Background: Mitochondrial dysfunction leads to excess production of reactive oxygen species (ROS) in renal tubular epithelial cells contributing to inflammation, fibrosis and progression of chronic kidney disease (CKD). Superoxide Dismutase 1 (SOD1), an antioxidative enzyme, and microtubule-associated protein 1A/1B light chain 3 (LC3), a marker of autophagy, are cellular defense mechanisms regulating ROS. Hypothesis: Autophagy and antioxidant mechanisms are reduced in progressive CKD. Animals: Snap-frozen and formalin-fixed paraffin-embedded (FFPE) kidney tissue from healthy cats (> 9 years) and those with stable and progressive CKD (n=7/group) euthanized for health reasons.
Methods: LC3 activation (LC3ii/LC3i) and SOD1 protein expression were semi-quantified by Western blot, normalizing to the loading control (β-actin) in kidney lysates. LC3 and SOD1 localization was assessed by immunohistochemistry using FFPE sections. Kruskal-Wallis with Dunn correction was used to determine significance between groups.
Results: Relative SOD1 protein expression was lower in progressive (median=0.54 [0.33, 0.83]) compared to stable (median=1.15 [0.97, 1.17], p=0.013) and control cats (median=1.49 [1.30, 1.75], p< 0.001); Figure 1). LC3 activation was not significantly different between the control (median=2.05 [1.95, 3.69]) and stable (median=1.94 [1.71, 2.56], p=0.32) group but those with progressive CKD (median=1.68 [1.36, 2.36]) trended towards reduced activation compared to control (p=0.06) group. LC3 and SOD1 (Figure 2) were localized to tubular epithelial cells. Conclusions and clinical importance: The prominent SOD1 antioxidant response in healthy tubular epithelial cells is diminished in CKD, with lower levels in progressive compared to stable cats. Better understanding factors limiting antioxidant defense in progressive CKD may provide novel therapeutic options.
