Systems toxicology of naturally aged polypropylene microplastics in zebrafish.

Microplastic (MP) pollution represents an emerging threat to aquatic ecosystems; yet, most studies remain restricted to isolated biomarker comparisons, without addressing how these pollutants reorganize organisms' systemic properties. In this study, we investigated the effects of naturally aged polypropylene microplastics (NAPP-MPs) - a highly abundant and environmentally relevant polymer - on adult Danio rerio, a recognized model organism in ecotoxicological studies. Fish were exposed to 4.5 mg/L of NAPP-MPs for 15 days and assessed using a biomarker panel that encompassed oxidative and nitrosative stress, antioxidant defenses, neurochemical and endocrine parameters, and digestive protease activities. We observed substantial particle bioaccumulation and consistent biochemical alterations, including increased production of reactive oxygen species, elevated levels of malondialdehyde and nitric oxide, activation of antioxidant enzymes such as superoxide dismutase and catalase, and enhancement of non-enzymatic antioxidant defenses, as measured by the DPPH method. Moreover, we detected elevated concentrations of dopamine, serotonin, and cortisol, as well as increased acetylcholinesterase activity and intensified actions of the digestive proteases trypsin and chymotrypsin. Beyond mean-level differences, we applied a multiscale analytical framework-including criticality analysis, signed networks, information theory, energy landscapes, and Bayesian causal modeling with bootstrap-which revealed that NAPP-MPs shift biochemical systems from adaptive critical regimes to less stable and fragmented states, characterized by loss of cohesion, reorganization of informational flows, and altered functional hierarchies across regulatory modules. Collectively, our results demonstrate that NAPP-MPs not only trigger discrete biochemical responses in zebrafish but also reshape the systemic architecture of homeostasis, undermining both stability and adaptive plasticity. Thus, our study advances the field by integrating innovative methodologies into MP ecotoxicology, providing a conceptual and methodological framework that broadens understanding of their risks and supports more realistic, complex environmental assessments.