Characterization of the endocannabinoid system in human neuronal cells and proteomic analysis of anandamide-induced apoptosis.
Anandamide (AEA) is an endogenous agonist of type 1 cannabinoid receptors (CB1R) that, along with metabolic enzymes of AEA and congeners, compose the "endocannabinoid system." Here we report the biochemical, morphological, and functional characterization of the endocannabinoid system in human neuroblastoma SH-SY5Y cells that are an experimental model for neuronal cell damage and death, as well as for major human neurodegenerative disorders. We also show that AEA dose-dependently induced apoptosis of SH-SY5Y cells. Through proteomic analysis, we further demonstrate that AEA-induced apoptosis was paralleled by an approximately 3 to approximately 5-fold up-regulation or down-regulation of five genes; IgG heavy chain-binding protein, stress-induced phosphoprotein-1, and triose-phosphate isomerase-1, which were up-regulated, are known to act as anti-apoptotic agents; actin-related protein 2/3 complex subunit 5 and peptidylprolyl isomerase-like protein 3 isoform PPIL3b were down-regulated, and the first is required for actin network formation whereas the second is still function-orphan. Interestingly, only the effect of AEA on BiP was reversed by the CB1R antagonist SR141716, in SH-SY5Y cells as well as in human neuroblastoma LAN-5 cells (that express a functional CB1R) but not in SK-NBE cells (which do not express CB1R). Silencing or overexpression of BiP increased or reduced, respectively, AEA-induced apoptosis of SH-SY5Y cells. In addition, the expression of BiP and of the BiP-related apoptotic markers p53 and PUMA was increased by AEA through a CB1R-dependent pathway that engages p38 and p42/44 mitogen-activated protein kinases. Consistently, this effect of AEA was minimized by SR141716. In conclusion, we identified BiP as a key protein in neuronal apoptosis induced by AEA.
Pasquariello
Nicoletta
N
Dipartimento di Scienze Biomediche, Università degli Studi di Teramo, 64100 Teramo, Italy.
Catanzaro
Giuseppina
G
Marzano
Valeria
V
Amadio
Daniele
D
Barcaroli
Daniela
D
Oddi
Sergio
S
Federici
Giorgio
G
Urbani
Andrea
A
Finazzi Agrò
Alessandro
A
Maccarrone
Mauro
M
D017209:Apoptosis / Q000187:drug effects*; D001095:Arachidonic Acids / Q000494:pharmacology*; D063385:Cannabinoid Receptor Modulators / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D063388:Endocannabinoids; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D008954:Models, Biological; D009419:Nerve Tissue Proteins / Q000096:biosynthesis*; D009474:Neurons / Q000378:metabolism*; D010880:Piperidines / Q000494:pharmacology; D053284:Polyunsaturated Alkamides / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology; D043884:Receptor, Cannabinoid, CB1 / Q000819:agonists / Q000378:metabolism; D000077285:Rimonabant
D001095:Arachidonic Acids; D063385:Cannabinoid Receptor Modulators; D063388:Endocannabinoids; D009419:Nerve Tissue Proteins; D010880:Piperidines; D053284:Polyunsaturated Alkamides; D011720:Pyrazoles; D043884:Receptor, Cannabinoid, CB1; D000077285:Rimonabant; C078814:anandamide
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