Investigation of Tartaric Acid Enantiomer‐Specific Nephrotoxicity and Citrate Synthase Inhibition in Madin–Darby Canine Kidney Cells In Vitro

OBJECTIVE: To investigate and compare the enantiomer-specific nephrotoxicity of tartaric acid (TTA) in Madin-Darby canine kidney (MDCK) cells and human kidney (HK-2) cells. Secondary aims were to determine the enantiomeric-specific TTA accumulation to induce these effects through distinct influx and efflux pathways via organic anion transporter (OAT)-1 and OAT-4. Subsequently, a critical objective was to determine the mitochondrial mechanisms of toxicity via citrate synthase inhibition and assess supplemental substrate mitigation within this reaction. DESIGN: In vitro concentration-response experiments and transporter transfection studies using racemic and enantiomeric forms of TTA in canine and human immortalized renal cell lines over a 48-h exposure period. SETTING: School of Pharmacy biomedical research laboratory and tissue culture facility. ANIMALS/SAMPLES: Immortalized MDCK and HK-2 proximal tubule epithelial cell lines. INTERVENTIONS: MDCK and HK-2 cells were treated with 5-50 mmol/L of racemic or enantiomeric TTA for 48 h. Cotreatment of 0.08 mmol/L oxaloacetate (OAA) was used to assess the mitigation of citrate synthase inhibition. Additional transporter transfection experiments were conducted using probenecid (PBD) as a cotreatment to gain a greater understanding of TTA accumulation and toxicity prevention. MEASUREMENTS AND MAIN RESULTS: Cellular ATP levels and citrate synthase activity were measured. L-Tartaric acid (L-TTA) caused significant ATP depletion (8.5%-24.8%) and citrate synthase inhibition (36.5%-45.0%) in MDCK cells but not in HK-2 cells, suggesting species-specific toxicity. OAT-4 transfection reduced ATP loss by 8.0%-11.2% after L-TTA exposure. Cotreatment with PBD reduced cytotoxic L-TTA effects by 11.7%, suggesting that OAT-1 uptake and insufficient OAT-4 efflux are central to TTA accumulation. Cotreatment with OAA rescued ATP loss by 15.6%. CONCLUSIONS: L-TTA is a species- and enantiomer-specific nephrotoxicant in dogs resulting from a lack of transporter-mediated efflux and subsequent citrate synthase inhibition. OAT-1 inhibition or OAT-4 transfection mitigates toxicity, supporting a transporter-driven mechanism for TTA accumulation.