Subchronic exposure to fit-for-purpose polystyrene nanoplastics impairs gut and liver health at low doses with non-monotonic effects in a diet-dependent manner.

Microplastic pollution is a growing global concern, yet the human health effects of exposure to microplastics, particularly in interaction with dietary factors, are not well understood. In this study, we investigated the impact of polyethylene microplastics (PE-MPs) on intestinal homeostasis in mice fed either a normal (ND) or Western (WD) diet for 90 days. Mice were orally exposed to PE-MPs (10 mg/kg body weight/day) throughout the experimental period. Under ND conditions, PE-MP exposure induced subtle changes in gut microbiota composition and short-chain fatty acid (SCFA) production, including a significant decrease in propionate, without overt intestinal inflammation. In contrast, under WD conditions, PE-MP exposure markedly exacerbated the diet-induced phenotype. WD-fed mice exposed to PE-MPs exhibited increased body weight gain, decreased expression of tight-junction related-genes and elevated faecal levels of the inflammatory marker lipocalin-2 and secretory IgA. These effects were accompanied by decreased abundances of Akkermansiaceae and Saccharimonadaceae, enrichment of Peptostreptococcaceae, and reduced caecal levels of propionate, isobutyrate, isovalerate and caproate. Together, these findings indicate that PE-MP exposure exacerbates WD effects, resulting in altered gut microbiota composition and metabolic activity, and increased intestinal inflammation. This study emphasises the importance of taking nutritional status into account when evaluating the toxicity of MPs and sheds new light on how this emerging class of food contaminant interacts with dietary stressors to shape host-microbiome relationships.

ABSTRACT Intestinal paracellular permeability was analyzed ex vivo by incubation of tissue segments at 0°C with the fluorescent dyes FM1‐43FX (FM) or TRITC‐dextran 3 kDa lysine‐fixable (TD3L) and confocal microscopy in (i) healthy mice and (ii) mice submitted to chronic stress or lipid diets. In the small intestine of healthy mice, FM staining was restricted to the apical surface of enterocytes but fully penetrated around Goblet cells, enteroendocrine cells, tuft cells, and apoptotic cells. The same cell types were similarly labeled in the colon when located on the tissue surface but not within the crypts. TD3L exhibited a comparable labeling pattern but also showed moderate staining of the basolateral surface of enterocytes at the tips of small intestinal villi, and also substantial penetration around colonic epithelial cells at the surface or top of crypts. The study reveals patterns of permeability likely corresponding to the “leak” pathway of paracellular transport through the intestinal epithelium, because transcellular endocytosis is blocked at 0°C. This pathway is found around specific cell populations involved in the luminal detection of food, antigens, microbes, or their secretions. These trigger immune, neural, and tissue responses that maintain intestinal homeostasis. Chronic stress induced by glucocorticoid exposure increased FITC‐dextran 4 kDa permeability in vivo. Using FM, increased paracellular permeability was also detected ex vivo and selectively localized in the colon of stressed mice. A single oral administration of a lipid‐rich food also increased ex vivo permeability around jejunal enterocytes. Pathophysiological increases in paracellular permeability are therefore detectable using the FM methodology.

Perfluorooctanoic acid (PFOA) is a persistent organic pollutant recognised for its environmental presence and bioaccumulation, despite regulatory restrictions under the Stockholm Convention. While systemic effects of PFOA have been studied extensively, its interaction with the first biological barrier encountered after oral exposure, the gastrointestinal barrier, remains poorly characterised. In this study, we evaluated the fate, uptake and effects of PFOA in human intestinal cell culture models after acute (24 h) and repeated (11-day) exposures. Using radiolabelled PFOA profiling, we found that PFOA was not metabolised by Caco-2 cells nor by HT29-MTX mucus-producing cells, in monocultures or in co-culture. Mass balance studies revealed higher basolateral passage of PFOA in Caco-2 monocultures compared to Caco-2/HT29-MTX co-cultures, suggesting that the mucus in the co-culture may trap PFOA and limit its passage. These observations led us to conduct further toxicological evaluations in the Caco-2 monoculture model. Acute exposure, while having no effect on cell viability, increased cellular ATP at highest exposure concentration. Repeated exposure led to a progressive concentration-dependent decrease in trans-epithelial electrical resistance, indicating a compromised barrier function. This effect was not linked to changes in tight junction gene expression, but rather attributed to cell death. High-resolution chemical imaging revealed intracellular accumulation of PFOA in Caco-2 cell cytosol after repeated exposure. These findings highlight the potential of repeated exposure to PFOA, even at low concentrations, to impair intestinal barrier integrity, which may have implications for systemic absorption and toxicity.

Nanoplastics (NPLs), the presence in the environment of which was considered only “highly plausible” until recently, have become the focus of environmental and ecotoxicological studies. However, up to know, little is known about the potential risks of NPLs to human health. In this review, we provide an overview of the evidence of a specific impact of NPLs on human digestive health reported to date. We focus on the different sources of oral exposure to NPLs, including food packaging, food and beverages. We then summarize the toxicological effects of such exposure on the digestive ecosystem in vitro and in vivo in rodents. Importantly, gut toxicity results should be assessed with the type of NPL model materials employed, due to the critical influence of their chemical and physical properties. As a result, we thoroughly describe NPLs with their source, chemical composition and physicochemical behavior to emphasize the lack of NPL characterization and/or model materials. Finally, we propose avenues for interdisciplinary studies at the interface of toxicology and chemistry, with a view to achieving appropriate scientific assessments of the risks to gut health posed by NPLs, and improvements in their management.

Silver biodistribution and gut toxicity of two different colloidal silver products were evaluated in mice after oral exposure. Biophysics-based methodologies provided novel insights into (nano)silver uptake, fate and toxicological effects.

Perfluorooctanoic acid (PFOA) is a synthetic perfluorinated chemical classified as a persistent organic pollutant. PFOA has been linked to many toxic effects, including liver injury. Many studies report that PFOA exposure alters serum and hepatic lipid metabolism. However, lipidomic pathways altered by PFOA exposure are largely unknown and only a few lipid classes, mostly triacylglycerol (TG), are usually considered in lipid analysis. Here, we performed a global lipidomic analysis on the liver of PFOA-exposed (high-dose and short-duration) and control mice by combining three mass spectrometry (MS) techniques: liquid chromatography with tandem mass spectrometry (LC–MS/MS), matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), and time-of-flight secondary ion mass spectrometry (TOF-SIMS). Among all hepatic lipids identified by LC–MS/MS analysis, more than 350 were statistically impacted (increased or decreased levels) after PFOA exposure, as confirmed by multi-variate data analysis. The levels of many lipid species from different lipid classes, most notably phosphatidylethanolamine (PE), phosphatidylcholine (PC), and TG, were significantly altered. Subsequent lipidomic analysis highlights the pathways significantly impacted by PFOA exposure, with the glycerophospholipid metabolism being the most impacted, and the changes in the lipidome network, which connects all the lipid species together. MALDI-MSI displays the heterogeneous distribution of the affected lipids and PFOA, revealing different areas of lipid expression linked to PFOA localization. TOF-SIMS localizes PFOA at the cellular level, supporting MALDI-MSI results. This multi-modal MS analysis unveils the lipidomic impact of PFOA in the mouse liver after high-dose and short-term exposure and opens new opportunities in toxicology.

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.

Hundreds of consumer and commercial products containing silver nanoparticles (AgNPs) are currently used in food, personal-care products, pharmaceutical, and many other applications. Human exposure to AgNPs includes oral intake, inhalation, and dermal contact. The aim of this review was to focus on oral intake, intentional and incidental of AgNPs where well-known antimicrobial characteristics that might affect the microbiome and mucus in the gastrointestinal tract (GIT). This critical review summarizes what is known regarding the impacts of AgNPs on gut homeostasis. It is fundamental to understand the forms of AgNPs and their physicochemical characterization before and during digestion. For example, lab-synthesized AgNPs differ from "real" ingestable AgNPs used as food additives and dietary supplements. Similarly, the gut environment alters the chemical and physical state of Ag that is ingested as AgNPs. Emerging research on in vitro and in vivo rodent and human indicated complex multi-directional relationships among AgNPs, the intestinal microbiota, and the epithelial mucus. It may be necessary to go beyond today's descriptive approach to a modeling-based ecosystem approach that might quantitatively integrate spatio-temporal interactions among microbial groups, host factors (e.g., mucus), and environmental factors, including lifestyle-based stressors. It is suggested that future research (1) utilize more representative AgNPs, focus on microbe/mucus interactions, (2) assess the effects of environmental stressors for longer and longitudinal conditions, and (3) be integrated using quantitative modeling.

Neonatal period is characterized by an immature intestinal barrier. Scattered evidence suggests that early life stressful events induce long lasting alterations of intestinal homeostasis mimicking Irritable Bowel Syndrome (IBS). Those observations highlighting defect of intestinal barrier by early life stress questioned its potential role as a risk factor for gastrointestinal disorders such as colitis and infections. In this study, we aimed to analyze if maternal separation (MS) in mice mimicks IBS main features. We next addressed whether MS could trigger or exacerbate colitis in genetically predisposed mice and/or enhance susceptibility to gastrointestinal infections in wild type mice. MS induced main features of IBS in adult wild type male mice i.e. intestinal hyperpermeability, visceral hypersensitivity, microbiota dysbiosis, bile acid malabsorption and low grade inflammation in intestine associated with a defect of Paneth cells and the ILC3 population. This breach in mucosal barrier functions in adults was associated with a systemic IgG response against commensal E. coli and increased IFNγ secretion by splenocytes. However, in IL10-/- mice, MS did not trigger nor worsen colitis. Furthermore, wild type mice submitted to MS did not show increase susceptibility to gastrointestinal infections (S. Typhimurium, L. monocytogenes or T. gondii) compared to controls. Altogether, our results identify MS in mice as a good experimental model for IBS mimicking all the main features. In addition, early life stress, even though it has long lasting consequences on intestinal homeostasis, does not constitute a facilitating factor to colitis in predisposed individuals nor to gastrointestinal infections in wild type mice.

Synthetic chemicals (environmental pollutants, food additives) are widely used for many industrial purposes and consumer-related applications, which implies, through manufactured products, diet, and environment, a repeated exposure of the general population with growing concern regarding health disorders. The gastrointestinal tract is the first physical and biological barrier against these compounds, and thus their first target. Mounting evidence indicates that the gut microbiota represents a major player in the toxicity of environmental pollutants and food additives; however, little is known on the toxicological relevance of the mucus/pollutant interplay, even though mucus is increasingly recognized as essential in gut homeostasis. Here, we aimed at describing how environmental pollutants (heavy metals, pesticides, and other persistent organic pollutants) and food additives (emulsifiers, nanomaterials) might interact with mucus and mucus-related microbial species; that is, “mucophilic” bacteria such as mucus degraders. This review highlights that intestinal mucus, either directly or through its crosstalk with the gut microbiota, is a key, yet underestimated gut player that must be considered for better risk assessment and management of environmental pollution.

There is an increasing awareness of the deleterious effects attributed to mycotoxins during their fate within the gut, particularly for deoxynivalenol (DON), zearalenone (ZEN), ochratoxin A (OTA), fumonisin B1 (FB1), aflatoxin B1 (AFB1), and patulin (PAT). Evidence indicates that disruption of the epithelial barrier is well established. However, intestinal barrier function on its luminal side involves two other partners, mucus and microbiota, which have rarely been considered in the context of mycotoxin exposure. The current review aimed at providing a summary of DON, ZEN, OTA, FB1, AFB1, and PAT effects on intestinal barrier function, with special focus on mucus and microbiota. DON, ZEN, OTA, FB1, AFB1, and PAT are known to markedly affect epithelial cell integrity and functions. Regarding mucus, DON is the most documentated mycotoxin. In vivo, toxicological impact of DON generally has only been assessed through goblet cell number. Evaluation of the mycotoxins/mucus interplay considering other indicators such as composition, thickness, and penetrability of mucus, mucin O-glycosylation thus warrants further attention. With respect to microbiota, few short-term studies to date have been reported indicating deleterious effects. However, long-term exposure to mycotoxins may also produce significant changes in microbiota composition and metabolic activity, which requires further experimentation. In conclusion, mucus and microbiota are key targets for dietary mycotoxins although assessment of induced effects is preliminary. A significant research effort is now underway to determine the adverse consequences of mycotoxins on mucus and microbiota considered as individual but also as tightly connected gut players.

We report here the complete genome sequence of Lactococcus lactis subsp. lactis strain A12, a strain isolated from sourdough. The circular chromosome and the four plasmids reveal genes involved in carbohydrate metabolism that are potentially required for the persistence of this strain in such a complex ecosystem.

Stressful events during the neonatal period result in alterations of intestinal homeostasis and predispose to brain-gut axis related disorders in adulthood. Maternal separation (MS) induces visceral hypersensitivity of 50-days old female and male C3H/HeN mice but selectively increases intestinal permeability of male. Considering that gut microbial colonization overlaps with MS, we addressed in this study subsequent consequences on their fecal microbiota. Methodology. Relative abundances of 21 microbial communities were compared using the GUt-Low-Density-Array approach and analysed using multivariate approaches. Results. Microbial profiles of adult male and female clustered separately regardless of MS in early life. Alteration in response to MS was more pronounced in male than female. Among the  microbial determinants associated with MS, a diminution of Roseburia spp. was observed in both sexes, whereas diminution of Bifidobacterium spp. and increase of Clostridium butyricum were male-specific. Divergent responses according to sex were observed regarding relative abundances of Methanobrevibacter smithii, Enterobacteriaceae and Eubacterium hallii. Conclusion. Sexual dimorphism highlighted in this study reflects previously observed host functional dimorphisms. Identification of stages at which each event occurs early in life may help to understand whether theses microbial alterations are a cause or a consequence of adverse outcomes in adulthood.

Background & Aims.Human colorectal cancer (CRC) often originates with loss of the tumor suppressor APC and many believe this loss occurs in stem cells.Despite the wellcharacterized multi-step model of CRC, much less is known about the specific genomic changes that occur in early onset adenomas.We engineered an inducible mouse model of colonic adenoma formation using the conditional loss of heterozygosity of Apc in Lrig1+ colonic stem/progenitor cells, yielding highly penetrant, multiple distal colonic adenomas.In this study, we examined the transcriptomic and genomic changes present in these adenomas between sibling mice and within a single mouse, and assessed the prevalence of intertumoral genetic heterogeneity.Methods.Adult Lrig1-CreERT2/+;Apc-flox/+ mice were given 2mg tamoxifen (i.p. on 3 consecutive days) that consistently led to multiple, dysplastic adenomas in the distal colon 100 days later.Whole tumors (n=24) from 18 mice, as well as adjacent normal colon and wildtype colon, were excised and both mRNA and exome DNA were sequenced.Changes in gene expression levels were identified using the R package DESeq2 and high-quality somatic mutations using Seurat, a GATK module.Results.Adenomas resulting from inducible loss of Apc in Lrig1+ stem/progenitor cells were histologically variegated and genetically hyper-mutated.Each tumor contained over 100 high-frequency, high-impact mutations with an abundance of G to T transversions.Each tumor contained a unique profile of mutated tumor suppressor genes, such as Pi3kca, Nf1 and Lrig1, and DNA repair genes, such as Msh4, Msh6, and Pold1 .We identified 40 genes that were independently mutated in more than one tumor and also occurred in greater than 10% of TCGA CRCs.A number of genes mutated are predicted to impact cell morphology and migration.We also identified >1,000 differentially expressed genes in tumors, including downregulation of Msh3 and Msh4, as well as some unexpected hits such as Dkk2 upregulation and Caecam1 downregulation.Interestingly, and in contrast to their highly variable genomes, exhibited substantial transcriptome similarity to TCGA CRCs, for example, CRCs with highlevel MYC module activation.Conclusions.We show that colonic adenomas from Lrig1-CreERT2/+;Apc-flox/+ mice are surprisingly heterogeneous.Based on the abundance of G to T transversions, we speculate the loss of Apc may lead to defects in DNA repair, and a dramatic increase in mutation rate.Our studies also show that Apc loss results in mutations affecting cell morphology and migration, implying that events leading to tumor growth and invasion may happen early in tumorigenesis.

The whole set of putative glucansucrases from Leuconostoc citreum LBAE-E16 and LBAE-C11 was retrieved from the draft genome sequence of these two sourdough strains previously suggested as alternan producers. Four and five putative glycoside hydrolase family 70 (GH70) encoding genes were identified in the genome sequence of strain C11 and E16, respectively. Some putative genes have high sequence identity to known Leuconostoc dextransucrases. Molecular and biochemical data confirmed that L. citreum C11 could be considered as a new alternan-producing strain, unlike strain E16. In the latter, two new putative glucansucrases with unusual structural features were retrieved. In particular, the GSE16-5 gene encodes for a protein of 2063 amino acids with a theoretical molecular mass of 229 kDa that shares 61% identity with the alternansucrase (ASR) of L. citreum NRRL B-1355, due to the presence of seven APY repeats identified in the C-terminal peptide sequence. Cloning and expression of the corresponding coding sequence revealed synthesis of a low molecular weight (10(4) Da) linear dextran polymer with glucosyl residues only linked by α-1,6 linkages. This novel GH70 enzyme may thus be viewed as a natural chimeric enzyme resulting from the addition of the ASR C-terminal region in a dextransucrase.