Publications

E.M. Rock, S.M. Ayoub, C.L. Limebeer, A. Gene, K.L. Wills, M.V. DeVuono, R. Smoum, V. Di Marzo, A.H. Lichtman, R. Mechoulam, and L.A. Parker. 2020. “Acute naloxone-precipitated morphine withdrawal elicits nausea-like somatic behaviors in rats in a manner suppressed by N-oleoylglycine.” Psychopharmacology, 237, 2, Pp. 375-384. Publisher's Version Abstract
Rationale: Acute naloxone-precipitated morphine withdrawal (MWD) produces a conditioned place aversion (CPA) in rats even after one or two exposures to high-dose (20 mg/kg, sc) morphine followed 24-h later by naloxone (1 mg/kg, sc). However, the somatic withdrawal reactions produced by acute naloxone-precipitated MWD in rats have not been investigated. A recently discovered fatty acid amide, N-oleoylglycine (OlGly), which has been suggested to act as a fatty acid amide hydrolase (FAAH) inhibitor and as a peroxisome proliferator-activated receptor alpha (PPARα) agonist, was previously shown to interfere with a naloxone-precipitated MWD-induced CPA in rats. Objectives: The aims of these studies were to examine the somatic withdrawal responses produced by acute naloxone-precipitated MWD and determine whether OlGly can also interfere with these responses. Results: Here, we report that following two exposures to morphine (20 mg/kg, sc) each followed by naloxone (1 mg/kg, sc) 24 h later, rats display nausea-like somatic reactions of lying flattened on belly, abdominal contractions and diarrhea, and display increased mouthing movements and loss of body weight. OlGly (5 mg/kg, ip) interfered with naloxone-precipitated MWD-induced abdominal contractions, lying on belly, diarrhea and mouthing movements in male Sprague–Dawley rats, by both a cannabinoid 1 (CB1) and a PPARα mechanism of action. Since these withdrawal reactions are symptomatic of nausea, we evaluated the potential of OlGly to interfere with lithium chloride (LiCl)-induced and MWD-induced conditioned gaping in rats, a selective measure of nausea; the suppression of MWD-induced gaping reactions by OlGly was both CB1 and PPARα mediated. Conclusion: These results suggest that the aversive effects of acute naloxone-precipitated MWD reflect nausea, which is suppressed by OlGly. © 2019, Springer-Verlag GmbH Germany, part of Springer Nature.
S.M. Ayoub, R. Smoum, M. Farag, H. Atwal, S.A. Collins, E.M. Rock, C.L. Limebeer, F. Piscitelli, F.A. Iannotti, A.H. Lichtman, F. Leri, V. Di Marzo, R. Mechoulam, and L.A. Parker. 2020. “Oleoyl alanine (HU595): a stable monomethylated oleoyl glycine interferes with acute naloxone precipitated morphine withdrawal in male rats.” Psychopharmacology, 237, 9, Pp. 2753-2765. Publisher's Version Abstract
Rationale: Oleoyl glycine, a little studied fatty acid amide similar in structure to anandamide, interferes with nicotine addiction in mice and acute naloxone-precipitated morphine withdrawal (MWD) in rats. Because endogenous oleoyl glycine is subject to rapid enzymatic deactivation, we evaluated the potential of more stable analogs to interfere with opiate withdrawal. Objectives: The potential of monomethylated oleoyl glycine (oleoyl alanine, HU595) to interfere with somatic and aversive effects of acute naloxone-precipitated MWD, its duration, and mechanism of action was assessed in male Sprague Dawley rats. The potential of dimethylated oleoyl glycine (HU596) to interfere with the aversive effects of naloxone-precipitated MWD was also investigated. Results: Oleoyl alanine (HU595) interfered with somatic and aversive effects produced by naloxone-precipitated MWD at equivalent doses (1 and 5 mg/kg, i.p.) as we have reported for oleoyl glycine; however, oleoyl alanine produced a longer lasting (60 min) interference, yet did not produce rewarding or aversive effects on its own and did not modify locomotor activity. HU596 was not effective. The interference with aversive effects of naloxone-precipitated MWD by oleoyl alanine was prevented by both a PPARα antagonist and a CB1 receptor antagonist. Accordingly, the compound was found to inhibit FAAH and activate PPARα in vitro. Finally, oleoyl alanine also reduced acute naloxone-precipitated MWD anhedonia, as measured by decreased saccharin preference. Conclusions: Oleoyl alanine (also an endogenous fatty acid) may be a more stable and effective treatment for opiate withdrawal than oleoyl glycine. © 2020, Springer-Verlag GmbH Germany, part of Springer Nature.
F. Piscitelli, F. Guida, L. Luongo, F.A. Iannotti, S. Boccella, R. Verde, A. Lauritano, R. Imperatore, R. Smoum, L. Cristino, A.H. Lichtman, L.A. Parker, R. Mechoulam, S. Maione, and V. Di Marzo. 2020. “Protective Effects of N-Oleoylglycine in a Mouse Model of Mild Traumatic Brain Injury.” ACS Chemical Neuroscience, 11, 8, Pp. 1117-1128. Publisher's Version Abstract
Traumatic brain injury (TBI) is one of the main causes of death in young people for which currently no efficacious treatment exists. Recently, we have reported that mice with mild-TBI with a specific injury in the insula showed elevated levels of a little investigated N-acyl amino acid, N-oleoylglycine (OlGly). N-acyl amino acids have recently experienced an increased interest because of their important biological activities. They belong to the endocannabinoidome family of lipids with structural similarities with the endocannabinoids (eCBs). The aim of this study was to test the neuroprotective and antihyperalgesic actions of OlGly in a model of mouse mild-TBI (mTBI) and its effect on levels of eCBs and N-acylethanolamines at the end of treatment. Following mTBI, mice were administered a daily injection of OlGly (10-50-100 mg/kg i.p.) for 14 days. Treatment with OlGly normalized motor impairment and behavior in the light/dark box test, ameliorated TBI-induced thermal hyperalgesia and mechanical allodynia, and normalized aggressiveness and depression. Moreover, levels of eCBs and some N-acylethanolamines underwent significant changes 60 days after TBI, especially in the prefrontal cortex and hypothalamus, and OlGly reversed some of these changes. In conclusion, our findings reveal that OlGly ameliorates the behavioral alterations associated with mTBI in mice, while concomitantly modulating eCB and eCB-like mediator tone. © 2020 American Chemical Society.
E.M. Rock, M.T. Sullivan, S.A. Collins, H. Goodman, C.L. Limebeer, R. Mechoulam, and L.A. Parker. 2020. “Evaluation of repeated or acute treatment with cannabidiol (CBD), cannabidiolic acid (CBDA) or CBDA methyl ester (HU-580) on nausea and/or vomiting in rats and shrews.” Psychopharmacology, 237, 9, Pp. 2621-2631. Publisher's Version Abstract
Rationale: When acutely administered intraperitoneally, the non-psychoactive cannabinoid cannabidiol (CBD), its acidic precursor cannabidiolic acid (CBDA) and a stable methyl ester of CBDA (HU-580) reduce lithium chloride (LiCl)–induced conditioned gaping in male rats (a selective preclinical model of acute nausea) via activation of the serotonin 1A (5-HT1A) receptor. Objectives: To utilise these compounds to manage nausea in the clinic, we must determine if their effectiveness is maintained when injected subcutaneously (s.c) and when repeatedly administered. First, we compared the effectiveness of each of these compounds to reduce conditioned gaping following repeated (7-day) and acute (1-day) pretreatments and whether these anti-nausea effects were mediated by the 5-HT1A receptor. Next, we assessed whether the effectiveness of these compounds can be maintained when administered prior to each of 4 conditioning trials (once per week). We also evaluated the ability of repeated CBD (7 days) to reduce LiCl-induced vomiting in Suncus murinus. Finally, we examined whether acute CBD was equally effective in male and female rats. Results: Both acute and repeated (7 day) s.c. administrations of CBD (5 mg/kg), CBDA (1 μg/kg) and HU-580 (1 μg/kg) similarly reduced LiCl-induced conditioned gaping, and these effects were blocked by 5HT1A receptor antagonism. When administered over 4 weekly conditioning trials, the anti-nausea effectiveness of each of these compounds was also maintained. Repeated CBD (5 mg/kg, s.c.) maintained its anti-emetic efficacy in S. murinus. Acute CBD (5 and 20 mg/kg, s.c.) administration reduced LiCl-induced conditioned gaping similarly in male and female rats. Conclusion: When administered repeatedly (7 days), CBD, CBDA and HU-580 did not lose efficacy in reducing nausea and continued to act via agonism of the 5-HT1A receptor. When administered across 4 weekly conditioning trials, they maintained their effectiveness in reducing LiCl-induced nausea. Repeated CBD also reduced vomiting in shrews. Finally, CBD’s anti-nausea effects were similar in male and female rats. This suggests that these cannabinoids may be useful anti-nausea and anti-emetic treatments for chronic conditions, without the development of tolerance. © 2020, Springer-Verlag GmbH Germany, part of Springer Nature.
T. Stark, M. Di Bartolomeo, R. Di Marco, E. Drazanova, C.B.M. Platania, F.A. Iannotti, J. Ruda-Kucerova, C. D'Addario, L. Kratka, V. Pekarik, F. Piscitelli, Z. Babinska, J. Fedotova, G. Giurdanella, S. Salomone, A. Sulcova, C. Bucolo, C.T. Wotjak, Jr Starcuk, Z., F. Drago, R. Mechoulam, V. Di Marzo, and V. Micale. 2020. “Altered dopamine D3 receptor gene expression in MAM model of schizophrenia is reversed by peripubertal cannabidiol treatment.” Biochemical Pharmacology, 177. Publisher's Version Abstract
Gestational methylazoxymethanol acetate (MAM) treatment produces offspring with adult phenotype relevant to schizophrenia, including positive- and negative-like symptoms, cognitive deficits, dopaminergic dysfunction, structural and functional abnormalities. Here we show that adult rats prenatally treated with MAM at gestational day 17 display significant increase in dopamine D3 receptor (D3) mRNA expression in prefrontal cortex (PFC), hippocampus and nucleus accumbens, accompanied by increased expression of dopamine D2 receptor (D2) mRNA exclusively in the PFC. Furthermore, a significant change in the blood perfusion at the level of the circle of Willis and hippocampus, paralleled by the enlargement of lateral ventricles, was also detected by magnetic resonance imaging (MRI) techniques. Peripubertal treatment with the non-euphoric phytocannabinoid cannabidiol (30 mg/kg) from postnatal day (PND) 19 to PND 39 was able to reverse in MAM exposed rats: i) the up-regulation of the dopamine D3 receptor mRNA (only partially prevented by haloperidol 0.6 mg/kg/day); and ii) the regional blood flow changes in MAM exposed rats. Molecular modelling predicted that cannabidiol could bind preferentially to dopamine D3 receptor, where it may act as a partial agonist according to conformation of ionic-lock, which is highly conserved in GPCRs. In summary, our results demonstrate that the mRNA expression of both dopamine D2 and D3 receptors is altered in the MAM model; however only the transcript levels of D3 are affected by cannabidiol treatment, likely suggesting that this gene might not only contribute to the schizophrenia symptoms but also represent an unexplored target for the antipsychotic activity of cannabidiol. © 2020 Elsevier Inc.
P. Pacher, N.M. Kogan, and R. Mechoulam. 2020. “Beyond THC and endocannabinoids.” Annual Review of Pharmacology and Toxicology, 60, Pp. 637-659. Publisher's Version Abstract
Research in the cannabinoid field, namely on phytocannabinoids, the endogenous cannabinoids anandamide and 2-arachidonoyl glycerol and their metabolizing and synthetic enzymes, the cannabinoid receptors, and anandamide-like cannabinoid compounds, has expanded tremendously over the last few years. Numerous endocannabinoid-like compounds have been discovered. The Cannabis plant constituent cannabidiol (CBD) was found to exert beneficial effects in many preclinical disease models ranging from epilepsy, cardiovascular disease, inflammation, and autoimmunity to neurodegenerative and kidney diseases and cancer. CBD was recently approved in the United States for the treatment of rare forms of childhood epilepsy. This has triggered the development of many CBD-based products for human use, often with overstated claims regarding their therapeutic effects. In this article, the recently published research on the chemistry and biological effects of plant cannabinoids (specifically CBD), endocannabinoids, certain long-chain fatty acid amides, and the variety of relevant receptors is critically reviewed. Copyright © 2020 by Annual Reviews. All rights reserved.
Y.F. Zhu, K. Linher-Melville, M.J. Niazmand, M. Sharma, A. Shahid, K.L. Zhu, N. Parzei, J. Sidhu, C. Haj, R. Mechoulam, and G. Singh. 2020. “An evaluation of the anti-hyperalgesic effects of cannabidiolic acid-methyl ester in a preclinical model of peripheral neuropathic pain.” British Journal of Pharmacology, 177, 12, Pp. 2712-2725. Publisher's Version Abstract
Background and Purpose: Chronic neuropathic pain (NEP) is associated with growing therapeutic cannabis use. To promote quality of life without psychotropic effects, cannabinoids other than Δ9-tetrahydrocannabidiol, including cannabidiol and its precursor cannabidiolic acid (CBDA), are being evaluated. Due to its instability, CBDA has been understudied, particularly as an anti-nociceptive agent. Adding a methyl ester group (CBDA-ME) significantly enhances its stability, facilitating analyses of its analgesic effects in vivo. This study examines early treatment efficacy of CBDA-ME in a rat model of peripherally induced NEP and evaluates sex as a biological variable. Experimental Approach: After 14 consecutive days of intraperitoneal CBDA-ME administration at 0.01, 0.1 and 1 μg·kg−1, commencing 1 day after surgically implanting a sciatic nerve-constricting cuff to induce NEP, the anti-nociceptive efficacy of this cannabinoid was assessed in male and female Sprague–Dawley rats relative to vehicle-treated counterparts. In females, 2 and 4 μg·kg−1 daily doses of CBDA-ME were also evaluated. Behavioural tests were performed for hind paw mechanical and thermal withdrawal thresholds once a week for 8 weeks. At endpoint, in vivo electrophysiological recordings were obtained to characterize soma threshold changes in primary sensory neurons. Key Results: In males, CBDA-ME elicited a significant concentration-dependent chronic anti-hyperalgesic effect, also influencing both nociceptive and non-nociceptive mechanoreceptors, which were not observed in females at any of the concentrations tested. Conclusion and Implications: Initiating treatment of a peripheral nerve injury with CBDA-ME at an early stage post-surgery provides anti-nociception in males, warranting further investigation into potential sexual dimorphisms underlying this response. © 2020 The British Pharmacological Society
I.D. Small, M. Schallenberg-Rüdinger, M. Takenaka, H. Mireau, and O. Ostersetzer-Biran. 2020. “Plant organellar RNA editing: what 30 years of research has revealed.” Plant Journal, 101, 5, Pp. 1040-1056. Publisher's Version Abstract
The central dogma in biology defines the flow of genetic information from DNA to RNA to protein. Accordingly, RNA molecules generally accurately follow the sequences of the genes from which they are transcribed. This rule is transgressed by RNA editing, which creates RNA products that differ from their DNA templates. Analyses of the RNA landscapes of terrestrial plants have indicated that RNA editing (in the form of C-U base transitions) is highly prevalent within organelles (that is, mitochondria and chloroplasts). Numerous C→U conversions (and in some plants also U→C) alter the coding sequences of many of the organellar transcripts and can also produce translatable mRNAs by creating AUG start sites or eliminating premature stop codons, or affect the RNA structure, influence splicing and alter the stability of RNAs. RNA-binding proteins are at the heart of post-transcriptional RNA expression. The C-to-U RNA editing process in plant mitochondria involves numerous nuclear-encoded factors, many of which have been identified as pentatricopeptide repeat (PPR) proteins that target editing sites in a sequence-specific manner. In this review we report on major discoveries on RNA editing in plant organelles, since it was first documented 30 years ago. © 2019 The Authors The Plant Journal © 2019 John Wiley & Sons Ltd
O. Ostersetzer-Biran and L. Klipcan. 2020. “Aminoacyl-tRNA synthetases and translational quality control in plant mitochondria.” Mitochondrion, 54, Pp. 15-20. Publisher's Version Abstract
Gene expression involves the transfer of information stored in the DNA to proteins by two sequential key steps: transcription and translation. Aminoacyl-tRNA synthetases (aaRSs), an ancient group of enzymes, are key to these processes as they catalyze the attachment of each of the 20 amino acids to their corresponding tRNA molecules. Yet, in addition to the 20 canonical amino acids, plants also produce numerous non-proteogenic amino acids (NPAAs), some of which are erroneously loaded into tRNAs, translated into non-functional or toxic proteins and may thereby disrupt essential cellular processes. While many studies have been focusing on plant organelle RNA metabolism, mitochondrial translation still lags behind its characterization in bacterial and eukaryotic systems. Notably, plant mitochondrial aaRSs generally have a dual location, residing also within the chloroplasts or cytosol. Currently, little is known about how mitochondrial aaRSs distinguish between amino acids and their closely related NPAAs. The organelle translation machineries in plants seem more susceptible to NPAAs due to protein oxidation by reactive oxygen species (ROS) and high rates of protein turnover. We speculate that plant organellar aaRSs have acquired high-affinities to their cognate amino acid substrates to reduce cytotoxic effects by NPAAs. © 2020 Elsevier B.V. and Mitochondria Research Society
A. Fluksman, E. Steinberg, N. Orehov, E. Shai, A. Lahiani, J. Katzhendler, C. Marcinkiewicz, P. Lazarovici, and O. Benny. 2020. “Integrin α2β1-Targeted Self-Assembled Nanocarriers for Tumor Bioimaging.” ACS Applied Bio Materials, 3, 9, Pp. 6059-6070. Publisher's Version Abstract
Recent developments in near-infrared (NIR) dyes and imaging modalities enable tumor fluorescent images in preclinical and clinical settings. However, NIR dyes have several drawbacks, and therefore, there is an unmet diagnostic need for NIR dye encapsulation in appropriate pharmaceutical nanocarriers with targeting abilities for the purpose of achieving effective diagnosis and image-guided surgeries. Because integrin receptors are established diagnostic targets, the cyclic Arg-Gly-Asp (RGD) peptides, recognizing the αVβ3 integrin, have been extensively investigated for radiology and bioimaging of tumors. However, the Lys(Arg)-Thr-Ser [K(R)TS] cyclic peptides, selective for collagen receptors α1β1/α2β1 integrins, which are overexpressed in many tumors, were not yet investigated and therefore used here for tumor bioimaging with a unique α2β1-integrin-targeted nanocarrier, encapsulating the indocyanine green NIR dye. We synthesized three kinds of peptides: two cyclic RTS peptides functional only in the cyclic conformation and a linear peptide lacking the cyclic cysteine constrained RTS loop. We used them for the preparation of integrin-targeted self-assembled nanocarriers (ITNCs), referred to as OF5 and OF27, and a nontargeted control nanocarrier, referred to as OF70. Their selective association was demonstrated with α2β1 integrin expressing cell cultures and three-dimensional tumor spheroids and by competition with a α2β1 selective disintegrin. Cytotoxicity experiments in vitro demonstrated the safety of the ITNCs. The targeting potential and the biodistribution of the ITNCs, applied intravenously in A431 tumor-bearing nude mice, were evaluated in vivo using NIR bioimaging. Time-dependent biodistributions indicated that the ITNC OF27 showed higher fluorescent signals in main tissues, with no cytotoxic effects to major organs, and presented higher accumulation in tumors. Cumulatively, these results highlight the potential of the ITNC OF27 as an optical and innovative pharmaceutical bioimaging system, suitable for integrin α2β1 receptor in vivo tumor targeting and visualization in the NIR region. © 2020 American Chemical Society.
Liquid nanovehicles are gaining interest in drug delivery because of the high solubilization capacity of bioactives at their interface and enhanced permeation of compounds across physiological membranes. However, the dermal application of liquid nanovehicles is still limited. The goal of this research is to develop a dermal delivery system based on embedding of liquid nanovehicles into polymeric films, which will allow controlled release of the nanodroplets with the solubilized drug. In this study, we describe the incorporation of empty and curcumin-loaded nanodomains into polymeric film. The novel technology results in formation of homogeneous, transparent and elastic films with high (up to 85 wt%) loading capacity of nanodomains. The fundamental structural characterizations show that nanodomain structures embedded in the dry film are spontaneously reformed during the dermal application with similar droplets size of 10 nm. Ex-vivo release studies were performed on Franz diffusion cells and demonstrated a significant permeation of curcumin through the pig skin. This novel film technology can serve as a “solid platform reservoir” for liquid nanovehicles which enables controlled release of nanodroplets with solubilized bioactive. © 2020 Elsevier B.V.
R. Bulvik, R. Breuer, M. Dvir-Ginzberg, E. Reich, N. Berkman, and S.B. Wallach-Dayan. 2020. “Sirt1 deficiency, specifically in fibroblasts, decreases apoptosis resistance and is associated with resolution of lung-fibrosis.” Biomolecules, 10, 7, Pp. 1-12. Publisher's Version Abstract
In contrast to normal regenerating tissue, resistance to Fas-and FasL-positive T cell-induced apoptosis were detected in myofibroblasts from fibrotic-lungs of humans and mice following bleomycin (BLM) exposure. In this study we show, decreased FLIP expression in lung-tissues with resolution of BLM-induced fibrosis and in isolated-lung fibroblasts, with decreased resistance to apoptosis. Using a FLIP-expression vector or a shFLIP-RNA, we further confirmed the critical need for FLIP to regain/lose susceptibility of fibrotic-lung myofibroblast to Fas-induced apoptosis. Our study further show that FLIP is regulated by SIRT1 (Sirtuin 1) deacetylase. Chimeric mice, with SIRT1-deficiency in deacetylase domain (H355Y-Sirt1y/y), specifically in mesenchymal cells, were not only protected from BLM-induced lung fibrosis but, as assessed following Ku70 immunoprecipitation, had also decreased Ku70-deacetylation, decreasedKu70/FLIP complex, and decreased FLIP levels in their lung myofibroblasts. In addition, myofibroblasts isolated from lungs of BLM-treated miR34a-knockout mice, exposed to a miR34a mimic, which we found here to downregulate SIRT1 in the luciferase assay, had a decreased Ku70-deacetylation indicating decrease in SIRT1 activity. Thus, SIRT1 may mediate, miR34a-regulated, persistent FLIP levels by deacetylation of Ku70 in lung myofibroblasts, promoting resistance to cell-death and lung fibrosis. © 2020 by the authors. Licensee MDPI, Basel, Switzerland.
A. Drori, A. Gammal, S. Azar, L. Hinden, R. Hadar, D. Wesley, A. Nemirovski, G. Szanda, M. Salton, B. Tirosh, and J. Tam. 2020. “Cb1r regulates soluble leptin receptor levels via chop, contributing to hepatic leptin resistance.” eLife, 9, Pp. 1-26. Publisher's Version Abstract
The soluble isoform of leptin receptor (sOb-R), secreted by the liver, regulates leptin bioavailability and bioactivity. Its reduced levels in diet-induced obesity (DIO) contribute to hyperleptinemia and leptin resistance, effects that are regulated by the endocannabinoid (eCB)/ CB1R system. Here we show that pharmacological activation/blockade and genetic overexpression/ deletion of hepatic CB1 R modulates sOb-R levels and hepatic leptin resistance. Interestingly, peripheral CB1 R blockade failed to reverse DIO-induced reduction of sOb-R levels, increased fat mass and dyslipidemia, and hepatic steatosis in mice lacking C/EBP homologous protein (CHOP), whereas direct activation of CB1 R in wild-type hepatocytes reduced sOb-R levels in a CHOP-dependent manner. Moreover, CHOP stimulation increased sOb-R expression and release via a direct regulation of its promoter, while CHOP deletion reduced leptin sensitivity. Our findings highlight a novel molecular aspect by which the hepatic eCB/CB1R system is involved in the development of hepatic leptin resistance and in the regulation of sOb-R levels via CHOP. © 2020, eLife Sciences Publications Ltd. All rights reserved.
S. Azar, S. Udi, A. Drori, R. Hadar, A. Nemirovski, K.V. Vemuri, M. Miller, D. Sherill-Rofe, Y. Arad, D. Gur-Wahnon, X. Li, A. Makriyannis, D. Ben-Zvi, Y. Tabach, I.Z. Ben-Dov, and J. Tam. 2020. “Reversal of diet-induced hepatic steatosis by peripheral CB1 receptor blockade in mice is p53/miRNA-22/SIRT1/PPARα dependent.” Molecular Metabolism, 42. Publisher's Version Abstract
Objective: The endocannabinoid (eCB) system is increasingly recognized as being crucially important in obesity-related hepatic steatosis. By activating the hepatic cannabinoid-1 receptor (CB1R), eCBs modulate lipogenesis and fatty acid oxidation. However, the underlying molecular mechanisms are largely unknown. Methods: We combined unbiased bioinformatics techniques, mouse genetic manipulations, multiple pharmacological, molecular, and cellular biology approaches, and genomic sequencing to systematically decipher the role of the hepatic CB1R in modulating fat utilization in the liver and explored the downstream molecular mechanisms. Results: Using an unbiased normalized phylogenetic profiling analysis, we found that the CB1R evolutionarily coevolves with peroxisome proliferator-activated receptor-alpha (PPARα), a key regulator of hepatic lipid metabolism. In diet-induced obese (DIO) mice, peripheral CB1R blockade (using AM6545) induced the reversal of hepatic steatosis and improved liver injury in WT, but not in PPARα−/− mice. The antisteatotic effect mediated by AM6545 in WT DIO mice was accompanied by increased hepatic expression and activity of PPARα as well as elevated hepatic levels of the PPARα-activating eCB-like molecules oleoylethanolamide and palmitoylethanolamide. Moreover, AM6545 was unable to rescue hepatic steatosis in DIO mice lacking liver sirtuin 1 (SIRT1), an upstream regulator of PPARα. Both of these signaling molecules were modulated by the CB1R as measured in hepatocytes exposed to lipotoxic conditions or treated with CB1R agonists in the absence/presence of AM6545. Furthermore, using microRNA transcriptomic profiling, we found that the CB1R regulated the hepatic expression, acetylation, and transcriptional activity of p53, resulting in the enhanced expression of miR-22, which was found to specifically target SIRT1 and PPARα. Conclusions: We provide strong evidence for a functional role of the p53/miR-22/SIRT1/PPARα signaling pathway in potentially mediating the antisteatotic effect of peripherally restricted CB1R blockade. © 2020 The Authors
S. Udi, L. Hinden, M. Ahmad, A. Drori, M.R. Iyer, R. Cinar, M. Herman-Edelstein, and J. Tam. 2020. “Dual inhibition of cannabinoid CB1 receptor and inducible NOS attenuates obesity-induced chronic kidney disease.” British Journal of Pharmacology, 177, 1, Pp. 110-127. Publisher's Version Abstract
Background and Purpose: Obesity, an important risk factor for developing chronic kidney disease (CKD), affects the kidneys by two main molecular signalling pathways: the endocannabinoid/CB1 receptor system, whose activation in obesity promotes renal inflammation, fibrosis, and injury, and the inducible NOS (iNOS), which generates ROS resulting in oxidative stress. Hence, a compound that inhibits both peripheral CB1 receptors and iNOS may serve as an effective therapeutic agent against obesity-induced CKD. Experimental Approach: Here, we describe the effect of a novel peripherally restricted, orally bioavailable dual CB1 receptor/iNOS antagonist, MRI-1867 (3 mg·kg−1), in ameliorating obesity-induced CKD, and compared its metabolic and renal efficacies to a stand-alone peripheral CB1 receptor antagonist (JD5037; 3 mg·kg−1), iNOS antagonist (1400W; 10 mg·kg−1), and pair feeding. Mice with high-fat diet-induced obesity were treated orally with these compounds or vehicle (Veh) for 28 days. Standard diet-fed mice treated with Veh served as controls. Key Results: Enhanced expression of CB1 receptors and iNOS in renal tubules was found in human kidney patients with obesity and other CKDs. The hybrid inhibitor ameliorated obesity-induced kidney morphological and functional changes via decreasing kidney inflammation, fibrosis, oxidative stress, and renal injury. Some of these features were independent of the improved metabolic profile mediated via inhibition of CB1 receptors. An additional interesting finding is that these beneficial effects on the kidney were partially associated with modulating renal adiponectin signalling. Conclusions and Implications: Collectively, our results highlight the therapeutic relevance of blocking CB1 receptors and iNOS in ameliorating obesity-induced CKD. © 2019 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society

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