Perfluorononanoic acid (PFNA), a persistent long-chain per- and polyfluoroalkyl substance (PFAS), is increasingly recognized as a potential metabolic disruptor; however, the cellular and molecular mechanisms underlying its obesogenic activity remain poorly understood. In the present study, we investigated the effects of low nanomolar PFNA concentrations on 3T3-L1 adipocyte differentiation and function, with particular emphasis on the exposure timing during adipogenesis process and the molecular mechanisms underlying PFNA-induced adipose dysfunction. PFNA exposure during the proliferative preadipocyte phase, more than during differentiation period, induced a persistent senescence-like phenotype in fully differentiated adipocytes, characterized by senescence-associated-β- galactosidase activity and an increase inthe p53/p21 checkpoint axis. These effects were accompanied by activation of the unfolded protein response (UPR), as indicated by upregulation of ER stress-associated markers. Early PFNA exposure, also promoted adipogenesis and pro-inflammatory signaling. Interestingly, pharmacological inhibition with tauroursodeoxycholic acid (TUDCA) markedly prevented PFNA-induced cellular dysfunction, identifying ER stress as a key upstream mediator linking PFNA exposure to downstream senescence-like, oxidative, adipogenic and inflammatory alterations. Collectively, these findings identify the proliferative preadipocyte phase as a particularly sensitive window for low-concentration PFNA exposure and suggest that PFNA-induced senescence and ER stress represent interconnected mechanisms contributing to persistent adipocyte dysfunction and its potential obesogenic effects.
Mammalian kidneys, although exposed to a diverse array of environmental pollutants and commonly used drugs that are dose-limited by nephrotoxicity, have a significant but limited capacity for repair and regeneration. Primary cultures of human proximal tubular (hPT) cells were used to investigate the cellular response to exposures to one of two environmental toxicants (mercuric chloride [HgCl2] or S-(1,2-dichlorovinyl)-L-cysteine [DCVC; key metabolite of trichloroethylene] or one of three nephrotoxic drugs (cisplatin [CDDP], polymyxin B [PmxB], tenofovir disoproxil fumarate [TDF]) at concentrations that cause no to modest cellular injury. Cellular phenotype was first characterized by measuring protein expression of key drug metabolism enzymes and plasma membrane transporters. To enable identification of biomarkers when the hPT cells are still capable of repair and regeneration, release of Kidney Injury Molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) from exposed hPT cells was determined. Results validated that for each of the 3 concentrations of the 5 toxicants or drugs tested at the 3 time points, there was either no significant pattern of or modest cellular injury. Untargeted metabolomics was conducted to identify high resolution mass spectrometry features representing metabolites that differ after exposures. Pathway analysis was then conducted to map these metabolites into altered pathways. Prostaglandin metabolism from arachidonate, multiple fatty acid metabolism pathways, several amino acid metabolism pathways, and pentose phosphate and related vitamin metabolism pathways were the most common pathways altered by all 5 chemicals. Detection of these altered pathways can be used as indicators to identify renal injury and promote repair.
Plant protection products (PPPs) contain one or more active substances (AS) and several co-formulants. While AS and PPPs are subjected to different approval requirements, the data requirements for PPPs are generally less comprehensive. New approach methodologies (NAMs) have the potential to detect early biological effects, provide mechanistic insight, and support the derivation of points of departure. In this study, in vitro metabolomics was used to assess a currently authorised PPP, its ASmix (i.e. tebuconazole (Teb) + prothioconazole (Pro)), and the ASmix combined with a co-formulant of concern (i.e. Teb + Pro + N,N-dimethyldecan-1-amide (NDA); ASmix + NDA). HepaRG cells were exposed for 24 h to seven concentrations and intracellular metabolites were analysed by liquid chromatography mass spectrometry. The targeted metabolomics dataset comprised of 236 unique intracellular metabolites. All treatments increased short-chain triacylglycerols (TAGs) and ceramides while decreasing carnitine, potentially indicating patterns associated with hepatic steatosis and apoptosis. Principal component analysis was used to derive multivariate metabolomics-based points of departure of 1.92 mg/L (PPP), 9.32 mg/L (ASmix+NDA), and 22.5 mg/L (ASmix). These values were then translated into acceptable operator exposure levels (AOELs) using quantitative in vitro-in vivo extrapolation and uncertainty factors. ASmix-based mixture risk assessment resulted in a hazard index (HI) below 1 indicating no operator risk. However, when PPP-based AOELs were applied, HI exceeded 1, suggesting insufficient protection for operators even with protective equipment. Overall, this NAM-based approach successfully predicts AS and PPP-based mixture effects and can be a valuable tool to refine operator safety assessment in PPP regulation where indicated by a tiered testing strategy.
Atrazine (ATR) is a widely used herbicide that can affect liver metabolism. ATR disrupts hepatic lipid metabolism, and iron homeostasis is closely associated with hepatic lipid regulation. The BMP9/SMAD1/5 signaling pathway plays an important role in maintaining iron homeostasis. Therefore, we investigated whether ATR-induced disruption of BMP9/SMAD1/5 signaling contributes to hepatic lipid accumulation by disturbing iron homeostasis. The research results show that in the liver of rats and L02 cells, ATR can cause lipid accumulation by down-regulating lipid metabolism-related proteins. ATR alters the expression of iron metabolism-related proteins, leading to iron accumulation in the liver and L02 cells. In addition, ATR exposure can inhibit the BMP9/SMAD1/5 pathway, causing iron overload in the liver. ATR-induced iron overload further exacerbated hepatic lipid metabolic dysfunction. The activation of the BMP9/SMAD1/5 signaling pathway can reduce intracellular iron accumulation induced by ATR and alleviate the increase in TC and TG levels. Overall, these findings suggested that ATR can induce iron overload by inhibiting the BMP9/SMAD1/5 pathway, thereby contributing to hepatic lipid accumulation. This provides new insights into the toxic mechanisms of ATR.
