Human gasdermin D and MLKL disrupt mitochondria, endocytic traffic and TORC1 signalling in budding yeast.

Gasdermin D (GSDMD) and mixed lineage kinase domain-like protein (MLKL) are the pore-forming effectors of pyroptosis and necroptosis, respectively, with the capacity to disturb plasma membrane selective permeability and induce regulated cell death. The budding yeast has long been used as a simple eukaryotic model for the study of proteins associated with human diseases by heterologous expression. In this work, we expressed in yeast both GSDMD and its N-terminal domain (GSDMD(NT)) to characterize their cellular effects and compare them to those of MLKL.

Circulating Tumor DNA as a Marker of Minimal Residual Disease After Radical Resection of Colorectal Liver Metastases.

Purpose: Prognostic tools to estimate the risk of relapse for patients with liver-limited metastatic colorectal cancer (LL-mCRC) undergoing resection with curative intent are needed. Circulating tumor DNA (ctDNA) as a surrogate of postsurgical minimal residual disease is a promising marker in localized CRC. We explored the role of postoperative ctDNA as a marker of minimal residual disease in patients with radically resected LL-mCRC.

Neuroserpin Inclusion Bodies in a FENIB Yeast Model.

FENIB (familial encephalopathy with neuroserpin inclusion bodies) is a human monogenic disease caused by point mutations in the gene, characterized by the intracellular deposition of polymers of neuroserpin (NS), which leads to proteotoxicity and cell death. Despite the different cell and animal models developed thus far, the exact mechanism of cell toxicity elicited by NS polymers remains unclear. Here, we report that human wild-type NS and the polymerogenic variant G392E NS form protein aggregates mainly localized within the endoplasmic reticulum (ER) when expressed in the yeast .

Heterologous Expression and Auto-Activation of Human Pro-Inflammatory Caspase-1 in and Comparison to Caspase-8.

Caspases are a family of cysteine proteases that play an essential role in inflammation, apoptosis, cell death, and development. Here we delve into the effects caused by heterologous expression of human caspase-1 in the yeast and compare them to those of caspase-8. Overexpression of both caspases in the heterologous model led to their activation and caused mitochondrial hyperpolarization, damage to different organelles, and cell death.

Modeling human disease in yeast: recreating the PI3K-PTEN-Akt signaling pathway in Saccharomyces cerevisiae.

The yeast Saccharomyces cerevisiae is a model organism that has been thoroughly exploited to understand the universal mechanisms that govern signaling pathways. Due to its ease of manipulation, humanized yeast models that successfully reproduce the function of human genes permit the development of highly efficient genetic approaches for molecular studies. Of special interest are those pathways related to human disease that are conserved from yeast to mammals.

Gaussian mixture models based 2D-3D registration of bone shapes for orthopedic surgery planning.

In orthopedic surgery, precise kinematics assessment helps the diagnosis and the planning of the intervention. The correct placement of the prosthetic component in the case of knee replacement is necessary to ensure a correct load distribution and to avoid revision of the implant. 3D reconstruction of the knee kinematics under weight-bearing conditions becomes fundamental to understand existing in vivo loads and improve the joint motion tracking.

Cannabidiol, a non-psychoactive cannabinoid compound, inhibits proliferation and invasion in U87-MG and T98G glioma cells through a multitarget effect.

In the present study, we found that CBD inhibited U87-MG and T98G cell proliferation and invasiveness in vitro and caused a decrease in the expression of a set of proteins specifically involved in growth, invasion and angiogenesis. In addition, CBD treatment caused a dose-related down-regulation of ERK and Akt prosurvival signaling pathways in U87-MG and T98G cells and decreased hypoxia inducible factor HIF-1α expression in U87-MG cells. Taken together, these results provide new insights into the antitumor action of CBD, showing that this cannabinoid affects multiple tumoral features and molecular pathways.

Molecular mechanisms involved in the antitumor activity of cannabinoids on gliomas: role for oxidative stress.

Cannabinoids, the active components of Cannabis sativa, have been shown to exert antiproliferative and proapoptotic effects on a wide spectrum of tumor cells and tissues. Of interest, cannabinoids have displayed great potency in reducing the growth of glioma tumors, one of the most aggressive CNS tumors, either in vitro or in animal experimental models curbing the growth of xenografts generated by subcutaneous or intrathecal injection of glioma cells in immune-deficient mice. Cannabinoids appear to be selective antitumoral agents as they kill glioma cells without affecting the viability of non-transformed cells.

Expression and function of the endocannabinoid system in glial cells.

In the last few years the role and significance of the glia in CNS function and pathology have been drastically reassessed. Glial cells physiology appears very different in healthy versus pathological brain and the recent identification of cannabinoid receptors and their endogenous ligands in glia has triggered a number of studies exploring the role of (endo)cannabinoid system in glia functionality and disease. (Endo)cannabinoids exert their effects in these cells directly affecting some important peculiar functions of the glia and actively promoting biochemical signals ending in a pro-survival fate for these cells.

Functional lipidomics. Calcium-independent activation of endocannabinoid/endovanilloid lipid signalling in sensory neurons by protein kinases C and A and thrombin.

N-arachidonoylethanolamine (anandamide, AEA), is a full agonist at both cannabinoid CB(1) receptors and "transient receptor potential vanilloid" type 1 (TRPV1) channels, and N-palmitoylethanolamine (PEA) potentiates these effects. In neurons of the rat dorsal root ganglia (DRG), TRPV1 is activated and/or sensitised by AEA as well as upon activation of protein kinases C (PKC) and A (PKA). We investigated here the effect on AEA levels of PKC and PKA activators in DRG neurons.

Identification of endocannabinoids and related compounds in human fat cells.

Objective: Recently, an activation of the endocannabinoid system during obesity has been reported. More particularly, it has been demonstrated that hypothalamic levels of both endocannabinoids, 2-arachidonoylglycerol and anandamide (N-arachidonoylethanolamine), are up-regulated in genetically obese rodents. Circulating levels of both endocannabinoids were also shown to be higher in obese compared with lean women.

Overactivity of the intestinal endocannabinoid system in celiac disease and in methotrexate-treated rats.

The endocannabinoid system is upregulated in both human inflammatory bowel diseases and experimental models of colitis. In this study, we investigated whether this upregulation is a marker also of celiac disease-induced atrophy. The levels of the cannabinoid CB(1) receptor, of the endocannabinoids, anandamide, and 2-arachidonoyl-glycerol (2-AG), and of the anti-inflammatory mediator palmitoylethanolamide (PEA) were analyzed in bioptic samples from the duodenal mucosa of celiac patients at first diagnosis assessed by the determination of antiendomysial antibodies and histological examination.

Peripheral blood dendritic cells and monocytes are differently regulated in the elderly.

Dendritic cells (DCs) are the single most central player in all immune responses. To assess whether DC alterations may contribute to the immune dysregulation that affects the elderly, we investigated the effects of ageing on DCs. We analyzed the number, phenotype and function of peripheral blood DCs from 70 healthy subjects aged 20-92 years by using flow cytometric methods that allow cell characterization directly in whole blood samples.

Up-regulation of anandamide levels as an endogenous mechanism and a pharmacological strategy to limit colon inflammation.

Direct stimulation of cannabinoid CB1 receptors exerts a protective function in animal models of inflammatory bowel diseases (IBDs). However, it is not known whether endocannabinoids are up-regulated during IBDs in animals or humans, nor whether pharmacological elevation of endocannabinoid levels can be exploited therapeutically in these disorders. In this study we addressed these questions.

Elevation of endocannabinoid levels in the ventrolateral periaqueductal grey through inhibition of fatty acid amide hydrolase affects descending nociceptive pathways via both cannabinoid receptor type 1 and transient receptor potential vanilloid type-1 receptors.

In the ventrolateral periaqueductal gray (PAG), activation of excitatory output neurons projecting monosynaptically to OFF cells in the rostral ventromedial medulla (RVM) causes antinociceptive responses and is under the control of cannabinoid receptor type-1 (CB1) and vanilloid transient receptor potential vanilloid type 1 (TRPV1) receptors. We studied in healthy rats the effect of elevation of PAG endocannabinoid [anandamide and 2-arachidonoylglycerol (2-AG)] levels produced by intra-PAG injections of the inhibitor of fatty acid amide hydrolase URB597 [cyclohexylcarbamic acid-3'-carbamoyl-biphenyl-3-yl ester] on 1) nociception in the "plantar test" and 2) spontaneous and tail-flick-related activities of RVM neurons. Depending on the dose or time elapsed since administration, URB597 (0.

Decreased endocannabinoid levels in the brain and beneficial effects of agents activating cannabinoid and/or vanilloid receptors in a rat model of multiple sclerosis.

Recent studies have addressed the changes in endocannabinoid ligands and receptors that occur in multiple sclerosis, as a way to explain the efficacy of cannabinoid compounds to alleviate spasticity, pain, tremor, and other signs of this autoimmune disease. Using Lewis rats with experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, we recently found a decrease in cannabinoid CB1 receptors mainly circumscribed to the basal ganglia, which could be related to the motor disturbances characteristic of these rats. In the present study, using the same model, we explored the potential changes in several neurotransmitters in the basal ganglia that might be associated with the motor disturbances described in these rats, but we only found a small increase in glutamate contents in the globus pallidus.

Fatty acid amide hydrolase controls mouse intestinal motility in vivo.

Background & Aims: Fatty acid amide hydrolase (FAAH) catalyzes the hydrolysis both of the endocannabinoids (which are known to inhibit intestinal motility) and other bioactive amides (palmitoylethanolamide, oleamide, and oleoylethanolamide), which might affect intestinal motility. The physiologic role of FAAH in the gut is largely unexplored. In the present study, we evaluated the possible role of FAAH in regulating intestinal motility in mice in vivo.

Effect of repeated systemic administration of selective inhibitors of endocannabinoid inactivation on rat brain endocannabinoid levels.

Several selective inhibitors of endocannabinoid inactivation via either the fatty acid amide hydrolase (FAAH) or the putative endocannabinoid transporter have been developed so far. Here, we have studied the effect in rats of a subchronic intraperitoneal treatment with three recently developed selective inhibitors of endocannabinoid uptake (VDM-11, UCM-707 and OMDM-2) or with a selective FAAH inhibitor (N-arachidonoyl-serotonin, AA-5-HT), on the brain levels of anandamide and 2-arachidonoylglycerol (2-AG) measured by means of isotope dilution LC-MS 1, 5 and 12 h after the last treatment. OMDM-2 was the most efficacious compound at enhancing the levels of anandamide at all time points, with a maximal effect (1.

Cisplatin increases brain 2-arachidonoylglycerol (2-AG) and concomitantly reduces intestinal 2-AG and anandamide levels in the least shrew.

The chemotherapeutic agent cisplatin may produce emesis via release of several neurotransmitters such as serotonin (5-HT), substance P and/or dopamine as well as production of prostaglandins (PGs). Administration of synthetic 2-arachidonoylglycerol (2-AG) but not of anandamide, which are two putative endocannabinoids, causes vomiting via its downstream metabolites such as arachidonic acid (AA) and PGs in the least shrew (Cryptotis parva). We report here that cisplatin (0, 5, 10 and 20 mg/kg, i.

Involvement of the cannabimimetic compound, N-palmitoyl-ethanolamine, in inflammatory and neuropathic conditions: review of the available pre-clinical data, and first human studies.

The endogenous cannabimimetic compound, and anandamide analogue, N-palmitoyl-ethanolamine (PEA), was shown to exert potent anti-inflammatory and analgesic effects in experimental models of visceral, neuropathic and inflammatory pain by acting via several possible mechanisms. However, only scant data have been reported on the regulation of PEA levels during pathological conditions in animals or, particularly, humans. We review the current literature on PEA and report the results of three separate studies indicating that its concentrations are significantly increased during three different inflammatory and neuropathic conditions, two of which have been assessed in humans, and one in a mouse model.

Pharmacological modulation of the endocannabinoid system in a viral model of multiple sclerosis.

Theiler's virus infection of the central nervous system (CNS) induces an immune-mediated demyelinating disease in susceptible mouse strains and serves as a relevant infection model for human multiple sclerosis (MS). Cannabinoids have been shown to exert beneficial effects on animal models of MS and evidence suggests that the endocannabinoid system plays a role in the tonic control of spasticity. In this study we show that OMDM1 [(R)-N-oleoyl-(1'-hydroxybenzyl)-2'-ethanolamine] and OMDM2 [(S)-N-oleoyl-(1'-hydroxybenzyl)-2'-ethanolamine], two selective inhibitors of the putative endocannabinoid transporter and hence of endocannabinoid inactivation, provide an effective therapy for Theiler murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD).

Lipopolysaccharide-induced pulmonary inflammation is not accompanied by a release of anandamide into the lavage fluid or a down-regulation of the activity of fatty acid amide hydrolase.

The effect of lipopolysaccharide inhalation upon lung anandamide levels, anandamide synthetic enzymes and fatty acid amide hydrolase has been investigated. Lipopolysaccharide exposure produced a dramatic extravasation of neutrophils and release of tumour necrosis factor alpha into the bronchoalveolar lavage (BAL) fluid, which was not accompanied by epithelial cell injury. The treatment, however, did not change significantly the levels of anandamide and the related compound palmitoylethanolamide in the cell-free fraction of the BAL fluid.

Endocannabinoids link feeding state and auditory perception-related gene expression.

Singing by adult male zebra finches is a learned behavior important for courtship, kin recognition, and nest defense (Zann, 1996) and is inhibited by both brief periods of limited food availability and systemic injection of cannabinoids. These similar effects on singing, combined with increasing evidence for endocannabinoid involvement in feeding behavior, led us to evaluate a possible shared mechanism. We found that limited food availability both reduces singing in a cannabinoid antagonist-reversible manner and increases levels of the endocannabinoid 2-arachidonyl glycerol in various brain regions including the caudal telencephalon, an area that contains auditory telencephalon including the L2 subfield of L (L2) and caudal medial nidopallium (NCM).