Coupled Lipidomics and Digital Pathology as an Effective Strategy to Identify Novel Adverse Outcome Pathways in Eisenia fetida Exposed to MoS₂ Nanosheets and Ionic Mo

Abstract

Molybdenum disulfide (MoS2) nanosheets are increasingly applied in several fields, but effective and accurate strategies to fully characterize potential risks to soil ecosystems are lacking. We introduce a coelomocyte-based in vivo exposure strategy to identify novel adverse outcome pathways (AOPs) and molecular endpoints from nontransformed (NTMoS2) and ultraviolet-transformed (UTMoS2) MoS2 nanosheets (10 and 100 mg Mo/L) on the earthworm Eisenia fetida using nontargeted lipidomics integrated with transcriptomics. Machine learning-based digital pathology analysis coupled with phenotypic monitoring was further used to establish the correlation between lipid profiling and whole organism effects. As an ionic control, Na2MoO4 exposure significantly reduced (61.2–79.5%) the cellular contents of membrane-associated lipids (glycerophospholipids) in earthworm coelomocytes. Downregulation of the unsaturated fatty acid synthesis pathway and leakage of lactate dehydrogenase (LDH) verified the Na2MoO4-induced membrane stress. Compared to conventional molybdate, NTMoS2 inhibited genes related to transmembrane transport and caused the differential upregulation of phospholipid content. Unlike NTMoS2, UTMoS2 specifically upregulated the glyceride metabolism (10.3–179%) and lipid peroxidation degree (50.4–69.4%). Consequently, lipolytic pathways were activated to compensate for the potential energy deprivation. With pathology image quantification, we report that UTMoS2 caused more severe epithelial damage and intestinal steatosis than NTMoS2, which is attributed to the edge effect and higher Mo release upon UV irradiation. Our results reveal differential AOPs involving soil sentinel organisms exposed to different Mo forms, demonstrating the potential of liposome analysis to identify novel AOPs and furthermore accurate soil risk assessment strategies for emerging contaminants.

Earthworm Coelomocyte Internalization of MoS2 Nanosheets: Multiplexed Imaging, Molecular Profiling, and Computational Modeling

“…Previous lipidomics-based analyses have reported similar results that ionic-Mo species can induce earthworm coelomocyte membrane stress by inhibiting membrane-related glycerophospholipid synthesis and downregulating the fatty acid synthesis pathway. 58 Compared to conventional molybdate, DEGs in the P-NSMoS 2 group were associated with cytoskeletal protein and actin filament binding processes, which phenotypically induced potential cytoskeletal stress and actin-mediated endocytosis (Figures 4A and 5F). Additionally, the Wnt pathway was activated by P-NSMoS 2 exposure.…”

Photoaged Tire Wear Particles Leading to the Oxidative Damage on Earthworms (Eisenia fetida) by Disrupting the Antioxidant Defense System: The Definitive Role of Environmental Free Radicals

“…fetida showed a significant increase at 1 and 10% photoaged TWPs (Figure e), compared to the blank group, which further strengthens this conclusion. The accumulation of ROS is frequently accompanied by oxidative damage to cellular components . Consistently, MDA, reflecting the intensity of lipid peroxidation, significantly increased at 1 and 10% photoaged TWPs (Figure f).…”

“…Comparatively, the photoaged TWP at a relatively low concentration (0.1%) resulted in muscle pigmentation and the disruption of the epidermis structure (Figure g 5 ), indicating that exposure to the photoaged TWP may enhance the damage to earthworms. Meanwhile, the epithelium is the primary barrier against environmental stress . Disruption of epithelial integrity may further lead to the damage of muscle protein, which was also observed in the group associated with the photoaged TWP with concentrations of ≥10 g·kg –1 (Figure g 6–7 ).…”

“…52 The present observations suggested that the accumulation of H 2 O 2 , exogenous and SOD catalyzed, has exceeded the clearing capacity of CAT and POD, inhibiting its activity. 53 Moreover, GSH, which plays a crucial role in antioxidation and detoxification in organisms, can be converted by GSSG under the role of glutathione reductase (GR). Although no significant difference was observed in the activity of GSH between the photoaged TWP and the blank group (Figure S8), an increase in the activity of GSSG was evident in the photoaged TWP, as illustrated in Figure 4d.…”

Exploring the Relationship Among Lipid Profile Changes, Growth, and Reproduction in Folsomia candida Exposed to Teflubenzuron Over Time

“…In the context of untargeted metabolomics, lipids are one of the main molecular classes that are involved in many metabolic pathways and are closely associated with development, growth, and energy storage (Olzmann & Carvalho, 2019). Therefore, metabolomics can be used in risk assessment to detect alteration in organisms exposed to sublethal doses of chemicals in a short period of time, from several hours to a week of exposure (Jeong & Simpson, 2019;Kariuki et al, 2017;Sun et al, 2023). For instance, Jeong and Simpson (2019) demonstrate that alterations in the metabolome of Daphnia magna, exposed to sublethal concentrations of propranolol (18 μg L −1 water) for 6 h, serve as an early warning sign for observed changes in reproduction, decreased growth, and an increase in neonate abnormality after 2 days of exposure.…”