Exposure to Trimethyltin Chloride Induces Pyroptosis and Immune Dysfunction in Grass Carp CIK Cells by Activating the NF-κB Pathway Through Oxidative Stress.

Trimethyltin chloride (TMT) is a highly toxic organotin pollutant frequently found in aquatic environments, posing a significant threat to the ecological system. The kidney plays a vital role in the body's detoxification processes, and TMT present in the environment tends to accumulate in the kidneys. However, it remained unclear whether exposure to different doses of TMT could induce pyroptosis and immune dysfunction in grass carp kidney cells (CIK cells). For this purpose, after assessing the half-maximal inhibitory concentration (IC50) of TMT on CIK cells, we established a model for exposure of CIK cells at varying concentrations of TMT. CIK cells were treated with various doses of TMT (2.5, 5, 10 µM) for 24 h. Oxidative stress levels were measured using kits and fluorescence methods, whereas the expression of related genes was verified through western blot and quantitative real-time PCR (qRT-PCR). The results indicated that TMT exposure led to oxidative stress, with increased levels of ROS, H2O2, MDA, and GSH, and inhibited activities of T-AOC, SOD, and CAT. It activated the NF-κB pathway, leading to the upregulation of NF-κB p65, NF-κB p50, GSDMD, NLRP3, ASC, and Caspase-1. Furthermore, TMT exposure also resulted in increased expression of cytokines (IL-18, IL-6, IL-2, IL-1ß, and TNF-α) and decreased expression of antimicrobial peptides (LEAP2, HEPC, and ß-defensin). In summary, exposure to TMT induces dose-dependent oxidative stress that activates the NF-κB pathway, leading to pyroptosis and immune dysfunction in grass carp CIK cells.

Glyphosate induces autophagy in hepatic L8824 cell line through NO-mediated activation of RAS/RAF/MEK/ERK signaling pathway and energy metabolism disorders.

Glyphosate is an herbicide commonly used worldwide, and its substantial use causes widespread pollution with runoff. However, research on glyphosate toxicity has mostly remained at the embryonic level and existing studies are limited. In the present study, we investigated whether glyphosate can induce autophagy in hepatic L8824 cells by regulating energy metabolism and rat sarcoma (RAS)/rapidly accelerated fibrosarcoma (RAF)/mitogen-activated extracellular signal-regulated kinase (MEK)/extracellular regulated protein kinases (ERK) signaling by activating nitric oxide (NO). First, we selected 0, 50, 200, and 500 µg/mL as the challenge doses, according to the inhibitory concentration of 50% (IC50) of glyphosate. The results showed that glyphosate exposure increased the enzyme activity of inducible nitric oxide synthase (iNOS), which in turn increased the NO content. The activity and expression of enzymes related to energy metabolism, such as hexokinase (HK)1, HK2, phosphofructokinase (PFK), phosphokinase (PK), succinate dehydrogenase (SDH), and nicotinamide adenine dinucleotide with hydrogen (NADH), were inhibited, and the RAS/RAF/MEK/ERK signaling pathway was activated. This led to the negative expression of mammalian target of rapamycin (mTOR) and P62 in hepatic L8824 cells and the activation of the autophagy marker genes microtubule-associated proteins light chain 3 (LC3) and Beclin1 to induce autophagy. The above results were dependent on glyphosate concentration. To verify whether autophagy can be excited by the RAS/RAF/MEK/ERK signaling pathway, we treated L8824 cells with the ERK inhibitor U0126 and found that the autophagy gene LC3 was reduced due to the inhibition of ERK, thus demonstrating the reliability of the results. In conclusion, our results demonstrate that glyphosate can induce autophagy in hepatic L8824 cells by activating NO, thus regulating energy metabolism and the RAS/RAF/MEK/ERK signaling pathway.