Acute kidney disease in mice is associated with early cardiovascular dysfunction

Abstract

Acute kidney injury (AKI) and chronic kidney disease (CKD) are major health concerns due to their increasing incidence and high mortality. They are interconnected syndromes; AKI without recovery evolves into acute kidney disease (AKD), which can indicate an AKI-to-CKD transition. Both AKI and CKD are associated with a risk of long-term cardiovascular complications, but whether vascular and cardiac dysfunctions can occur as early as the AKD period has not been studied extensively. In a mouse model of kidney injury (KI) with non-recovery, we performed vasoreactivity and echocardiography analyses on days 15 (D15) and 45 (D45) after KI. We determined the concentrations of two major gut-derived protein-bound uremic toxins known to induce cardiovascular toxicity-indoxyl sulfate (IS) and para-cresyl sulfate (PCS)-and the levels of inflammation and contraction markers on D7, D15, and D45. Mice with KI showed acute tubular and interstitial kidney lesions on D7 and D15 and chronic glomerulosclerosis on D45. They showed significant impairment of aorta relaxation and systolic-diastolic heart function, both on D15 and D45. Such dysfunction was associated with downregulation of the expression of two contractile proteins, αSMA and SERCA2a, with a more pronounced effect on D15 than on D45. KI was also followed by a rapid increase in IS and PCS serum concentrations and the expression induction of pro-inflammatory cytokines and endothelial adhesion molecules in serum and cardiovascular tissues. Therefore, these results highlight that AKD leads to early cardiac and vascular dysfunctions. How these dysfunctions could be managed to prevent cardiovascular events deserves further study.

Keywords: Acute kidney injury-to-chronic kidney disease transition; cardiac dysfunction; pro-inflammatory cytokines; protein-bound uremic toxins; vascular dysfunction.

Graphical abstract

Figure 1.

Protocol design detailing the functional explorations and the biological and tissue experiments conducted at D0, D7, D15 and D45.

Figure 2.

Histological kidney lesions in the glomerular (A, B), tubular (C, D), and interstitial (E, F) compartments of mice with KI on D7, D15, and D45 after KI. Representative images (G-I) (scale bar = 100 µm) with Masson trichrome (1, 2, and 4) and Periodic acid-Schiff (3) staining. Data are expressed as the mean ± SEM (n = 4). *p < 0.05 vs. Sham, **p < 0.01 vs. Sham, §p < 0.05, §§p < 0.01. Abbreviations: KI, kidney injury.

Figure 3.

Vascular reactivity in the KI and Sham groups on D15 and D45 after KI. Vascular reactivity in the KI and Sham groups on D15 (A, B) and D45 (C, D) after KI and comparison of the difference in relaxation (Δ relaxation Sham-KI) between the two groups on D15 and D45 (E). Changes in ESAo by echocardiography on D15 and D45 after KI (F). Data are expressed as the mean ± SEM for vascular reactivity and data are shown as box and whisker plot for ESAo (n = 14). *p < 0.05 vs. Sham, **p < 0.01 vs. Sham, ***p < 0.001 vs. Sham, ****p < 0.0001 vs. Sham. Abbreviations: Ach, acetylcholine; ESAo, systolic expansion rate of the aorta; KI, kidney injury.

Figure 4.

Echocardiography measurements in the KI and Sham groups on D15 and D45 after KI. A-F. Changes in echocardiographic parameters: FS (A), IVRT (B), Tei index (C), and LV mass (D) on D15 and D45 after KI. Data are shown as box and whisker plots (n = 7). *p < 0.05 vs. Sham, §p < 0.05. Abbreviations: Ach, acetylcholine; FS, fractional shortening; IVRT, isovolumic relaxation time; LV, left ventricular; KI, kidney injury.

Figure 5.

mRNA levels of pro-inflammatory cytokines in aortas and left ventricles on D7, D15 and D45 after KI. mRNA levels (fold-change relative to the Sham group on D7) for IL-1β (A, B), IL-6 (C, D), and TNFα (E, F) in aortic tissue (A, C, and E) and the left ventricle (B, D and F) on D7, D15, and D45 after KI. Data are shown as box and whisker plots (n = 7). **p < 0.01 vs. Sham, ****p < 0.0001 vs. Sham, §§p < 0.01, §§§§p < 0.0001. Protein expression of the different cardiac and aortic inflammatory markers are shown in the supplementary materials. Abbreviations: IL, interleukin; TNF, tumor necrosis factor; KI, kidney injury.

Figure 6.

Immunostaining-based quantification and representative images for ICAM-1 (A, B) and VCAM-1 (C, D) in the aorta on D7, D15, and D45 after KI (magnification = 40× and scale bar = 25 µm). Data are shown as box and whisker plots (n = 7). *p < 0.05 vs. Sham, ****p < 0.0001 vs. Sham, §§§p < 0.001, §§§§p < 0.0001. Abbreviations: ICAM-1, intercellular adhesion molecule 1; VCAM-1, vascular cell adhesion molecule 1; KI, kidney injury.

Figure 7.

mRNA levels of endothelial adhesion molecules in aortas and left ventricles on D7, D15 and D45 after KI. mRNA levels (fold-change relative to Sham D7) for ICAM-1 (A) and VCAM-1 (B) in the aorta on D7, D15, and D45 following KI. Data are shown as box and whisker plots (n = 7). **p < 0.01 vs. Sham, §§p < 0.01, §§§p < 0.001. Protein expression of the different cardiac and aortic endothelial markers are shown in the supplementary materials. Abbreviations: ICAM-1, intercellular adhesion molecule 1; VCAM-1, vascular cell adhesion molecule 1; KI, kidney injury.

Figure 8.

mRNA levels and Western-blot analysis of aortic αSMA (A, C, E, and G) and heart SERCA2a (B, D, F, and H) on D7, D15, and D45 after KI. A-B. mRNA levels (fold-change difference relative to Sham D7) for αSMA (A) and SERCA2a (B) in the aorta and heart on D7, D15, and D45 after KI. C-H. Western-blot analysis of aortic αSMA (C, E, and G) and heart SERCA2a (D, F, and H) levels on D7, D15, and D45 after KI. GAPDH was used as an internal control. Data are shown as box and whisker plots for mRNA levels (n = 7) and expressed as the mean ± SEM for Western-blot analysis (n = 3 independent mice in each group). *p < 0.05 vs. Sham; ****p < 0.0001 vs. Sham, §§§p < 0.001, §§§§p < 0.0001. Abbreviations: αSMA, alpha-smooth muscle actin; SERCA2a, sarco/endoplasmic reticulum Ca²+-ATPase 2a; KI, kidney injury.