The increasing prevalence of oxidative stress-related disorders underscores the need for safe and effective nanomaterials with strong antioxidant properties and good biocompatibility. In this study, copper oxide nanoparticles (Cu2O NPs) were successfully prepared by a simple chemical method and evaluated for their biocompatibility and antioxidant properties. The prepared NPs were characterized by transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and dynamic light scattering (DLS). Results showed that Cu2O NPs are monodispersed with a spherical shape and an average diameter of 88 ± 6 nm. In vitro biocompatibility studies on 3T3 murine fibroblasts revealed that Cu2O NPs exhibited no significant cytotoxic effects at lower concentrations (0.07-7.15 μg/mL), indicating their potential use in biomedical applications within this range. Increasing the concentration from 7.12 μg/mL to 100 μg/mL resulted in a pronounced reduction in cell viability and oxidative stress induction, as assessed using the Almar Blue test and the intracellular CM-H2CDFDA probe, respectively. DPPH scavenging and TMB kinetic assays demonstrated significant antioxidant activity (IC50 = 7.05 μg/mL) and catalytic properties of Cu2O NPs. This study provides a novel and systematic correlation between physicochemical properties, catalytic activity, and biological response, identifying a potential range of biocompatibility for the biomedical application of Cu2O nanoparticles.

In recent decades, anthropogenic activities have substantially increased the release of chemical contaminants in the environment and continuous exposure to chemical stressors has become a pervasive condition for living organisms [...].

The oxidative potential (OP) of PM2.5 was investigated during two measurement campaigns in 2024 (winter and summer) done simultaneously at an urban background and a traffic site. The research provides unprecedented chemical detail for this region, integrating elemental analysis (ED-XRF), carbonaceous fractions (OC/EC, WSOC), major ions, organic tracers (sugars/levoglucosan), and water-soluble organic nitrogen (WSON). The OP was quantified by using two assays: dithiothreitol (OPDTT) and ascorbic acid (OPAA). Source apportionment revealed competing trends of sources limiting spatial and seasonal variabilities and distinct drivers for the two OP assays. Traffic emerged as the primary contributor to OPAA at both sites, while OPDTT was influenced by traffic, secondary organic aerosols (SOAs), biomass burning, and resuspension/construction. Sea spray, nitrate, and construction-related emissions significantly impacted OPDTT but had a negligible effect on OPAA. Primary biological aerosols (fungal spores) influenced the OPAA. Seasonal variations showed dominance of traffic and biomass burning in winter (50-60% of OP), whereas sulfate and SOA became prominent during summer, for OPDTT. OPAA peaked in summer, while OPDTT peaked in winter. Traffic-induced SOA exhibits a higher redox activity relative to its mass contribution, opposite to sulfate. Season-dependent mitigation strategies could be useful to effectively reduce the oxidative burden of PM2.5.

Nanoplastics (NPs) are pervasive pollutants whose detection and collection from the environment is challenging due to their extremely small size, which limits the effectiveness of characterization techniques. This study analyses the impact of crystallization kinetics on the autofluorescence behavior of unlabeled polyethylene terephthalate (PET) NPs produced by mechanically fragmenting plastic waste in conditions that simulate natural degradation. This process yields particles with properties similar to those found in nature. By studying crystallization kinetics, it is possible to restore polymer crystallinity, effectively replicating the increased crystallinity observed in environmentally degraded polymers. Additionally, the enhanced crystallinity leads to a stronger fluorescence signal allowing the resulting nanoparticles to be used without additional fluorescent staining, a novel feature that overcomes the limitations of conventional dye labelling.The ecotoxicological relevance of the obtained label-free PET nanoplastics is validated through in vivo exposure on the freshwater zooplanctonic model species Daphnia magna at two different temperatures. Confocal microscopy indicates a prevalent accumulation in the gastrointestinal tract. Recovery results reveal a clear decline in organism survival that is dependent on both dose and temperature, indicating that higher temperatures amplify the toxic effects of PET nanoparticles. This highlights the potential for global warming to exacerbate the ecological impacts of microplastics.

Bacterial cellulose (BC) hydrogel is a promising skin wound healing biomaterial due to its unique properties, including a moist environment that facilitates tissue healing. To enhance its antimicrobial efficacy, BC dressings were loaded with penicillin and streptomycin. FT-IR analysis confirmed successful drug binding, while SEM revealed a nanofibrous and porous hydrogel structure. In vitro studies using 3T3 mouse fibroblasts demonstrated biocompatibility, and scratch wound assays achieved complete closure across all tested concentrations. Antibacterial activity, assessed via agar diffusion against Pseudomonas aeruginosa and Staphylococcus aureus, showed a concentration-dependent increase in inhibition zones, highlighting the potential of BC-Pen/Strep hydrogels as effective antimicrobial wound dressings.

Trolox, a water-soluble analog of vitamin E, is widely used as a reference antioxidant in in vitro biochemical assays. However, its intracellular redox behavior is known to vary depending on both concentration and oxidative context. In this study, we investigated the dose-dependent antioxidant and prooxidant effects of Trolox in two cellular models, HeLa cells and 3T3 cells exposed for 1 h to increasing concentrations (2–160 µM), under both basal conditions and oxidative stress induced by hydrogen peroxide. Intracellular oxidative changes were assessed using the oxidative stress-sensitive fluorescent probe CM-H2DCFDA. Under basal conditions, Trolox exerted slight dose-dependent antioxidant behavior in 3T3 cells on the basal production of ROS in concentrations ranging from 2 µM to 160 µM. In contrast, in HeLa cells Trolox displayed a biphasic activity: antioxidant at low doses (≤10 µM) and a switch to prooxidant behavior at higher concentrations. Under H2O2-induced stress, in HeLa cells Trolox retained antioxidant activity at low concentrations (≤10 µM), but this effect gradually declined at higher doses, disappearing around 80 µM and shifting to a slight prooxidant effect at 160 µM. Confocal microscopy confirmed the spectrofluorimetric results. Conversely, 3T3 cells exhibited an early shift toward prooxidant activity already at 10 µM. These findings highlight that the Trolox redox activity is determined not only by concentration but also by cell-specific intracellular environment and redox state. The study suggests caution against generalized antioxidant use of Trolox and highlights the need for specific dose–response evaluations in specific cell types and biological settings.

Water scarcity for crop production and the need to ensure environmental protection lead to the reuse of treated wastewater for irrigation. However, the use of wastewater raises concerns about its quality and related toxicological and ecotoxicological risks. Ecotoxicity tests on soil organisms can offer an integrated assessment of the environmental adverse effects of bioavailable toxic substances in wastewater for irrigation. This work aims to investigate the assessment of ecotoxicity in agricultural soils irrigated with treated wastewater through a multi-biomarker approach in earthworms. In particular, molecular and cellular biomarkers (lysosomal membrane stability, glutathione peroxidase, glutathione reductase, GSH/GSSG, metallothionein, acethylcholinesterase) were measured, combined with acute and chronic toxicity tests on Eisenia fetida exposed to soil samples from agricultural fields irrigated with treated wastewater. Three experimental fields in Apulia, Italy, were irrigated with varying types of treated wastewater. Acute toxicity tests on Eisenia fetida exposed to the irrigated soils showed no significant mortality in any soils, while chronic toxicity was observed in fields irrigated with secondary-treated wastewater, but not in fields irrigated with tertiary-treated wastewater. Biomarkers indicated reduced lysosomal membrane stability, increased oxidative stress, and reduced acetylcholinesterase activity in worms exposed to secondary-treated wastewater. The results showed how the combined use of toxicity assays and biomarker analyses in soil bioindicator organisms allows the toxicity of soils irrigated with treated wastewater to be assessed, in order to provide an integrated measure of chemical pollutants bioavailable and their biological effects.

Nanoplastics pose significant environmental and public health risks, prompting the need for sensitive, cost-effective, and rapid assays for ecotoxicity assessment. The present work proposes the use of a portable smartphone-based platform to enhance traditional Daphnia magna acute toxicity assays by integrating behavior analysis and heart rate measurements. The aim is to improve sensitivity in detecting toxic effects of nanoplastics. In particular, the study focused on nano-sized carboxylated polystyrene (PS) nanoparticles. Two variability factors that could influence biological effects of nanoplastics, the particle size and the age of the organisms, were considered. Results demonstrated that the application of the proposed integrated approach allowed the detection of early subtle effects such as a significant impact on the heart rate and behavior of Daphnia magna under short-term exposure to PS carboxylated nanoparticles. In particular, a stimulation of heart rate was observed for both neonates and adults either for 40 nm or 200 nm particles after 48 h exposure, presumably attributable to an interference of carboxylated PS NPs with adrenergic-type receptors. Behavioral alterations were detectable for 40 nm particles but not for 200 nm ones consisting of a decrease in velocity and alterations of trajectories. Obtained results demonstrated the suitability of the proposed smartphone platform for friendly and real-time integration of behavioral analysis with physiological outcome measurements during acute exposure of Daphnia magna to nano-sized carboxylated PS NPs, expanding the sensitivity of the traditional acute toxicity tests. It offers a novel, cost-effective, and field-applicable method for environmental monitoring of nanoparticle toxicity and impact.

Plastic pollution has become a major environmental and public health issue due to rising global production. Nanoplastics (NPs) are especially concerning due to their widespread presence and potential health risks. This study aims to determine the impact of the exposure to polyethylene terephthalate (PET) NPs on fibroblast cells using the murine NIH-3T3 cells as experimental model. This is a relevant cellular model for several biological fields of application, including cell migration in wound healing and tissue regeneration. The PET NPs used represented an environmentally realistic PET NPs model since they were produced by a fast top down approach in a process close to the mechanical abrasion of microplastics occurring in the environment. They were characterized by an intrinsic autofluorescence which enables their use in studies of NPs interactions with biological systems without the need for additional fluorescent dyes. Additionally, the Hansen solubility parameters (HSP) of the PET NPs and the culture medium were determined to better understand their interaction. PET NPs were internalized by fibroblasts in a dose-dependent manner, localizing in the cytoplasm. While they caused only a slight reduction in cell viability (within 20% inhibition at 10-100 μg/mL) after 24 h exposure, they significantly impaired fibroblast migration, as demonstrated by the scratch assay, indicating possible interference in tissue repair. The exposure of the cells to PET NPs induced a significant dose-dependent ROS increase suggesting the induction of intracellular oxidative stress as possible mechanisms underlying the observed migration impairment. These findings highlight the potential risks of PET NPs to fibroblasts, emphasizing the need for further research into their impact on cellular functions and mechanisms.

Hemocytes are the circulating immune-competent cells in bivalve mollusks and play a key role in several important functions of cell-mediated innate immunity. During the early stages of the immune response, hemocytes actively migrate to the site of infection. This inherent motility is a fundamental characteristic of these cells. It represents a key cellular function that integrates multiple processes, such as cell adhesion, cell signaling, cytoskeletal dynamics, and changes in cell volume. Therefore, alterations in cell motility following exposure to drugs or pollutants can serve as a useful toxicological endpoint. Despite the fundamental role of cell motility in cellular physiology, it has been poorly investigated from a toxicological perspective. This work proposes a novel in vitro method for the rapid and sensitive assessment of the toxicity and ecotoxicity of pollutants, based on evaluating the hemocyte motility of Mytilus galloprovincialis. We developed a cell motility assay on hemocytes adhering to the bottom of a 96-well polystyrene microplate. Following exposure to increasing concentrations of drugs, cell trajectories, and velocities were quantified by cell tracking under time-lapse microscopy, allowing us to measure the effects on hemocyte motility. Due to the ease of hemocyte collection from the animals in a relatively non-invasive manner, the proposed method offers an alternative test for screening the effects and mechanisms of action of pollutants and drugs. It aligns with the 3Rs (Replacement, Reduction, and Refinement) criteria, addressing ethical concerns and contributing to the reduction of vertebrate in vivo animal testing.

In recent years, significant academic and commercial interest has focused on collagen derived from horse tendons, with potential applications across diverse sectors such as medicine, pharmaceuticals, and cosmetics. Nano collagen, with its enhanced wound penetration, improved cell contact, and heightened cellular regeneration and repair capabilities due to its high surface area, holds promise for a wide range of applications. In this study, we present a novel method for producing nano collagen from the equine tendon. Our approach is characterized by its speed, affordability, simplicity and environmentally friendly nature, with precise temperature-control to prevent collagen denaturation. We conducted a comprehensive characterization of the obtained samples, including assessments of morphology, chemical and thermal properties, particle size distribution and biocompatibility. Importantly, our results indicate improvements in thermal stability, and surface roughness of nano collagen, while preserving its molecular weight. These advancements expand the potential applications of nano collagen in various fields.

Microplastics (MPs) and nanoplastics (NPs) have emerged as significant environmental pollutants with potential detrimental effects on ecosystems and human health. Several studies indicate their interaction with enzymes; this topic represents a multifaceted research field encompassing several areas of interest from the toxicological and ecotoxicological impact of MPs and NPs on humans and wildlife to the biodegradation of plastics by microbial enzymes. This review aims to provide a critical analysis of the state-of-the-art knowledge of the interaction of MPs and NPs on the enzyme carbonic anhydrase (CA), providing recent insights, analyzing the knowledge gaps in the field, and drawing future perspectives of the research and its application. CA is a widespread and crucial enzyme in various organisms; it is critical for various physiological processes in animals, plants, and bacteria. It catalyzes the reversible hydration of CO2, which is essential for respiration, acid–base balance, pH homeostasis, ion transport, calcification, and photosynthesis. Studies demonstrate that MPs and NPs can inhibit CA activity with mechanisms including adsorption to the enzyme surface and subsequent conformational changes. In vitro and in silico studies highlight the role of electrostatic and hydrophobic interactions in these processes. In vivo studies present mixed results, which are influenced by factors like particle type, size, concentration, and organism type. Moreover, the potentiality of the esterase activity of CA for plastic degradation is discussed. The complexity of the interaction between CA and MPs/NPs underscores the need for further research to fully understand the ecological and health impacts of MPs and NPs on CA activity and expression and glimpses of the potentiality and perspectives in this field.

Osteoarthritis (OA) stands as a prevalent and progressively debilitating clinical condition globally, impacting joint structures and leading to their gradual deterioration through inflammatory mechanisms. While both non-modifiable and modifiable factors contribute to its onset, numerous aspects of OA pathophysiology remain elusive despite considerable research strides. Presently, diagnosis heavily relies on clinician expertise and meticulous differential diagnosis to exclude other joint-affecting conditions. Therapeutic approaches for OA predominantly focus on patient education for self-management alongside tailored exercise regimens, often complemented by various pharmacological interventions primarily targeting pain alleviation. However, pharmacological treatments typically exhibit short-term efficacy and local and/or systemic side effects, with prosthetic surgery being the ultimate resolution in severe cases. Thus, exploring the potential integration or substitution of conventional drug therapies with natural compounds and extracts emerges as a promising frontier in enhancing OA management. These alternatives offer improved safety profiles and possess the potential to target specific dysregulated pathways implicated in OA pathogenesis, thereby presenting a holistic approach to address the condition’s complexities.

Aquatic invertebrates play a pivotal role in (eco)toxicological assessments because they offer ethical, cost-effective and repeatable testing options. Additionally, their significance in the food chain and their ability to represent diverse aquatic ecosystems make them valuable subjects for (eco)toxicological studies. To ensure consistency and comparability across studies, international (eco)toxicology guidelines have been used to establish standardised methods and protocols for data collection, analysis and interpretation. However, the current standardised protocols primarily focus on a limited number of aquatic invertebrate species, mainly from Arthropoda, Mollusca and Annelida. These protocols are suitable for basic toxicity screening, effectively assessing the immediate and severe effects of toxic substances on organisms. For more comprehensive and ecologically relevant assessments, particularly those addressing long-term effects and ecosystem-wide impacts, we recommended the use of a broader diversity of species, since the present choice of taxa exacerbates the limited scope of basic ecotoxicological studies. This review provides a comprehensive overview of (eco)toxicological studies, focusing on major aquatic invertebrate taxa and how they are used to assess the impact of chemicals in diverse aquatic environments. The present work supports the use of a broad-taxa approach in basic environmental assessments, as it better represents the natural populations inhabiting various ecosystems. Advances in omics and other biochemical and computational techniques make the broad-taxa approach more feasible, enabling mechanistic studies on non-model organisms. By combining these approaches with in vitro techniques together with the broad-taxa approach, researchers can gain insights into less-explored impacts of pollution, such as changes in population diversity, the development of tolerance and transgenerational inheritance of pollution responses, the impact on organism phenotypic plasticity, biological invasion outcomes, social behaviour changes, metabolome changes, regeneration phenomena, disease susceptibility and tissue pathologies. This review also emphasises the need for harmonised data-reporting standards and minimum annotation checklists to ensure that research results are findable, accessible, interoperable and reusable (FAIR), maximising the use and reusability of data. The ultimate goal is to encourage integrated and holistic problem-focused collaboration between diverse scientific disciplines, international standardisation organisations and decision-making bodies, with a focus on transdisciplinary knowledge co-production for the One-Health approach.

Transitional waters are fragile ecosystems with high ecological values, representing the breeding and resting sites for rare and threatened species. They warrant particular attention in regards to protection, as they experience numerous anthropogenic threats. The present review aims to analyze the recent literature on Aphanius fasciatus, currently considered one of the most strictly estuarine-dependent fish species, thus affected by the degradation of lagoon habitats, and to discuss its suitability as a sentinel species for protection of the quality of transitional water environments. The analysis and discussion highlight the potential applicability of the molecular, cellular, and physiological responses of this species as diagnostic tools for detecting the subtle effects induced by environmental pollution on the biota in transitional water environments. Moreover, the suitability of the responses of this species is suggested in the wider framework of the One Health perspective, which considers human and animal health and the environmental state to be highly interconnected, sharing common aspects. To date, omics technologies show great potential in reacquiring novel knowledge on the responses of the organisms to environmental changes and to the alterations of the environmental health status. Therefore, considering the relevant potential of this organism as a sentinel species, many efforts are required in the near future to improve the quantity and quality of the omics tools that refer to A. fasciatus.

Throughout the cold and the warm periods of 2020, chemical and toxicological characterization of the water-soluble fraction of size segregated particulate matter (PM) (<0.49, 0.49–0.95, 0.95–1.5, 1.5–3.0, 3.0–7.2 and >7.2 μm) was conducted in the urban agglomeration of Thessaloniki, northern Greece. Chemical analysis of the water-soluble PM fraction included water-soluble organic carbon (WSOC), humic-like substances (HULIS), and trace elements (V, Cr, Mn, Fe, Ni, Cu, Zn, As, Cd and Pb). The bulk (sum of all size fractions) concentrations of HULIS were 2.5 ± 0.5 and 1.2 ± 0.3 μg m−3, for the cold and warm sampling periods, respectively with highest values in the <0.49 μm particle size fraction. The total HULIS-C/WSOC ratio ranged from 17 to 26% for all sampling periods, confirming that HULIS are a significant part of WSOC. The most abundant water-soluble metals were Fe, Zn, Cu, and Mn. The oxidative PM activity was measured abiotically using the dithiothreitol (DTT) assay. In vitro cytotoxic responses were investigated using mitochondrial dehydrogenase (MTT). A significant positive correlation was found between OPmDTT, WSOC, HULIS and the MTT cytotoxicity of PM. Multiple Linear Regression (MLR) showed a good relationship between OPMDTT, HULIS and Cu.

The work aimed to study the induction of morphological alterations in M. galloprovincialis in the field and its suitability to be integrated into a sensitive, simple, and cost-effective cell-based multimarker approach for the detection of the stress status induced by pollution in coastal marine environments in view of ecotoxicological biomonitoring and assessment application. Cellular morphometric alterations was paralleled by the analysis of standardized biomarkers such as lysosomal membrane destabilization, and genotoxocity biomarkers such as micronuclei and binuclated cells frequencies were investigated. The study was carried out by means of a transplanting experiment in the field, using caged organisms from an initial population exposed in the field in two multi-impacted coastal sites of the central Mediterranean area, Bagnoli in the eastern Tyrrhenian Sea and Augusta-Melilli-Priolo in the western Ionian Sea. Capo Miseno (NA) for the Tyrrhenian area and Brucoli (ME) for the Ionian area were chosen as control sites. Hemocyte enlargement and filopodial elongation increased frequencies were observed in organisms exposed to the impacted sites. These morphometric alterations showed strong agreement with the lysosomal membrane destabilization and biomarkers of genotoxicity, suggesting their usefulness in detecting the pollutant-induced stress syndrome related to genotoxic damage.

Recent years have seen a significant increase in the scientific literature related to various methods for analyzing oxidative potential (OP) of atmospheric particulate matter (PM). The presence of several types of PM, differing chemical and physical properties, released by both anthropogenic and natural sources, leads to numerous health issues in living organisms and represents an attractive target for air quality monitoring. Therefore, several studies have focused on developing rapid and self-operative tests, employing different target molecules to assess OP of atmospheric aerosols as well as unique approaches to overcome some of the most common laboratory-related issues in this kind of analysis. This work provides an overview of online and automated systems, as well as a broad picture of the state-of-art of the various devices and methods developed on this topic over the last two decades. Moreover, representative studies on this subject will be discussed, analyzing the advantages and drawbacks of the developed automated techniques.

Two advanced oxidation processes (AOPs), namely ozone/H2O2 and UV/H2O2, were tested at pilot scale as zero-liquid-discharge alternative treatments for the removal of microbiological (bacteria and viruses), chemical (compounds of emerging concern (CECs)) and genotoxic responses from tertiary municipal wastewater for indirect potable reuse (IPR). The AOP treated effluents were further subjected to granular activated carbon (GAC) adsorption and UV disinfection, following the concept of multiple treatment barriers. As a reference, a consolidated advanced wastewater treatment train consisting of ultrafiltration, UV disinfection, and reverse osmosis (RO) was also employed. The results showed that, for the same electrical energy applied, the ozone/H2O2 treatment was more effective than the UV/H2O2 treatment in removing CECs. Specifically, the ozone/H2O2 treatment, intensified by high pressure and high mixing, achieved an average CECs removal efficiency higher than UV/H2O2 (66.8% with respect to 18.4%). The subsequent GAC adsorption step, applied downstream the AOPs, further improved the removal efficiency of the whole treatment trains, achieving rates of 98.5% and 96.8% for the ozone/H2O2 and UV/H2O2 treatments, respectively. In contrast, the ultrafiltration step of the reference treatment train only achieved a removal percentage of 22.5%, which increased to 99% when reverse osmosis was used as the final step. Microbiological investigations showed that all three wastewater treatment lines displayed good performance in the complete removal of regulated and optional parameters according to both national and the European Directive 2020/2184. Only P. aeruginosa resulted resistant to all treatments with a higher removal by UV/H2O2 when higher UV dose was applied. In addition, E. coli STEC/VTEC and enteric viruses, were found to be completely removed in all tested treatments and no genotoxic activity was detected even after a 1000-fold concentration. The obtained results suggest that the investigated treatments are suitable for groundwater recharge to be used as a potable water source being such a procedure an IPR. The intensified ozone/H2O2 or UV/H2O2 treatments can be conveniently incorporated into a multi-barrier zero-liquid-discharge scheme, thus avoiding the management issues associated with the retentate of the conventional scheme that uses reverse osmosis. By including the chemical cost associated with using 11-12 mg/L of H2O2 in the cost calculations, the overall operational cost (energy plus chemical) required to achieve 50% average CECs removal in tertiary effluent for an hypothetical full-scale plant of 250 m3/h (or 25,000 inhabitants) was 0.183 €/m3 and 0.425 €/m3 for ozone/H2O2 and UV/H2O2 treatment train, respectively.