The Non-Psychoactive Plant Cannabinoid, Cannabidiol Affects Cholesterol Metabolism-Related Genes in Microglial Cells
Cannabidiol (CBD) is a non-psychoactive plant cannabinoid that is clinically used in a 1:1 mixture with the psychoactive cannabinoid Δ9-tetrahydrocannabinol (THC) for the treatment of neuropathic pain and spasticity in multiple sclerosis. Our group previously reported that CBD exerts anti-inflammatory effects on microglial cells. In addition, we found that CBD treatment increases the accumulation of the endocannabinoid N-arachidonoyl ethanolamine (AEA), thus enhancing endocannabinoid signaling. Here we proceeded to investigate the effects of CBD on the modulation of lipid-related genes in microglial cells. Cell viability was tested using FACS analysis, AEA levels were measured using LC/MS/MS, gene array analysis was validated with real-time qPCR, and cytokine release was measured using ELISA. We report that CBD significantly upregulated the mRNAs of the enzymes sterol-O-acyl transferase (Soat2), which synthesizes cholesteryl esters, and of sterol 27-hydroxylase (Cyp27a1). In addition, CBD increased the mRNA of the lipid droplet-associated protein, perilipin2 (Plin2). Moreover, we found that pretreatment of the cells with the cholesterol chelating agent, methyl-β-cyclodextrin (MBCD), reversed the CBD-induced increase in Soat2 mRNA but not in Plin2 mRNA. Incubation with AEA increased the level of Plin2, but not of Soat2 mRNA. Furthermore, MBCD treatment did not affect the reduction by CBD of the LPS-induced release of the proinflammatory cytokine IL-1β. CBD treatment modulates cholesterol homeostasis in microglial cells, and pretreatment with MBCD reverses this effect without interfering with CBD’s anti-inflammatory effects. The effects of the CBD-induced increase in AEA accumulation on lipid-gene expression are discussed.
KeywordsCholesteryl ester Cannabidiol Cholesterol Diacylglycerol-O-acyl transferase Lipid droplet N-arachidonoyl ethanolamine Protein tyrosine phosphatase non receptor 22 Sterol-O-acyl transferase Sterol 27-hydroxylase Microglia
We would like to thank Prof. Daniel Geschwind and Dr. Giovanni Coppola, from the Program in Neurogenetics, Department of Neurology, University of California, Los Angeles, CA, for running the microarray analysis. NR and AJ were supported by the Israeli Ministry of Immigrant Absorption, The Center for Absorption in Science. This work was supported by The Dr. Miriam and Sheldon G. Adelson Center for the Biology of Addictive Disease.
- Bisogno T, Hanus L, De Petrocellis L, Tchilibon S, Ponde DE, Brandi I, Moriello AS, Davis JB, Mechoulam R, Di Marzo V (2001) Molecular targets for cannabidiol and its synthetic analogues: effect on vanilloid VR1 receptors and on the cellular uptake and enzymatic hydrolysis of anandamide. Br J Pharmacol 134:845–852PubMedCrossRefGoogle Scholar
- Biswas KK, Sarker KP, Abeyama K, Kawahara K, Iino S, Otsubo Y, Saigo K, Izumi H, Hashiguchi T, Yamakuchi M, Yamaji K, Endo R, Suzuki K, Imaizumi H, Maruyama I (2003) Membrane cholesterol but not putative receptors mediates anandamide-induced hepatocyte apoptosis. Hepatology 38:1167–1177PubMedCrossRefGoogle Scholar
- De Petrocellis L, Ligresti A, Moriello AS, Allara M, Bisogno T, Petrosino S et al (2011) Effects of cannabinoids and cannabinoid-enriched Cannabis extracts on TRP channels and endocannabinoid metabolic enzymes. Br J Pharmacol. doi: 10.1111/j.1476-5381.2010.01166.x
- Gentleman RC, Carey VJ, Bates DM, Bolstad B, Dettling M, Dudoit S, Ellis B, Gautier L, Ge Y, Gentry J, Hornik K, Hothorn T, Huber W, Iacus S, Irizarry R, Leisch F, Li C, Maechler M, Rossini AJ, Sawitzki G, Smith C, Smyth G, Tierney L, Yang JY, Zhang J (2004) Bioconductor: open software development for computational biology and bioinformatics. Genome Biol 5:R80PubMedCrossRefGoogle Scholar
- Juknat A, Pietr M, Kozela E, Rimmerman N, Levy R, Coppola G, Geschwind D, Vogel Z (2011) Differential transcriptional profiles mediated by exposure to the cannabinoids cannabidiol and Δ9-tetrahydrocannabinol in BV-2 microglial cells. Br J Pharmacol (accepted)Google Scholar
- Kozela E, Pietr M, Juknat A, Rimmerman N, Levy R, Vogel Z (2010) Cannabinoids Delta(9)-tetrahydrocannabinol and cannabidiol differentially inhibit the lipopolysaccharide-activated NF-kappaB and interferon-beta/STAT proinflammatory pathways in BV-2 microglial cells. J Biol Chem 285:1616–1626PubMedCrossRefGoogle Scholar
- Kozela E, Lev N, Kaushansky N, Eilam R, Rimmerman N, Levy R, Ben-Nun A, Juknat A Vogel Z (2011) Cannabidiol inhibits pathogenic T-cells, decreases spinal microglial activation and ameliorates multiple sclerosis-like disease in C57BL/6 mice. Br J Pharmacol. doi: 10.1111/j.1476-5381.2011.01379.x
- Ligresti A, Cascio MG, Pryce G, Kulasegram S, Beletskaya I, De Petrocellis L, Saha B, Mahadevan A, Visintin C, Wiley JL, Baker D, Martin BR, Razdan RK, Di Marzo V (2006) New potent and selective inhibitors of anandamide reuptake with antispastic activity in a mouse model of multiple sclerosis. Br J Pharmacol 147:83–91PubMedCrossRefGoogle Scholar
- Rimmerman N, Bradshaw HB, Kozela E, Levy R, Juknat A, Vogel Z (2011) Compartmentalization of endocannabinoids into lipid rafts in a microglial cell line which is devoid of caveolin-1. Br J Pharmacol. doi: 10.1111/j.14765381.2011.01380.x
- Wu HY, Chang AC, Wang CC, Kuo FH, Lee CY, Liu DZ, Jan TR (2010) Cannabidiol induced a contrasting pro-apoptotic effect between freshly isolated and precultured human monocytes. Toxicol Appl Pharmacol. doi: 10.1016/j.physletb.2003.10.071