Abstract Body: Background: N-(1,3-dimethylbutyl)-N’-phenyl-p-phenylenediamine quinone (6-PPDq), an ozone-derived product of the tire antioxidant 6-PPD, is an emerging airborne toxicant. Although human exposure likely occurs at ng-to-µg levels, its cardiopulmonary effects remain poorly understood.

Hypothesis/Aim: We hypothesized that 6-PPDq disrupts cardiopulmonary homeostasis by impairing mitochondrial function. We investigated effects on cell viability, mitochondrial respiration, and iPSC-derived cardiomyocyte contractility.

Methods: Lung tissue from COPD patients and healthy controls was analyzed for 6-PPDq by LC-MS/MS. A549 epithelial cells, airway smooth muscle (ASM) cells, and AC16 cardiomyocytes were studied in 2D/3D culture. Cytotoxicity was assessed by MTT after 6-PPDq exposure (1–1000 ng/mL). Mitochondrial function and membrane potential were measured using Seahorse XF Cell Mito Stress Test and JC-10 dye. Cell injury and barrier integrity were evaluated by LDH release and TEER. Cardiomyocyte contractility was measured using IonOptix, and mitochondrial fusion/fission proteins were analyzed by immunoblotting after 24 h.

Results: 6-PPDq levels were ~2-fold higher in COPD lungs than controls. In vitro, 6-PPDq reduced viability in A549 and ASM cells after 48 h and in AC16 cells within 24 h (p<0.001). It reduced spare respiratory capacity at 24 h and membrane potential at 12 h (p<0.05), increased LDH release (p<0.01), and caused a dose-dependent decline in TEER. In 3D spheroids, 6-PPDq altered morphology and reduced viability. It also disrupted mitochondrial fusion/fission protein expression in A549 and AC16 cells and reduced contractility in iPSC-derived cardiomyocytes (p<0.05).

Conclusion: 6-PPDq accumulates in lung tissue and disrupts mitochondrial bioenergetics, impairs barrier integrity, alters mitochondrial dynamics, and reduces contractility, highlighting its cardiopulmonary toxicity and the need for risk assessment.