Radiocontrast Agent Diatrizoic Acid Induces Mitophagy and Oxidative Stress via Calcium Dysregulation

• Published in: International journal of molecular sciences 2019-08, Vol.20 (17), p.4074
• Main Authors: Ward, Dakota B, Brown, Kathleen C, Valentovic, Monica A
• Format: Article
• Language: English
• Subjects: Acidification; Acute Kidney Injury - metabolism; Apoptosis - drug effects; ATP synthase; Blotting, Western; Calcium; Calcium - metabolism; Carbonyl compounds; Carbonyls; Cell Line; Cell membranes; Cell Survival - drug effects; Cell viability; Contrast Media - adverse effects; contrast-induced acute kidney injury; Cyanides; Cytotoxicity; Diatrizoate - adverse effects; diatrizoic acid; Electron transport chain; Endoplasmic reticulum; Endoplasmic Reticulum Stress - drug effects; HK-2 cells; Homeostasis; Humans; Mitochondria; Mitochondrial Membranes - drug effects; Mitochondrial Membranes - metabolism; mitophagy; Mitophagy - drug effects; Oligomycin; Oxidative phosphorylation; Oxidative stress; Oxidative Stress - drug effects; Oxygen consumption; Phosphorylation; Protein folding; proximal tubule cytotoxicity; Rotenone; Seahorse XFe; Superoxide Dismutase - metabolism
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms20174074

Contrast-induced acute kidney injury (CI-AKI) is the third most common cause of hospital associated kidney damage. Potential mechanisms of CI-AKI may involve diminished renal hemodynamics, inflammatory responses, and direct cytotoxicity. The hypothesis for this study is that diatrizoic acid (DA) induces direct cytotoxicity to human proximal tubule (HK-2) cells via calcium dysregulation, mitochondrial dysfunction, and oxidative stress. HK-2 cells were exposed to 0-30 mg I/mL DA or vehicle for 2-24 h. Conversion of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and trypan blue exclusion indicated a decrease in mitochondrial and cell viability within 2 and 24 h, respectively. Mitochondrial dysfunction was apparent within 8 h post exposure to 15 mg I/mL DA as shown by Seahorse XF cell mito and Glycolysis Stress tests. Mitophagy was increased at 8 h by 15 mg I/mL DA as confirmed by elevated LC3BII/I expression ratio. HK-2 cells pretreated with calcium level modulators BAPTA-AM, EGTA, or 2-aminophenyl borinate abrogated DA-induced mitochondrial damage. DA increased oxidative stress biomarkers of protein carbonylation and 4-hydroxynonenol (4HNE) adduct formation. Caspase 3 and 12 activation was induced by DA compared to vehicle at 24 h. These studies indicate that clinically relevant concentrations of DA impair HK-2 cells by dysregulating calcium, inducing mitochondrial turnover and oxidative stress, and activating apoptosis.