PMID- 25880134 OWN - NLM STAT- MEDLINE DCOM- 20160307 LR - 20221207 IS - 1742-2094 (Electronic) IS - 1742-2094 (Linking) VI - 12 DP - 2015 Mar 15 TI - Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells. PG - 52 LID - 10.1186/s12974-015-0273-0 [doi] LID - 52 AB - BACKGROUND: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear. METHODS: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (TMOG) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. TMOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified TMOG were carried out using immunoblotting. RESULTS: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4(+)CD25(-) accessory T cells. This subtype of CD4(+)CD25(-)CD69(+)LAG3(+) T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated TMOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in TMOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19(+) B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions. CONCLUSIONS: Our data suggests that CBD exerts its immunoregulatory effects via induction of CD4(+)CD25(-)CD69(+)LAG3(+) cells in MOG35-55-activated APC/TMOG co-cultures. This is accompanied by EGR2-dependent anergy of stimulated TMOG cells as well as a switch in their intracellular STAT3/STAT5 activation balance leading to the previously observed decrease in Th17 activity. FAU - Kozela, Ewa AU - Kozela E AD - The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel. ewa.kozela@weizmann.ac.il. FAU - Juknat, Ana AU - Juknat A AD - The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel. Ana.juknat@weizmann.ac.il. FAU - Kaushansky, Nathali AU - Kaushansky N AD - Neurobiology Department, Weizmann Institute of Science, Rehovot, Israel. nathali.kaushansky@weizmann.ac.il. FAU - Ben-Nun, Avraham AU - Ben-Nun A AD - Immunology Department, Weizmann Institute of Science, Rehovot, Israel. avi.ben-nun@weizmann.ac.il. FAU - Coppola, Giovanni AU - Coppola G AD - Neurology Department, UCLA, Los Angeles, CA, USA. gcoppola@ucla.edu. FAU - Vogel, Zvi AU - Vogel Z AD - The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel. vogelz@post.tau.ac.il. AD - Neurobiology Department, Weizmann Institute of Science, Rehovot, Israel. vogelz@post.tau.ac.il. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20150315 PL - England TA - J Neuroinflammation JT - Journal of neuroinflammation JID - 101222974 RN - 0 (Antigens, CD) RN - 0 (Cytokines) RN - 0 (Early Growth Response Protein 2) RN - 0 (Egr2 protein, mouse) RN - 0 (Myelin-Oligodendrocyte Glycoprotein) RN - 0 (Peptide Fragments) RN - 0 (Psychotropic Drugs) RN - 0 (STAT3 Transcription Factor) RN - 0 (STAT5 Transcription Factor) RN - 0 (Stat3 protein, mouse) RN - 0 (myelin oligodendrocyte glycoprotein (35-55)) RN - 19GBJ60SN5 (Cannabidiol) RN - 0 (Lymphocyte Activation Gene 3 Protein) RN - 0 (Lag3 protein, mouse) SB - IM MH - Animals MH - Antigen-Presenting Cells/drug effects MH - Antigens, CD/metabolism MH - Cannabidiol/*pharmacology MH - Cell Line MH - Coculture Techniques MH - Cytokines/*metabolism MH - Early Growth Response Protein 2/genetics/*metabolism MH - Female MH - Flow Cytometry MH - Gene Expression Regulation/drug effects MH - Lymphocyte Activation/*drug effects MH - Mice MH - Mice, Inbred C57BL MH - Myelin-Oligodendrocyte Glycoprotein/pharmacology MH - Peptide Fragments/pharmacology MH - Psychotropic Drugs/*pharmacology MH - STAT3 Transcription Factor/metabolism MH - STAT5 Transcription Factor/genetics/metabolism MH - Signal Transduction/drug effects MH - T-Lymphocytes/*drug effects MH - Lymphocyte Activation Gene 3 Protein PMC - PMC4363052 EDAT- 2015/04/17 06:00 MHDA- 2016/03/08 06:00 PMCR- 2015/03/15 CRDT- 2015/04/17 06:00 PHST- 2015/01/06 00:00 [received] PHST- 2015/02/26 00:00 [accepted] PHST- 2015/04/17 06:00 [entrez] PHST- 2015/04/17 06:00 [pubmed] PHST- 2016/03/08 06:00 [medline] PHST- 2015/03/15 00:00 [pmc-release] AID - 10.1186/s12974-015-0273-0 [pii] AID - 273 [pii] AID - 10.1186/s12974-015-0273-0 [doi] PST - epublish SO - J Neuroinflammation. 2015 Mar 15;12:52. doi: 10.1186/s12974-015-0273-0.