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Perfluorooctanoic acid (PFOA) is a persistent organic pollutant that accumulates in the human body, leading to major health issues. Upon oral uptake, the gastrointestinal tract is the first biological barrier against PFOA. However, the localization of PFOA and its impact on the intestinal wall are largely unknown. Here we achieve a breakthrough in the knowledge of intestinal absorption, intracellular fate and toxicity of PFOA using in vitro assays combined with novel analytical imaging techniques. For the first time, we localized PFOA in the cytosol of Caco-2 cells after acute exposure using high spatial resolution mass spectrometry imaging, and we estimated the PFOA cytosolic concentration. Knowing that PFOA enters and accumulates in the intestinal cells, we also performed common toxicity assays assessing cell metabolic activity, membrane integrity, oxidative stress response, and cell respiration. This study integrating powerful analytical techniques with widely used toxicology assays provides insightful information to better understand potential negative impacts of PFOA and opens new opportunities in toxicology and life science in general., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Due to its amphiphilic structure, lignin has the potential to stabilize emulsions via adsorption at the oil/water interface. By converting lignin into nanoparticles, we can significantly enhance its emulsion-stabilizing capabilities through a Pickering-type stabilization mechanism. Two essential elements may be modified to fine-tune emulsion stability: the size of the lignin nanoparticles (LNPs) and the physicochemical nature of the lipid phase. In this context, we highlight the behavior and utility of unmodified LNPs in the preparation of Pickering emulsions made up of water and a complex bio-based pharmaceutical-grade wax that can be used for the formulation of lipid carriers. As a proof-of-concept, we employ the developed Pickering emulsions to encapsulate indocyanine green (ICG), an FDA-approved dye commonly used in medical imaging applications. We demonstrate that ultra-small LNPs are well-suited for the colloidal stabilization of pharmaceutical wax ester micro beads. This stabilization does not require any lignin modification. Additionally, we present evidence that our new lipid/lignin hybrid carrier has potential as a new drug delivery system., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2024. Published by Elsevier B.V.)

Nelfinavir is an HIV protease inhibitor that has been widely prescribed as a component of highly active antiretroviral therapy, and has been reported to exert in vitro antiviral activity against SARS-CoV-2. We here assessed the effect of Nelfinavir in a SARS-CoV-2 infection model in hamsters. Despite the fact that Nelfinavir, [50 mg/kg twice daily (BID) for four consecutive days], did not reduce viral RNA load and infectious virus titres in the lung of infected animals, treatment resulted in a substantial improvement of SARS-CoV-2-induced lung pathology. This was accompanied by a dense infiltration of neutrophils in the lung interstitium which was similarly observed in non-infected hamsters. Nelfinavir resulted also in a marked increase in activated neutrophils in the blood, as observed in non-infected animals. Although Nelfinavir treatment did not alter the expression of chemoattractant receptors or adhesion molecules on human neutrophils, in vitro migration of human neutrophils to the major human neutrophil attractant CXCL8 was augmented by this protease inhibitor. Nelfinavir appears to induce an immunomodulatory effect associated with increasing neutrophil number and functionality, which may be linked to the marked improvement in SARS-CoV-2 lung pathology independent of its lack of antiviral activity. Since Nelfinavir is no longer used for the treatment of HIV, we studied the effect of two other HIV protease inhibitors, namely the combination Lopinavir/Ritonavir (Kaletra™) in this model. This combination resulted in a similar protective effect as Nelfinavir against SARS-CoV2 induced lung pathology in hamsters., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Environmental stressors are assessed through methods that quantify their impacts on a wide range of metrics including species density, growth rates, reproduction, behaviour and physiology, as on host-pathogen interactions and immunocompetence. Environmental stress may induce additional sublethal effects, like mutations and epigenetic signatures affecting offspring via germline mediated transgenerational inheritance, shaping phenotypic plasticity, increasing disease susceptibility, tissue pathologies, changes in social behaviour and biological invasions. The growing diversity of pollutants released into aquatic environments requires the development of a reliable, standardised and 3R (replacement, reduction and refinement of animals in research) compliant in vitro toolbox. The tools have to be in line with REACH regulation 1907/2006/EC, aiming to improve strategies for potential ecotoxicological risks assessment and monitoring of chemicals threatening human health and aquatic environments. Aquatic invertebrates' adult stem cells (ASCs) are numerous and can be pluripotent, as illustrated by high regeneration ability documented in many of these taxa. This is of further importance as in many aquatic invertebrate taxa, ASCs are able to differentiate into germ cells. Here we propose that ASCs from key aquatic invertebrates may be harnessed for applicable and standardised new tests in ecotoxicology. As part of this approach, a battery of modern techniques and endpoints are proposed to be tested for their ability to correctly identify environmental stresses posed by emerging contaminants in aquatic environments. Consequently, we briefly describe the current status of the available toxicity testing and biota-based monitoring strategies in aquatic environmental ecotoxicology and highlight some of the associated open issues such as replicability, consistency and reliability in the outcomes, for understanding and assessing the impacts of various chemicals on organisms and on the entire aquatic environment. Following this, we describe the benefits of aquatic invertebrate ASC-based tools for better addressing ecotoxicological questions, along with the current obstacles and possible overhaul approaches., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

The aim of the present study was the assessment of the sub-chronic effects of silver (AgNPs) and gold nanoparticles (AuNPs) of 40 nm primary size either stabilised with citrate (CIT) or coated with polyethylene glycol (PEG) on the freshwater invertebrate Gammarus fossarum. Silver nitrate (AgNO 3 ) was used as a positive control in order to study the contribution of silver ions potentially released from AgNPs on the observed effects. A multibiomarker approach was used to assess the long-term effects of AgNPs and AuNPs 40 nm on molecular, cellular, physiological and behavioural responses of G. fossarum. Specimen of G. fossarum were exposed for 15 days to 0.5 and 5 µgL -1 of CIT and PEG AgNPs and AuNPs 40 nm in the presence of food. A significant uptake of both Ag and Au was observed in exposed animals but was under the toxic threshold leading to mortality of G. fossarum. Silver nanoparticles (CIT-AgNPs and PEG-AgNPs 40 nm) led to an up-regulation of Na + K + ATPase gene expression. An up-regulation of Catalse and Chitinase gene expressions due to exposure to PEG-AgNPs 40 nm was also observed. Gold nanoparticles (CIT and PEG-AuNPs 40 nm) led to an increase of CuZnSOD gene expression. Furthermore, both AgNPs and AuNPs led to a more developed digestive lysosomal system indicating a general stress response in G. fossarum. Both AgNPs and AuNPs 40 nm significantly affected locomotor activity of G. fossarum while no effects were observed on haemolymphatic ions and ventilation., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Colloidal silver products are sold for a wide range of disinfectant and health applications. This has increased the potential for human exposure to silver nanoparticles (AgNPs) and ions (Ag + ), for which oral ingestion is considered to be a major route of exposure. Our objective was to evaluate and compare the toxicity of two commercially available colloidal silver products on two human intestinal epithelial models under realistic exposure conditions. Mesosilver™ and AgC were characterized and a concentration range between 0.1 and 12 μg/mL chosen. Caco-2 cells vs. co-culture of Caco-2 and mucus-secreting HT29-MTX cells (90/10) were used. Repeated exposure was carried out to determine cell viability over 18 days of cell differentiation in 24-well plates. Selected concentrations (0.1, 1, and 3 μg/mL) were tested on cells cultured in E-plates and Transwells with the same repeated exposure regimen, to determine cell impedance, and cell viability and trans-epithelial electrical resistance (TEER), respectively. Silver uptake, intracellular localisation, and translocation were determined by CytoViva™, HIM-SIMS, and ICP-MS. Genotoxicity was determined on acutely-exposed proliferating Caco-2 cells by γH2AX immunofluorescence staining. Repeated exposure of a given concentration of AgC, which is composed solely of ionic silver, generally exerted more toxic effects on Caco-2 cells than Mesosilver™, which contains a mix of AgNPs and ionic silver. Due to its patchy structure, the presence of mucus in the Caco-2/HT29-MTX co-culture only slightly mitigated the deleterious effects on cell viability. Increased genotoxicity was observed for AgC on proliferating Caco-2 cells. Silver uptake, intracellular localisation, and translocation were similar. In conclusion, Mesosilver™ and AgC colloidal silver products show different levels of gut toxicity due to the forms of distinct silver (AgNPs and/or Ag + ) contained within. This study highlights the applicability of high-resolution (chemical) imaging to detect and localize silver and provides insights into its uptake mechanisms, intracellular fate and cellular effects., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Because of their antimicrobial properties, silver nanoparticles are increasingly incorporated in food-related and hygiene products, which thereby could lead to their ingestion. Although their cytotoxicity mediated by oxidative stress has been largely studied, their effects on inflammation remain controversial. Moreover, the involvement of silver ions (originating from Ag0 oxidation) in their mode of action is still unclear. In this context, the present study aims at assessing the impact of silver nanoparticles on the secretion of the pro-inflammatory chemokine interleukin-8 by Caco-2 cells forming an in vitro model of the intestinal mucosal barrier. Silver nanoparticles induced a vectorized secretion of interleukin-8 towards the apical compartment, which is found in the medium 21 h after the incubation. This secretion seems mediated by Nrf2 signalling pathway that orchestrates cellular defense against oxidative stress. The soluble silver fraction of silver nanoparticles suspensions led to a similar amount of secreted interleukin-8 than silver nanoparticles, suggesting an involvement of silver ions in this interleukin-8 secretion., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Organic-inorganic perovskite solar cells (PSCs) are promising candidates as photovoltaic cells. Recently, they have attracted significant attention due to certified power conversion efficiencies exceeding 23%, low-cost engineering, and superior electrical/optical characteristics. These PSCs extensively utilize a perovskite-structured composite with a hybrid of Pb-based nanomaterials. Operation of them may cause the release of Pb-based nanoparticles. However, limited information is available regarding the potential toxicity of Pb-based PSCs on various organisms. This study conducted a battery of in vitro and in vivo toxicity bioassays for three quintessential Pb-based PSCs (CH 3 NH 3 PbI 3 , NHCHNH 3 PbBr 3 , and CH 3 NH 3 PbBr 3 ) using progressively more complex forms of life. For all species tested, the three different perovskites had comparable toxicities. The viability of Caco-2/TC7 cells was lower than that of A549 cells in response to Pb-based PSC exposure. Concentration-dependent toxicity was observed for the bioluminescent bacterium Vibrio fischeri, for soil bacterial communities, and for the nematode Caenorhabditis elegans. Neither of the tested Pb-based PSCs particles had apparent toxicity to Pseudomonas putida. Among all tested organisms, V. fischeri showed the highest sensitivity with EC 50 values (30 min of exposure) ranging from 1.45 to 2.91 mg L -1 . Therefore, this study recommends that V. fischeri should be preferably utilized to assess. PSC toxicity due to its increased sensitivity, low costs, and relatively high throughput in a 96-well format, compared with the other tested organisms. These results highlight that the developed assay can easily predict the toxic potency of PSCs. Consequently, this approach has the potential to promote the implementation of the 3Rs (Replacement, Reduction, and Refinement) principle in toxicology and decrease the dependence on animal testing when determining the safety of novel PSCs., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The specific and unique properties of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs), make them of high interest for different scientific and industrial applications. Their increasing use will inevitably lead to their release in the environment and aquatic ecosystems where they may represent a threat to aquatic organisms. Being a widespread and important component of the aquatic macroinvertebrate assemblage, amphipods and more specifically Gammarus fossarum will certainly be exposed to AgNPs and AuNPs. For these reasons, G. fossarum was selected as model organism for this study. The aim of the present work was the evaluation of the influence of both size (20, 40 and 80 nm) and surface coating (citrate CIT, polyethylene glycol PEG) on the acute toxicity of AgNPs and AuNPs on G. fossarum. We investigated the effects of AgNPs and AuNPs on the uptake by G. fossarum, NP tissue distribution and the expression of stress related genes by the use of ICP-MS, NanoSIMS50, Cytoviva®, and Rt-qPCR, respectively. Ag and Au bioaccumulation revealed a significant surface-coating dependence, with CIT-AgNPs and CIT-AuNPs showing the higher bio-accumulation potential in G. fossarum as compared to PEG-NPs. Opposite to that, no size-dependent effects on the bioaccumulation potential was observed. SIMS imaging and CytoViva® revealed an influence of the type of metal on the tissue distribution after uptake, with AgNPs detected in the cuticle and the gills of G. fossarum, while AuNPs were detected in the gut area. Furthermore, AgNPs were found to up-regulate CuZnSOD gene expression while AuNPs led to its down-regulation. Modulation of SOD may indicate generation of reactive species of oxygen and a possible activation of antioxidant defence in order to prevent and defend the organism from oxidative stress. However, further investigations are still needed to better define the mechanisms underlying the observed AgNPs and AuNPs effects., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The use of silver nanomaterials in everyday products, such as cosmetics, textiles, certain types of packaging, etc. is increasing, leading to their release into the environment, including aquatic ecosystems. This last point initiated this investigation on the toxicological effects of Ag nanoparticles (Ag NPs) in the aquatic model organism Danio rerio. For this purpose, zebrafish larvae were exposed to 20nm bare Ag NPs at different concentrations and AgNO 3 , used as a positive control for Ag + ions toxicity, at the beginning of their foraging behaviour to determine adverse effects on fitness parameters. We used secondary ion mass spectrometry (SIMS) to determine the localization of Ag and transcriptomics (microarray) to determine the toxicity at the level of gene expression in fish larvae. Exposure to Ag NPs did not result in adverse effects on survival and growth of the fish. However, SIMS analysis showed that Ag NPs mainly concentrate around liver blood vessels and in the interstitial tissue between the intestine and the liver. Gene expression profiles revealed that AgNO 3 and Ag NPs impacted common pathways, suggesting similar targets, such as the phototransduction system. However, the Ag NPs showed a broader set of genes impacted following the exposure, including the circadian clock regulation and the photoreception, suggesting specific particle-related effects in addition to those induced by ions., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The P-glycoprotein (P-gp, ABCB1) and multidrug resistance associated protein 1 (MRP1), important members of the ABC (ATP-binding cassette) transporters, protect cells and organisms via efflux of xenobiotics and are responsible for the phenomenon of multidrug or multixenobiotic resistance (MXR). In this study we first evaluated, in vitro, the interaction of silver nanoparticles (Ag NPs, 20, 23 and 27nm), Ag 200nm particles and Ag ions (AgNO3) with MXR efflux transporters using MDCKII and the P-gp over-expressing MDCKII-MDR1 cells and calcein-AM as a substrate of the transporters. Next the in vivo modulation of MXR activity was studied in Daphnia magna juveniles with the model P-gp and MRP1 inhibitors verapamil-HCl and MK571, respectively. The common environmental contaminants perfluorooctane sulfonate and bisphenol A, previously observed to interfere with the P-gp in vitro, also inhibited the efflux of calcein in vivo. Small-sized Ag NPs (with biomolecules present on the surface) and AgNO3 inhibited the MXR activity in daphnids and MDCKII-MDR1 cells, but abcb1 gene expression remained unchanged. Both Ag NPs and dissolved ions contributed to the effects. This study provides evidence of the interference of Ag NPs and AgNO3 with the MXR activity both in vitro and in D. magna, and should be taken into account when Ag NP toxicity is assessed., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Amphipods are one of the most important components of freshwater ecosystems. Among them, gammarids are the most widespread group in Europe and are often used as bioindicators and model organisms in ecotoxicology. However, their use, especially of Gammarus fossarum for the study of the environmental impact of nanoparticles, has been rather limited so far. G. fossarum was selected to assess effects of well-characterized chemically synthesized silver nanoparticles (AgNPs 20nm and 200nm) and "green" laboratory synthetized (from plant leaf extracts) AgNPs (AgNPs 23nm and 27nm). AgNO3 was used as a positive control to compare AgNPs effects and silver ions effects. A multibiomarker approach was used to investigate the sub-lethal effects of AgNPs on physiological and behavioural responses of G. fossarum. Two different experiments were carried out. In a preliminary experiment, two populations of G. fossarum (G.f1 and G.f2) were tested for sensitivity differences and the most sensitive one was exposed, in a final experiment, to sub-lethal concentrations of AgNO3 and the most toxic AgNPs. AgNO3 and AgNPs 23nm led to a significant decrease in survival rates, osmoregulation and locomotor activity. Ag internalisation, performed with Secondary Ion Mass Spectrometry (SIMS), showed the presence of silver in gills of G.f2 exposed to AgNPs 23 and 27nm. This study highlighted the influence of method of synthesis on ion release, uptake and toxic effects of AgNPs on G. fossarum. Osmoregulation appeared to be an effective biomarker indicating the physiological health status of G. fossarum. Locomotor activity, which was the most impacted response, reflects the potential effects of released ions from AgNPs 23nm at the population level as locomotion is necessary for foraging, finding mates and escaping from predators. Therefore, we propose G. fossarum as a suitable model for environmental nanotoxicology, providing information both at individual and population levels., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Hexabromocyclododecane (HBCD) is a brominated flame retardant known for its low acute toxicity as observed in animal experiments. However, HBCD exposure can affect liver functioning and thyroid hormone (TH) status. As exact mechanisms are unknown and only limited toxicological data exists, a gel-based proteomic approach was undertaken. In a eu- and hypothyroid female rat model, rats were exposed to 3 and 30 mg/kg bw/day HBCD for 7 days via their diet, and exposure was related to a range of canonical endpoints (hormone status, body weight) available for these animals. Alterations in the liver proteome under HBCD exposure were determined in comparison with patterns of control animals, for both thyroid states. This revealed significantly changed abundance of proteins involved in metabolic processes (gluconeogenesis/glycolysis, amino acid metabolism, lipid metabolism), but also in oxidative stress responses, in both euthyroid and hypothyroid rats. The results provide a more detailed picture on the mechanisms involved in these alterations, e.g. at the protein level changes of the proposed influence of HBCD on the lipid metabolism. Present results show that proteomic approaches can provide further mechanistic insights in toxicological studies., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Release of chemicals and fluctuation in oxygen content in the aquatic environment represent hazards for fish health. The present study aims at assessing the genotoxic impact of low concentration exposures to waterborne uranium (U), dietary methyl mercury (MeHg) and hyperoxia in zebrafish by using the RAPD-PCR quantitative method. A significant increase of the number hybridization sites was observed in fish exposed to 30μgU/L and 100μgU/L and hyperoxia. In fish exposed to MeHg (13.5μg Hg/g, dry weight) no change in the number of hybridization sites were found, however, the frequency of PCR products showed significant variation. The mechanisms of toxicity leading to DNA damage in fish exposed to waterborne uranium, mercury and hyperoxia are discussed and the results from the literature given by the comet assay, micronucleus test and RAPD-PCR method compared. The study provides new data regarding the genotoxic effects of MeHg, hyperoxia and low U concentrations (30μgU/L) in fish. The present work highlights the use of the RAPD-PCR as a sensitive method in the assessment of chemically-induced DNA damage in animals., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Genotoxic effects of a polymetallic pollution gradient displayed by the Lot River and one of its tributary have been assessed on zebrafish Danio rerio. Three methods were compared: RAPD-PCR, the comet assay, and 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodG) formation. The fishes were exposed for 14 days to waters collected from three stations: Joanis, a site polluted by cadmium (Cd) and zinc (Zn) (mean concentrations: 15 μg Cd/L and 550 μg Zn/L), Bouillac (mean concentrations: 0.55 μg Cd/L and 80 μg Zn/L), and Boisse-Penchot, a reference station (mean concentrations: <0.05 μg Cd/L and 7 μg Zn/L). The quantitative RAPD-PCR methodology proved to be sensitive enough to unmask metal genotoxicity after 3 and 7 days of exposure to Joanis water and after 14 days to Bouillac water, whereas the comet assay only detected DNA damages at the most contaminated station (Joanis). The 8-oxodG quantification was not sensitive enough to be used in zebrafish under these environmental conditions., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Genotoxic effects of cadmium on zebra fish Danio rerio have been assessed by random amplified polymorphic DNA and real time PCR, followed by a comparison of the melting temperature patterns between each amplification reaction. Fish were exposed to two concentrations of cadmium chloride dissolved in the medium (1.9+/-0.6 microg Cdl(-1), C(1); 9.6+/-2.9 microg Cdl(-1), C(2)) for 21 days. A discriminative RAPD probe, OPB11, was first selected producing differential band patterns between control and metal-exposed genomic DNAs. RAPD-PCR showed an increase in the relative hybridization efficiency of OPB11 on the genomic DNAs coming from fish exposed to both Cd concentrations as compared to the control condition. In addition, the RAPD-PCR melting temperature patterns showed that with the OPB11 probe, the frequency of PCR products whose fusion temperature belongs to the [86-87 degrees C] interval decreased with Cd contamination, whereas an increase of frequency for the [78-80 degrees C] and [85-86 degrees C] intervals was correlated with Cd exposure., ((c) 2009 Elsevier Inc. All rights reserved.)

Brain arteriovenous malformations (BAVMs) are a rare but potentially devastating hemorrhagic disease. Transforming growth factor-beta signaling is required for proper vessel development, and defective transforming growth factor-beta superfamily signaling has been implicated in BAVM pathogenesis. We hypothesized that expression of the transforming growth factor-beta activating integrin, alphavbeta8, is reduced in BAVMs and that decreased beta8 expression leads to defective neoangiogenesis. We determined that beta8 protein expression in perivascular astrocytes was reduced in human BAVM lesional tissue compared with controls and that the angiogenic response to focal vascular endothelial growth factor stimulation in adult mouse brains with local Cre-mediated deletion of itgb8 and smad4 led to vascular dysplasia in newly formed blood vessels. In addition, common genetic variants in ITGB8 were associated with BAVM susceptibility, and ITGB8 genotypes associated with increased risk of BAVMs correlated with decreased beta8 immunostaining in BAVM tissue. These three lines of evidence from human studies and a mouse model suggest that reduced expression of integrin beta8 may be involved in the pathogenesis of sporadic BAVMs.

The neurotoxic compound methylmercury (MeHg) is a commonly encountered pollutant in the environment, and constitutes a hazard for human health through fish eating. To study the impact of MeHg on mitochondrial structure and function, we contaminated the model fish species Danio rerio with food containing 13 microg of MeHg per gram, an environmentally relevant dose. Mitochondria from contaminated zebrafish muscles presented structural abnormalities under electron microscopy observation. In permeabilized muscle fibers, we observed, a strong inhibition of both state 3 mitochondrial respiration and functionally isolated maximal cytochrome c oxidase (COX) activity after 49 days of MeHg exposure. However, the state 4 respiratory rate remained essentially unchanged. This suggested a defect at the level of ATP synthesis. Accordingly, we measured a dramatic decrease in the rate of ATP release by skinned muscle fibers using either pyruvate and malate or succinate as respiratory substrates. However, the amount and the assembly of the ATP synthase were identical in both control and contaminated muscle mitochondrial fractions. This suggests that MeHg induced a decoupling of mitochondrial oxidative phosphorylation in the skeletal muscle of zebrafish. Western blot analysis showed a 30% decrease of COX subunit IV levels, a 50% increase of ATP synthase subunit alpha, and a 40% increase of the succinate dehydrogenase Fe/S protein subunit in the contaminated muscles. This was confirmed by the analysis of gene expression levels, using RT-PCR. Our study provides a basis for further analysis of the deleterious effect of MeHg on fish health via mitochondrial impairment.

Trophic interactions between pulmonary epithelial and mesenchymal cell types, known as the epithelial-mesenchymal trophic unit (EMTU), are crucial in lung development and lung disease. Transforming growth factor (TGF)-beta is a key factor in mediating these interactions, but it is expressed in a latent form that requires activation to be functional. Using intact fetal tracheal tissue and primary cultures of fetal tracheal epithelial cells and fibroblasts, we demonstrate that a subset of integrins, alpha(v)beta(6) and alpha(v)beta(8), are responsible for almost all of the TGF-beta activation in the EMTU. Both alpha(v)beta(8) and alpha(v)beta(6) contribute to fetal tracheal epithelial activation of TGF-beta, whereas only alpha(v)beta(8) contributes to fetal tracheal fibroblast activation of TGF-beta. Interestingly, fetal tracheal epithelial alpha(v)beta(8)-mediated TGF-beta activation can be enhanced by phorbol esters, likely because of the increased activity of MT1-MMP, an essential co-factor in alpha(v)beta(8)-mediated activation of TGF-beta. Autocrine alpha(v)beta(8)-mediated TGF-beta activation by fetal tracheal fibroblasts results in suppression of both transcription and secretion of hepatocyte growth factor, which is sufficient to affect phosphorylation of the airway epithelial hepatocyte growth factor receptor, c-Met, as well as airway epithelial proliferation in a co-culture model of the EMTU. These findings elucidate the function and complex regulation of integrin-mediated activation of TGF-beta within the EMTU.

Brain hemorrhage is a severe complication of both neoplastic and nonneoplastic brain disease. Mice deficient in the alpha(v)beta8 integrin display defective brain vessel formation resulting in hemorrhage and perinatal death, but the mechanism of brain hemorrhage is unknown. Because the alpha(v)beta8 integrin is expressed by astrocytes and not expressed by endothelium, paracrine interactions between astrocytes and endothelial cells could contribute to the maintenance of brain vessel integrity. We have investigated the mechanisms underlying astrocytic-endothelial paracrine signaling and have found that integrin-mediated activation of transforming growth factor (TGF)-beta by astrocytes influences endothelial cell function. Thus, we identified the integrin alpha(v)beta8 in human perivascular glial cell processes surrounding developing blood vessels. Human astrocytic alpha(v)beta8 was a major cell surface receptor for latent TGF-beta, and alpha(v)beta8-dependent activation of TGF-beta was the major mechanism of TGF-beta activation in primary cultures of astrocytes or freshly dissociated fetal brain cells. This activation of TGF-beta was sufficient to inhibit endothelial migration in fibrin gels and to alter expression of genes affecting proteolytic and angiogenic pathways. Taken together, our data suggest that astrocytic alpha(v)beta8 acts as a central regulator of brain vessel homeostasis through regulation of TGF-beta activation and expression of TGF-beta-responsive genes that promote vessel differentiation and stabilization, most notably plasminogen activator inhibitor-1 and thrombospondin-1.

Transforming growth factor (TGF)-beta is a potent multifunctional cytokine that is an essential regulator of epithelial proliferation. Because TGF-beta is expressed almost entirely in a latent state in vivo, a major source of regulation of TGF-beta function is its activation. A subset of integrins, alphavbeta8 and alphavbeta6, which are expressed in the human airway, has recently been shown to activate latent TGF-beta in vitro, suggesting a regulatory role for integrins in TGF-beta function in vivo. Here we have developed a novel, biologically relevant experimental model of human airway epithelium using intact human bronchial tissue. We have used this model to determine the function of integrin-mediated activation of TGF-beta in the airway. In human bronchial fragments cultured in vitro, authentic epithelial-stromal interactions were maintained and integrin and TGF-beta expression profiles correlated with profiles found in normal lung. In addition, in this model, we found that either the integrin alphavbeta8 or TGF-beta could inhibit airway epithelial cell proliferation. Furthermore, we found that one mechanism of integrin-alphavbeta8-dependent inhibition of cell proliferation was through activation of TGF-beta because anti-beta8 antibody blocked the majority (76%) of active TGF-beta released from bronchial fragments. These data provide compelling evidence for a functional role for integrin-mediated activation of TGF-beta in control of human airway epithelial proliferation in vivo.