Cannabidiol protects against hepatic ischemia/reperfusion injury by attenuating inflammatory signaling and response, oxidative/nitrative stress, and cell death.

Ischemia/reperfusion (I/R) is a pivotal mechanism of liver damage after liver transplantation or hepatic surgery. We have investigated the effects of cannabidiol (CBD), the nonpsychotropic constituent of marijuana, in a mouse model of hepatic I/R injury. I/R triggered time-dependent increases/changes in markers of liver injury (serum transaminases), hepatic oxidative/nitrative stress (4-hydroxy-2-nonenal, nitrotyrosine content/staining, and gp91phox and inducible nitric oxide synthase mRNA), mitochondrial dysfunction (decreased complex I activity), inflammation (tumor necrosis factor α (TNF-α), cyclooxygenase 2, macrophage inflammatory protein-1α/2, intercellular adhesion molecule 1 mRNA levels; tissue neutrophil infiltration; nuclear factor κB (NF-κB) activation), stress signaling (p38MAPK and JNK), and cell death (DNA fragmentation, PARP activity, and TUNEL). CBD significantly reduced the extent of liver inflammation, oxidative/nitrative stress, and cell death and also attenuated the bacterial endotoxin-triggered NF-κB activation and TNF-α production in isolated Kupffer cells, likewise the adhesion molecule expression in primary human liver sinusoidal endothelial cells stimulated with TNF-α and attachment of human neutrophils to the activated endothelium. These protective effects were preserved in CB(2) knockout mice and were not prevented by CB(1/2) antagonists in vitro. Thus, CBD may represent a novel, protective strategy against I/R injury by attenuating key inflammatory pathways and oxidative/nitrative tissue injury, independent of classical CB(1/2) receptors

Cannabidiol attenuates cardiac dysfunction, oxidative stress, fibrosis, and inflammatory and cell death signaling pathways in diabetic cardiomyopathy.

Objectives In this study, we have investigated the effects of cannabidiol (CBD) on myocardial dysfunction, inflammation, oxidative/nitrative stress, cell death, and interrelated signaling pathways, using a mouse model of type I diabetic cardiomyopathy and primary human cardiomyocytes exposed to high glucose. Background Cannabidiol, the most abundant nonpsychoactive constituent of Cannabis sativa (marijuana) plant, exerts anti-inflammatory effects in various disease models and alleviates pain and spasticity associated with multiple sclerosis in humans. Methods Left ventricular function was measured by the pressure-volume system. Oxidative stress, cell death, and fibrosis markers were evaluated by molecular biology/biochemical techniques, electron spin resonance spectroscopy, and flow cytometry. Results Diabetic cardiomyopathy was characterized by declined diastolic and systolic myocardial performance associated with increased oxidative-nitrative stress, nuclear factor-kappa B and mitogen-activated protein kinase (c-Jun N-terminal kinase, p-38, p38 alpha) activation, enhanced expression of adhesion molecules (intercellular adhesion molecule-1, vascular cell adhesion molecule-1), tumor necrosis factor-alpha, markers of fibrosis (transforming growth factor-beta, connective tissue growth factor, fibronectin, collagen-1, matrix metalloproteinase-2 and -9), enhanced cell death (caspase 3/7 and poly[adenosine diphosphate-ribose] polymerase activity, chromatin fragmentation, and terminal deoxynucleotidyl transferase dUTP nick end labeling), and diminished Akt phosphorylation. Remarkably, CBD attenuated myocardial dysfunction, cardiac fibrosis, oxidative/nitrative stress, inflammation, cell death, and interrelated signaling pathways. Furthermore, CBD also attenuated the high glucose-induced increased reactive oxygen species generation, nuclear factor-kappa B activation, and cell death in primary human cardiomyocytes. Conclusions Collectively, these results coupled with the excellent safety and tolerability profile of CBD in humans, strongly suggest that it may have great therapeutic potential in the treatment of diabetic complications, and perhaps other cardiovascular disorders, by attenuating oxidative/nitrative stress, inflammation, cell death and fibrosis. (J Am Coll Cardiol 2010;56:2115-25) (C) 2010 by the American College of Cardiology Foundation

Mining a human transcriptome database for chemical modulators of NRF2

Nuclear factor erythroid-2 related factor 2 (NRF2) encoded by the NFE2L2 gene is a transcription factor critical for protecting cells from chemically-induced oxidative stress. We developed computational procedures to identify chemical modulators of NRF2 in a large database of human microarray data. A gene expression biomarker was built from statistically-filtered gene lists derived from microarray experiments in primary human hepatocytes and cancer cell lines exposed to NRF2-activating chemicals (oltipraz, sulforaphane, CDDO-Im) or in which the NRF2 suppressor Keap1 was knocked down by siRNA. Directionally consistent biomarker genes were further filtered for those dependent on NRF2 using a microarray dataset from cells after NFE2L2 siRNA knockdown. The resulting 143-gene biomarker was evaluated as a predictive tool using the correlation-based Running Fisher algorithm. Using 59 gene expression comparisons from chemically-treated cells with known NRF2 activating potential, the biomarker gave a balanced accuracy of 93%. The biomarker was comprised of many well-known NRF2 target genes (AKR1B10, AKR1C1, NQO1, TXNRD1, SRXN1, GCLC, GCLM), 69% of which were found to be bound directly by NRF2 using ChIP-Seq. NRF2 activity was assessed across ~9840 microarray comparisons from ~1460 studies examining the effects of ~2260 chemicals in human cell lines. A total of 260 and 43 chemicals were found to activate or suppress NRF2, respectively, most of which have not been previously reported to modulate NRF2 activity. Using a NRF2-responsive reporter gene in HepG2 cells, we confirmed the activity of a set of chemicals predicted using the biomarker. The biomarker will be useful for future gene expression screening studies of environmentally-relevant chemicals.Toxicolog

HNO Protects the Myocardium against Reperfusion Injury, Inhibiting the mPTP Opening via PKCε Activation

Donors of nitroxyl (HNO), the one electron-reduction product of nitric oxide (NO. ), posi-tively modulate cardiac contractility/relaxation while limiting ischemia-reperfusion (I/R) injury. The mechanisms underpinning HNO anti-ischemic effects remain poorly understood. Using isolated perfused rat hearts subjected to 30 min global ischemia/1 or 2 h reperfusion, here we tested whether, in analogy to NO., HNO protection requires PKCε translocation to mitochondria and KATP channels activation. To this end, we compared the benefits afforded by ischemic preconditioning (IPC; 3 cycles of I/R) with those eventually granted by the NO. donor, diethylamine/NO, DEA/NO, and two chemically unrelated HNO donors: Angeli’s salt (AS, a prototypic donor) and isopropyla-mine/NO (IPA/NO, a new HNO releaser). All donors were given for 19 min before I/R injury. In control I/R hearts (1 h reperfusion), infarct size (IS) measured via tetrazolium salt staining was 66 ± 5.5% of the area at risk. Both AS and IPA/NO were as effective as IPC in reducing IS [30.7 ± 2.2 (AS), 31 ± 2.9 (IPA/NO), and 31 ± 0.8 (IPC), respectively)], whereas DEA/NO was significantly less so (36.2 ± 2.6%, p < 0.001 vs. AS, IPA/NO, or IPC). IPA/NO protection was still present after 120 min of reperfusion, and the co-infusion with the PKCε inhibitor (PKCV1-2500 nM) prevented it (IS = 30 ± 0.5 vs. 61 ± 1.8% with IPA/NO alone, p < 0.01). Irrespective of the donor, HNO anti-ischemic effects were insensitive to the KATP channel inhibitor, 5-OH decanoate (5HD, 100 μM), that, in contrast, abrogated DEA/NO protection. Finally, both HNO donors markedly enhanced the mitochondrial permeability transition pore (mPTP) ROS threshold over control levels (≅35–40%), an action again insensitive to 5HD. Our study shows that HNO donors inhibit mPTP opening, thus limiting myo-cyte loss at reperfusion, a beneficial effect that requires PKCε translocation to the mitochondria but not mitochondrial K+ channels activation. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Novel dithiolethione-modified nonsteroidal anti-inflammatory drugs in human hepatoma HepG2 and colon LS180 cells

PURPOSE: Nonsteroidal anti-inflammatory drugs (NSAID) are promising chemopreventive agents against colon and other cancers. However, the molecular basis mediated by NSAIDs for chemoprevention has not been fully elucidated. Environmental carcinogens induce DNA mutation and cellular transformation; therefore, we examined the effect of NSAIDs on carcinogenesis mediated by the aryl hydrocarbon receptor signaling pathway. In this study, we investigated the activities of a new class of NSAIDs containing dithiolethione moieties (S-NSAID) on both arms of carcinogenesis. EXPERIMENTAL DESIGN: We investigated the effects of the S-NSAIDs, S-diclofenac and S-sulindac, on carcinogen activation and detoxification mechanisms in human hepatoma HepG2 and human colonic adenocarcinoma LS180 cells. RESULTS: We found that S-diclofenac and S-sulindac inhibited the activity and expression of the carcinogen activating enzymes, cytochromes P-450 (CYP) CYP1A1, CYP1B1, and CYP1A2. Inhibition was mediated by transcriptional regulation of the aryl hydrocarbon receptor (AhR) pathway. The S-NSAIDs down-regulated carcinogen-induced expression of CYP1A1 heterogeneous nuclear RNA, a measure of transcription rate. Both compounds blocked carcinogen-activated AhR from binding to the xenobiotic responsive element as shown by chromatin immunoprecipitation. S-diclofenac and S-sulindac inhibited carcinogen-induced CYP enzyme activity through direct inhibition as well as through decreased transcriptional activation of the AhR. S-sulindac induced expression of several carcinogen detoxification enzymes of the glutathione cycle including glutathione S-transferase A2, glutamate cysteine ligase catalytic subunit, glutamate cysteine ligase modifier subunit, and glutathione reductase. CONCLUSIONS: These results indicate that S-diclofenac and S-sulindac may serve as effective chemoprevention agents by favorably balancing the equation of carcinogen activation and detoxification mechanism

Dithiolethiones inhibit NF-\u3baB Activity via covalent modification in human estrogen receptor-negative breast cancer

The NF-\u3baB transcription factor family influences breast cancer outcomes by regulating genes involved in tumor progression, angiogenesis, and metastasis. Dithiolethiones, a class of naturally occurring compounds with cancer chemoprevention effects that have become clinically available, have been found to inhibit NF-\u3baB activity. However, the mechanism of this inhibition has not been identified, and the influence of dithiolethines on NF-\u3baB pathway in breast cancer cells has not been examined. Here, we investigated the chemical and biochemical effects of dithiolethione on NF-\u3baB and downstream effector molecules in estrogen receptor-negative breast cancer cells and murine tumor xenografts. The dithiolethiones ACS-1 and ACS-2 inhibited NF-\u3baB transcriptional activity. Interestingly, this inhibition was not due to H2S release or protein phosphatase 2A activation, which are key properties of dithiolethiones, but occurred via a covalent reaction with the NF-\u3baB p50 and p65 subunits to inhibit DNA binding. Dithiolethione-mediated inhibition of NF-\u3baB-regulated genes resulted in the inhibition of interleukin (IL)-6, IL-8, urokinase-type plasminogen activator, and VEGF production. ACS-1 also inhibited matrix metalloproteinase-9 activity, cellular migration, and invasion, and ACS-2 reduced tumor burden and resulted in increased tumor host interactions. Together, our findings suggest that dithiolethiones show potential clinical use for estrogen negative breast cancer as a chemotherapeutic or adjuvant therapy