy (transgenic). A, top: H&E staining 24 h post-IRI. A, bottom: staining for apoptotic nuclei with TUNEL assay 24 h post-IRI. Each picture is representative of n = 10. B: TUNEL-positive nuclei per 100 nuclei examined in an average of five high-power fields are reported. Tubule dilation and tubule cast score were assessed in hematoxylin-eosin stained sections and scored on a scale of 0 to 4. For each of the 2 groups (transgenic and wild type), n = 10. Bars represent means, and the error bars represent SD. *P < 0.05.