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De Filippis D.,University of Naples Federico II | De Filippis D.,Endocannabinoid Research Group | Luongo L.,The Second University of Naples | Luongo L.,Endocannabinoid Research Group | And 10 more authors.
Molecular Pain | Year: 2011

The aim of this study was to obtain evidences of a possible analgesic role for palmitoylethanolamide (PEA) in chronic granulomatous inflammation sustained by mast cell (MC) activation in rats at 96 hours. PEA (200-400-800 μg/mL), locally administered at time 0, reduced in a concentration-dependent manner the expression and release of NGF in comparison with saline-treated controls. PEA prevented nerve formation and sprouting, as shown by histological analysis, reduced mechanical allodynia, evaluated by Von Frey filaments, and inhibited dorsal root ganglia activation. These results were supported by the evidence that MCs in granuloma were mainly degranulated and closely localized near nerve fibres and PEA significantly reduced MC degranulation and nerves fibre formation. These findings are the first evidence that PEA, by the modulation of MC activation, controls pain perception in an animal model of chronic inflammation, suggesting its potential use for the treatment of all those painful conditions in which MC activation is an initial key step. © 2011 De Filippis et al; licensee BioMed Central Ltd.


Romano B.,University of Naples Federico II | Romano B.,Endocannabinoid Research Group | Romano B.,University of Aberdeen | Borrelli F.,University of Naples Federico II | And 7 more authors.
Phytomedicine | Year: 2014

Purpose Colon cancer is a major public health problem. Cannabis-based medicines are useful adjunctive treatments in cancer patients. Here, we have investigated the effect of a standardized Cannabis sativa extract with high content of cannabidiol (CBD), here named CBD BDS, i.e. CBD botanical drug substance, on colorectal cancer cell proliferation and in experimental models of colon cancer in vivo. Methods Proliferation was evaluated in colorectal carcinoma (DLD-1 and HCT116) as well as in healthy colonic cells using the MTT assay. CBD BDS binding was evaluated by its ability to displace [ 3H]CP55940 from human cannabinoid CB1 and CB2 receptors. In vivo, the effect of CBD BDS was examined on the preneoplastic lesions (aberrant crypt foci), polyps and tumours induced by the carcinogenic agent azoxymethane (AOM) as well as in a xenograft model of colon cancer in mice. Results CBD BDS and CBD reduced cell proliferation in tumoral, but not in healthy, cells. The effect of CBD BDS was counteracted by selective CB 1 and CB2 receptor antagonists. Pure CBD reduced cell proliferation in a CB1-sensitive antagonist manner only. In binding assays, CBD BDS showed greater affinity than pure CBD for both CB1 and CB2 receptors, with pure CBD having very little affinity. In vivo, CBD BDS reduced AOM-induced preneoplastic lesions and polyps as well as tumour growth in the xenograft model of colon cancer. Conclusions CBD BDS attenuates colon carcinogenesis and inhibits colorectal cancer cell proliferation via CB1 and CB2 receptor activation. The results may have some clinical relevance for the use of Cannabis-based medicines in cancer patients. © 2013 Elsevier GmbH.


Aviello G.,University of Naples Federico II | Aviello G.,Endocannabinoid Research Group | Borrelli F.,University of Naples Federico II | Borrelli F.,Endocannabinoid Research Group | And 13 more authors.
Journal of Molecular Medicine | Year: 2011

The hallucinogenic compound, salvinorin A, is a potent κ-opioid receptor (KOR) agonist. However, other target(s) than the KOR, such as the cannabinoid CB1 receptor, have been proposed to explain its multiple pharmacological actions. Here, we have evaluated the effect of salvinorin A in lipopolysaccharide (LPS)-stimulated macrophages as well as in models of inflammation in vivo. Salvinorin A (0.1-10 pM) reduced LPS-stimulated nitrite, TNF-α and IL-10 (but not IL-1β) levels as well as iNOS (but not COX-2) LPS-induced hyperexpression. The effect of salvinorin A on nitrite levels was reverted by the opioid antagonist naloxone, the KOR antagonist nor-binaltorphimine and by the CB1 antagonist rimonabant Salvinorin A also prevented KOR and CB1 hyperexpression induced by LPS. In vivo, salvinorin A reduced the LPS- and the carrageenan-induced paw oedema and formalin-induced inflammatory pain, in a nor-binaltorphimine and rimonabant-sensitive manner. It is concluded that salvinorin A-via KORs and CB1 receptors-exerts ultrapotent actions on macrophages and also shows moderate antinflammatory effects in vivo. © 2011 Springer-Verlag.


Marczylo E.L.,Systems Toxicology Group | Amoako A.A.,Endocannabinoid Research Group | Konje J.C.,Endocannabinoid Research Group | Gant T.W.,Systems Toxicology Group | And 2 more authors.
Epigenetics | Year: 2012

Recent work has suggested that environmental chemicals, including those contained in cigarette smoke, can have adverse effects on the exposed individuals as well as their future progeny. The mechanisms underlying transmission of environmentally-induced phenotypes through the germ line are not well understood. However, a predominant process appears to be the establishment of permanent heritable epigenetic alterations, and a number of studies have implicated microRNAs in such processes. Here, we show that cigarette smoke induces specific differences in the spermatozoal microRNA content of human smokers compared with non-smokers, and that these altered microRNAs appear to predominantly mediate pathways vital for healthy sperm and normal embryo development, particularly cell death and apoptosis. microRNA-mediated perturbation of such pathways may explain how harmful phenotypes can be induced in the progeny of smokers. © 2012 Landes Bioscience.


El-Talatini M.R.,Endocannabinoid Research Group | Taylor A.H.,Endocannabinoid Research Group | Taylor A.H.,Preterm Birth Research Group | Konje J.C.,Endocannabinoid Research Group
Fertility and Sterility | Year: 2010

Objective: To further investigate the relationship between plasma anandamide (AEA), sex steroids, and gonadotrophins to improve our understanding of how AEA may be involved in human fertility. Design: Cross-sectional and longitudinal study. Setting: University Hospital of Leicester NHS Trust, Leicester Royal Infirmary. Patient(s): Healthy premenopausal and postmenopausal volunteers. Intervention(s): UPLC-MS/MS-measured plasma AEA and ELISA-measured serum FSH, LH, estradiol, and progesterone levels at five different phases of the menstrual cycle and postmenopause. Main Outcome Measure(s): Plasma AEA, serum steroids and gonadotrophins. Result(s): Changes in AEA levels were similar in the two cohorts. The mean ± SEM levels in the early follicular phase (0.89 ± 0.06) for the cross-sectional cohort and the longitudinal cohort (0.73 ± 0.03) were higher than those in the late follicular phase (0.77 ± 0.09 cross-sectional; 0.63 ± 0.08 longitudinal). The highest AEA levels were measured at ovulation (1.38 ± 0.14 and 1.33 ± 0.16) and the lowest level was measured in the late luteal phase (0.66 ± 0.07 and 0.56 ± 0.06). There was a statistically significant positive correlation between AEA, estradiol (P=0.0015), LH (P<0.0001) and FSH levels but not progesterone (P=0.022). Conclusion(s): Peak plasma AEA occurred at ovulation and positively correlated with estradiol and gonadotrophin levels suggesting that these may be involved in the regulation of AEA levels. © 2010.


Cipriano M.,University of Naples Federico II | Cipriano M.,Umeå University | Esposito G.,University of Rome La Sapienza | Negro L.,University of Naples Federico II | And 7 more authors.
CNS and Neurological Disorders - Drug Targets | Year: 2015

Aβ-induced astrogliosis can worsen the eziopathogenesis of Alzheimer disease (AD) by the release of proinflammatory and pro-oxidant mediators. Activated glial cells may release also pro-angiogenic molecules. The role of angiogenesis in AD is still controversial: although angiogenesis brings oxygen and nutrients to injured tissue, it may also exacerbate reactive gliosis. Moreover, by altering blood-brain barrier permeability pro-angiogenic mediators promote passage of inflammatory/immune-competent cells into the brain, thereby exacerbating gliosis. The release of proangiogenic factors during astrogliosis may thus be a key-step in controlling AD progression. The endogenous fatty acid amide, palmitoylethanolamide (PEA), is a pleiotropic mediator exerting anti-inflammatory, antinociceptive and antiangiogenic effects in several in vitro and in vivo models of chronic-degenerative disease. In this study, we investigated the effects of PEA in AD angiogenesis and neuroinflammation by using conditioned medium from untreated and Aβ-treated C6 rat astroglioma cells and HUVEC human endothelial cells. PEA (10-8-10-6 M) concentration-dependently reduced expression of pro-inflammatory and pro-angiogenic markers in Aβ (1 μg/mL)-stimulated C6 cells. Moreover, culture medium from PEA-treated C6 cells reduced HUVEC cell proliferation as compared to cells treated with conditioned medium from Aβ-treated C6 cells. Immunocytochemical analysis revealed that PEA treatment inhibited nuclear levels of mitogen-activated protein kinase 1 (the main pro-angiogenic pathway) and cytoplasmic vascular endothelial growth factor in HUVEC cells receiving C6 conditioned medium. Finally, the peroxisome proliferator-activated receptor alpha inhibitor GW6471, added to Aβ-treated C6 cells blocked all PEA effects in this model, suggesting that PEA acts through a proliferator-activated receptor alpha-dependent mechanism on astroglial cells. Collectively, these data support the potential therapeutic utility of PEA in AD. © 2015 Bentham Science Publishers.


Pessina F.,University of Siena | Capasso R.,University of Naples Federico II | Capasso R.,National Research Council Italy | Capasso R.,Endocannabinoid Research Group | And 16 more authors.
Journal of Urology | Year: 2015

Purpose PEA is an endogenous mediator released together with the endocannabinoid anandamide from membrane phospholipids. It is a plant derived compound with analgesic and anti-inflammatory properties. We verified whether the pathophysiology of experimental cystitis involves changes in the levels of PEA and of some of its targets, ie CB1 and CB2 receptors, and PPARα. We also determined whether exogenously administered PEA could be proposed as a preventive measure for cystitis. Materials and Methods Cystitis was induced by cyclophosphamide in female rats. Nociceptive responses, voiding episodes, gross damage, myeloperoxidase activity, bladder weight, bladder PEA and endocannabinoid levels (measured by liquid chromatography-mass spectrometry) and the expression of PEA targets (measured by quantitative reverse transcriptase-polymerase chain reaction) were recorded. Results Cyclophosphamide induced pain behavior, bladder inflammation and voiding dysfunction associated with increased bladder levels of PEA, up-regulation of CB1 receptor mRNA expression, down-regulation of PPARα mRNA and no change in CB2 receptor mRNA expression. Exogenously administered, ultramicronized PEA attenuated pain behavior, voids and bladder gross damage. The CB1 antagonist rimonabant and the PPARα antagonist GW6471 counteracted the beneficial effect of PEA on gross damage. Also, GW6471 further decreased voiding episodes in rats treated with PEA. Conclusions The current study provides strong evidence for a protective role of PEA as well as an alteration in bladder levels of PEA and of some of its targets in cyclophosphamide induced cystitis. © 2015 American Urological Association Education and Research, Inc.


Amoako A.A.,Endocannabinoid Research Group | Amoako A.A.,Leeds Teaching Hospitals NHS Trust | Marczylo T.H.,Public Health England | Elson J.,The London Clinic | And 3 more authors.
Fertility and Sterility | Year: 2014

Objective: To determine whether changes in seminal plasma concentrations of the endogenous lipid signaling molecules palmitoylethanolamide (PEA) and oleoylethanolamide (OEA) have significant effects on sperm quality. Design: Biochemical and physiological studies of human seminal plasma and spermatozoa. Setting: Academic tertiary care medical center. Patient(s): Ninety men attending an infertility clinic for semen analysis. Intervention(s): Palmitoylethanolamide and OEA extracted from seminal plasma were quantified by ultra high-performance liquid chromatography (HPLC)-tandem mass spectrometry. Patient sperm from semen with normal parameters were exposed in vitro to PEA or OEA to determine effects on sperm motility, viability, and mitochondrial activity. Main Outcome Measure(s): The relationship between seminal plasma concentrations of PEA and OEA and sperm quality and the effect of these compounds on sperm motility, viability, and mitochondria activity in vitro. Result(s): Palmitoylethanolamide and OEA concentrations in seminal plasma were lower in men with asthenozoospermia and oligoasthenoteratozospermia compared with men with normal semen parameters. Palmitoylethanolamide and OEA rapidly and significantly improved sperm motility and maintained viability without affecting mitochondria activity in vitro. Conclusion(s): Maintenance of normal PEA and OEA tone in human seminal plasma may be necessary for the preservation of normal sperm function and male fertility. Exocannabinoids found in Cannabis, such as delta-9-tetrahydrocannabinol and cannabidiol, could compete with these endocannabinoids upsetting their finely balanced, normal functioning and resulting in male reproductive failure. ©2014 by American Society for Reproductive Medicine.


Janssen F.J.,Leiden University | Deng H.,Leiden University | Baggelaar M.P.,Leiden University | Allara M.,Endocannabinoid Research Group | And 8 more authors.
Journal of Medicinal Chemistry | Year: 2014

sn-1-Diacylglycerol lipase α (DAGL-α) is the main enzyme responsible for the production of the endocannabinoid 2-arachidonoylglycerol in the central nervous system. Glycine sulfonamides have recently been identified by a high throughput screening campaign as a novel class of inhibitors for this enzyme. Here, we report on the first structure-activity relationship study of glycine sulfonamide inhibitors and their brain membrane proteome-wide selectivity on serine hydrolases with activity-based protein profiling (ABPP). We found that (i) DAGL-α tolerates a variety of biaryl substituents, (ii) the sulfonamide is required for inducing a specific orientation of the 2,2-dimethylchroman substituent, and (iii) a carboxylic acid is essential for its activity. ABPP revealed that the sulfonamide glycine inhibitors have at least three off-targets, including α/β-hydrolase domain 6 (ABHD6). Finally, we identified LEI-106 as a potent, dual DAGL-α/ABHD6 inhibitor, which makes this compound a potential lead for the discovery of new molecular therapies for diet-induced obesity and metabolic syndrome. © 2014 American Chemical Society.


de Filippis D.,University of Naples Federico II | de Filippis D.,Endocannabinoid Research Group | Esposito G.,University of Rome La Sapienza | Cirillo C.,University of Naples Federico II | And 10 more authors.
PLoS ONE | Year: 2011

Enteric glial cells (EGC) actively mediate acute and chronic inflammation in the gut; EGC proliferate and release neurotrophins, growth factors, and pro-inflammatory cytokines which, in turn, may amplify the immune response, representing a very important link between the nervous and immune systems in the intestine. Cannabidiol (CBD) is an interesting compound because of its ability to control reactive gliosis in the CNS, without any unwanted psychotropic effects. Therefore the rationale of our study was to investigate the effect of CBD on intestinal biopsies from patients with ulcerative colitis (UC) and from intestinal segments of mice with LPS-induced intestinal inflammation. CBD markedly counteracted reactive enteric gliosis in LPS-mice trough the massive reduction of astroglial signalling neurotrophin S100B. Histological, biochemical and immunohistochemical data demonstrated that S100B decrease was associated with a considerable decrease in mast cell and macrophages in the intestine of LPS-treated mice after CBD treatment. Moreover the treatment of LPS-mice with CBD reduced TNF-α expression and the presence of cleaved caspase-3. Similar results were obtained in ex vivo cultured human derived colonic biopsies. In biopsies of UC patients, both during active inflammation and in remission stimulated with LPS+INF-γ, an increased glial cell activation and intestinal damage were evidenced. CBD reduced the expression of S100B and iNOS proteins in the human biopsies confirming its well documented effect in septic mice. The activity of CBD is, at least partly, mediated via the selective PPAR-gamma receptor pathway. CBD targets enteric reactive gliosis, counteracts the inflammatory environment induced by LPS in mice and in human colonic cultures derived from UC patients. These actions lead to a reduction of intestinal damage mediated by PPARgamma receptor pathway. Our results therefore indicate that CBD indeed unravels a new therapeutic strategy to treat inflammatory bowel diseases. © 2011 De Filippis et al.

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