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. 2009 Jun 26;297(2):H874–H886. doi: 10.1152/ajpheart.00311.2009

Fig. 8.

Fig. 8.

EMMPRIN stimulates IL-18 expression via PI3K, Akt, and ERK. A: EMMPRIN stimulates PI3K activation. Quiescent SMCs were incubated with function-blocking antibodies or transduced with Ad.mEMMPRIN before EMMPRIN (5 μg/ml for 30 min) addition. Normal IgG and empty virus served as respective controls. PI3K ELISA was performed as described under materials and methods. A, top: immunoblot analysis of the same samples with anti-p85 antibody. *P < 0.001 vs. untreated; †P < 0.05 vs. EMMPRIN (n = 6 experiments). B: EMMPRIN activates Akt. Quiescent SMCs were treated as A but for 1 h, and Akt activation was analyzed by immunoblotting using pAkt antibodies. A representative of 3 independent experiments is shown. C: EMMPRIN stimulates ERK activity. Quiescent SMCs treated as in A, but for 1 h, were analyzed for ERK activity using immunecomplex kinase assays. Elk served as a substrate (n = 3 experiments). α-Tubulin served as a control. D: adenoviral transduction of dnPI3Kp85 blocks EMMPRIN-mediated PI3K activation. SMCs were transduced with Ad.dnPI3K or Ad.GFP before EMMPRIN addition. PI3K ELISA was performed as described under materials and methods. D, top: immunoblot analysis of the same samples with anti-p85 antibody. *P < 0.01 vs. untreated; †P < 0.05 vs. EMMPRIN + Ad.GFP. E: adenoviral transduction of dnPI3K or dnAkt blunts EMMPRIN-mediated Akt activation. SMCs were transduced with Ad.dnPI3Kp85 or dnAkt before EMMPRIN addition. Akt activation was analyzed as in B (n = 3 experiments). F: EMMPRIN stimulates ERK activation via PI3K and Akt. SMCs were either traduced with Ad.dnPI3Kp85 or Ad.dnAkt or treated with PD-98059 (10 μM in DMSO for 1 h) before EMMPRIN addition. ERK activation was analyzed as in C (n = 3 experiments). G: EMMPRIN induces IL-18 expression via PI3K, Akt, and ERK. SMCs were transduced with Ad.nPI3Kp85, dnAkt, or GFP or treated with PD-98059 before EMMPRIN (5 μg/ml) or Fc (1.5 μg/ml) addition for 24 h. IL-18 levels in culture supernatants were analyzed by ELISA. *P < 0.001 vs. untreated; †P < at least 0.05 vs. EMMPRIN + respective controls (n = 6 experiments).