Figure 4

Cannabinoid CB ₂ agonist treatment opposes IFN-γ-induced phosphorylation of JAK/STAT1 in microglial cells. A, B, Primary microglial cells were seeded in 6-well tissue-culture plates (5 × 10⁵/well) and treated with IFN-γ (100 U/mL) in the presence or absence of CB₂ agonist (JWH-015) at the indicated doses for 30 min. Cell lysates were prepared from these cells and subjected to Western immunoblotting using antibodies against phospho-JAK1 (Tyr1022/1023) and JAK2 (Tyr1007/1008), or total JAK1 and JAK2, as indicated. Densitometric quantification of all Western immunoblots results are summarized by the histograms below, representative of Western immunoblots from two independent experiments. Dose-dependent reductions in phospho-JAK1/total JAK1 and phosphor-JAK2/total JAK2 correlated with JWH-015 treatments, becoming significant (** p < 0.05) at doses greater than or equal to 1.25 μM and 0.62 μM for JAK1 and JAK2, respectively. C, In parallel experiments, cell lysates were subjected to Western immunoblotting using anti-phospho-STAT1 (Ser727) or anti-total STAT1 antibody as indicated. Dose-dependent reductions in phospho-Stat1/total Stat1 correlated with JWH-015 treatments, becoming significant (** p < 0.05) at doses greater than or equal to 0.62 μM.