Methylglyoxal-induced neurotoxic effects in primary neuronal-like cells transdifferentiated from human mesenchymal stem cells: Impact of low concentrations.

In the last decades, advanced glycation end-products (AGEs) have aroused the interest of the scientific community due to the increasing evidence of their involvement in many pathophysiological processes including various neurological disorders and cognitive decline age related. Methylglyoxal (MG) is one of the reactive dicarbonyl precursors of AGEs, mainly generated as a by-product of glycolysis, whose accumulation induces neurotoxicity. In our study, MG cytotoxicity was evaluated employing a human stem cell-derived model, namely, neuron-like cells (hNLCs) transdifferentiated from mesenchymal stem/stromal cells, which served as a source of human based species-specific "healthy" cells. MG increased ROS production and induced the first characteristic apoptotic hallmarks already at low concentrations (≥10 µM), decreased the cell growth (≥5-10 µM) and viability (≥25 µM), altered Glo-1 and Glo-2 enzymes (≥25 µM), and markedly affected the neuronal markers MAP-2 and NSE causing their loss at low MG concentrations (≥10 µM). Morphological alterations started at 100 µM, followed by even more marked effects and cell death after few hours (5 h) from 200 µM MG addition. Substantially, most effects occurred as low as 10 µM, concentration much lower than that reported from previous observations using different in vitro cell-based models (e.g., human neuroblastoma cell lines, primary animal cells, and human iPSCs). Remarkably, this low effective concentration approaches the level range measured in biological samples of pathological subjects. The use of a suitable cellular model, that is, human primary neurons, can provide an additional valuable tool, mimicking better the physiological and biochemical properties of brain cells, in order to evaluate the mechanistic basis of molecular and cellular alterations in CNS.

MAM-2201 acute administration impairs motor, sensorimotor, prepulse inhibition, and memory functions in mice: a comparison with its analogue AM-2201.

RATIONALE: 1-[(5-fluoropentyl)-1H-indol-3-yl](4-methyl-1-naphthalenyl) methanone (MAM-2201) is a potent synthetic cannabinoid receptor agonist illegally marketed in "spice" products and as "synthacaine" for its psychoactive effects. It is a naphthoyl-indole derivative which differs from its analogue 1-[(5-Fluoropentyl)-1H-indol-3-yl](1-naphthylenyl) methanone (AM-2201) by the presence of a methyl substituent on carbon 4 (C-4) of the naphthoyl moiety. Multiple cases of intoxication and impaired driving have been linked to AM-2201 and MAM-2201 consumption. OBJECTIVES: This study aims to investigate the in vitro (murine and human cannabinoid receptors) and in vivo (CD-1 male mice) pharmacodynamic activity of MAM-2201 and compare its effects with those induced by its desmethylated analogue, AM-2201. RESULTS: In vitro competition binding studies confirmed that MAM-2201 and AM-2201 possess nanomolar affinity for both CD-1 murine and human CB1 and CB2 receptors, with preference for the CB1 receptor. In agreement with the in vitro binding data, in vivo studies showed that MAM-2201 induces visual, acoustic, and tactile impairments that were fully prevented by pretreatment with CB1 receptor antagonist/partial agonist AM-251, indicating a CB1 receptor mediated mechanism of action. Administration of MAM-2201 also altered locomotor activity and PPI responses of mice, pointing out its detrimental effect on motor and sensory gating functions and confirming its potential use liability. MAM-2201 and AM-2201 also caused deficits in short- and long-term working memory. CONCLUSION: These findings point to the potential public health burden that these synthetic cannabinoids may pose, with particular emphasis on impaired driving and workplace performance.

Human Astrocyte Spheroids as Suitable In Vitro Screening Model to Evaluate Synthetic Cannabinoid MAM2201-Induced Effects on CNS.

There is growing concern about the consumption of synthetic cannabinoids (SCs), one of the largest groups of new psychoactive substances, its consequence on human health (general population and workers), and the continuous placing of new SCs on the market. Although drug-induced alterations in neuronal function remain an essential component for theories of drug addiction, accumulating evidence indicates the important role of activated astrocytes, whose essential and pleiotropic role in brain physiology and pathology is well recognized. The study aims to clarify the mechanisms of neurotoxicity induced by one of the most potent SCs, named MAM-2201 (a naphthoyl-indole derivative), by applying a novel three-dimensional (3D) cell culture model, mimicking the physiological and biochemical properties of brain tissues better than traditional two-dimensional in vitro systems. Specifically, human astrocyte spheroids, generated from the D384 astrocyte cell line, were treated with different MAM-2201 concentrations (1-30 µM) and exposure times (24-48 h). MAM-2201 affected, in a concentration- and time-dependent manner, the cell growth and viability, size and morphological structure, E-cadherin and extracellular matrix, CB1-receptors, glial fibrillary acidic protein, and caspase-3/7 activity. The findings demonstrate MAM-2201-induced cytotoxicity to astrocyte spheroids, and support the use of this human 3D cell-based model as species-specific in vitro tool suitable for the evaluation of neurotoxicity induced by other SCs.

Human Umbilical Cord Mesenchymal Stem Cell-Based in vitro Model for Neurotoxicity Testing.

Neurotoxicity (NT) testing for regulatory purposes is based on in vivo animal testing. There is general consensus, however, about the need for the development of alternative methodologies to allow researchers to more rapidly and cost effectively screen large numbers of chemicals for their potential to cause NT, or to investigate their mode of action. In vitro assays are considered an important source of information for making regulatory decisions, and human cell-based systems are recommended as one of the most relevant models in toxicity testing, to reduce uncertainty in the extrapolation of results from animal-based models. Human neuronal models range from various neuroblastoma cell lines to stem cell-derived systems, including those derived from mesenchymal stem/stromal cells (hMSC). hMSCs exhibit numerous advantages, including the fact that they can be obtained in high yield from healthy human adult tissues, can be cultured with a minimal laboratory setup and without genetic manipulations, are able of continuous and repeated self-renewal, are nontumorigenic, and can form large populations of stably differentiated cells representative of different tissues, including neuronal cells. hMSCs derived from human umbilical cord (hUC) in particular possess several prominent advantages, including a painless, non-invasive, and ethically acceptable collection procedure, simple and convenient preparation, and high proliferation capacity. In addition, hMSCs can be efficiently differentiated into neuron-like cells (hNLCs), which can then be used for the assessment of neuronal toxicity of potential neurotoxic compounds in humans. Here, we describe a step-by-step procedure to use hMSCs from the umbilical cord for in vitro neurotoxicity testing. First, we describe how to isolate, amplify, and store hMSCs derived from the umbilical cord. We then outline the steps to transdifferentiate these cells into hNLCs, and then use the hNLCs for neurotoxicity testing by employing multiple common cytotoxicity assays after treatment with test compounds. The approach follows the most updated guidance on using human cell-based systems. These protocols will allow investigators to implement an alternative system for obtaining primary NLCs of human origin, and support advancement in neurotoxicity research. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Isolation and maintenance of human mesenchymal stem/stromal cells (hMSCs) obtained from the umbilical cord lining membrane Basic Protocol 2: Transdifferentiation of hMSCs into neuron-like cells (hNLCs) and basic neurotoxicity assessment.

Three-dimensional spheroid cell culture of human MSC-derived neuron-like cells: New in vitro model to assess magnetite nanoparticle-induced neurotoxicity effects.

As nanoparticles (NPs) can access the brain and impact on CNS function, novel in vitro models for the evaluation of NPs-induced neurotoxicity are advocated. Three-dimensional spheroids of primary neuron-like cells (hNLCs) of human origin have been generated, from differentiation of human umbilical cord mesenchymal stem cells (MSCs). The study evaluated Fe3 O4 NP impact on the differentiation process by applying the challenge at complete 3D hNLC spheroid formation (after 4 days, T4) or at beginning of neurogenic induction/simultaneously 3D forming (T0). Different endpoints were monitored over time (up to 10 days): spheroid growth, size, morphology, ATP, cell death, neuronal markers (ß-Tub III, MAP-2, and NSE), NP uptake. At T0 application, a marked concentration- and time-dependent cell mortality occurred: effect started early (day 2) and low concentration (1 µg/ml) and exacerbated (80% mortality) after prolonged time (day 6) and increased concentrations (50 µg/ml). ATP was strikingly affected. All neuronal markers were downregulated, and spheroid morphology altered in a concentration-dependent manner (from ≥5 µg/ml) after day 2. Fe3 O4 NPs applied at complete 3D formation (T4) still induced adverse effects although less severe: cell mortality (20-60%) and ATP content decrease (10-40%) were observed in a concentration-dependent manner (from ≥ 5 µg/ml). A neuronal-specific marker effect and spheroid size reduction from 25 µg/ml without morphology alteration were evidenced. This finding provides additional information on neurotoxic effects of Fe3 O4 NPs in a new 3D hNLC spheroid model derived from MSCs that could find a consistent application as in a testing strategy serving in first step hazard identification for correct risk assessment.

Biomarkers for alcohol abuse/withdrawal and their association with clinical scales and temptation to drink. A prospective pilot study during 4-week residential rehabilitation.

A bulk of evidence in the field of translational medicine applied to clinical toxicology and rehabilitation has highlighted the possibility of using biomarkers as a support in the diagnosis of alcohol-related diseases and in monitoring of alcohol withdrawal. In a cohort of 55 subjects admitted to a 4-week residential rehabilitation period for alcohol detoxification, we applied a complementary approach correlating novel and conventional peripheral blood and urine parameters in combination with clinical and functional evaluation, contextually considered with the patient's history. Biomarkers of oxidative, inflammatory, hepatic, and neurochemical effects paralleled by alcohol craving and clinical scale measurements were determined at two specific time points, i.e., admission and discharge. Concerning the post-discharge assessment (i.e., relapse evaluation one month after discharge), a follow-up oral interview during a clinical examination was applied to evaluate alcohol abstinence.Selected biomarkers, i.e., MCP1, F2-IsoPs, and SOD1, were altered in chronic alcoholics at admission, and then showed a clearly changing trend during hospitalization. Our findings demonstrated that these specific non-traditional biomarkers, measured together with more conventional ones (e.g., CDT, EtG, IL8, ALT, AST, GGT), could represent novel key parameters for monitoring alcohol use disorders and withdrawal, being also suggestive of the complexity of the psychoneuroimmune response to alcohol. A general improvement in psychological functioning (i.e., decreases in anxiety, depression, and psychological distress) was also revealed during the 4-week rehabilitation treatment, paralleled by an increase of well-being and positive changes in terms of scores. Moreover, a positive association between SOD1 and drink craving at admission was evidenced. Notably, both SOD1 and well-being displayed a significant relation with lower risk of alcohol relapse one month after discharge, indicating that SOD1 is a good predictor of reduced relapse probability. This 4-week residential rehabilitation protocol represents a sound strategy enabling identification of alcohol use disorders and monitoring of alcohol addiction state and withdrawal. However, it has to be emphasized that results derived from this pilot study need to be extensively validated in large and independent cohorts of subjects.

How Do Inflammatory Mediators, Immune Response and Air Pollution Contribute to COVID-19 Disease Severity? A Lesson to Learn.

Inflammatory and immune processes are defensive mechanisms that aim to remove harmful agents. As a response to infections, inflammation and immune response contribute to the pathophysiological mechanisms of diseases. Coronavirus disease 2019 (COVID-19), whose underlying mechanisms remain not fully elucidated, has posed new challenges for the knowledge of pathophysiology. Chiefly, the inflammatory process and immune response appear to be unique features of COVID-19 that result in developing a hyper-inflammatory syndrome, and air pollution, the world's largest health risk factor, may partly explain the behaviour and fate of COVID-19. Understanding the mechanisms involved in the progression of COVID-19 is of fundamental importance in order to avoid the late stage of the disease, associated with a poor prognosis. Here, the role of the inflammatory and immune mediators in COVID-19 pathophysiology is discussed.

Developmental Neurotoxicity Screening for Nanoparticles Using Neuron-Like Cells of Human Umbilical Cord Mesenchymal Stem Cells: Example with Magnetite Nanoparticles.

Metallic nanoparticles (NPs), as iron oxide NPs, accumulate in organs, cross the blood-brain barrier and placenta, and have the potential to elicit developmental neurotoxicity (DNT). Human stem cell-derived in vitro models may provide more realistic platforms to study NPs effects on neural cells, and to obtain relevant information on the potential for early or late DNT effects in humans. Primary neuronal-like cells (hNLCs) were generated from mesenchymal stem cells derived from human umbilical cord lining and the effects caused by magnetite (Fe3O4NPs, 1-50 µg/mL) evaluated. Neuronal differentiation process was divided into stages: undifferentiated, early, mid- and fully-differentiated (from day-2 to 8 of induction) based on different neuronal markers and morphological changes over time. Reduction in neuronal differentiation induction after NP exposure was observed associated with NP uptake: ß-tubulin III (ß-Tub III), microtubule-associated protein 2 (MAP-2), enolase (NSE) and nestin were downregulated (10-40%), starting from 25 µg/mL at the early stage. Effects were exacerbated at higher concentrations and persisted up to 8 days without cell morphology alterations. Adenosine triphosphate (ATP) and caspase-3/7 activity data indicated Fe3O4NPs-induced cell mortality in a concentration-dependent manner and increases of apoptosis: effects appeared early (from day-3), started at low concentrations (≥5 µg/mL) and persisted. This new human cell-based model allows different stages of hNLCs to be cultured, exposed to NPs/chemicals, and analyzed for different endpoints at early or later developmental stage.

In vitro evaluation of magnetite nanoparticles in human mesenchymal stem cells: comparison of different cytotoxicity assays.

This work was aimed at defining the suitable test for evaluating Fe3O4 NPs cytotoxicity after short-term exposure in human mesenchymal stem cells (hMSCs) using different viability tests, namely NRU, MTT and TB assays, paralleled by cell morphology analyses for cross checking. MTT and NRU data (culture medium with/without hMSCs plus Fe3O4NPs) indicated artificial/false increments in cell viability after Fe3O4NPs. These observations did not fit with the morphological analyses showing reduced cell density, loss of monolayer features, and morphological alterations at Fe3O4NPs ≥50 µg/ml. Fe3O4NPs alone induced a substantial increased absorbance at the wavelength required for MTT and NRU. A significant death (25%) of hMSC at Fe3O4NPs ≥10 µg/ml, with a maximum effect (45%) at 300 µg/ml after 24 h, exacerbated after 48 h, was observed when applying TB test. These results paralleled the effects on cell morphology. The optical properties and stability of Fe3O4NP suspension (tendency to agglomerate in a specific culture medium) represent factors that limit in vitro result interpretation. These findings suggest the non applicability of the spectrophotometric assays for hMSC culture conditions, while TB is an accurate method for determining cell viability after Fe3O4NP exposure in this model. In relation to NPs safety assessment: cell-based assays must be considered on case-by-case basis; selection of relevant cell models is also important for predictive toxicological studies; application of a testing strategy is fundamental for understanding the toxicity pathways driving cellular responses.

In vitro toxicity screening of magnetite nanoparticles by applying mesenchymal stem cells derived from human umbilical cord lining.

Despite the growing interest in nanoparticles (NPs), their toxicity has not yet been defined and the development of new strategies and predictive models are required. Human stem cells (SCs) offer a promising and innovative cell-based model. Among SCs, mesenchymal SCs (MSCs) derived from cord lining membrane (CL) may represent a new species-specific tool for establishing efficient platforms for primary screening and toxicity/safety testing of NPs. Superparamagnetic iron oxide NPs, including magnetite (Fe3 O4 NPs), have aroused great public health and scientific concerns despite their extensive uses. In this study, CL-MSCs were characterized and applied for in vitro toxicity screening of Fe3 O4 NPs. Cytotoxicity, internalization/uptake, differentiation and proliferative capacity were evaluated after exposure to different Fe3 O4 NP concentrations. Data were compared with those obtained from bone marrow (BM)-MSCs. We observed, at early passages (P3), that: (1) cytotoxicity occurred at 10 µg/mL in CL-MSCs and 100 µg/mL in BM-MSCs (no differences in toxicity, between CL- and BM-MSCs, were observed at higher dosage, 100-300 µg/mL); (2) cell density decrease and monolayer features loss were affected at ≥50 µg/mL in CL-MSCs only; and (3) NP uptake was concentration-dependent in both MSCs. After 100 µg/mL Fe3 O4 NP exposures, the capacity of proliferation was decreased (P5-P9) in CL-MSCs without morphology alteration. Moreover, a progressive decrease of intracellular Fe3 O4 NPs was observed over culture time. Antigen surface expression and multilineage differentiation were not influenced. These findings suggest that CL-MSCs could be used as a reliable cell-based model for Fe3 O4 NP toxicity screening evaluation and support the use of this approach for improving the confidence degree on the safety of NPs to predict health outcomes.

Organoids are promising tools for species-specific in vitro toxicological studies.

Organoids are three-dimensional self-aggregating structures generated from stem cells (SCs) or progenitor cells in a process that recapitulates molecular and cellular stages of early organ development. The differentiation process leads to the appearance of specialized mature cells and is connected with changes in the organoid internal structure rearrangement and self-organization. The formation of organ-specific structures in vitro with highly ordered architecture is also strongly influenced by the extracellular matrix. These features make organoids as a powerful model for in vitro toxicology. Nowadays this technology is developing very quickly. In this review we present, from a toxicological and species-specific point of view, the state of the art of organoid generation from adult SCs and pluripotent SCs: embryonic SCs or induced pluripotent SCs. The current culture organoid techniques are discussed for their main advantages, disadvantages and limitations. In the second part of the review, we concentrated on the characterization of species-specific organoids generated from tissue-specific SCs of different sources: mammary (bovine), epidermis (canine), intestinal (porcine, bovine, canine, chicken) and liver (feline, canine).

Cytotoxic Effects of 3,4-Catechol-PV (One Major MDPV Metabolite) on Human Dopaminergic SH-SY5Y Cells.

3,4-Methylenedioxypyrovalerone (MDPV), one of the most commonly abused synthetic cathinones, has caused several intoxications and deaths despite its short presence on the market. Apart from its effects on the monoamine systems in the brain, recent in vitro investigations have revealed cytotoxicity. In this study, the effects of increasing concentrations (10-1000 µM) of 3,4-Catechol-PV, one of major MDPV metabolites, on cell viability, morphology, and apoptosis have been evaluated after acute exposure (24-48 h) in human neuroblastoma SH-SY5Y cells-undifferentiated and differentiated to a more mature neuronal-like phenotype. Results indicated the following: (i) Cell viability: concentration-dependent decrease (15-55%) in differentiated SH-SY5Y after 24 h, with no exacerbation after 48 h (LC50 values 1028 and 951 µM, respectively); marked concentration-dependent decrease after 48 h (20-63%) in undifferentiated SH-SY5Y (LC50 553.9 µM) with mild effect (18-22% cell death) after 24 h at ≥ 500 µM only; the lowest toxic concentrations were 500 and 100 µM after 24 h, for undifferentiated and differentiated SH-SY5Y, respectively, and 10 µM after 48 h. (ii) Concentration- and time-dependent alterations of cell morphology in both SH-SY5Y types characterized by several intracellular cytoplasmic vesicles (undifferentiated more susceptible (effect at ≥ 50 µM) than differentiated cells (effect at ≥ 100 µM)), loss of the typical cell shape, neurite retraction, and cell density decrease. (iii) Activation of caspase-3 enzyme in differentiated and undifferentiated cells after 48 h. These findings suggest the potential involvement of 3,4-Catechol-PV in MDPV-induced neurotoxicity and support the use of this human cellular model as a species-specific in vitro tool to clarify the neurotoxicity mechanisms of synthetic cathinones and metabolites.

Neuron-Like Cells Generated from Human Umbilical Cord Lining-Derived Mesenchymal Stem Cells as a New In Vitro Model for Neuronal Toxicity Screening: Using Magnetite Nanoparticles as an Example.

The wide employment of iron nanoparticles in environmental and occupational settings underlines their potential to enter the brain. Human cell-based systems are recommended as relevant models to reduce uncertainty and to improve prediction of human toxicity. This study aimed at demonstrating the in vitro differentiation of the human umbilical cord lining-derived-mesenchymal stem cells (hCL-MSCs) into neuron-like cells (hNLCs) and the benefit of using them as an ideal primary cell source of human origin for the neuronal toxicity of Fe3O4NPs (magnetite-nanoparticles). Neuron-like phenotype was confirmed by: live morphology; Nissl body staining; protein expression of different neuronal-specific markers (immunofluorescent staining), at different maturation stages (i.e., day-3-early and day-8-full differentiated), namely ß-tubulin III, MAP-2, enolase (NSE), glial protein, and almost no nestin and SOX-2 expression. Synaptic makers (SYN, GAP43, and PSD95) were also expressed. Fe3O4NPs determined a concentration- and time-dependent reduction of hNLCs viability (by ATP and the Trypan Blue test). Cell density decreased (20-50%) and apoptotic effects were detected at ≥10 µg/mL in both types of differentiated hNLCs. Three-day-differentiated hNLCs were more susceptible (toxicity appeared early and lasted for up to 48 h) than 8-day-differentiated cells (delayed effects). The study demonstrated that (i) hCL-MSCs easily differentiated into neuronal-like cells; (ii) the hNCLs susceptibility to Fe3O4NPs; and (iii) human primary cultures of neurons are new in vitro model for NP evaluation.

Human 3D Cultures as Models for Evaluating Magnetic Nanoparticle CNS Cytotoxicity after Short- and Repeated Long-Term Exposure.

Since nanoparticles (NPs) can translocate to the brain and impact the highly vulnerable central nervous system (CNS), novel in vitro tools for the assessment of NP-induced neurotoxicity are advocated. In this study, two types of CNS spheroids have been developed from human D384 astrocyte- and SH-SY5Y neuronal-like cells, and optimized in combination with standard assays (viability readout and cell morphology) to test neurotoxic effects caused by Fe3O4NPs, as NP-model, after short- (24­48 h; 1­100µg/ml) and long-term repeated exposure (30days; 0.1­25µg/ml). Short-term exposure of 3D-spheroids to Fe3O4NP induced cytotoxicity at 10 µg/mL in astrocytes and 25 µg/mL neurons. After long-term repeated dose regimen, spheroids showed concentration- and time-dependent cell mortality at 10 µg/mL for D384 and 0.5 µg/mL for SH-SY5Y, indicating a higher susceptibility of neurons than astrocytes. Both spheroid types displayed cell disaggregation after the first week of treatment at ≥0.1 µg/mL and becoming considerably evident at higher concentrations and over time. Recreating the 3D-spatial environment of the CNS allows cells to behave in vitro more closely to the in vivo situations, therefore providing a model that can be used as a stand-alone test or as a part of integrated testing strategies. These models could drive an improvement in the species-relevant predictivity of toxicity testing.

Blood MCP-1 levels are increased in chronic obstructive pulmonary disease patients with prevalent emphysema.

Background and aims: Chronic obstructive pulmonary disease (COPD) is a heterogeneous disease characterized by different phenotypes with either bronchial airways alterations or emphysema prevailing. As blood biomarkers could be clinically useful for COPD stratification, we aimed at investigating the levels of blood biomarkers in COPD patients differentiated by phenotype: prevalent chronic airway disease versus emphysema. Methods: In 23 COPD patients with prevalent airway disease (COPD-B), 22 COPD patients with prevalent emphysema (COPD-E), 9 control smokers (CSs), and 18 control nonsmokers (CNSs), we analyzed the expression levels of interleukin (IL)-1α, IL-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, epidermal growth factor (EGF), monocyte chemotactic protein (MCP)-1, and vascular endothelial growth factor by enzyme-linked immunosorbent assay in plasma/serum; glutathione peroxidase and superoxide dismutase (SOD)-1 by immunochemical kits in plasma; and free F2-isoprostanes (F2-IsoPs) by gas chromatography in plasma. Results: F2-IsoPs level was increased in COPD-B and COPD-E compared with CSs and CNSs; in addition, CS showed higher levels than CNSs; SOD1 level was lower in COPD-B and COPD-E than that in CNSs. Interestingly, MCP-1 level was higher only in COPD-E versus CSs and CNSs; EGF and IL-8 levels were higher in COPD-B and COPD-E versus CNSs; IL-6 level was increased in all three smoking groups (COPD-B, COPD-E, and CSs) versus CNS; IFN-γ and IL-1α levels were higher in CSs than in CNSs; and IL-1α level was also higher in CSs versus COPD-B and COPD-E. In all subjects, F2-IsoPs level correlated positively and significantly with MCP-1, IL-2, IL-1ß, IFN-γ, and TNF-α and negatively with SOD1. When correlations were restricted to COPD-E and COPD-B groups, F2-IsoPs maintained the positive associations with IFN-γ, TNF-α, and IL-2. Conclusion: We did not find any specific blood biomarkers that could differentiate COPD patients with prevalent airway disease from those with prevalent emphysema. The MCP-1 increase in COPD-E, associated with the imbalance of oxidant/antioxidant markers, may play a role in inducing emphysema.

A Review of the Mycotoxin Enniatin B.

Mycotoxin enniatin B (ENN B) is a secondary metabolism product by Fusarium fungi. It is a well-known antibacterial, antihelmintic, antifungal, herbicidal, and insecticidal compound. It has been found as a contaminant in several food commodities, particularly in cereal grains, co-occurring also with other mycotoxins. The primary mechanism of action of ENN B is mainly due to its ionophoric characteristics, but the exact mechanism is still unclear. In the last two decades, it has been a topic of great interest since its potent mammalian cytotoxic activity was demonstrated in several mammalian cell lines. Moreover, the co-exposure in vitro with other mycotoxins enhances its toxic potential through synergic effects, depending on the concentrations tested. Despite its clear cytotoxic effect, European Food Safety Authority stated that acute exposure to ENNs, such as ENN B, does not indicate concern for human health, but a concern might be the chronic exposure. However, given the lack of relevant toxicity data, no firm conclusion could be drawn and a risk assessment was not possible. In fact, very few studies have been carried out in vivo and, in these studies, no adverse effects were observed. So, research on toxicological effects induced by ENN B is still on-going. Recently, some studies are dealing with new advances regarding ENN B. This review summarizes the information on biochemical and biological activity of ENN B, focusing on toxicological aspects and on the latest advances in research on ENN B.

Human Co-culture Model of Neurons and Astrocytes to Test Acute Cytotoxicity of Neurotoxic Compounds.

Alternative methods and their use in planning and conducting toxicology experiments have become essential for modern toxicologists, thus reducing or replacing living animals. Although in vitro human co-culture models allow the establishment of biologically relevant cell-cell interactions that recapitulate the tissue microenvironment and better mimic its physiology, the number of publications is limited specifically addressing this scientific area and utilizing this test method which could provide an additional valuable model in toxicological studies. In the present study, an in vitro model based on central nervous system (CNS) cell co-cultures was implemented using a transwell system combining human neuronal cells (SH-SY5Y cell line) and glial cells, namely astrocytes (D384 cell line), to investigate neuroprotection of D384 on SH-SY5Y and vice versa. The model was applied to test acute (24-48 hours) cytotoxicity of 3 different neurotoxicants: (1) methyl mercury (1-2.5 µM), (2) Fe3O4 nanoparticles (1-100 µg/mL), and (3) methylglyoxal (0.5-1 mM). Data were compared to mono-cultures evaluating the mitochondrial function and cell morphology. The results clearly showed that all compounds tested affected the mitochondrial activity and cell morphology in both mono-culture and co-culture conditions. However, astrocytes, when cultured together with neurons, diminish the neurotoxicant-induced cytotoxic effects that occurred in neurons cultured alone, and astrocytes become more resistant in the presence of neurons. This human CNS co-culture system seems a suitable cell model to feed high-throughput acute screening platforms and to evaluate both human neuronal and astrocytic toxicity and neuroprotective effects of new and emerging materials (eg, nanomaterials) and new products with improved sensitivity due to the functional neuron-astrocyte metabolic interactions.