Severe neurological nicotine intoxication by e-cigarette liquids: Systematic literature review.

Electronic cigarettes are a popular, easily purchased, alternative source of nicotine that is considered safer than conventional tobacco. However, Intentional or accidental exposure to e-liquid substances, mainly nicotine, can lead to serious, potentially fatal toxicity. Emergency and critical care physicians should keep in mind acute intoxication of this poison with a biphasic toxic syndrome. We highlight its potentially fatal outcome and suggest monitoring the adverse effects of nicotine according to a multimodal protocol integrating somatosensory evoked potentials, electroencephalography and neuroimaging data with anamnestic report and toxicological and laboratory data.

Ethanol Toxicity During Brain Development: Alterations of Excitatory Synaptic Transmission in Immature Organotypic Hippocampal Slice Cultures.

BACKGROUND: The developing brain is particularly vulnerable to alcohol: Drinking during pregnancy can lead to a number of physical, learning, and behavioral disorders in the newborn. It has been demonstrated that immature and mature brain tissues display a differential sensitivity to ethanol (EtOH) toxicity and that cerebral structure and function are diversely impaired according to the stage of synaptic maturation. METHODS: Rat organotypic hippocampal slice cultures were exposed for 7 days to EtOH (100 to 300 mM) after 2 days (immature) or 10 days (mature) of culture in vitro; EtOH was then removed from the medium, and 24 hours later, slices were analyzed by fluorescence microscopy, Western blotting, electrophysiology, and electron microscopy to explore the molecular mechanisms of EtOH toxicity in the developing hippocampus. RESULTS: EtOH withdrawal elicited a selective CA1 pyramidal cell injury in mature slices, but not in immature slices. A significant increase in the expression of pre- and postsynaptic proteins in mature slices revealed that slice maturation is presumably associated with the development of new synapses. Incubation with chronic EtOH for 7 days and its removal from the medium induced a significant decrease in GluA1 and GluA2 expression levels; a significant reduction in the expression of synaptophysin and GluN2A was observed only after EtOH withdrawal. Whole-cell patch-clamp recordings showed that incubation with EtOH for 7 days induced a significant decrease in spontaneous excitatory postsynaptic current (sEPSC) frequency in CA1 pyramidal cells of immature slices and a trend toward a decrease in sEPSC amplitude. Electron microscopy revealed a disorganization of neurotubuli in immature slices after chronic exposure to EtOH. CONCLUSIONS: These results indicate that prolonged incubation with EtOH and its subsequent withdrawal from the medium induce an impairment of excitatory synaptic transmission and possibly an incorrect formation of neuronal circuits in developing hippocampus in vitro, which is suggestive of mechanisms that may lead to mental retardation in fetal alcohol spectrum disorders.

Analytical quality by design: Development and control strategy for a LC method to evaluate the cannabinoids content in cannabis olive oil extracts.

Cannabidiol (CBD) and Δ9-tetrahydrocannabinol (Δ9-THC) are considered as the most interesting cannabinoids in Cannabis sativa L. for the clinical practice. Since 2013, the Italian law allows pharmacists to prepare and dispense cannabis extracts to patients under medical prescription, and requires the evaluation of CBD and Δ9-THC content in cannabis extracts before sale. Cannabis olive oil extracts are prepared from dried female cannabis inflorescences, but a standard protocol is still missing. In this study, a fast RP-HPLC/UV method has been developed to quantify CBD and Δ9-THC in cannabis olive oil extracts. The analytical quality by design strategy has been applied to the method development, setting critical resolution and total analysis time as critical method attributes (CMAs), and selecting column temperature, buffer pH and flow rate as critical method parameters. Information from Doehlert Design in response surface methodology combined to Monte-Carlo simulations led to draw the risk of failure maps and to identify the method operable design region. The method was validated according to the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines and then implemented in routine analysis. A control strategy based on system control charts was planned to monitor the developed method performances. Evaluation data were recorded over a period of one year of routine use, and both the CMAs showed values within the specifications in every analysis performed. Hence, a new risk evaluation for the future performances of the method was achieved by using a Bayesian approach based on the routine use data, computing the future distribution of the two CMAs. Finally, a study focusing on the monitoring of CBD and Δ9-THC concentrations in cannabis olive oil extracts was carried out. The developed method was applied to 459 extracts. The statistical analysis of the obtained results highlighted a wide variability in terms of concentrations among different samples from the same starting typology of cannabis, underlining the compelling need of a standardised procedure to harmonise the preparation of the extracts.

Galenic Preparations of Therapeutic Cannabis sativa Differ in Cannabinoids Concentration: A Quantitative Analysis of Variability and Possible Clinical Implications.

Introduction: Magistral preparations of therapeutic cannabis are extracted from standardized products imported from Holland or from the Florence Military Pharmaceutical Chemical Works, but extraction protocols differ among galenic laboratories. This study assessed the inter-laboratory variability in concentrations of cannabidiol (CBD), cannabinol (CBN), tetrahydrocannabinol (THC), and tetrahydrocannabinolic acid (THCA) among different magistral oil preparations. Methods: 219 samples of Bediol, Bedrobinol, Bedrolite or FM-2 70 or 100 mg/ml in oil were collected from 3 laboratories. Concentrations of CBD, CBN, THC, and THCA were quantified by high-pressure liquid chromatography; inter-laboratories variability was assessed using the Kruskal-Wallis test. Results: A significant variability in CBD and THC concentrations was found for Bediol 70 mg/ml samples from 2 laboratories [for CBD: median 5.4 (range 4.8-6.6) vs. 6.1 (4.9-7.2) mg/ml, p = 0.033; for THC: 3.6 (3.1-3.9) vs. 4.0 (2.6-5.1) mg/ml, p = 0.020]. As for Bediol 100 mg/ml, a significant variability emerged in THC concentrations among the three considered laboratories [5.7 (-) vs. 4.2 (1.5-4.8) vs. 5.2 (4.2-6.9), p = 0.030]. No significant inter-laboratory variability emerged for Bedrocan and Bedrolite. Concentrations of CBD, CBN, and THC were

Group I metabotropic glutamate receptors stimulate the activity of poly(ADP-ribose) polymerase in mammalian mGlu1-transfected cells and in cortical cell cultures.

Group I metabotropic glutamate (mGlu) receptors (i.e. mGlu1 and mGlu5) coupled to phospholipase C have been widely investigated for their possible role in excitotoxic and post-ischemic neuronal death. Recently, phospholipase C has been shown to directly stimulate the activity of poly(ADP-ribose) polymerase (PARP), a nuclear enzyme involved in DNA repair that has been proposed to play a key role in necrotic cell death. In this study, we investigated whether the stimulation of group I mGlu receptors leads to an increase in PARP activity, as detected by flow cytometry, immunodot blot and immunocytochemistry, both in baby hamster kidney cells transfected with mGlu1a or mGlu5a receptors and in cultured cortical cells. Our results show that the group I mGlu receptor agonist DHPG elicited a significant increase in PARP activity that was completely abolished by the administration of the mGlu1 antagonist 3-MATIDA and partially prevented, in cortical neurons, by the mGlu5 antagonist MPEP. To evaluate whether this pathway is involved in post-ischemic neuronal death, we used a sublethal model of oxygen-glucose deprivation in mixed cortical cell cultures. DHPG exacerbated neuronal death, and this effect was significantly prevented by the application of the PARP inhibitor DPQ. This novel pathway may contribute to the effects of mGlu1 receptors in the mechanisms leading to post-ischemic neuronal death.

Neutrophils CD11b and fibroblasts PGE(2) are elevated in Alzheimer's disease.

To evaluate whether inflammation-like mechanisms present in the brain of Alzheimer's disease (AD) patients are reflected in the periphery, the expression of CD11b in peripheral blood neutrophils and the expression and activity of inflammatory markers in cultured skin fibroblasts were examined. We found significantly higher levels of CD11b in neutrophils from sporadic AD patients than in controls and this elevation was positively correlated with disease severity and progression rate of mental decline. Cultured skin fibroblasts from familial (FAD) and sporadic AD patients and from controls were immunopositive for both isoforms of cyclooxygenase with no differences between groups. In unstimulated culture, the production of prostaglandin-E2 in the medium was significantly higher in fibroblasts from sporadic AD and FAD patients than in controls, and this elevation was reverted by the addition of 25 microM of ibuprofen. Our findings provide further evidence of the presence of inflammatory and immuno-related markers in the periphery of AD patients and support those studies indicating the beneficial effects of anti-inflammatory therapy in AD.

Carbon monoxide modulates the response of human basophils to FcepsilonRI stimulation through the heme oxygenase pathway.

We report the effects of exogenous and endogenous carbon monoxide (CO) on the immunological activation of human basophils. Hemin (1-100 microM), a heme oxygenase substrate analogue, significantly increased the formation of bilirubin from partially purified human basophils, thus indicating that these cells express heme oxygenase. This effect was reversed by preincubating the cells for 30 min with Zn-protoporphyrin IX (100 microM), a heme oxygenase inhibitor. Hemin (100 microM) also decreased immunoglobulin G anti-Fcepsilon (anti-IgE)-induced activation of basophils, measured by the expression of a membrane granule-associated protein, identified as cluster differentiation protein 63 (CD63), and by histamine release. These effects were reversed by Zn-protoporphyrin IX (100 microM), by oxyhemoglobin (HbO(2)), a CO scavenger (100 microM), and by 1H-[1,2,4]oxadiazolo[4,3-a] quinoxalin-1-one (ODQ), an inhibitor of the soluble guanylyl cyclase (100 microM). Exposure of basophils to exogenous CO (10 microM for 30 min) also decreased their activation, while nitrogen (N(2)) was ineffective. HbO(2) and ODQ reversed the inhibition, reversing both membrane protein CD63 expression and histamine release to basal values. Both hemin and exogenous CO significantly raised cGMP levels in basophils and blunted the rise of calcium levels caused by immunological activation. This study suggests that CO increases cGMP formation, which in turn induces a fall in intracellular Ca(2+) concentration, thereby resulting in the inhibition of human basophil activation.

Inhibitory effects of relaxin on human basophils activated by stimulation of the Fc epsilon receptor. The role of nitric oxide.

This study investigates whether relaxin (RLX), a hormone previously shown to inhibit mast cell function and to stimulate endogenous nitric oxide (NO) biosynthesis, counteracts the activation of isolated human basophils stimulated with anti-IgE or phorbol ester, and, if so, whether NO is involved. RLX reduced dose-dependently the expression of the activation marker CD63, the release of histamine and the rise of intracellular Ca2+ levels which triggers granule release by stimulated basophils. RLX also blunts the ultrastructural signs of anaphylactic granule release. The effects of RLX appear to depend upon activation of Ca2+/calmodulin-dependent NO synthase and endogenous NO production. They were reproduced by the NO donor sodium nitroprusside (SNP) and were reverted by the NO synthase inhibitor N(omega)-monomethyl-L-arginine, or by the NO scavenger oxyhemoglobin, or by blocking the NO physiological target guanylate cyclase with ODQ. In conclusion, RLX appears to play a role in down-regulating basophil function upon immunologic and nonimmunologic activation.

Poly(ADP-ribose) polymerase as a key player in excitotoxicity and post-ischemic brain damage.

Poly(ADP-ribose) polymerases (PARPs) are a group of protein-modifying and nucleotide-polymerizing enzymes able to catalyze the transfer of multiple ADP-ribose units from NAD to substrate proteins. In the human genome, 16 different genes encoding for members of this emerging family of enzymes have been identified. Known family members are PARP-1, PARP-2, PARP-3, vPARP, tankyrase 1 and tankyrase 2, each of them with a possible specific role in cell biology. The most studied member of the family is PARP-1, which is abundantly present in the nucleus and is involved in the maintenance of genomic stability. In pathological conditions, highly reactive radical species may cause DNA damage and PARP-1 hyperactivation. This may lead to necrotic cell death through massive NAD consumption. We show that following middle cerebral artery occlusion, rats treated with PARP inhibitors displayed reduced brain infarct volumes. Similarly, PARP inhibitors reduced neuronal death induced by oxygen-glucose deprivation (OGD) or excitotoxins in primary cultures of murine cortical cells. On the contrary, PARP inhibitors did not attenuate the OGD-induced selective loss of CA1 pyramidal cells in rat organotypic hippocampal slices. In addition, they were not neuroprotective against transient bilateral carotid occlusion in gerbils. We observed that post-ischemic brain damage was predominally necrotic in cultured cortical cells, whereas a caspase-dependent apoptotic process was responsible for the CA1 pyramidal cell loss in hippocampal slices. Hence, it appears reasonable to propose PARP inhibitors as useful therapeutic agents in pathological brain conditions were necrosis predominates.

Gastrointestinal bleeding and massive liver damage in neuroleptic malignant syndrome.

BACKGROUND: Neuroleptic malignant syndrome (NMS) is a rare side effect of antipsychotic therapy characterized by fever, muscular rigidity, altered mental status, increased level of serum creatinine phosphokinase, and increased number of white blood cells. The mortality rate of patients with NMS remains elevated. METHODS: We examined the clinical records of patients diagnosed with severe NMS admitted to the Clinical Toxicology Unit, Florence University Hospital, between 1990 and 2004. RESULTS: Eight patients presented with this neurological disorder. All were treated with supportive therapy, which included dantrolene, levodopa/benserazide, benzodiazepines, metamizole and/or paracetamol, and antibiotics. Five survived and three died. Of the three deceased, two had large hemorrhages in the gastrointestinal tract, and one had massive liver damage and diffuse hemorrhages throughout the body. CONCLUSION: Our results suggest that gastrointestinal bleeding is a frequent cause of death in NMS patients. Bleeding may occur as a consequence of commonly accepted medical treatments (especially the use of cyclooxygenase inhibitors as antipyretic agents) and NMS-induced changes in blood coagulation status. To increase the survival rate of these patients, it is necessary to avoid using drugs that may facilitate gastrointestinal lesions and to utilize procedures known to decrease the risk of bleeding.

Differential role of poly(ADP-ribose) polymerase-1in apoptotic and necrotic neuronal death induced by mild or intense NMDA exposure in vitro.

Overactivation of the nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) plays a key role in the mechanisms responsible for neuronal death. In the present study, we examined the effects of the PARP-1 inhibitor 3,4-dihydro-5-[4-1(1-piperidinyl)buthoxy]-1(2H)-isoquinolinone (DPQ) in two models of N-methyl-d-aspartate (NMDA)-induced neurotoxicity. The exposure of mixed cultured cortical cells to 300 microM NMDA for 10 min induced a caspase-dependent type of apoptotic neuronal death. Conversely, exposure to 2 mM NMDA for 10 min led to the appearance of morphological features of necrosis, with no increase in caspase-3 activity and depletion in adenosine triphosphate (ATP) levels. DPQ (10 microM) reduced the NMDA-induced PARP activation, restored ATP to near control levels and significantly attenuated neuronal injury only in the severe NMDA exposure model. Similar results were obtained when pure neuronal cortical cultures were used. PARP-1 activation thus appears to play a preferential role in necrotic than in caspase-dependent apoptotic neuronal death.

Novel isoquinolinone-derived inhibitors of poly(ADP-ribose) polymerase-1: pharmacological characterization and neuroprotective effects in an in vitro model of cerebral ischemia.

Excessive activation of poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear enzyme catalyzing the transfer of ADP-ribose units from NAD to acceptor proteins, induces cellular energy failure by NAD and ATP depletion and has been proposed to play a causative role in a number of pathological conditions, including ischemia/reperfusion injury. In this study, we used an in vitro enzyme activity assay to characterize a series of newly synthesized isoquinolinone derivatives as potential PARP-1 inhibitors. Several compounds displayed powerful inhibitory activity: thieno[2,3-c]isoquinolin-5-one (TIQ-A) displayed a submicromolar IC50 of 0.45 +/- 0.1 microM, whereas the 5-hydroxy and 5-methoxy TIQ-A derivatives had IC50 values of 0.39 +/- 0.19 and 0.21 +/- 0.12 microM, respectively. We then examined the neuroprotective effects of the newly characterized compounds in cultured mouse cortical cells exposed to 60 min of oxygen and glucose deprivation (OGD). When PARP-1 inhibitors were present in the incubation medium during OGD and the subsequent 24-h recovery period, they significantly attenuated neuronal injury. TIQ-A provided neuroprotection even when added to the culture 30 min after OGD and was able to reduce the early activation of PARP induced by OGD as detected by flow cytometry. When the IC50 values observed in the PARP-1 activity assay for selected compounds were compared with their IC50 values for the neuroprotective activity, a significant correlation (r = 0.93, P < 0.01) was observed. Our results suggest that TIQ-A and its derivatives are a new class of neuroprotectants that may be helpful in studies aimed at understanding the involvement of PARP-1 in physiology and pathology.