Structure-Dependent Effects of Phthalates on Intercellular and Intracellular Communication in Liver Oval Cells

Humans are exposed to phthalates released from plastics, cosmetics, or food on a daily basis. Phthalates have low acute liver toxicity, but their chronic exposures could induce molecular and cellular effects linked to adverse health outcomes, such as liver tumor promotion or chronic liver diseases. The alternation of gap junctional intercellular communication (GJIC) and MAPK-Erk1/2 pathways in liver progenitor or oval cells can disrupt liver tissue homeostatic mechanisms and affect the development and severity of these adverse outcomes. Our study with 20 different phthalates revealed their structurally dependent effects on liver GJIC and MAPK-Erk1/2 signaling in rat liver WB-F344 cell line with characteristics of liver oval cells. The phthalates with a medium-length side chain (3-6 C) were the most potent dysregulators of GJIC and activators of MAPK-Erk1/2. The effects occurred rapidly, suggesting the activation of non-genomic (non-transcriptional) mechanisms directly by the parental compounds. Short-chain phthalates (1-2 C) did not dysregulate GJIC even after longer exposures and did not activate MAPK-Erk1/2. Longer chain (>= 7 C) phthalates, such as DEHP or DINP, moderately activated MAPK-Erk1/2, but inhibited GJIC only after prolonged exposures (>12 h), suggesting that GJIC dysregulation occurs via genomic mechanisms, or (bio)transformation. Overall, medium-chain phthalates rapidly affected the key tissue homeostatic mechanisms in the liver oval cell population via non-genomic pathways, which might contribute to the development of chronic liver toxicity and diseases

Microcystin-LR Does Not Alter Cell Survival and Intracellular Signaling in Human Bronchial Epithelial Cells

Changes in ecological and environmental factors lead to an increased occurrence of cyanobacterial water blooms, while secondary metabolites-producing cyanobacteria pose a threat to both environmental and human health. Apart from oral and dermal exposure, humans may be exposed via inhalation and/or swallowing of contaminated water and aerosols. Although many studies deal with liver toxicity, less information about the effects in the respiratory system is available. We investigated the effects of a prevalent cyanotoxin, microcystin-LR (MC-LR), using respiratory system-relevant human bronchial epithelial (HBE) cells. The expression of specific organic-anion-transporting polypeptides was evaluated, and the western blot analysis revealed the formation and accumulation of MC-LR protein adducts in exposed cells. However, MC-LR up to 20 mu M neither caused significant cytotoxic effects according to multiple viability endpoints after 48-h exposure, nor reduced impedance (cell layer integrity) over 96 h. Time-dependent increase of putative MC-LR adducts with protein phosphatases was not associated with activation of mitogen-activated protein kinases ERK1/2 and p38 during 48-h exposure in HBE cells. Future studies addressing human health risks associated with inhalation of toxic cyanobacteria and cyanotoxins should focus on complex environmental samples of cyanobacterial blooms and alterations of additional non-cytotoxic endpoints while adopting more advanced in vitro models

Applicability of Scrape Loading-Dye Transfer Assay for Non-Genotoxic Carcinogen Testing

Dysregulation of gap junction intercellular communication (GJIC) is recognized as one of the key hallmarks for identifying non-genotoxic carcinogens (NGTxC). Currently, there is a demand for in vitro assays addressing the gap junction hallmark, which would have the potential to eventually become an integral part of an integrated approach to the testing and assessment (IATA) of NGTxC. The scrape loading-dye transfer (SL-DT) technique is a simple assay for the functional evaluation of GJIC in various in vitro cultured mammalian cells and represents an interesting candidate assay. Out of the various techniques for evaluating GJIC, the SL-DT assay has been used frequently to assess the effects of various chemicals on GJIC in toxicological and tumor promotion research. In this review, we systematically searched the existing literature to gather papers assessing GJIC using the SL-DT assay in a rat liver epithelial cell line, WB-F344, after treating with chemicals, especially environmental and food toxicants, drugs, reproductive-, cardio- and neuro-toxicants and chemical tumor promoters. We discuss findings derived from the SL-DT assay with the known knowledge about the tumor-promoting activity and carcinogenicity of the assessed chemicals to evaluate the predictive capacity of the SL-DT assay in terms of its sensitivity, specificity and accuracy for identifying carcinogens. These data represent important information with respect to the applicability of the SL-DT assay for the testing of NGTxC within the IATA framework

Cyanobacteria and microcystin contamination in untreated and treated drinking water in Ghana

© 2017, Page Press Publications. All rights reserved. Although cyanobacterial blooms and cyanotoxins represent a worldwide-occurring phenomenon, there are large differences among different countries in cyanotoxin-related human health risk assessment, management practices and policies. While national standards, guideline values and detailed regulatory frameworks for effective management of cyanotoxin risks have been implemented in many in­dustrialized countries, the extent of cyanobacteria occurrence and cyanotoxin contamination in certain geographical regions is underreported and not very well understood. Such regions include major parts of tropical West and Central Africa, a region constisting of more than 25 countries occupying an area of 12 million km2, with a total population of 500 milion people. Only few studies focusing on cyanotoxin occurrence in this region have been published so far, and reports dealing specifically with cyanotoxin contamination in drinking water are extremely scarce. In this study, we report seasonal data on cyanobactcria and microcystin (MC) contamination in drinking water reservoirs and adjacent treatment plants located in Ghana, West Africa. During January-June 2005, concentrations of MCs were monitored in four treatment plants supplying drinking water to major metropolitan areas in Ghana: the treatment plants Barekese and Owabi, which serve Kumasi Metropolitan Area, and the plants Kpong and Weija, providing water for Accra-Tema Metropolitan Area. HPLC analyses showed that 65% samples of raw water at the intake of the treatment plants contained intracellular MCs (maximal detected concentration was 8.73 fig L-1), whereas dissolved toxins were detected in 33% of the samples. Significant reduction of cyanobacterial cell counts and MC concentrations was achieved during the entire monitoring period by the applied conventional water treatment methods (alum flocculation, sedimentation, rapid sand filtration and chlorination), and MC concentration in the final treated water never exceeded 1 fig L-1 (WHO guideline limit for MC-LR in drinking water). However, cyanobacterial cells (93-3,055 cell mL-1) were frequently found in the final treated water and intracellular MCs were detected in 17% of the samples (maximal concentration 0.61 (μg L-1), while dissolved MCs were present in 14% of the final treated water samples (maximal concentration 0.81 μg L-1). It indicates a borderline efficiency of the water treatment, thus MC concentrations in drinking water might exceed the WHO guideline limit if the treatment efficiency gets compromised. In addition, MC concentrations found in the raw water might represent significant human health risks for people living in areas with only a limited access to the treated or underground drinking water

Treatment of cylindrospermopsin by hydroxyl and sulfate radicals: Does degradation equal detoxification?

Drinking water treatment ultimately aims to provide safe and harmless drinking water. Therefore, the suitability of a treatment process should not only be assessed based on reducing the concentration os a pollutant concentration but, more importantly, on reducing its toxicity. Hence, the main objective of this study was to answer whether the degradation of a highly toxic compound of global concern for drinking water equals its detoxification. We, therefore, investigated the treatment of cylindrospermopsin (CYN) by •OH and SO4-• produced in Fenton and Fenton-like reactions. Although SO4-• radicals removed the toxin more effectively, both radical species substantially degraded CYN. The underlying degradation mechanisms were similar for both radical species and involved hydroxylation, dehydrogenation, decarboxylation, sulfate group removal, ring cleavage, and further fragmentation. The hydroxymethyl uracil and tricyclic guanidine moieties were the primary targets. Furthermore, the residual toxicity, assessed by a 3-dimensional human in vitro liver model, was substantially reduced during the treatment by both radical species. Although the results indicated that some of the formed degradation products might still be toxic, the overall reduction of the toxicity together with the proposed degradation pathways allowed us to conclude: “Yes, degradation of CYN equals its detoxification!”

Treatment of cylindrospermopsin by hydroxyl and sulfate radicals: Does degradation equal detoxification

Drinking water treatment ultimately aims to provide safe and harmless drinking water. Therefore, the suitability of a treatment process should not only be assessed based on reducing the concentration os a pollutant concentration but, more importantly, on reducing its toxicity. Hence, the main objective of this study was to answer whether the degradation of a highly toxic compound of global concern for drinking water equals its detoxification. We, therefore, investigated the treatment of cylindrospermopsin (CYN) by center dot OH and SO4-center dot produced in Fenton and Fenton-like reactions. Although SO4-center dot radicals removed the toxin more effectively, both radical species substantially degraded CYN. The underlying degradation mechanisms were similar for both radical species and involved hydroxylation, dehydrogenation, decarboxylation, sulfate group removal, ring cleavage, and further fragmentation. The hydroxymethyl uracil and tricyclic guanidine moieties were the primary targets. Furthermore, the residual toxicity, assessed by a 3-dimensional human in vitro liver model, was substantially reduced during the treatment by both radical species. Although the results indicated that some of the formed degradation products might still be toxic, the overall reduction of the toxicity together with the proposed degradation pathways allowed us to conclude: "Yes, degradation of CYN equals its detoxification!"

Branched Poly(ethylene imine)s as Anti‐algal and Anti‐cyanobacterial Agents with Selective Flocculation Behavior to Cyanobacteria over Algae

Poly(ethylene imine)s (PEIs) have been widely studied for biomedical applications, including antimicrobial agents against potential human pathogens. The interactions of branched PEIs (B‐PEIs) with environmentally relevant microorganisms whose uncontrolled growth in natural or engineered environments causes health, economic, and technical issues in many sectors of water management are studied. B‐PEIs are shown to be potent antimicrobials effective in controlling the growth of environmentally relevant algae and cyanobacteria with dual‐functionality and selectivity. Not only did they effectively inhibit growth of both algae and cyanobacteria, mostly without causing cell death (static activity), but they also selectively flocculated cyanobacteria over algae. Thus, unmodified B‐PEIs provide a cost‐effective and chemically facile framework for the further development of effective and selective antimicrobial agents useful for control of growth and separation of algae and cyanobacteria in natural or engineered environments.Poly(ethylene imine)s (B‐PEIs) are shown as effective antimicrobial agents against environmentally relevant microorganisms (algae CR – Chlamydomonas reinhardtii, cyanobacterium SE – Synechococcus elongatus). B‐PEIs have dual‐functionalities and can 1) inhibit growth of both algae and cyanobacteria and 2) selectively flocculate cyanobacteria over algae. These molecules provide a cost‐effective and chemically facile framework for the further development of selective anti‐algal and anti‐cyanobacterial agents.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/146449/1/mabi201800187_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/146449/2/mabi201800187.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/146449/3/mabi201800187-sup-0001-SupMat.pd

Tumor promoting properties of a cigarette smoke prevalent polycyclic aromatic hydrocarbon as indicated by the inhibition of gap junctional intercellular communication via phosphatidylcholine-specific phospholipase C

Inhibition of gap junctional intercellular communication (GJIC) and the activation of intracellular mitogenic pathways are common hallmarks of epithelial derived cancer cells. We previously determined that the 1-methyl and not the 2-methyl isomer of anthracene, which are prominent cigarette smoke components, activated extracellular receptor kinase, and inhibited GJIC in WB-F344 rat liver epithelial cells. Using these same cells, we show that an immediate upstream response to 1-methylanthracene was a rapid ( LT 1 min) release of arachidonic acid. Inhibition of phosphatidylcholine-specific phospholipase C prevented the inhibition of GJIC by 1-methylanthracene. In contrast, inhibition of phosphatidylinositol specific phospholipase C, phospholipase A(2), diacylglycerol lipase, phospholipase D, protein kinase C, and tyrosine protein kinases had no effect on 1-methylanthracene-induced inhibition of GJIC. Inhibition of protein kinase A also prevented inhibition of GJIC by 1-methylanthracene. Direct measurement of phosphatidylcholine-specific phospholipase C and sphingomyelinase indicated that only phosphatidylcholine-specific phospholipase C was activated in response to 1-methylanthracene, while 2-methylanthracene had no effect. 1-methylanthracene also activated p38-mitogen activated protein kinase; however, like extracellular kinase, its activation was not involved in 1-methylanthracene-induced regulation of GJIC, and this activation was independent of phosphatidylcholine-specific phospholipase C. Although mitogen activated protein kinases were activated, Western blot analyzes indicated no change in connexin43 phosphorylation status. Our results indicate that phosphatidylcholine-specific phospholipase C is an important enzyme in the induction of a tumorigenic phenotype, namely the inhibition of GJIC; whereas mitogen activated protein kinases triggered in response to 1-methylanthracene, were not involved in the deregulation of GJIC