Figure 6.
Disruption of lipid rafts inhibits the antiproliferative effects of GPR55 activation. (A) Mz-ChA-1 cells were pretreated with lipid raft disrupters 0.1 mM β–methylcyclodextrin, or 1 μg/mL filipin III for 1 hr prior to the addition of O-1602 (10−5 M). Cell viability was determined by MTS assays. Data are expressed as average ± SEM (*p<0.05) compared to basal treatment (n=7). (B) Mz-ChA-1 cells were treated with AEA or O-1602 (both at 10−5 M) for 4 hr and detergent-resistant lipid rafts were isolated on a discontinuous 5%-40% sucrose gradient. The resulting fractions were analyzed by immunoblotting using GPR55 and Gα12-specific antibodies. (D) Co-localization of GPR55 immunoreactivity and lipid raft staining after AEA and O-1602 treatment. Mz-ChA-1 cells were treated with AEA and O-1602 (both at 10−5 M) and the lipid rafts were stained with Alexa Fluor 488-conjugated cholera toxin B subunit (green) as well as GPR55 immunoreactivity (red). Co-localization is indicated by yellow areas. Nuclei were counterstained with DAPI (blue). Scale bar represents 10 μm.