Glutaminolysis impairment and immunometabolic dysregulation in U937 cells: Key mechanisms in occupational and environmental skin exposure to UV and benzo[a]pyrene

Abstract

Dermal exposure to polycyclic aromatic hydrocarbons (PAHs) and UV irradiation in occupational and environmental settings poses a health risk by inducing skin toxicity, including immunomodulatory effects. This study investigated the effects of benzo[a]pyrene (B[a]P), a well-characterized PAH, at three concentrations (0.04 nM, 4 nM, and 4 µM) and UV irradiation on human monocytic U937 cells, employing both single and combined exposure scenarios. An integrated metabolomics and toxicological approach was utilized to assess cellular responses, with a focus on understanding the immunometabolic effects of these exposures. Our findings revealed that only the highest B[a]P concentration in combination with UV irradiation resulted in significant metabolic dysregulation and impaired cellular function. Notably, we observed a pronounced downregulation of glutaminolysis, a critical metabolic pathway for cellular energy production and biosynthesis. This was evidenced by decreased levels of glutamate and key intermediates within the tricarboxylic acid cycle (e.g., succinate, fumarate, malate, and citrate), as well as reduced levels of glycine, a precursor for glutathione synthesis. In parallel, toxicological assays revealed increased levels of oxidative stress markers, lipid peroxidation, and enhanced DNA damage. Furthermore, the combined exposure led to alterations in tryptophan metabolism and dysregulation of lipid species, particularly sphingolipids and phosphatidylinositols. These findings lead us to propose the hypothesis that metabolic disruption, specifically the impairment of glutaminolysis, initiated a cascade of events, including increased oxidative stress, lipid peroxidation, and ultimately, ferroptosis in our study. Our results indicate that the combined exposure to UV irradiation and B[a]P can induce immunometabolic reprogramming and significantly contribute to the pathogenesis of inflammatory skin diseases.

Keywords: Ferroptosis; Glutaminolysis; Immunometabolism; Lipid peroxidation; Lipids; Metabolomics; Monocytes; Polycyclic aromatic hydrocarbons; Skin; UV.

Fig. 1

A a schematic overview of the pathways influenced by glutamate in our study. Detected metabolites are highlighted in black, while undetected metabolites are shown in gray. B the log2 fold change of significantly altered metabolites (p < 0.05) between the treatment and control. Statistical significance was determined using one-way ANOVA followed by post hoc Fisher's LSD test. GABA γ-aminobutyric acid, GMP guanosine monophosphate, AMP adenosine monophosphate, NAA N-acetyl-L-aspartic acid, MTA methylthioadenosine, 5-oxoproline pyroglutamic acid

Fig. 2

A a schematic overview of the pathways influenced by tryptophan in our study. Detected metabolites are highlighted in black, while undetected metabolites are shown in gray. B the log2 fold change of significantly altered metabolites (p < 0.05) between the treatment and control. Statistical significance was determined using one-way ANOVA followed by post hoc Fisher's LSD test. AhR aryl hydrocarbon receptor, NAD +  nicotinamide adenine dinucleotide

Fig. 3

A a schematic overview of the altered lipid pathways in our study. Detected lipids are highlighted in black, while undetected metabolites are shown in gray. The lipids marked with * were quantified using the targeted approach. B the log2 fold change of significantly altered lipids of the untargeted approach (p < 0.05) between the treatment and control. Statistical significance was determined using one-way ANOVA followed by post hoc Fisher's LSD test. C the log2 fold change of the significantly regulated lipids of the targeted approach. Statistical significance was determined using one-way ANOVA followed by post hoc Fisher's LSD test. TAG triacylglycerol, TCA tricarboxylic acid cycle, PUFA polyunsaturated fatty acid

Fig. 4

Effects of exposures on A oxidative status in terms of GSH/GSSG ratio, ROS, MDA, NQO1, B viability/cytotoxicity in terms of LDH, MTT, MMP, and (C) DNA damage, normalized to the control condition. Data are presented as mean ± SEM (n = 4 technical, n = 4 biological replicates; 3 technical replicates were performed for the COMET analyses). Statistical significance was determined by ANOVA with post hoc Scheffé test (p < 0.005). GSH/GSSG, reduced/oxidized glutathione; ROS reactive oxygen species, MDA malondialdehyde; NQO1 NAD(P)H:quinone oxidoreductase 1, LDH lactate dehydrogenase, MTT 3-(4,5-dimethylthiazol-2-yl)−2,5-diphenyltetrazolium bromide, MMP mitochondrial membrane potential