Fig. 5.

LPS-induced accumulation of SHP-2 in lipid rafts is impaired in BTLA^−/− DCs. (A) BTLA^−/− BMDCs and WT BMDCs were stimulated with LPS (100 ng/mL) for indicated time periods. Whole-cell lysates were subjected to immunoblotting with antibodies against phosphorylated SHP-1 (p-SHP-1), SHP-1, phosphorylated SHP-2 (p-SHP-2), and SHP-2. Shown are representative blots of three independent experiments. (B) BTLA^−/− BMDCs and WT BMDCs were stimulated with LPS (1 μg/mL) for 30 min. Cell lysates were fractionated by sucrose density gradient centrifugation, and immunoblotting with anti-SHP-1 antibody and anti-SHP-2 antibody was performed. Lipid raft fraction containing glycosphingolipid GM1 was labeled with HRP-conjugated cholera toxin B subunit. (C and D) BTLA^−/− BMDCs and WT BMDCs were treated with SSG (C), NSC87877 (D), or vehicle for 30 min and then stimulated with LPS (100 μg/mL) for 12 h. The levels of TNF-α and IL-12 in the supernatants were measured by ELISA. Data are mean ± SD, n = 4. **P < 0.01.