ABSTRACT
Synthetic cannabinoids (SCs) are a chemically diverse group of new psychoactive substances (NPSs) that target the endocannabinoid system, triggering a plethora of actions (e.g., elevated mood sensation, relaxation, appetite stimulation) that resemble, but are more intense than, those induced by cannabis. Although some of these effects have been explored for therapeutic applications, anticipated stronger psychoactive effects than cannabis and reduced risk perception have increased the recreational use of SCs, which have dominated the NPS market in the United States and Europe over the past decade. However, rising SC-related intoxications and deaths represent a major public health concern and embody a major challenge for policy makers. Here, we review the pharmacology and toxicology of SCs. A thorough characterization of SCs' pharmacodynamics and toxicodynamics is important to better understand the main mechanisms underlying acute and chronic effects of SCs, interpret the clinical/pathological findings related to SC use, and improve SC risk awareness.
Subject(s)
Cannabinoids , Humans , Cannabinoids/pharmacology , Cannabinoid Receptor Agonists/pharmacology , EndocannabinoidsABSTRACT
Dimethyl fumarate (DMF) is an old drug used for psoriasis treatment that has recently been repurposed to treat relapse-remitting multiple sclerosis, mostly due to its neuro- and immunomodulatory actions. However, mining of a pharmacovigilance database recently ranked DMF as the second pharmaceutical most associated with cognitive adverse events. To our best knowledge, the signaling mechanisms underlying its therapeutic and neurotoxic outcomes remain mostly undisclosed. This work thus represents the first-hand assessment of DMF-induced metabolic changes in undifferentiated SH-SY5Y human neuroblastoma cells, through an untargeted metabolomic approach using gas chromatography-mass spectrometry (GC-MS). The endometabolome was analyzed following 24 h and 96 h of exposure to two pharmacologically relevant DMF concentrations (0.1 and 10 µM). None of these conditions significantly reduced metabolic activity (MTT reduction assay). Our data showed that 24 h-exposure to DMF at both concentrations tested mainly affected metabolic pathways involved in mitochondrial activity (e.g., citric acid cycle, de novo triacylglycerol biosynthesis), and the synthesis of catecholamines and serotonin by changing the levels of their respective precursors, namely phenylalanine (0.68-fold decrease for 10 µM DMF vs vehicle), and tryptophan (1.36-fold increase for 0.1 µM DMF vs vehicle). Interestingly, taurine, whose levels can be modulated via Nrf2 signaling (DMF's primary target), emerged as a key mediator of DMF's neuronal action, displaying a 3.86-fold increase and 0.27-fold decrease for 10 µM DMF at 24 h and 96 h, respectively. A 96 h-exposure to DMF seemed to mainly trigger pathways associated with glucose production (e.g., gluconeogenesis, glucose-alanine cycle, malate-aspartate shuttle), possibly related to the metabolism of DMF into monomethyl fumarate and its further conversion into glucose via activation of the citric acid cycle. Overall, our data contribute to improving the understanding of the events associated with neuronal exposure to DMF.
Subject(s)
Dimethyl Fumarate , Neuroblastoma , Humans , Dimethyl Fumarate/toxicity , Dimethyl Fumarate/therapeutic use , NF-E2-Related Factor 2/metabolism , Neuroblastoma/metabolism , Neurons/metabolism , Glucose/metabolismABSTRACT
This is the first study to analyze the anti-inflammatory and antinociceptive effect of withanicandrin, isolated from Datura Ferox leaves, and the possible mechanism of action involved in adult zebrafish (ZFa). To this end, the animals were treated intraperitoneally (i. p.) with withanicandrin (4; 20 and 40â mg/kg; 20â µL) and subjected to locomotor activity and acute toxicity. Nociception tests were also carried out with chemical agents, in addition to tests to evaluate inflammatory processes induced by κ-Carrageenan 1.5 % and a Molecular Docking study. As a result, withanicandrin reduced nociceptive behavior by capsaicin at a dose of 40â mg/kg and by acid saline at doses of 4 and 40â mg/kg, through neuromodulation of TRPV1 channels and ASICs, identified through blocking the antinociceptive effect of withanicandrin by the antagonists capsazepine and naloxone. Furthermore, withanicandrin caused an anti-inflammatory effect through the reduction of abdominal edema, absence of leukocyte infiltrate in the liver tissue and reduction of ROS in thel liver tissue and presented better affinity energy compared to control morphine (TRPV1) and ibuprofen (COX-1 and COX-2).
Subject(s)
Analgesics , Zebrafish , Animals , Analgesics/pharmacology , Analgesics/chemistry , Analgesics/isolation & purification , Acid Sensing Ion Channels/metabolism , Molecular Docking Simulation , Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/isolation & purification , Carrageenan , Structure-Activity Relationship , Dose-Response Relationship, Drug , TRPV Cation Channels/antagonists & inhibitors , TRPV Cation Channels/metabolism , Edema/drug therapy , Edema/chemically induced , Plant Leaves/chemistry , Molecular StructureABSTRACT
Mitochondria play a critical role in the regulation of several biological processes (e.g., programmed cell death, inflammation, neurotransmission, cell differentiation). In recent years, accumulating findings have evidenced that cannabinoids, a group of endogenous and exogenous (synthetic and plant-derived) psychoactive compounds that bind to cannabinoid receptors, may modulate mitochondrial function and dynamics. As such, mitochondria have gained increasing interest as central mediators in cannabinoids' pharmacological and toxicological signatures. Here, we review the mechanisms underlying the cannabinoids' modulation of mitochondrial activity and dynamics, as well as the potential implications of such mitochondrial processes' disruption on cell homeostasis and disease. Interestingly, cannabinoids may target different mitochondrial processes (e.g., regulation of intracellular calcium levels, bioenergetic metabolism, apoptosis, and mitochondrial dynamics, including mitochondrial fission and fusion, transport, mitophagy, and biogenesis), by modulating multiple and complex signaling pathways. Of note, the outcome may depend on the experimental models used, as well as the chemical structure, concentration, and exposure settings to the cannabinoid, originating equivocal data. Notably, this interaction seems to represent not only an important feature of cannabinoids' toxicological signatures, with potential implications for the onset of distinct pathological conditions (e.g., cancer, neurodegenerative diseases, metabolic syndromes), but also an opportunity to develop novel therapeutic strategies for such pathologies, which is also discussed in this review.
Subject(s)
Cannabinoids , Cannabinoids/pharmacology , Cannabinoids/therapeutic use , Cannabinoids/analysis , Mitochondria/metabolism , Synaptic Transmission , Mitophagy , Energy MetabolismABSTRACT
Research on the health services' response to substance use disorders and respective comorbidities holds major relevance due to the increasing prevalence of these ailments. We thus invite contributions to a new Collection of articles launched by BMC Health Services Research titled "Health services for substance use disorders".
Subject(s)
Substance-Related Disorders , Humans , Substance-Related Disorders/epidemiology , Substance-Related Disorders/therapy , Health Services , ComorbidityABSTRACT
The effects of a cooling strategy following repeated high-intensity running (RHIR) on soccer kicking performance in a hot environment (>30ºC) were investigated in youth soccer players. Fifteen academy under-17 players participated. In Experiment 1, players completed an all-out RHIR protocol (10×30 m, with 30s intervals). In Experiment 2 (cross-over design), participants performed this running protocol under two conditions: (1) following RHIR 5 minutes of cooling where ice packs were applied to the quadriceps/hamstrings, (2) a control condition involving passive resting. Perceptual measures [ratings of perceived exertion (RPE), pain and recovery], thigh temperature and kick-derived video three-dimensional kinematics (lower limb) and performance (ball speed and two-dimensional placement indices) were collected at baseline, post-exercise and intervention. In Experiment 1, RHIR led to small-to-large impairments (p < 0.03;d = -0.42--1.83) across perceptual, kinematic and performance measures. In experiment 2, RPE (p < 0.01; Kendall's W = 0.30) and mean radial error (p = 0.057; η2 = 0.234) increased only post-control. Significant small declines in ball speed were also observed post-control (p < 0.05; d = 0.35). Post-intervention foot centre-of-mass velocity was moderately faster in the cooling compared to control condition (p = 0.04; d = 0.60). In youth soccer players, a short cooling period was beneficial in counteracting declines in kicking performance, in particular ball placement, following intense running activity in the heat.
Subject(s)
Athletic Performance , Running , Soccer , Adolescent , Humans , Biomechanical Phenomena , Hot Temperature , Cross-Over StudiesABSTRACT
Immunohistochemical staining of cell and molecular targets in brain samples is a powerful tool that can provide valuable information on neurological mechanisms. However, post-processing of photomicrographs acquired after 3,3'-Diaminobenzidine (DAB) staining is particularly challenging due to the complexity associated with the size, samples number, analyzed targets, image quality, and even the subjectivity inherent to the analysis by different users. Conventionally, this analysis relies on the manual quantification of distinct parameters (e.g., the number and size of cells and the number and length of cell branching) in a large set of images. These represent extremely time-consuming and complex tasks, defaulting the processing of high amounts of information. Here we describe an improved semi-automatic method to quantify glial fibrillary acidic protein (GFAP)-labelled astrocytes in immunohistochemistry images of rat brains, at magnifications as low as 20×. This method is a straightforward adaptation of the Young & Morrison method, using ImageJ's plugin Skeletonize, coupled with intuitive data processing in datasheet-based software. It allows swifter and more efficient post-processing of brain tissue samples, regarding astrocyte size and number quantification, the total area occupied, as well as astrocyte branching and branch length (indicators of astrocyte activation), thus contributing to better understand the possible inflammatory response developed by astrocytes.
Subject(s)
Astrocytes , Brain , Rats , Animals , Astrocytes/metabolism , Immunohistochemistry , Glial Fibrillary Acidic Protein/metabolism , Brain/metabolism , Head , NeurogenesisABSTRACT
Citrus canker, caused by the bacterium Xanthomonas citri (Xcc), is one of the most devastating diseases for the citrus industry. Xylose is a constituent of the cell wall of plants, and the ability of Xcc to use this carbohydrate may play a role in virulence. Xcc has two genes codifying for xylose isomerase (XI), a bifunctional enzyme that interconverts D-xylose into D-xylulose and D-glucose into D-fructose. The aim of this work was to investigate the functional role of the two putative XI ORFs, XAC1776 (xylA1) and XAC4225 (xylA2), in Xcc pathogenicity. XI-coding genes of Xcc were deleted, and the single mutants (XccΔxylA1 or XccΔxylA2) or the double mutant (XccΔxylA1ΔxylA2) remained viable. The deletion of one or both XI genes (xylA1 and/or xylA2) increased the aggressiveness of the mutants, causing disease symptoms. RT-qPCR analysis of wild strain and xylA deletion mutants grown in vivo and in vitro revealed that the highest expression level of hrpX and xylR was observed in vivo for the double mutant. The results indicate that XI depletion increases the expression of the hrp regulatory genes in Xcc. We concluded that the intracellular accumulation of xylose enhances Xcc virulence.
Subject(s)
Citrus , Xanthomonas , Virulence/genetics , Xylose/metabolism , Citrus/metabolism , Plant Diseases/microbiologyABSTRACT
Synthetic cathinones, such as 3,4-methylenedioxypyrovalerone (MDPV), are widely abused due to their psychostimulant effects. As they are chiral molecules, studies of their stereochemical stability (racemization can occur in certain temperatures and acidic/basic environments) and of their biological and/or toxicity effects (enantiomers might display different properties) are of great relevance. In this study, the liquid chromatography (LC) semi-preparative enantioresolution of MDPV was optimized to collect both enantiomers with high recovery rates and enantiomeric ratio (e.r.) values. The absolute configuration of the MDPV enantiomers was determined by electronic circular dichroism (ECD) with the aid of theoretical calculations. The first eluted enantiomer was identified as S-(-)-MDPV and the second eluted enantiomer was identified as R-(+)-MDPV. A racemization study was performed by LC-UV, showing enantiomers' stability up to 48 h at room temperature and 24 h at 37 °C. Racemization was only affected by higher temperatures. The potential enantioselectivity of MDPV in cytotoxicity and in the expression of neuroplasticity-involved proteins-brain-derived neurotrophic factor (BDNF) and cyclin-dependent kinase 5 (Cdk5)-was also evaluated using SH-SY5Y neuroblastoma cells. No enantioselectivity was observed.
Subject(s)
Central Nervous System Stimulants , Neuroblastoma , Humans , Synthetic Cathinone , Stereoisomerism , Chromatography, Liquid , Pyrrolidines/chemistry , Benzodioxoles/chemistryABSTRACT
The fermented beverage industry is always pursuing alternatives to make products that delight consumers with special or unique characteristics. The identification and improvement of new yeast strains emerge as an opportunity; however, wild strains usually have a limitation in maltose fermentation and/or off-flavors production. Here we report the production of a Blond-style ale beer using a bioethanol isolated strain (LBGA-287) with flavor complexity approved in sensorial panels. LBGA-287 also showed an increase in maltose consumption, growth and fermentation rates when compared to the commercial yeast. Using qPCR analysis, genes related to the (i) efficiency of fermentation (ii) production of aromas/off-flavors, and (iii) metabolization of carbohydrates were found as differentially expressed in the isolated strains when compared to industrial yeast. This suggests that LBGA-287 could have an important impact on beer production, improving brewing efficiency, quality and diversity of this beverage, and most importantly satisfying the final consumer.
Subject(s)
Beer , Saccharomyces cerevisiae , Beer/analysis , Ethanol/analysis , Fermentation , Fermented Beverages , Saccharomyces cerevisiae/geneticsABSTRACT
Synthetic cathinones are among the most popular new psychoactive substances, being abused for their stimulant properties, which are similar to those of amphetamine and 3,4-methylenedioxymethamphetamine (MDMA). Considering that the liver is a likely target for cathinones-induced toxicity, and for their metabolic activation/detoxification, we aimed to determine the hepatotoxicity of three commonly abused synthetic cathinones: butylone, α-methylamino-butyrophenone (buphedrone) and 3,4-dimethylmethcathinone (3,4-DMMC). We characterized their cytotoxic profile in primary rat hepatocytes (PRH) and in the HepaRG and HepG2 cell lines. PRH was the most sensitive cell model, showing the lowest EC50 values for all three substances (0.158 mM for 3,4-DMMC; 1.21 mM for butylone; 1.57 mM for buphedrone). Co-exposure of PRH to the synthetic cathinones and CYP450 inhibitors (selective and non-selective) proved that hepatic metabolism reduced the toxicity of buphedrone but increased that of butylone and 3,4-DMMC. All compounds were able to increase oxidative stress, disrupting mitochondrial homeostasis and inducing apoptotic and necrotic features, while also increasing the occurrence of acidic vesicular organelles in PRH, compatible with autophagic activation. In conclusion, butylone, buphedrone and 3,4-DMMC have hepatotoxic potential, and their toxicity lies in the interference with a number of homeostatic processes, while being influenced by their metabolic fate.
Subject(s)
3,4-Methylenedioxyamphetamine/analogs & derivatives , Butyrophenones/toxicity , Chemical and Drug Induced Liver Injury/etiology , Methylamines/toxicity , Propiophenones/toxicity , 3,4-Methylenedioxyamphetamine/administration & dosage , 3,4-Methylenedioxyamphetamine/toxicity , Animals , Autophagy/drug effects , Butyrophenones/administration & dosage , Cell Line, Tumor , Chemical and Drug Induced Liver Injury/pathology , Designer Drugs/administration & dosage , Designer Drugs/toxicity , Dose-Response Relationship, Drug , Hep G2 Cells , Hepatocytes/drug effects , Hepatocytes/pathology , Humans , Male , Methylamines/administration & dosage , Oxidative Stress/drug effects , Propiophenones/administration & dosage , Rats , Rats, WistarABSTRACT
OBJECTIVES: This study evaluated the capacity of cardiac risk stratification protocols on simple complications that occur during activities of a cardiovascular rehabilitation program. DESIGN: Observational longitudinal cohort study. SETTING: Outpatient clinic of cardiovascular rehabilitation. SUBJECT: Patients diagnosed with cardiovascular disease and/or risk factors. INTERVENTIONS: Not applicable. MAIN MEASURES: The relationship between the cardiac risk classes of seven risk stratification protocols and the occurrence of simple complications (such angina, abnormal changes in blood pressure, arrhythmias, fatigue, muscle pain, pallor) was assessed using the chi-square test, and when statistical significance was observed, sensitivity, specificity and accuracy were determined. RESULTS: About 76 patients were analyzed. The American Association of Cardiovascular and Pulmonary Rehabilitation (AACVPR) protocol showed a statistically significant relationship between simple complications and cardiac risk classes (P-value = 0.046), however the results of sensitivity (0.53), specificity (0.52), and accuracy (0.53) were not significant. The other protocols analyzed were not significant: American College of Sports Medicine (P-value = 0.801), Brazilian Society of Cardiology (P-value = 0.734), American Heart Association (P-value = 0.957), Pashkow (P-value = 0.790), Society French Cardiology (P-value = 0.314), and Spanish Society of Cardiology (P-value = 0.078). CONCLUSION: The AACVPR protocol showed a significant relationship between the risk classes and the occurrence of simple complications, however, the low values obtained for sensitivity, specificity and accuracy show that it is not useful for this purpose. CLINICAL TRIALS REGISTRATION: NCT03446742.
Subject(s)
Cardiac Rehabilitation , Cardiovascular Diseases/complications , Aged , Clinical Protocols , Female , Humans , Longitudinal Studies , Male , Middle Aged , Predictive Value of Tests , Risk Assessment , Risk Factors , United StatesABSTRACT
The endocannabinoid (eCB) system is a complex system comprising endogenous cannabinoids (eCBs), their synthesis and degradation enzymes, and cannabinoid receptors. These elements crucially regulate several biological processes during neurodevelopment, such as proliferation, differentiation, and migration. Recently, eCBs were also reported to have an epigenetic action on genes that play key functions in the neurotransmitter signaling, consequently regulating their expression. In turn, epigenetic modifications (e.g. DNA methylation, histone modifications) may also modulate the function of eCB system's elements. For example, the expression of the cnr gene in the central nervous system may be epigenetically regulated (e.g. DNA methylation, histone modifications), thus altering the function of the cannabinoid receptor type-1 (CB1R). Considering the importance of the eCB system during neurodevelopment, it is thus reasonable to expect that alterations in this interaction between the eCB system and epigenetic modifications may give rise to neurodevelopmental disorders. Here, we review key concepts related to the regulation of neuronal function by the eCB system and the different types of epigenetic modifications. In particular, we focus on the mechanisms involved in the intricate interplay between both signaling systems and how they control cell fate during neurodevelopment. Noteworthy, such mechanistic understanding assumes high relevance considering the implications of the dysregulation of key neurogenic processes towards the onset of neurodevelopment-related disorders. Moreover, considering the increasing popularity of cannabis and its synthetic derivatives among young adults, it becomes of utmost importance to understand how exogenous cannabinoids may epigenetically impact neurodevelopment.
Subject(s)
Endocannabinoids/metabolism , Epigenesis, Genetic , Neurogenesis/genetics , Animals , Cannabinoids/pharmacology , Humans , Neurodevelopmental Disorders/genetics , Neurodevelopmental Disorders/metabolism , Receptors, Cannabinoid/metabolism , Signal TransductionABSTRACT
New phenylethylamine derivatives are among the most commonly abused new psychoactive substances. They are synthesized and marketed in lieu of classical amphetaminic stimulants, with no previous safety testing. Our study aimed to determine the in vitro hepatotoxicity of two benzofurans [6-(2-aminopropyl)benzofuran (6-APB) and 5-(2-aminopropyl)benzofuran (5-APB)] that have been misused as 'legal highs'. Cellular viability was assessed through the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay, following 24-h drug exposure of human hepatoma HepaRG cells (EC50 2.62 mM 5-APB; 6.02 mM 6-APB), HepG2 cells (EC50 3.79 mM 5-APB; 8.18 mM 6-APB) and primary rat hepatocytes (EC50 964 µM 5-APB; 1.94 mM 6-APB). Co-incubation of primary hepatocytes, the most sensitive in vitro model, with CYP450 inhibitors revealed a role of metabolism, in particular by CYP3A4, in the toxic effects of both benzofurans. Also, 6-APB and 5-APB concentration-dependently enhanced oxidative stress (significantly increased reactive species and oxidized glutathione, and decreased reduced glutathione levels) and unsettled mitochondrial homeostasis, with disruption of mitochondrial membrane potential and decline of intracellular ATP. Evaluation of cell death mechanisms showed increased caspase-8, -9, and -3 activation, and nuclear morphological changes consistent with apoptosis; at concentrations higher than 2 mM, however, necrosis prevailed. Concentration-dependent formation of acidic vesicular organelles typical of autophagy was also observed for both drugs. Overall, 5-APB displayed higher hepatotoxicity than its 6-isomer. Our findings provide new insights into the potential hepatotoxicity of these so-called 'safe drugs' and highlight the putative risks associated with their use as psychostimulants.
Subject(s)
Benzofurans/toxicity , Designer Drugs/toxicity , Hepatocytes/drug effects , Propylamines/toxicity , Animals , Autophagy/drug effects , Cells, Cultured , Chemical and Drug Induced Liver Injury/pathology , Cytochrome P-450 Enzyme Inhibitors/toxicity , Cytochrome P-450 Enzyme System/metabolism , Hep G2 Cells , Hepatocytes/metabolism , Humans , Isomerism , Male , Membrane Potential, Mitochondrial/drug effects , Oxidative Stress/drug effects , Rats, Wistar , Reactive Oxygen Species/metabolismABSTRACT
Drug-induced Mood- and Cognition-related adverse events (MCAEs) are often only detected during the clinical trial phases of drug development, or even after marketing, thus posing a major safety concern and a challenge for both pharmaceutical companies and clinicians. To fill some gaps in the understanding and elucidate potential biological mechanisms of action frequently associated with MCAEs, we present a unique workflow linking observational population data with the available knowledge at molecular, cellular, and psychopharmacology levels. It is based on statistical analysis of pharmacovigilance reports and subsequent signaling pathway analyses, followed by evidence-based expert manual curation of the outcomes. Our analysis: (a) ranked pharmaceuticals with high occurrence of such adverse events (AEs), based on disproportionality analysis of the FDA Adverse Event Reporting System (FAERS) database, and (b) identified 120 associated genes and common pathway nodes possibly underlying MCAEs. Nearly two-thirds of the identified genes were related to immune modulation, which supports the critical involvement of immune cells and their responses in the regulation of the central nervous system function. This finding also means that pharmaceuticals with a negligible central nervous system exposure may induce MCAEs through dysregulation of the peripheral immune system. Knowledge gained through this workflow unravels putative hallmark biological targets and mediators of drug-induced mood and cognitive disorders that need to be further assessed and validated in experimental models. Thereafter, they can be used to substantially improve in silico/in vitro/in vivo tools for predicting these adversities at a preclinical stage.
Subject(s)
Adverse Drug Reaction Reporting Systems , Affect/drug effects , Brain/drug effects , Cognition/drug effects , Cognitive Dysfunction/chemically induced , Data Mining , Mood Disorders/chemically induced , Pharmacovigilance , Brain/metabolism , Brain/physiopathology , Cognitive Dysfunction/genetics , Cognitive Dysfunction/metabolism , Cognitive Dysfunction/psychology , Gene Expression Regulation , Gene Regulatory Networks , Humans , Mood Disorders/genetics , Mood Disorders/metabolism , Mood Disorders/psychology , Protein Interaction Maps , Risk Assessment , Signal TransductionABSTRACT
Cannabinoids comprise a broad group of psychoactive substances that activate endogenous cannabinoid (EC) receptors (ie, CB1 R and CB2 R), altering neurotransmitter release in the brain. The importance of their regulatory role in different biological processes has prompted the development of synthetic cannabinoids (SCs), substantially more potent than tetrahydrocannabinol (THC, the main psychoactive substance of cannabis). Although SCs were primarily designed given their therapeutic applications, their recreational use has become a major public health concern due to several reports of severe intoxications and deaths. SCs have favored increased popularity over recent years due to their intensified psychoactive effects, compared with THC, turning regular cannabis users into SCs. Among cannabinoid users (mainly young people), pregnant women and women of child-bearing potential (WoCBP) comprise particular risk groups, due to the potential onset of neurodevelopment disorders in the offspring (eg, schizophrenia and autism spectrum disorders). Understanding the role played by cannabinoids, and the potential action of emerging SCs in the regulation of the neuronal function, especially during neuronal development, thus assumes critical relevance. Here, we review the mechanistic regulation of neuronal processes, namely during neuronal development, by the endocannabinoid system. Most important, we further develop on the potential of SCs to modulate such mechanisms and subsequently disrupt proper neurodevelopment.
Subject(s)
Brain/drug effects , Brain/embryology , Cannabinoids/pharmacology , Fetal Development/drug effects , Illicit Drugs/pharmacology , Animals , Brain/physiopathology , Female , Humans , Male , Pregnancy , RatsABSTRACT
Recreational use of synthetic cannabinoids (SCs) before and during pregnancy poses a major public health risk, due to the potential onset of neurodevelopmental disorders in the offspring. Herein, we report the assessment of the neurotoxic potential of two commonly abused SCs, THJ-2201 and 5F-PB22, particularly focusing on how they affect neuronal differentiation in vitro. Differentiation ratios, total neurite length, and neuronal marker expression were assessed in NG108-15 neuroblastoma x glioma cells exposed to the SCs at non-toxic, biologically relevant concentrations (≤1 µM), either in acute or repeated exposure settings. Both SCs enhanced differentiation ratios and total neurite length of NG108-15 cells near two-fold compared to vehicle-treated cells, in a CB1R activation-dependent way, as the CB1R blockade with a specific antagonist (SR141718) abrogated SC-induced effects. Interestingly, repeated 5F-PB22 exposure was required to reach effects similar to a single THJ-2201 dose. Cell viability and proliferation, mitochondrial membrane potential, and intracellular ATP levels were also determined. The tested SCs increased mitochondrial tetramethyl rhodamine ethyl ester (TMRE) accumulation after 24 h at biologically relevant concentrations but did not affect any of the other toxicological parameters. Overall, we report firsthand the CB1R-mediated enhancement of neurodifferentiation by 5F-PB22 and THJ-2201 at biologically relevant concentrations.
Subject(s)
Cannabinoid Receptor Agonists/pharmacology , Cell Differentiation , Glioma/pathology , Indazoles/pharmacology , Indoles/pharmacology , Naphthalenes/pharmacology , Neuroblastoma/pathology , Quinolines/pharmacology , Receptor, Cannabinoid, CB1/metabolism , Animals , Cell Survival , Glioma/drug therapy , Glioma/metabolism , Mice , Mitochondria/drug effects , Mitochondria/metabolism , Neuroblastoma/drug therapy , Neuroblastoma/metabolism , Rats , Tumor Cells, CulturedABSTRACT
Beta-thalassaemia (BT) is classified according to blood transfusion requirement as minor (BTMi), intermedia (BTI) and major (BTM). BTM is the most severe form, requiring regular transfusions while transfusion need is only occasional in BTI. Differential gene expression between patients has not been assessed so far. Here, we evaluated the global gene expression profiles during differentiation of human erythroid cells of two patients carrying the same mutation [CD39, (C â T)], though displaying different phenotypes (BTI and BTM). Considering the role of reactive oxygen species (ROS) in the pathophysiology of thalassaemia, we focused on differentially expressed genes involved in metabolic pathways triggered by ROS, such as inflammation and apoptosis, and, from these, we selected the Apurinic/Apyrimidinic Endodeoxyribonuclease 1 (APEX1) and High Mobility Group Box1 (HMGB1) genes, whose role in BT is not well established. An in-depth expression analysis of transcriptional and protein levels in patients carrying a range of mutations associated with BT phenotypes indicated that APEX1 was increased in both BTI and BTM. Furthermore, higher amounts of HMGB1 was found in the plasma of BTI patients. Our findings suggest that these proteins have important roles in BT and could represent new targets for further studies aiming to improve the management of the disease.
Subject(s)
DNA-(Apurinic or Apyrimidinic Site) Lyase/genetics , HMGB1 Protein/genetics , Oxidative Stress , Signal Transduction , Transcriptome , beta-Thalassemia/genetics , beta-Thalassemia/metabolism , Adult , Apoptosis , Apyrase/metabolism , Biomarkers , Case-Control Studies , Cell Differentiation/genetics , Computational Biology/methods , DNA-(Apurinic or Apyrimidinic Site) Lyase/metabolism , Erythroid Cells/cytology , Erythroid Cells/metabolism , Female , Gene Expression Profiling , Gene Expression Regulation , Gene Regulatory Networks , HMGB1 Protein/metabolism , Humans , Male , Middle Aged , Phenotype , Reactive Oxygen Species/metabolism , beta-Thalassemia/diagnosisABSTRACT
3-Methylmethcathinone (3-MMC or metaphedrone) has become one of the most popular recreational drugs worldwide after the ban of mephedrone, and was recently deemed responsible for several intoxications and deaths. This study aimed at assessing the hepatotoxicity of 3-MMC. For this purpose, Wistar rat hepatocytes were isolated by collagenase perfusion, cultured and exposed for 24 h at a concentration range varying from 31 nM to 10 mM 3-MMC. The modulatory effects of cytochrome P450 (CYP) inhibitors on 3-MMC hepatotoxicity were evaluated. 3-MMC-induced toxicity was perceived at the lysosome at lower concentrations (NOEC 312.5 µM), compared to mitochondria (NOEC 379.5 µM) and cytoplasmic membrane (NOEC 1.04 mM). Inhibition of CYP2D6 and CYP2E1 diminished 3-MMC cytotoxicity, yet for CYP2E1 inhibition this effect was only observed for concentrations up to 1.3 mM. A significant concentration-dependent increase of intracellular reactive species was observed from 10 µM 3-MMC on; a concentration-dependent decrease in antioxidant glutathione defences was also observed. At 10 µM, caspase-3, caspase-8, and caspase-9 activities were significantly elevated, corroborating the activation of both intrinsic and extrinsic apoptosis pathways. Nuclear morphology and formation of cytoplasmic acidic vacuoles suggest prevalence of necrosis and autophagy at concentrations higher than 10 µM. No significant alterations were observed in the mitochondrial membrane potential, but intracellular ATP significantly decreased at 100 µM. Our data point to a role of metabolism in the hepatotoxicity of 3-MMC, which seems to be triggered both by autophagic and apoptotic/necrotic mechanisms. This work is the first approach to better understand 3-MMC toxicology.
Subject(s)
Apoptosis/drug effects , Autophagy/drug effects , Hepatocytes/drug effects , Methamphetamine/analogs & derivatives , Oxidative Stress/drug effects , Psychotropic Drugs/toxicity , Animals , Cell Survival/drug effects , Cells, Cultured , Dose-Response Relationship, Drug , Hepatocytes/metabolism , Hepatocytes/pathology , Humans , Male , Methamphetamine/toxicity , Primary Cell Culture , Rats, WistarABSTRACT
The feasibility of bacterial cellulose (BC) as a novel substrate for retinal pigment epithelium (RPE) culture was evaluated. Thin (41.6 ± 2.2 µm of average thickness) and heat-dried BC substrates were surface-modified via acetylation and polysaccharide adsorption, using chitosan and carboxymethyl cellulose. All substrates were characterized according to their surface chemistry, wettability, energy, topography, and also regarding their permeability, dimensional stability, mechanical properties, and endotoxin content. Then, their ability to promote RPE cell adhesion and proliferation in vitro was assessed. All surface-modified BC substrates presented similar permeation coefficients with solutes of up to 300 kDa. Acetylation of BC decreased it's swelling and the amount of endotoxins. Surface modification of BC greatly enhanced the adhesion and proliferation of RPE cells. All samples showed similar stress-strain behavior; BC and acetylated BC showed the highest elastic modulus, but the latter exhibited a slightly smaller tensile strength and elongation at break as compared to pristine BC. Although similar proliferation rates were observed among the modified substrates, the acetylated ones showed higher initial cell adhesion. This difference may be mainly due to the moderately hydrophilic surface obtained after acetylation.