Exposure to imidacloprid induce oxidative stress, mitochondrial dysfunction, inflammation, apoptosis and mitophagy via NF-kappaB/JNK pathway in grass carp hepatocytes

Imidacloprid (IMI) is a neonicotinoid compound widely used in agriculture production, causing surface water pollution and threatening non-target organisms. The aim of this study was to analyze the effects of IMI on grass carp (Ctenopharyngodon idellus) liver cell (L8824) injury. The L8824 cells were exposed to different doses of IMI (65 mg/L, 130 mg/L and 260 mg/L) for 24 h. Our results demonstrated that exposure IMI significantly suppressed the activity of anti-oxidant enzymes (SOD, CAT and T-AOC) and accumulated oxidase (MDA) levels, and promoting reactive oxygen species (ROS) generation in L8824 cells. Additionally, mitochondrial membrane potential (ΔΨ m), mitochondria-derived ROS and ATP content and the MitoTracker Green indicated that IMI aggravated mitochondrial dysfunction, thereby inducing inflammation and enhancing pro-inflammatory genes (NF-kappaB, TNFα, IL-1β and IL-6) expressions. However, the addition of 2 mM N-acetyl-l-cysteine (NAC) can reverse these adverse effects of high-dose IMI- induced. Hence, ROS is the main factor of IMI-induced mitochondrial dysfunction and inflammation. We further found that exposure to IMI induced apoptosis, which is characterized by promoting release of cytochrome c (Cyt-C), and increasing the expression of Bcl-2-Associated X (BAX), cysteinyl aspartate specific proteinases (Caspase 9 and 3), decreasing Bcl-2 level. Immunofluorescent staining, qRT-PCR and Western Blot results indicated that IMI exposure also activated mitophagy, which was demonstrated by the expression of mitophagy-related genes (BNIP3, LC3B and P62). Conversely, scavenging JNK by SP600125(10 μM) alleviated the expression of mitochondrial apoptosis and mitophagy-related gene induced by high-dose IMI. Therefore, these results of study demonstrated that IMI-induced oxidative stress to regulate mitochondrial dysfunction, thus causing inflammation, mitochondrial apoptosis and mitophagy in grass carp hepatocytes through NF-kappaB/JNK pathway. [Display omitted] •IMI exposure induced cell damage through triggering oxidative stress in grass carp hepatocytes.•IMI caused mitochondrial dysfunction and inflammation by excessive ROS production in grass carp hepatocytes.•IMI caused mitochondrial apoptosis and BNIP3-mediated mitophagy via NF-kappaB/JNK pathway.