TGF-ß induces matrisome pathological alterations and EMT in patient-derived prostate cancer tumoroids.

Extracellular matrix (ECM) tumorigenic alterations resulting in high matrix deposition and stiffening are hallmarks of adenocarcinomas and are collectively defined as desmoplasia. Here, we thoroughly analysed primary prostate cancer tissues obtained from numerous patients undergoing radical prostatectomy to highlight reproducible structural changes in the ECM leading to the loss of the glandular architecture. Starting from patient cells, we established prostate cancer tumoroids (PCTs) and demonstrated they require TGF-ß signalling pathway activity to preserve phenotypical and structural similarities with the tissue of origin. By modulating TGF-ß signalling pathway in PCTs, we unveiled its role in ECM accumulation and remodelling in prostate cancer. We also found that TGF-ß-induced ECM remodelling is responsible for the initiation of prostate cell epithelial-to-mesenchymal transition (EMT) and the acquisition of a migratory, invasive phenotype. Our findings highlight the cooperative role of TGF-ß signalling and ECM desmoplasia in prompting prostate cell EMT and promoting tumour progression and dissemination.

Short-term exposure to polystyrene microplastics hampers the cellular function of gills in the Mediterranean mussel Mytilus galloprovincialis.

Plastic is undoubtedly the most useful and versatile polymeric material that man has developed in the last two centuries Despite the societal benefits, plastic is now a serious global issue because it is persistent and may bioaccumulate into aquatic biota as microplastics (MPs). This study was designed to evaluate the daily uptake and cellular effects due to a short-term (up to 72 h) exposure to 3 µm red polystyrene MPs (50 beads/mL) in the gills of the Mediterranean mussel Mytilus galloprovincialis, chosen as model species for its ecological and commercial relevance. After measuring the daily uptake of MPs and detecting their presence within the branchial epithelium at all the exposure time-points (T24, T48, T72), some cleaning mechanisms were observed by neutral and acid mucous secretions at mussel gills. The protonic Nuclear Magnetic Resonance (1H NMR)-based metabolomics, combined with chemometrics, allowed to comprehensively explore the time-dependent metabolic disorders triggered by MPs in mussel gills over the short-term trial. Specifically, the clear clustering between MP-treated mussel gills and those from control, together with the grouping for experimental time-points as depicted by the Principal Component Analysis (PCA), were due to changes in the amino acids and energy metabolism, disturbances in the osmoregulatory processes, as well as in the cholinergic neurotransmission. Moreover, as evidenced by enzymatic assays, even the oxidative defense systems and lipid metabolism were hampered by MP exposure. Overall, these findings provides the first insights into the early time-dependent mechanisms of toxicity of polystyrene MPs in marine mussels, and underline the potential environment and human health risk posed by MPs contamination.

Histomorphological Changes in Fish Gut in Response to Prebiotics and Probiotics Treatment to Improve Their Health Status: A Review.

The gastrointestinal tract (GIT) promotes the digestion and absorption of feeds, in addition to the excretion of waste products of digestion. In fish, the GIT is divided into four regions, the headgut, foregut, midgut, and hindgut, to which glands and lymphoid tissues are associated to release digestive enzymes and molecules involved in the immune response and control of host-pathogens. The GIT is inhabited by different species of resident microorganisms, the microbiota, which have co-evolved with the host in a symbiotic relationship and are responsible for metabolic benefits and counteracting pathogen infection. There is a strict connection between a fish's gut microbiota and its health status. This review focuses on the modulation of fish microbiota by feed additives based on prebiotics and probiotics as a feasible strategy to improve fish health status and gut efficiency, mitigate emerging diseases, and maximize rearing and growth performance. Furthermore, the use of histological assays as a valid tool for fish welfare assessment is also discussed, and insights on nutrient absorptive capacity and responsiveness to pathogens in fish by gut morphological endpoints are provided. Overall, the literature reviewed emphasizes the complex interactions between microorganisms and host fish, shedding light on the beneficial use of prebiotics and probiotics in the aquaculture sector, with the potential to provide directions for future research.

Use of canakinumab and remdesivir in moderate-severe COVID-19 patients: A retrospective analysis.

Objectives: The dysregulated immune response occurring upon COVID-19 infection can lead to tissue damage and organ failure. Different therapeutic strategies are needed to cope with the current and future outspread of COVID-19, including antiviral and anti-inflammatory agents. We describe the outcome of hospitalized patients treated with canakinumab and remdesivir plus the standard of care therapy. Methods: This observational study describes the outcome of the combination of canakinumab (450 mg for patients ≥40 and <60 kg, 600 mg for those ≥60 and <80 kg, or 750 mg for patients ≥80 kg) and 200 mg remdesivir intravenous infusion, plus standard of care (SOC), in 17 moderate-to-severe COVID-19 patients hospitalized in the "Annunziata" Hospital, Cosenza, Italy, between August and November 2021. Hematological markers, biochemical, and hemogasanalysis values at baseline versus day 7 of combination treatment were compared by paired t test after checking for normal distribution and correcting for multiple comparison. Results: The median age of patients was 64 years (range: 39-85), and the median hospitalization time (calculated on the 16 patients that were not transferred to intensive care unit) was of 12.5 days (range: 7-35 days); 15/17 patients (88%) did not experience complications. After 7 days of combination therapy, all the inflammatory parameters were significantly reduced with the exception of procalcitonin; moreover, hematological prognostic markers such neutrophil-to-lymphocyte ratio, CRP-to-lymphocyte ratio, and derived neutrophil-to-lymphocyte ratio reduced. Overall, 16/17 patients (94%) recovered after 14 days. Conclusions: Canakinumab and remdesivir treatment, in addition to SOC, in the early stage of moderate-to-severe COVID-19 showed promising outcomes in terms of safety and effectiveness potentially leading to a reduction in inflammatory and hematological prognostic markers after 7 days of treatment.

Single and combined effects of caffeine and salicylic acid on mussel Mytilus galloprovincialis: Changes at histomorphological, molecular and biochemical levels.

Caffeine (CAF) and salicylic acid (SA) are frequently detected in waterbody, though information on their biological impact is poor. This work assesses the effects of CAF (5 ng/L to 10 µg/L) and SA (0.05 µg/L to 100 µg/L) alone and combined as CAF+SA (5 ng/L+0.05 µg/L to 10 µg/L+100 µg/L) on mussel Mytilus galloprovincialis under 12-days exposure by histomorphology of digestive gland and oxidative stress defense at molecular and biochemical levels. Besides evaluating tissue accumulation, absence of histomorphological damage and haemocyte infiltration highlighted activation of defensive mechanisms. Up-regulation of Cu/Zn-sod, Mn-sod, cat and gst combined with increased catalase and glutathione S-transferase activity were found in CAF-exposed mussels, while SA reduced ROS production and mitochondrial activity. CAF+SA exposure induced differential responses, and the integrated biomarker response (IBR) revealed more pronounced effects of SA than CAF. These results enlarge knowledge on pharmaceuticals impact on non-target organisms, emphasizing the need for proper environmental risk assessment.

Cytoprotective and Antioxidant Effects of Hydrolysates from Black Soldier Fly (Hermetia illucens).

The black soldier fly (BSF), Hermetia illucens, has been recognized as one of the most promising insect species for its ability to valorize organic waste while producing a valuable larval biomass with a great potential as a sustainable source of nutrients, including proteins and bioactive molecules. In the present study, BSF larvae were used to produce and characterize the protein hydrolysates (BPHs) that were then evaluated for their potential biological activity in vitro. The BPHs obtained from the BSF larvae proteins by enzymatic digestion were characterized by Nuclear Magnetic Resonance (NMR) and polyacrylamide gel electrophoresis and assessed for their antioxidant activity (BPHs in the range of 0.1 to 1.5 mg/mL) in L-929 cells. Our findings show that BPHs can exert a dose-dependent cytoprotective role against H2O2-iduced oxidative stress in cells. This antioxidant activity relies on the reduction of ROS levels in challenged cells as measured by flow cytometry and fluorescence microscopy, together with the induction and nuclear translocation of Nrf2, as evaluated by qPCR and indirect immunofluorescence analysis, respectively. Overall, our findings on the remarkable biological activity of the BPHs obtained in a large-scale process strongly suggest the application of BPHs as ingredients promoting animal health in feed formulations.

Differential Cell Metabolic Pathways in Gills and Liver of Fish (White Seabream Diplodus sargus) Coping with Dietary Methylmercury Exposure.

Mercury (Hg) is a dangerous and persistent trace element. Its organic and highly toxic form, methylmercury (MeHg), easily crosses biological membranes and accumulates in biota. Nevertheless, understanding the mechanisms of dietary MeHg toxicity in fish remains a challenge. A time-course experiment was conducted with juvenile white seabreams, Diplodus sargus (Linnaeus, 1758), exposed to realistic levels of MeHg in feed (8.7 µg g-1, dry weight), comprising exposure (E; 7 and 14 days) and post-exposure (PE; 28 days) periods. Total Hg levels increased with time in gills and liver during E and decreased significantly in PE (though levels of control fish were reached only for gills), with liver exhibiting higher levels (2.7 times) than gills. Nuclear magnetic resonance (NMR)-based metabolomics revealed multiple and often differential metabolic changes between fish organs. Gills exhibited protein catabolism, disturbances in cholinergic neurotransmission, and changes in osmoregulation and lipid and energy metabolism. However, dietary MeHg exposure provoked altered protein metabolism in the liver with decreased amino acids, likely for activation of defensive strategies. PE allowed for the partial recovery of both organs, even if with occurrence of oxidative stress and changes of energy metabolism. Overall, these findings support organ-specific responses according to their sensitivity to Hg exposure, pointing out that indications obtained in biomonitoring studies may depend also on the selected organ.

Ameliorative Hematological and Histomorphological Effects of Dietary Trigonella foenum-graecum Seeds in Common Carp (Cyprinus carpio) Exposed to Copper Oxide Nanoparticles.

Different types of metal oxide nanoparticles (NPs) are being used for wastewater treatment worldwide but concerns have been raised regarding their potential toxicities, especially toward non-targeted aquatic organisms including fishes. Therefore, the present study aimed to evaluate the toxicity of copper oxide (CuO) NPs (1.5 mg/L; positive control group) in a total of 130 common carp (Cyprinus carpio), as well as the potential ameliorative effects of fenugreek (Trigonella foenum-graecum) seed extracts (100 mg/L as G-1 group, 125 mg/L as G-2 group, and 150 mg/L as G-3 group) administered to fish for 28 days. Significant changes were observed in the morphometric parameters: the body weight and length of the CuO-NP-treated fish respectively decreased from 45.28 ± 0.34 g and 14.40 ± 0.56 cm at day one to 43.75 ± 0.41 g and 13.57 ± 0.67 cm at day 28. Conversely, fish treated with T. foenum-graecum seed extract showed significant improvements in body weight and length. After exposure to CuO NPs, a significant accumulation of Cu was recorded in the gills, livers, and kidneys (1.18 ± 0.006 µg/kg ww, 1.38 ± 0.006 µg/kg ww, and 0.05 ± 0.006 µg/kg ww, respectively) of the exposed common carp, and significant alterations in fish hematological parameters and oxidative stress biomarkers (lipid peroxidation (LPO), glutathione (GSH), and catalase (CAT)) were also observed. However, supplementing diets with fenugreek extracts modulated the blood parameters and the oxidative stress enzymes. Similarly, histological observations revealed that sub-lethal exposure to CuO NPs caused severe histomorphological changes in fish gills (i.e., degenerative epithelium, fused lamellae, necrotic lamellae, necrosis of primary lamellae, complete degeneration, and complete lamellar fusion), liver (i.e., degenerative hepatocytes, vacuolization, damaged central vein, dilated sinusoid, vacuolated degeneration, and complete degeneration), and kidney (i.e., necrosis and tubular degeneration, abnormal glomerulus, swollen tubules, and complete degeneration), while the treatment with the fenugreek extract significantly decreased tissue damage in a dose-dependent manner by lowering the accumulation of Cu in the selected fish tissues. Overall, this work demonstrated the ameliorative effects of dietary supplementation with T. foenum-graecum seed extract against the toxicity of NPs in aquatic organisms. The findings of this study therefore provided evidence of the promising nutraceutical value of fenugreek and enhanced its applicative potential in the sector of fish aquaculture, as it was shown to improve the growth performance and wellness of organisms.

Toxicological impact of environmental microplastics and benzo[a]pyrene in the seaworm Hediste diversicolor under environmentally relevant exposure conditions.

Nowadays, marine ecosystems are under severe threat from the simultaneous presence of multiple stressors, including microplastics (MPs) and polycyclic aromatic hydrocarbons (PAHs) such as benzo[a]pyrene (B[a]P). In addition to their presence in various marine compartments, there are increasing concerns on the potential capacity of MPs to sorb, concentrate and transfer these pollutants in the environment. Although their ecotoxicological impacts are currently evident, few works have studied the combined effects of these contaminants. Therefore, the major purpose of this work was to assess the toxicity of environmental relevant concentrations of MPs (<30 µm) and B[a]P, alone and in mixture, in the seaworm Hediste diversicolor by exploring their accumulation and hazardous biological effects for 3 and 7 days. Environmental MPs were able to increase B[a]P in a time-dependent manner. The obtained results showed that individual treatments, as well as co-exposure to contaminants, caused cytotoxicity and genotoxicity in the cœlomic fluid cells, while oxidative stress effects were observed at tissue and gene levels associated with alteration in neurotransmission. Overall, our findings provide additional clues about MPs as organic pollutant vectors in the marine environment, and contribute to a clearer understanding of their toxicological risk to aquatic invertebrates.

Time- and dose-dependent biological effects of a sub-chronic exposure to realistic doses of salicylic acid in the gills of mussel Mytilus galloprovincialis.

Among nonsteroidal anti-inflammatory drugs (NSAIDs) commonly found in seawater and wastewater, salicylic acid (SA) represents one of the most persistent and hazardous compounds for aquatic organisms. This study was therefore designed to elucidate the biological effects of SA in mussel Mytilus galloprovincialis. During a sub-chronic exposure (12 days), mussels were exposed to five realistic concentrations of SA (C1: 0.05 µg/L; C2: 0.5 µg/L; C3: 5 µg/L; C4: 50 µg/L; C5: 100 µg/L) and gills, selected as the target organ, were collected at different time points (T3: 3 days; T5: 5 days; T12: 12 days). Exposure to SA induced no histological alterations in mussel gills, despite a relevant hemocyte infiltration was observed throughout the exposure as a defensive response to SA. Temporal modulation of glutathione S-transferase (GST), catalase (CAT), and superoxide dismutase (SOD) activities suggested the occurrence of antioxidant and detoxifying responses against SA exposure, while lipid peroxidation (LPO), except for a partial increase at T3, was prevented. Inhibition of the cholinergic system was also reported by reduced acetylcholinesterase (AChE) activity, mainly at T12. Overall, findings from this study contribute to enlarge the current knowledge on the cytotoxicity of SA, on non-target aquatic organisms, and might for the enhancement of new ecopharmacovigilance programs and optimization of the efficacy of wastewater treatment plants for mitigation of pharmaceutical pollution in coastal areas.

Comparison of cellular mechanisms induced by pharmaceutical exposure to caffeine and its combination with salicylic acid in mussel Mytilus galloprovincialis.

Urban and hospital-sourced pharmaceuticals are continuously discharged into aquatic environments, threatening biota. To date, their impact as single compounds has been widely investigated, whereas few information exists on their effects as mixtures. We assessed the time-dependent biological impact induced by environmental concentrations of caffeine alone (CAF; 5 ng/L to 10 µg/L) and its combination with salicylic acid (CAF+SA; 5 ng/L+0.05 µg/L to 10 µg/L+100 µg/L) on gills of mussel Mytilus galloprovincialis during a 12-day exposure. Although no histological alteration was observed in mussel gills, haemocyte infiltration was noticed at T12 following CAF+SA exposure, as confirmed by flow cytometry with increased hyalinocytes. Both the treatments induced lipid peroxidation and cholinergic neurotoxicity, which the antioxidant system was unable to counteract. We have highlighted the biological risks posed by pharmaceuticals on biota under environmental scenarios, contributing to the enhancement of ecopharmacovigilance programmes and amelioration of the efficacy of wastewater treatment plants.

Metabolomic disorders unveil hepatotoxicity of environmental microplastics in wild fish Serranus scriba (Linnaeus 1758).

Coastal areas are worldwide subject to large inputs of anthropogenic wastes that are discharged directly into inshore waters, where they will be weathered into small microplastics (MPs) of up to a size <20 µm. This study provides information about the presence of small environmental MPs (≤3 µm) in the liver of adult benthopelagic fish Serranus scriba (Linnaeus 1758), caught from three coastal regions in Tunisia distinguished by different patterns of human activity. Polymer composition in fish liver was identified using Raman microspectroscopy. Results revealed differences in the abundance, size distribution and presence of plastic additives over the investigated sites. Polyethylene-vinyl acetate (PEVA: 34% particles/g of tissue), high density polyethylene (HDPE: 24.4%) and the two smaller size classes, i.e. 3-1.2 µm and 1.2-0.45 µm, were the most abundant MPs types and size distribution found, respectively, in Bizerte channel (BC) site (Bizerte city, Tunisia). Moreover, at hepatic level data showed a significant site-dependent cytotoxicity expressed by changes in malondialdehyde (MDA) content, presence of reactive oxygen species (ROS) expressed by altered level of catalase (CAT) and glutathione-S-transferase (GST) activities and in the content of metallothioneins (MTs), as well as genotoxicity by changes in the amount of micronucleus (MN), and neurotoxicity by altered activity of acetylcholinesterase (AChE). A innovative metabolomics analysis was also performed to further investigate the distinct patterns of key metabolite changes in the liver of Serranus scriba. A total of 36 metabolites were significantly affected, mainly involved in energy, amino acid and osmolyte metabolism. These findings emphasised for the first time a close relationship between the source, abundance and size ranges of environmental MPs ≤ 3 µm and their hepatotoxicity in wild organisms.

Impact of environmental microplastics alone and mixed with benzo[a]pyrene on cellular and molecular responses of Mytilus galloprovincialis.

The hazard of microplastic (MP) pollution in marine environments is a current concern. However, the effects of environmental microplastics combined with other pollutants are still poorly investigated. Herein, impact of ecologically relevant concentrations of environmental MP alone (50 µg/L) or combined with B[a]P (1 µg/L) was assessed in mussel Mytilus galloprovincialis after a short-term exposure (1 and 3 days) to environmental MP collected from a north-Mediterranean beach. Raman Microspectroscopy (RMS) revealed bioaccumulation in mussel hemolymph of MP, characterized by polyethylene (PE), polyethylene terephthalate (PET), polypropylene (PP), polyethylene vinyl acetate (PEVA) and high-density polyethylene (HDPE), with abundance of MP sized 1.22-0.45 µm. An increase of B[a]P was detected in mussels after 3-day exposure, particularly when mixed with MP. Both contaminants induced cytotoxic and genotoxic effects on hemocytes as determined by lysosomal membrane stability (LMS), micronuclei frequency (FMN), and DNA fragmentation rate by terminal dUTP nick-end labeling (TUNEL). About apoptosis/DNA repair processes, P53 and DNA-ligase increased at 1-day exposure in all conditions, whereas after 3 days increase of bax, Cas-3 and P53 and decrease of Bcl-2 and DNA-ligase were revealed, suggesting a shift towards a cell apoptotic event in exposed mussels. Overall, this study provides new insights on the risk of MP for the marine ecosystem, their ability to accumulate xenobiotics and transfer them to marine biota, with potential adverse repercussion on their health status.

Histological endpoints and oxidative stress transcriptional responses in the Mediterranean mussel Mytilus galloprovincialis exposed to realistic doses of salicylic acid.

Despite the availability of analytic data, little is known about the toxicity of salicylic acid (SA) on aquatic non-target organisms. The present study aimed at evaluating the impact of SA through a short-term exposure of the Mediterranean mussel Mytilus galloprovincialis to five environmentally relevant concentrations of SA. A set of suitable biomarkers was applied at selected time-points on mussel digestive glands, including histological observations and expression of oxidative stress related genes. The obtained results showed a conspicuous hemocytic infiltration among mussel digestive tubules, as confirmed also by a flow cytometric approach that revealed an increase of halinocytes and granulocytes. Interestingly, a significant dose and time dependent decrease in the expression levels of oxidative stress related genes was found in mussels exposed to SA except for the glutathione S-transferase gene that was significantly up-regulated in a time-dependent manner confirming its important role against oxidant species and in the metabolism of pharmaceuticals.

Assessment of the effectiveness of a novel BioFilm-Membrane BioReactor oil-polluted wastewater treatment technology by applying biomarkers in the mussel Mytilus galloprovincialis.

Petrochemical industries and oil refineries are sources of hazardous chemicals into the aquatic environments, and often a leading cause of reduced oxygen availability, thus resulting in adverse effects in biota. This study is an expansion of our previous work on the assessment of the BioFilm-Membrane Bioreactor (BF-MBR) to mitigate the impact of oil-polluted wastewater on marine environments. Specifically, this study evaluated the reduction of selected chemical constituents (hydrocarbons and trace metals) and toxicity related to hypoxia and DNA damage to mussels Mytilus galloprovincialis, before and after treatment of oil-polluted wastewater with the BF-MBR. The application of a multidisciplinary approach provided evidence of the efficiency of BF-MBR to significantly reducing the pollutants load from oily contaminated seawaters. As result, the health status of mussels was preserved by a hypoxic condition due to oily pollutants, as evidenced by the modulation in the gene expression of HIF-1α and PHD and changes in the level of hypotaurine and taurine. Moreover, ameliorative effects in the energy metabolism were also found in mussel gills showing increased levels of glycogen, glucose and ATP, as well as a mitigated genotoxicity was revealed by the Micronucleus and Comet assays. Overall, findings from this study support the use of the BF-MBR as a promising treatment biotechnology to avoid or limiting the compromise of marine environments from oil pollution.

Embryotoxicity of polystyrene microplastics in zebrafish Daniorerio.

In the recent years, increasing scientific and societal concern has been raised over the presence and accumulation of plastic debris in the environment and the effects of microplastics (MPs) that can easily interact with biota. In order to elucidate the impact of MPs at the critical development stages of freshwater fish species, a fish embryo toxicity test was herein performed on the zebrafish Danio rerio, exposed to 10 µm polystyrene MPs at 200 particles/mL for 120 hpf. After exposure, accumulation of MPs in larvae was measured, survival, hatching and larvae development were monitored and the oxidant/anti-oxidant responses and cellular detoxification evaluated. No impact on survival of developing zebrafish was revealed, but a moderate delay in hatching was observed. Alterations in larvae development were recorded with zebrafish exhibiting serious deformities, mainly at the level of column and tail, as well as a compromised integrity of the visual structure of the eyes. Moreover, increased levels of gene transcription involved in the oxidative stress (sod1, sod2 and cat) and in cellular detoxification (gst and cyp) were also detected in MPs-exposed zebrafish larvae. Overall, this research work provides new insights on the ecotoxicological impact of polystyrene MPs on the critical developmental stages of a freshwater fish species, therefore enhancing the current knowledge of the environmental risk posed by MPs to the aquatic ecosystem.

Trace elements (Al, Cd, Cr, Cu, Fe, Mn, Ni, Pb and Zn) in Mytilus galloprovincialis and Tapes decussatus from Faro and Ganzirri Lakes (Sicily, Italy): Flow cytometry applied for hemocytes analysis.

BACKGROUND: Trace elements present in sessile molluscs, are important because they are used in human consumption and it has significantly increased in recent years. While their filtering of the water can lead to their build-up of organic and inorganic materials that can be sampled and analyzed, this can also lead to bioaccumulation of harmful substances, such as essential and non-essential elements, that can harm the human health if in taken in high concentrations or for a long period of time. METHODS: In the present study, the trace metal content (Al, Cd, Cr, Cu, Fe, Mn, Ni, Pb and Zn) of two sessile crustaceans, 20 Mytilus galloprovincialis (mussel) and 20 Tapes decussatus (clam) in Faro and Ganzirri Lakes (Messina, Sicily, Italy) were analyzed. Haemolymph samples were taken on both molluscs in order to analyze the haemocyte population by flow cytometric analysis. Unpaired t-tests were used to determine significant differences for the essential and non-essential metallic elements concentrations in the lake waters and in the tissues of M. galloprovincialis and T. decussatus and for hemocyte populations R1 (halinocytes) and R2 (granulocytes). RESULTS: The results suggested that that in Faro Lake, the tissue Al, Cr and Pb levels in M. galloprovincialis were higher than those for T. decussatus, in contrast to Mn, Fe, Ni, Cu, Zn and Cd, which were higher in T. decussatus. Unpaired t-tests showed that there were significantly higher proportions of halinocytes in M. galloprovincialis versus T. decussatus for both Faro Lake (41.8 % vs. 24.3 %; P < 0.001) and Ganzirri Lake (43.0 % vs. 22.4 %; P < 0.001). In contrast, while there were significantly higher proportions of granulocytes in Faro Lake (21.2 % vs. 9.1 %; P < 0.001), this difference was not seen for the granulocytes of M. galloprovincialis versus T. decussatus in Ganzirri Lake (9.6 % vs. 13.0 %). CONCLUSION: This study shows that M. galloprovincialis and T. decussatus can indeed bioaccumulate some of these metal, such that activation of the immune responses is specific to certain cell types. Future research must focus on the balance of trace elements in the consumption of these shellfish, and analyzes with more sophisticated tools can be used to diagnose the increased concentration of trace elements and the quantification of trace metals from shellfish to clams.

Acute and Sub-Chronic Effects of Microplastics (3 and 10 µm) on the Human Intestinal Cells HT-29.

Due to ingestion of contaminated foods, the human gastrointestinal tract is the most likely site of exposure to microplastics (MPs) with gut barrier dysfunction and intestinal inflammation. Aimed to assess the effects induced by MPs with different granulometry (polystyrene (PS) 3 and 10 µm), we performed an in vitro study by using the human intestinal cell line HT29. As a novelty, we assessed the sub-chronic exposure extending the treatment up to 48 days simulating the in vivo situation. In the range of 100-1600 particles mL-1, both the PS suspensions had moderate cytotoxicity after 24 h with percentages of mortality between 6.7 and 21.6 for the 10 µm and 6.1 and 29.6 for the 3 µm PS. Microscopic observation highlighted a more pronounced lysosomal membrane permeabilization in HT29 exposed to PS 3µm. Reactive oxygen species production was higher in cells exposed to PS 10 µm, but sub-chronic exposure highlighted the ability of the cells to partially neutralize this effect. Comet-assay confirmed the temporary oxidative damage that was PS-induced. Overall, considering the very fast turnover of intestinal cells, the increase in cell mortality, equal to 25% and 11% for 3 and 10 µm PS-MPs for each time point, could trigger intestinal disorders due to prolonged exposure.

Time-dependent metabolic disorders induced by short-term exposure to polystyrene microplastics in the Mediterranean mussel Mytilus galloprovincialis.

In the modern society, plastic has achieved a crucial status in a myriad of applications because of its favourable properties. Despite the societal benefits, plastic has become a growing global concern due to it is persistence and bioavailability as microplastics (MPs) to aquatic biota. In order to provide mechanistic insights into the early toxicity effects of MPs on aquatic invertebrates, a short-term (up to 72 h) exposure to 3 µm red polystyrene MPs (50 particles/mL) was conducted on marine mussels Mytilus galloprovincialis, selected as model organism for their ability to ingest MPs and their commercial relevance. The use of protonic Nuclear Magnetic Resonance (1H NMR)-based metabolomics, combined with chemometrics, enabled a comprehensive exploration at fixed exposure time-points (T24, T48, T72) of the impact of MPs accumulated in mussel digestive glands, chosen as the major site for pollutants storage and detoxification processes. In detail, 1H NMR metabolic fingerprints of MP-treated mussels were clearly separated from control and grouped for experimental time-points by a Principal Component Analysis (PCA). Numerous metabolites, including amino acids, osmolytes, metabolites involved in energy metabolism, and antioxidants, participating in various metabolic pathways significantly changed over time in MP-exposed mussel digestive glands related to control, reflecting also the fluctuations in MPs accumulation and pointing out the occurrence of disorders in amino acid metabolism, osmotic equilibrium, antioxidant defense system and energy metabolism. Overall, the present work provides the first insights into the early mechanisms of toxicity of polystyrene MPs in marine invertebrates.

Organ-Specific Metabolome Deciphering Cell Pathways to Cope with Mercury in Wild Fish (Golden Grey Mullet Chelon auratus).

Metabolomics is a powerful approach in evaluating the health status of organisms in ecotoxicological studies. However, metabolomics data reflect metabolic variations that are attributable to factors intrinsic to the environment and organism, and it is thus crucial to accurately evaluate the metabolome of the tissue/organ examined when it is exposed to no stressor. The metabolomes of the liver and gills of wild golden grey mullet (Chelon auratus) from a reference area were analyzed and compared by proton nuclear magnetic resonance (1H NMR)-based metabolomics. Both organs were characterized by amino acids, carbohydrates, osmolytes, nucleosides and their derivatives, and miscellaneous metabolites. However, similarities and differences were revealed in their metabolite profile and related to organ-specific functions. Taurine was predominant in both organs due to its involvement in osmoregulation in gills, and detoxification and antioxidant protective processes in liver. Environmental exposure to mercury (Hg) triggered multiple and often differential metabolic alterations in fish organs. Disturbances in ion-osmoregulatory processes were highlighted in the gills, whereas differential impairments between fish organs were pointed out in energy-producing metabolic pathways, protein catabolism, membrane stabilization processes, and antioxidant defense system, reflecting the induction of organ-specific adaptive and defensive strategies. Overall, a strict correlation between metabolites and organ-specific functions of fish gills and liver were discerned in this study, as well as organ-specific cytotoxicity mechanisms of Hg in fish.