Apoptosis in kidney tissue of senior and geriatric cats with chronic kidney disease

Abstract

Apoptosis, an important pathological event associated with kidney disease progression, is expected to be a therapeutic target in chronic kidney disease (CKD). However, its role in naturally occurring CKD in aged cats remains unclear. Therefore, here, we investigated kidney tissues from aged cats (≥10 years) with or without azotemic CKD to evaluate apoptotic events using a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay. The positive TUNEL signals of the renal cells were quantified and statistically analyzed for correlation with the severity of plasma creatinine (pCre) concentration, renal lesions (glomerulosclerosis, interstitial cell infiltration, peritubular capillaries, and interstitial fibrosis), and oxidative damage of the kidney tissue. Oxidative damage was evaluated using immunohistochemistry for 8-hydroxy-2'-deoxyguanosine (OHdG) and 4-hydroxynonenal (HNE). In the TUNEL assay, regardless of azotemia, positive nuclear signals were observed in the tubular epithelial and intraluminal cells, interstitial infiltrating cells, and glomerular cells. Quantitative TUNEL scores showed no significant differences between the azotemic and non-azotemic groups in any compartment of the kidney tissues. In the azotemic group, TUNEL scores did not correlate with pCre or renal lesion severity. However, the scores showed a significant positive correlation with the scores of 8-OHdG and 4-HNE. These findings suggest that apoptosis associated with oxidative damage in renal tissue is an initial pathological event that leads to CKD, rather than a change following CKD progression, in aged cats. Inhibiting apoptosis by antioxidant treatment may be a key strategy to prevent the development of CKD.

Keywords: aged cat; apoptosis; chronic kidney disease; oxidative damage.

Fig. 1.

Histopathological findings in kidneys from aged cats. (a) The cortex of a non-azotemic cat. (b) The cortex of an azotemic cat with chronic kidney disease. Tubulointerstitial lesions with fibrosis are observed in both cats. Masson’s trichrome stain. Scale bars, 50 µm.

Fig. 2.

Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL)-positive signals in kidneys from azotemic cats with chronic kidney disease. (a) Cortex. (b) Medulla. (c) A glomerulus. (d) Interstitial infiltrated cells. Arrows show TUNEL-positive apoptotic cells. TUNEL assay. Scale bars, 50 µm.

Fig. 3.

Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay and 8-hydroxy-2’-deoxyguanosine (OHdG) immunohistochemistry in the mirror sections. (a and c) TUNEL assay. (b and d) 8-OHdG immunohistochemistry. (a and b) The cortex of a non-azotemic cat. (c and d) The glomeruli of an azotemic cat with chronic kidney disease. The cells labeled with black arrows indicate positive signals for TUNEL but not for 8-OHdG. The cells labeled using black arrowheads indicate positive-signals for 8-OHdG, but not for TUNEL. Scale bars, 50 µm.

Fig. 4.

Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay and CD204 immunohistochemistry in the mirror sections from an azotemic cat with chronic kidney disease. (a) TUNEL assay. (b) CD204 immunohistochemistry. Tubular intraluminal cells (black arrow) indicate positive signals for TUNEL, but not for CD204. Scale bars, 50 µm.

Fig. 5.

Differences in histopathological parameters between the non-azotemic and azotemic chronic kidney disease groups. (a) Glomerulosclerosis. (b) Interstitial cell infiltration. (c) CD34-positive peritubular capillaries. (d) Interstitial fibrosis. Mann–Whitney U test. *: P<0.05, **: P<0.01.

Fig. 6.

Differences in terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) scores between the non-azotemic and azotemic chronic kidney disease groups. (a) TUNEL score for tubular epithelial cells (TECs) in cortex. (b) TUNEL score for TECs in medulla. No significant differences are detected in TUNEL scores. Mann–Whitney U test.

Fig. 7.

Differences in clinicopathological and pathological parameters between the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL)-positive and TUNEL-negative groups in the glomerular cell nuclei. (a) Creatinine. (b) Glomerulosclerosis. (c) Interstitial cell infiltration. (d) CD34-positive peritubular capillaries. (e) Interstitial fibrosis. (f) Cortical 8-hydroxy-2’-deoxyguanosine (OHdG). (g) Medullary 8-OHdG. (h) Glomerular 8-OHdG. (i) Cortical 4-hydroxynonenal (HNE). (j) Medullary 4-HNE. No significant differences are detected in any parameters. Mann–Whitney U test.