Endocannabinoid system modulation of the ocular immune response

Abstract

The endocannabinoid system (ECS) is an endogenous lipid signaling system, comprised of receptors, ligands, and their degradative enzymes. The cannabinoid 2 receptor (CB2R) is localized on immune cells and modulates immune function in health and disease. Activation of the CB2R has anti-inflammatory actions in several organ systems, however, the effects of ECS modulation during ocular inflammation has not been thoroughly investigated. Novel therapeutics for ocular inflammation are needed as current treatments can exhibit lack of efficacy, become refractory, and produce side effects. To address these problems, the work presented in this thesis examined the manipulation of the ECS during ocular inflammation and the effects of CB2R activation on the inflammatory response. To examine the effects of CB2R activation, in vivo and in vitro models of inflammation were generated using bacterial endotoxin (lipopolysaccharide, LPS). LPS interacts with the cell surface Toll-like receptor 4 (TLR4) to induce inflammation. Following systemic or intraocular injection of LPS, leukocyte-endothelial interactions and local blood flow were investigated in the iridial microcirculation as a marker of inflammation. Cannabinoid treatments were administered i.v. or topically to the eye. Pharmacological treatments and genetic knock out animals were used to confirm the receptor and cellular targets of cannabinoid agonists. Endocannabinoid levels, release of inflammatory mediators, generation of intracellular signaling molecules, and receptor expression were investigated during the inflammatory response and following treatment to elucidate mechanisms of action. This was further probed by in vitro assays investigating the mitogen activated protein kinase (MAPK) pathway. Taken together, the findings presented in this thesis demonstrated that activation of CB2R has anti-inflammatory properties during acute ocular inflammation with efficacy comparable to current clinical therapies. CB2R treatment decreases leukocyte-endothelial adhesion, improves local blood flow and diminishes release of inflammatory mediators. Additionally, I have proposed a mechanism of action for these anti-inflammatory effects based on my findings. My results suggest that CB2R agonists inhibit activation of the MAPK pathway by the LPS-TLR4 interactions in activated macrophages. This prevents activation of pro-inflammatory transcription factors and the subsequent release of the associated pro-inflammatory mediators. Overall, activation of the CB2R provides a potential therapeutic target for the treatment of ocular inflammation.