Fig. 5
From: Cannabis suppresses antitumor immunity by inhibiting JAK/STAT signaling in T cells through CNR2
Cnr2 deficiency promotes T-cell-mediated antitumor immunity. a Cnr2GFP (CD45.1+ CD45.2+) and Cnr2CKO (CD45.2+) OT-I CD8+ T cells were 1:1 mixed and intravenously injected into mice (CD45.1+) with B16-OVA tumor. Flow cytometric analysis shows the frequencies of Cnr2GFP and Cnr2CKO OT-I T cells from TILs in tumors (left). The representative ratio of wild-type to Cnr2CKO OT-I T cells in the tumor were evaluated on day 5 (right). b Flow cytometry analysis of the production of IFN‐γ in Cnr2GFP and Cnr2CKO OT-I T cells isolated from tumors stimulated with PMA and Ionomycin in vitro. Data are representative of three independent experiments. Statistical significance was assessed by two-tailed unpaired Student’s t-test, mean ± SD. *P < 0.05 and **P < 0.01. c Mice bearing B16-OVA tumors were treated with 1 × 106 Cnr2GFP or Cnr2CKO OT-I T cells. PBS was used as control. Tumor volume was measured every other day (mean ± SEM, **P < 0.01) (Left). The survival curves of the three groups were compared (right). Statistical significance was assessed by the two-way ANOVA (left), or log-rank (Mantel–Cox) test of survival curve (right). d Mice bearing B16-OVA tumors were treated with 1 × 106 Cnr2GFP, Cnr2CKO OT-I T cells, Cnr2GFP OT-I T cells plus THC and Cnr2CKO OT-I T cells plus THC. Tumor volume were measured every other day (mean ± SEM, **P < 0.01) (left). The survival curves of the three groups were compared (right). Statistical significance was assessed by the two-way ANOVA (left), or log-rank (Mantel–Cox) test of survival curve (right). e T cells were transduced for 48 h by lentivirus and positive cells were sorted by flow cytometry. The expression of CNR2 was validated by qPCR and Western blotting. f Mice bearing B16-OVA tumors were treated with 1 × 106 shLacZ or shCnr2 OT-I T cells. PBS was used as control. Tumor volume was measured every other day (mean ± SEM, **P < 0.01). The survival curves of the three groups were compared. Statistical significance was assessed by the two-way ANOVA, or log-rank (Mantel–Cox) test of survival curve. g The frequencies of OT-I T cells in tumors were shown. h Flow cytometry analysis of the production of IFN-γ in shLacZ and shCnr2 OT-I T cells isolated from tumors stimulated with PMA and Ionomycin in vitro. Statistical significance was assessed by two-tailed unpaired Student’s t-test, mean ± SD, **P < 0.01.
