Orexin-A/Hypocretin-1 Controls the VTA-NAc Mesolimbic Pathway via Endocannabinoid-Mediated Disinhibition of Dopaminergic Neurons in Obese Mice

Disinhibition of orexin-A/hypocretin-1 (OX-A) release occurs to several output areas of the lateral hypothalamus (LH) in the brain of leptin knockout obese ob/ob mice. In this study, we have investigated whether a similar increase of OX-A release occurs to the ventral tegmental area (VTA), an orexinergic LH output area with functional effects on dopaminergic signaling at the mesolimbic circuit. By confocal and correlative light and electron microscopy (CLEM) morphological studies coupled to molecular, biochemical, and pharmacological approaches, we investigated OX-A-mediated dopaminergic signaling at the LH-VTA-nucleus accumbens (NAc) pathway in obese ob/ob mice compared to wild-type (wt) lean littermates. We found an elevation of OX-A trafficking and release to the VTA of ob/ob mice and consequent orexin receptor-1 (OX1R)-mediated over-activation of dopaminergic (DA) neurons via phospholipase C (PLC)/diacylglycerol lipase (DAGL-α)-induced biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG). In fact, by retrograde signaling to cannabinoid receptor type 1 (CB1R) at inhibitory inputs to DA neurons, 2-AG inhibited GABA release thus inducing an increase in DA concentration in the VTA and NAc of ob/ob mice. This effect was prevented by the OX1R antagonist SB-334867 (30 mg/Kg, i.p.), or the CB1R antagonist AM251 (10 mg/Kg, i.p.) and mimicked by OX-A injection (40 μg/Kg, i.p.) in wt lean mice. Enhanced DA signaling to the NAc in ob/ob mice, or in OX-A-injected wt mice, was accompanied by β-arrestin2-mediated desensitization of dopamine D2 receptor (D2R) in a manner prevented by SB-334867 or the D2R antagonist L741 (1.5 mg/Kg, i.p.). These results further support the role of OX-A signaling in the control of neuroadaptive responses, such as compulsive reward-seeking behavior or binge-like consumption of high palatable food, and suggest that aberrant OX-A trafficking to the DA neurons in the VTA of ob/ob mice influences the D2R response at NAc, a main target area of the mesolimbic pathway, via 2-AG/CB1-mediated retrograde signaling

Mutant p53 improves cancer cells\u2019 resistance to endoplasmic reticulum stress by sustaining activation of the UPR regulator ATF6

Missense mutations in the TP53 gene are frequent in human cancers, giving rise to mutant p53 proteins that can acquire oncogenic properties. Gain of function mutant p53 proteins can enhance tumour aggressiveness by promoting cell invasion, metastasis and chemoresistance. Accumulating evidences indicate that mutant p53 proteins can also modulate cell homeostatic processes, suggesting that missense p53 mutation may increase resistance of tumour cells to intrinsic and extrinsic cancer-related stress conditions, thus offering a selective advantage. Here we provide evidence that mutant p53 proteins can modulate the Unfolded Protein Response (UPR) to increase cell survival upon Endoplasmic Reticulum (ER) stress, a condition to which cancer cells are exposed during tumour formation and progression, as well as during therapy. Mechanistically, this action of mutant p53 is due to enhanced activation of the pro-survival UPR effector ATF6, coordinated with inhibition of the pro-apoptotic UPR effectors JNK and CHOP. In a triple-negative breast cancer cell model with missense TP53 mutation, we found that ATF6 activity is necessary for viability and invasion phenotypes. Together, these findings suggest that ATF6 inhibitors might be combined with mutant p53-targeting drugs to specifically sensitise cancer cells to endogenous or chemotherapy-induced ER stress

Adjuvant vaginal interventional radiotherapy in early-stage non-endometrioid carcinoma of corpus uteri: a systematic review

Purpose: This systematic review focused on rare histological types of corpus uteri malignancy, including uterine carcinosarcoma (UCS), uterine clear cell carcinoma (UCCC), and uterine papillary serous carcinoma (UPSC), and it is proposed to assist with clinical decision-making. Adjuvant treatment decisions must be made based on available evidences. We mainly investigated the role of vaginal interventional radiotherapy (VIRt) in UCS, UCCC, and UPSC managements. Material and methods: A systematic research using PubMed and Cochrane library was conducted to identify full articles evaluating the efficacy of VIRt in early-stage UPSC, UCCC, and UCS. A search in ClinicalTrials.gov was performed in order to detect ongoing or recently completed trials as well as in PROSPERO for ongoing or recently completed systematic reviews. Survival outcomes and toxicity rates were obtained. Results: All studies were retrospective. For UCS, the number of evaluated patients was 432. The 2- to 5-year aver- age local control (LC) was 91% (range, 74.2-96%), disease-free survival (DFS) 88% (range, 82-94%), overall survival (OS) 79% (range, 53.8-84.3%), the average 5-year cancer-specific survival (CSS) was 70% (range, 70-94%), and G3-G4 toxicity was 0%. For UCCC, the number of investigated patients was 335 (UCCC – 124, mixed – 211), with an average 5-year LC of 100%, DFS of 83% (range, 82-90%), OS of 93% (range, 83-100%), and G3-G4 toxicity of 0%. For UPSC, the number of examined patients was 1,092 (UPSC – 866, mixed – 226). The average 5-year LC was 97% (range, 87.1-100%), DFS 84% (range, 74.7-95.6%), OS 93% (range, 71.9-100%), CSS 89% (range, 78.9-94%), and G3-G4 toxicity was 0%. Conclusions: These data suggest that in adequately selected early-stage UPSC and UCCC patients, VIRt alone may be suitable in women who underwent surgical staging and received adjuvant chemotherapy. In early-stage UCS, a multidisciplinary therapeutic approach has to be planned, considering high-rate of pelvic and distant relapses

Nonpsychotropic plant cannabinoids, cannabidivarin (CBDV) and cannabidiol (CBD), activate and desensitize transient receptor potential vanilloid 1 (TRPV1) channels in vitro: potential for the treatment of neuronal hyperexcitability

Epilepsy is the most common neurological disorder, with over 50 million people worldwide affected. Recent evidence suggests that the transient receptor potential cation channel subfamily V member 1 (TRPV1) may contribute to the onset and progression of some forms of epilepsy. Since the two nonpsychotropic cannabinoids cannabidivarin (CBDV) and cannabidiol (CBD) exert anticonvulsant activity in vivo and produce TRPV1-mediated intracellular calcium elevation in vitro, we evaluated the effects of these two compounds on TRPV1 channel activation and desensitization and in an in vitro model of epileptiform activity. Patch clamp analysis in transfected HEK293 cells demonstrated that CBD and CBDV dose-dependently activate and rapidly desensitize TRPV1, as well as TRP channels of subfamily V type 2 (TRPV2) and subfamily A type 1 (TRPA1). TRPV1 and TRPV2 transcripts were shown to be expressed in rat hippocampal tissue. When tested on epileptiform neuronal spike activity in hippocampal brain slices exposed to a Mg2+-free solution using multielectrode arrays (MEAs), CBDV reduced both epileptiform burst amplitude and duration. The prototypical TRPV1 agonist, capsaicin, produced similar, although not identical effects. Capsaicin, but not CBDV, effects on burst amplitude were reversed by IRTX, a selective TRPV1 antagonist. These data suggest that CBDV antiepileptiform effects in the Mg2+-free model are not uniquely mediated via activation of TRPV1. However, TRPV1 was strongly phosphorylated (and hence likely sensitized) in Mg2+-free solution-treated hippocampal tissue, and both capsaicin and CBDV caused TRPV1 dephosphorylation, consistent with TRPV1 desensitization. We propose that CBDV effects on TRP channels should be studied further in different in vitro and in vivo models of epilepsy

Activity of Antimicrobial Silver Polystyrene Nanocomposites

A simple technique based on doping polymers with in situ generated silver nanoparticles (Ag/PS films) has been developed. In particular, an antiseptic material has been prepared by dissolving silver 1,5-cyclooctadiene-hexafluoroacetylacetonate in amorphous polystyrene, and the obtained solid solution has been heated for ca. 10 s at a convenient temperature (180°C). Under such conditions the metal precursor decomposes producing silver atoms that diffuse into the polymer and clusterize. The antimicrobial characteristics of the resulting polystyrene-based material have been accurately evaluated toward Escherichia coli (E. coli) comparing the cytotoxicity effect of 10 wt.% and 30 wt.% (drastic and mild annealing) silver-doped polystyrene to the corresponding pure micrometric silver powder. Two different bacterial viability assays were performed in order to demonstrate the cytotoxic effect of Ag/PS films on cultured E. coli: (1) turbidimetric determination of optical density; (2) BacLight fluorescence-based test. Both methods have shown that silver-doped polystyrene (30 wt.%) provides higher antibacterial activity than pure Ag powder, under similar concentration and incubation conditions

Cannabinoid receptor type 1 and 2 expression in the skin of healthy dogs and dogs with atopic dermatitis.

OBJECTIVE: To determine the distribution of cannabinoid receptor type 1 (CB1) and cannabinoid receptor type 2 (CB2) in skin (including hair follicles and sweat and sebaceous glands) of clinically normal dogs and dogs with atopic dermatitis (AD) and to compare results with those for positive control samples for CB1 (hippocampus) and CB2 (lymph nodes). SAMPLE: Skin samples from 5 healthy dogs and 5 dogs with AD and popliteal lymph node and hippocampus samples from 5 cadavers of dogs. PROCEDURES: CB1 and CB2 were immunohistochemically localized in formalin-fixed, paraffin-embedded sections of tissue samples. RESULTS: In skin samples of healthy dogs, CB1 and CB2 immunoreactivity was detected in various types of cells in the epidermis and in cells in the dermis, including perivascular cells with mast cell morphology, fibroblasts, and endothelial cells. In skin samples of dogs with AD, CB1 and CB2 immunoreactivity was stronger than it was in skin samples of healthy dogs. In positive control tissue samples, CB1 immunoreactivity was detected in all areas of the hippocampus, and CB2 immunoreactivity was detected in B-cell zones of lymphoid follicles. CONCLUSIONS AND CLINICAL RELEVANCE: The endocannabinoid system and cannabimimetic compounds protect against effects of allergic inflammatory disorders in various species of mammals. Results of the present study contributed to knowledge of the endocannabinoid system and indicated this system may be a target for treatment of immune-mediated and inflammatory disorders such as allergic skin diseases in dogs

Role of 2-Arachidonoyl-Glycerol and CB1 Receptors in Orexin-A-Mediated Prevention of Oxygen-Glucose Deprivation-Induced Neuronal Injury

Orexin-A (OX-A) protects the brain against oxidative stress-mediated ischemic injury. Since the endocannabinoid 2-arachidonoylglycerol (2-AG) and cannabinoid type-1 (CB1) receptors were previously shown to mediate some of the effects of OX-A exerted through the orexin-1 receptor (OX-1R), we investigated the involvement of 2-AG in OX-A-induced neuroprotection following oxygen and glucose deprivation (OGD) in mouse cortical neurons. OGD-induced reactive oxygen species (ROS) accumulation and neuronal death were prevented by both OX-A and arachidonyl-2'-chloroethylamide (ACEA), a synthetic CB1 receptor agonist, in a manner sensitive to OX-1R and CB1 receptor antagonists, SB334867 and AM251. OX-A stimulated 2-AG biosynthesis in cortical neurons. In neurons isolated from monoacylglycerol lipase (MAGL, a 2-AG hydrolyzing enzyme) null mice, 10-fold higher 2-AG concentrations were found and OGD failed to induce ROS production and cell death, whereas AM251 restored these noxious effects. OX-A-induced neuroprotection was mediated by the phosphoinositide-3-kinase/Akt (PI3K/Akt) survival pathway since both OX-A and ACEA induced phosphorylation of Akt and prevented OGD-induced cytochrome c release from the mitochondria, in a manner counteracted by SB334867 or AM251. Administration of OX-A reduced infarct volume and elevated brain 2-AG levels in a mouse model of transient ischemia. These results suggest that 2-AG and CB1 receptor mediate OX-A prevention of ischemia-induced neuronal apoptosis