Cannabinoid/lysophosphatidylinositol-sensing G-protein coupled receptor 55 promotes intestinal tight junction assembly and its mechanistic insights

Intestinal tight junction disruption initiates progression of related diseases including inflammatory bowel disease (IBD) with no FDA-approved drug for tight junction recovery. To demonstrate the effect of pharmacological activation of the cannabinoid/lysophosphatidylinositol-sensing G-protein coupled receptor 55 (GPR55) by its specific synthetic agonist O1602 on intestinal barrier function, tight junction-dependent permeability, and its underlying mechanisms. We show that O1602 treatment increased transepithelial electrical resistance (TER) across intestinal epithelial-like T84 cell monolayers and suppressed 4-kDa FITC-dextran permeability. Neither CB1 inhibitor nor CB2 inhibitor has affected TER increases in response to O1602 treatment. O1602 was ineffective in enhancing intestinal barrier integrity in T84 monolayers treated with GPR55 antagonist or in GPR55 KD T84 monolayers, indicating that GPR55 agonism promotes intestinal barrier function and inhibits tight junction-dependent leak pathway permeability. In fact, O1602 treatment also prevented TNF-α-induced intestinal barrier disruption in IFN-γ-primed T84 and Caco-2BBe monolayers. The effect of O1602 treatment on enhancing TER across T84 cell monolayers was abolished by pre-treatment with inhibitors of PLC, CaMKKβ, AMPK, SIRT-1, ERK, PKA, β-arrestin, and mTOR. In addition, O1602 failed to promote TER increases in SIRT-1 KO T84 monolayers. Our data from western blot analysis, SIRT-1 activity assay, and immunofluorescence staining of tight junction proteins, coherently recapitulates that GPR55 agonism induces intestinal tight junction assembly via PLC/[Ca²⁺]_i/CaMKKβ/AMPK/SIRT-1/ERK-dependent mechanism. Hence, we furnish the first line of evidence supporting that GPR55 is the regulator of tight junction in intestinal epithelial monolayers and may serve as a novel class of therapeutic target for tight junction disruption-associated diseases.