Neuroprotective effects of cannabidiol in endotoxin-induced uveitis: critical role of p38 MAPK activation.
OBJECTIVE
Degenerative retinal diseases are characterized by inflammation and microglial activation. The nonpsychoactive cannabinoid, cannabidiol (CBD), is an anti-inflammatory in models of diabetes and glaucoma. However, the cellular and molecular mechanisms are largely unknown. We tested the hypothesis that retinal inflammation and microglia activation are initiated and sustained by oxidative stress and p38 mitogen-activated protein kinase (MAPK) activation, and that CBD reduces inflammation by blocking these processes.


METHODS
Microglial cells were isolated from retinas of newborn rats. Tumor necrosis factor (TNF)-alpha levels were estimated with ELISA. Nitric oxide (NO) was determined with a NO analyzer. Superoxide anion levels were determined by the chemiluminescence of luminol derivative. Reactive oxygen species (ROS) was estimated by measuring the cellular oxidation products of 2', 7'-dichlorofluorescin diacetate.


RESULTS
In retinal microglial cells, treatment with lipopolysaccharide (LPS) induced immediate NADPH oxidase-generated ROS. This was followed by p38 MAPK activation and resulted in a time-dependent increase in TNF-alpha production. At a later phase, LPS induced NO, ROS, and p38 MAPK activation that peaked at 2-6 h and was accompanied by morphological change of microglia. Treatment with 1 microM CBD inhibited ROS formation and p38 MAPK activation, NO and TNF-alpha formation, and maintained cell morphology. In addition, LPS-treated rat retinas showed an accumulation of macrophages and activated microglia, significant levels of ROS and nitrotyrosine, activation of p38 MAPK, and neuronal apoptosis. These effects were blocked by treatment with 5 mg/kg CBD.


CONCLUSIONS
Retinal inflammation and degeneration in uveitis are caused by oxidative stress. CBD exerts anti-inflammatory and neuroprotective effects by a mechanism that involves blocking oxidative stress and activation of p38 MAPK and microglia.
El-Remessy
A B
AB
Department of Ophthalmology, Medical College of Georgia, Augusta, GA 30912, USA.
Tang
Y
Y
Zhu
G
G
Matragoon
S
S
Khalifa
Y
Y
Liu
E K
EK
Liu
J-Y
JY
Hanson
E
E
Mian
S
S
Fatteh
N
N
Liou
G I
GI
D000818:Animals; D002185:Cannabidiol / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D004731:Endotoxins / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000201:enzymology / Q000473:pathology; D008297:Male; D017628:Microglia / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D008954:Models, Biological; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009569:Nitric Oxide / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D030421:Peroxynitrous Acid / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012160:Retina / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D013481:Superoxides / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D014605:Uveitis / Q000139:chemically induced* / Q000201:enzymology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D004731:Endotoxins; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D018696:Neuroprotective Agents; D014409:Tumor Necrosis Factor-alpha; D013481:Superoxides; D030421:Peroxynitrous Acid; D002185:Cannabidiol; D009569:Nitric Oxide; D019255:NADPH Oxidases; D048051:p38 Mitogen-Activated Protein Kinases