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. 2016 Sep 19;213(10):1973–1981. doi: 10.1084/jem.20151065

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© 2016 Drennan et al.

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Figure 3. — A20 regulates NKT1 and NKT2 sublineage activation and survival. qPCR analysis of A20 mRNA expression in bulk splenic iNKT (A) or NKT sublineage (B) cells after 24 h in vitro treatment with the indicated agents. Data are from two independent experiments. (C and D) CD69 and CXCR3 expression on NKT1^a, NKT1^b, NKT2, and NKT17 cells isolated from thymi of control (n = 6) and CD4-cre;A20^F/F mice (n = 6), respectively. Data are from two independent experiments. (E and F) α-CD3/CD28-induced cytokine production by in vitro–expanded control and A20-deficient NKT1 and NKT2 thymocytes (n = 3). Data are from two independent experiments. (G) FACS analysis of caspase 3 activity in NKT1, 2, and 17 sublineages isolated from thymus and spleen of control (n = 9) and CD4-cre;A20^F/F (n = 9) mice, respectively. Data are pooled from three independent experiments. (H and I) Frequency of control and A20-deficient iNKT cells recovered from CD45.1 congenic mice after in vivo treatment with vehicle (control, n = 16; CD4-cre;A20^F/F, n = 17), Z-VAD (10 mg/kg; CD4-cre;A20^F/F, n = 8), or Necrostatin-1 (10 mg/kg; CD4-cre;A20^F/F, n = 7), respectively. FACS dot blots show CD45.2⁺ iNKT cells isolated from chimeras and represents pooled data from two independent experiments. *, P < 0.05; **, P < 0.005; ***, P < 0.0005. n.s., not significant.