6PPD impairs growth performance by inducing intestinal oxidative stress and ferroptosis in zebrafish

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD), a tire-derived pollutant, has gained increasing attention due to its potential toxicity to aquatic organisms. Although previous studies have revealed that 6PPD impacts early developmental stages of larval fish, its effects on adult fish, particularly on key organs, remain unclear. In this study, we observed that adult zebrafish exposed to 6PPD exhibited reduced growth performance and increased fecal output. Histological examination with hematoxylin and eosin (H&E) staining revealed damage to the intestinal villi and a reduction in goblet cell numbers, indicating that 6PPD impairs growth performance by disrupting the digestive system. Comparative transcriptomic analysis revealed that 6PPD caused significant changes in the expression of 727 genes in the intestine, of which 280 genes were up-regulated and 447 genes were down-regulated. These genes were primarily associated with nutrient digestion and absorption, energy metabolism, immune response, and redox regulation. Mechanistically, 6PPD induced oxidative stress and triggered ferroptosis in the intestine, leading to structural damage of the intestinal villi. Treatment with the antioxidant N-acetylcysteine (NAC) alleviated 6PPD-induced oxidative stress and ferroptosis, thereby improving intestinal villi structure and promoting fish growth. This study provides insights into the mechanisms by which 6PPD impairs growth in adult zebrafish and highlights NAC as a potential therapeutic strategy to mitigate its toxicity.