Unexpected Variation of the Codeine/Morphine Ratio Following Fatal Heroin Overdose

Postmortem samples from 14 cases of suspected heroin overdose were subjected to a preliminary systematic toxicological analysis in order to highlight the presence of unknown exogenous compounds (e.g., drugs of abuse, alcohol) that may have played a role in the mechanism of death. This analysis unveiled histories of poly-drug use in seven of the cases under investigation. Moreover, the concentrations of morphine and codeine in the brain were also investigated, and the results were compared with the data obtained from the blood specimens. The concentration of morphine in blood ranged from 33 to 688 ng/mL, while the concentration of codeine ranged from 0 to 193 ng/mL. However, in the brain, the concentration of morphine was found to be between 85 and 396 ng/g, while the levels of codeine ranged from 11 to 160 ng/g. The codeine/morphine ratio in the blood ranged from 0.043 to 0.619; however, in the brain, the same ratio was found to be between 0.129 and 0.552. In most cases, a significantly higher codeine/morphine ratio was found in the brain, suggesting the accumulation of codeine in brain tissue due its high lipophilicity as compared with morphine

Detection and quantitation analysis of cocaine and metabolites in fixed liver tissue and formalin solutions.

This study reports the results of the detection and quantitation of cocaine and its metabolites in liver tissues fixed in formalin and in the formalin solutions in which the same tissues were fixed. Toxicological analyses were performed on formalin-fixed liver samples from four cases of death of cocaine abusers and on formalin solutions (10% buffered, pH 7) in which the samples were preserved. Analyses carried out at the time of autopsy on body fluids and tissues allowed identification of cocaine and the metabolite benzoylecgonine. Liver tissue samples were preserved in formalin solutions for four weeks before analysis. Results only showed the presence of benzoylecgonine in the studied materials. The mean levels of recovery of benzoylecgonine in fixed tissues were 12.31% in liver and 84.47% in formalin from liver. Results indicated that benzoylecgonine has good stability, even in biological specimens subjected to chemical fixation

Oxidative stress inhibits IFN-alpha-induced antiviral gene expression by blocking the JAK-STAT pathway.

Abstract BACKGROUND/AIMS: Unresponsiveness to IFN-alpha is common in chronic hepatitis C. Since conditions associated with an increased oxidative stress (advanced age, steatosis, fibrosis, iron overload, and alcohol consumption) reduce the likelihood of response, we hypothesized that oxidative stress may affect the antiviral actions of IFN-alpha. METHODS: We examined in a human hepatocellular carcinoma cell line (Huh-7) the effect of hydrogen peroxide (H2O2), as a generator of oxidative stress, on the IFN-alpha signaling pathway. RESULTS: Pretreatment of Huh-7 cells with 0.5-1 mM H2O2 resulted in the suppression of the IFN-alpha-induced antiviral protein MxA and of IRF-9 mRNA expression. The reduced expression of these genes was associated to H2O2 -mediated suppression of the IFN-alpha-induced assembly of signal transducer and activator of transcription (STAT) factors to specific promoter motifs on IFN-alpha-inducible genes. This was accomplished by preventing the IFN-alpha-induced tyrosine phosphorylation of STAT-1 and STAT-2 through the inactivation of the upstream receptor associated tyrosine kinases, JAK-1 and Tyk-2. The suppression was fast, occurring within 5mins of pretreatment with H2O2, and did not require protein synthesis. CONCLUSIONS: In conclusion, oxidative stress impairs IFN-alpha signaling and might cause resistance to the antiviral action of IFN-alpha in chronically HCV infected patients with high level of oxidative stress in the liver

Innate immune activating ligand SUMOylation affects tumor cell recognition by NK cells

Natural Killer cells are innate lymphocytes involved in tumor immunosurveillance. They express activating receptors able to recognize self-molecules poorly expressed on healthy cells but up-regulated upon stress conditions, including transformation. Regulation of ligand expression in tumor cells mainly relays on transcriptional mechanisms, while the involvement of ubiquitin or ubiquitin-like modifiers remains largely unexplored. Here, we focused on the SUMO pathway and demonstrated that the ligand of DNAM1 activating receptor, PVR, undergoes SUMOylation in multiple myeloma. Concurrently, we found that PVR is preferentially located in intracellular compartments in human multiple myeloma cell lines and malignant plasma cells and that inhibition of the SUMO pathway promotes its translocation to the cell surface, increasing tumor cell susceptibility to NK cell-mediated cytolysis. Our findings provide the first evidence of an innate immune activating ligand regulated by SUMOylation, and confer to this modification a novel role in impairing recognition and killing of tumor cells.Natural Killer cells are innate lymphocytes involved in tumor immunosurveillance. They express activating receptors able to recognize self-molecules poorly expressed on healthy cells but up-regulated upon stress conditions, including transformation. Regulation of ligand expression in tumor cells mainly relays on transcriptional mechanisms, while the involvement of ubiquitin or ubiquitin-like modifiers remains largely unexplored. Here, we focused on the SUMO pathway and demonstrated that the ligand of DNAM1 activating receptor, PVR, undergoes SUMOylation in multiple myeloma. Concurrently, we found that PVR is preferentially located in intracellular compartments in human multiple myeloma cell lines and malignant plasma cells and that inhibition of the SUMO pathway promotes its translocation to the cell surface, increasing tumor cell susceptibility to NK cell-mediated cytolysis. Our findings provide the first evidence of an innate immune activating ligand regulated by SUMOylation, and confer to this modification a novel role in impairing recognition and killing of tumor cells

Polymorphism in the corticotropin-releasing factor receptor 1 (CRF1-R) gene plays a role in shaping the high anxious phenotype of Marchigian Sardinian alcohol-preferring (msP) rats

Introduction: Marchigian Sardinian alcohol-preferring (msP) rats exhibit innate preference for alcohol along with anxious phenotype. In these animals, two single-nucleotide polymorphisms in position -1,836 and -2,097 from the first start codon of the CRF1-R transcript have been found. Materials and Methods: Here, we examined whether these point mutations account for the heightened anxiety-like behavior and stress responsiveness of msP rats. We rederived the msP rats to obtain two distinct lines carrying the wild-type (GG) and point mutations (AA), respectively. Results: CRF1-R gene expression analysis revealed significant dysregulation of the system in the extended amygdala of AA rats. At the behavioral level, using the elevated plus maze, we found that both AA and GG lines had higher basal anxiety compared to Wistar rats. In the defensive burying test, AA rats showed decreased burying behavior compared to the GG and the unselected Wistar lines. Freezing/immobility did not differ among AA and GG but was higher than that of Wistars. The selective CRF1-R antagonist antalarmin (0, 10, and 20 mg/kg) reduced burying behavior in Wistar animals. However, antalarmin (10 mg/kg) tended to increase rather than reducing this behavior when tested in the msP lines, an effect that appeared more marked in the GG as compared to the AA line. Conclusion: The present data suggest that rats with msP genetic background are more anxious and show different sensitivity to stress and CRF1-R blockade than Wistars. The point mutations occurring in the CRF1-R gene do not seem to influence basal anxiety while they appear to affect active responses to stress

Protection against alcohol-induced neuronal and cognitive damage by the PPARγ receptor agonist pioglitazone

Binge alcohol drinking has emerged as a typical phenomenon in young people. This pattern of drinking, repeatedly leading to extremely high blood and brain alcohol levels and intoxication is associated with severe risks of neurodegeneration and cognitive damage. Mechanisms involved in excitotoxicity and neuroinflammation are pivotal elements in alcohol-induced neurotoxicity. Evidence has demonstrated that PPARγ receptor activation shows anti-inflammatory and neuroprotective properties. Here we examine whether treatment with the PPARγ agonist pioglitazone is beneficial in counteracting neurodegeneration, neuroinflammation and cognitive damage produced by binge alcohol intoxication. Adult Wistar rats were subjected to a 4-day binge intoxication procedure, which is commonly used to model excessive alcohol consumption in humans. Across the 4-day period, pioglitazone (0, 30, 60mg/kg) was administered orally twice daily at 12-h intervals. Degenerative cells were detected by fluoro-jade B (FJ-B) immunostaining in brain regions where expression of pro-inflammatory cytokines was also determined. The effects of pioglitazone on cognitive function were assessed in an operant reversal learning task and the Morris water maze task. Binge alcohol exposure produced selective neuronal degeneration in the hippocampal dentate gyrus and the adjacent entorhinal cortex. Pioglitazone reduced FJ-B positive cells in both regions and prevented alcohol-induced expression of pro-inflammatory cytokines. Pioglitazone also rescued alcohol-impaired reversal learning in the operant task and spatial learning deficits in the Morris water maze. These findings demonstrate that activation of PPARγ protects against neuronal and cognitive degeneration elicited by binge alcohol exposure. The protective effect of PPARγ agonist appears to be linked to inhibition of pro-inflammatory cytokines

Liver X Receptors: Regulators of Cholesterol Metabolism, Inflammation, Autoimmunity, and Cancer

The interplay between cellular stress and immune response can be variable and sometimes contradictory. The mechanisms by which stress-activated pathways regulate the inflammatory response to a pathogen, in autoimmunity or during cancer progression remain unclear in many aspects, despite our recent knowledge of the signalling and transcriptional pathways involved in these diseases. In this context, over the last decade many studies demonstrated that cholesterol metabolism is an important checkpoint for immune homeostasis and cancer progression. Indeed, cholesterol is actively metabolized and can regulate, through its mobilization and/or production of active derivatives, many aspects of immunity and inflammation. Moreover, accumulation of cholesterol has been described in cancer cells, indicating metabolic addiction. The nuclear receptors liver-X-receptors (LXRs) are important regulators of intracellular cholesterol and lipids homeostasis. They have also key regulatory roles in immune response, as they can regulate inflammation, innate and adaptive immunity. Moreover, activation of LXRs has been reported to affect the proliferation and survival of different cancer cell types that show altered metabolic pathways and accumulation of cholesterol. In this minireview we will give an overview of the recent understandings about the mechanisms through which LXRs regulate inflammation, autoimmunity, and cancer, and the therapeutic potential for future treatment of these diseases through modulation of cholesterol metabolism

A 3D Human Posture Approach for Activity Recognition Based on Depth Camera

Human activity recognition plays an important role in the context of Ambient Assisted Living (AAL), providing useful tools to improve people quality of life. This work presents an activity recognition algorithm based on the extraction of skeleton joints from a depth camera. The system describes an activity using a set of few and basic postures extracted by means of the X-means clustering algorithm. A multi-class Support Vector Machine, trained with the Sequential Minimal Optimization is employed to perform the classification. The system is evaluated on two public datasets for activity recognition which have different skeleton models, the CAD-60 with 15 joints and the TST with 25 joints. The proposed approach achieves precision/recall performances of 99.8 % on CAD-60 and 97.2 %/91.7 % on TST. The results are promising for an applied use in the context of AAL

relA over-expression reduces tumorigenicity and activates apoptosis in human cancer cells

We previously demonstrated that bcl-2 over-expression increases the malignant behaviour of the MCF7 ADR human breast cancer cell line and enhances nuclear factor-kappa B (NF-k B) transcriptional activity. Here, we investigated the direct effect of increased NF-k B activity on the tumorigenicity of MCF7 ADR cells by over-expressing the NF-k B subunit relA/p65. Surprisingly, our results demonstrated that over-expression of relA determines a considerable reduction of the tumorigenic ability in nude mice as indicated by the tumour take and the median time of tumour appearance. In vitro studies also evidenced a reduced cell proliferation and the activation of the apoptotic programme after relA over-expression. Apoptosis was associated with the production of reactive oxygen species, and the cleavage of the specific substrate Poly-ADP-ribose-polymerrase. Our data indicate that there is no general role for NF-k B in the regulation of apoptosis and tumorigenicity. In fact, even though inhibiting NF-k B activity has been reported to be lethal to tumour cells, our findings clearly suggest that an over-induction of nuclear NF-k B activity may produce the same effect. © 2001 Cancer Research Campaign http://www.bjcancer.co

PPL-138 (BU10038):A bifunctional NOP/mu partial agonist that reduces cocaine self-administration in rats

The search for new and effective treatments for cocaine use disorder (CUD) is a priority. We determined whether PPL-138 (BU10038), a compound with partial agonist activity at both nociceptin opioid peptide (NOP) and mu-opioid receptors, reduces cocaine consumption, reinstatement, and whether the compound itself produces reinforcing effects in rats. Using an intermittent access (IntA) cocaine self-administration procedure, we found that PPL-138 (0.1 and 0.3 mg/kg) effectively decreased the total number of cocaine infusions and burst-like cocaine intake in both male and female rats. Responses for food in an IntA model of food self-administration were not altered for either sex, although locomotor activity was increased in female but not male rats. Blockade of NOP receptors with the selective antagonist J-113397 (5 mg/kg) did not prevent the PPL-138-induced suppression of cocaine self-administration, whereas blockade of mu-opioid receptors by naltrexone (1 mg/kg) reversed such effect. Consistently, treatment with morphine (1, 3, and 10 mg/kg) dose-dependently reduced IntA cocaine self-administration measures. PPL-138 also reduced reinstatement of cocaine seeking at all doses examined. Although an initial treatment with PPL-138 (2.5, 10, and 40 μg/kg/infusion) appeared rewarding, the compound did not maintain self-administration behavior. Animals treated with PPL-138 showed initial suppression of cocaine self-administration, which was eliminated following repeated daily dosing. However, suppression of cocaine self-administration was retained when subsequent PPL-138 treatments were administered 48 h apart. These findings demonstrate that the approach of combining partial NOP/mu-opioid activation successfully reduces cocaine use, but properties of PPL-138 seem to depend on the timing of drug administration.</p