Reconstitution of PDE4B2 expression prevents cAMP-induced apoptosis. (A) Expression of PDE4B2-WT (wild type) and PDE4B2-PI (phosphodiesterase inactive) in retrovirally infected DHL6 cells. The expression of the indicated Flag-tagged constructs was analyzed with an anti-Flag immunoblot. Size markers are indicated on the left in kilodaltons. (B) cAMP induction in DHL6-eGFP, DHL6-PDE4B2-WT, and DHL6-PDE4B2-PI cells following forskolin treatment. The indicated cells were incubated with 40 μM forskolin for 1 hour, and intracellular cAMP levels were analyzed by ELISA. (C) Growth inhibition following forskolin treatment. DHL6-eGFP, DHL6-PDE4B2-WT, and DHL6-PDE4B2-PI cells were incubated with 40 μM forskolin for 24 hours. Data represent the ratio of proliferation of treated cells to cells cultured with vehicle alone for the same time period. Cell proliferation was measured by MTS assay. (D) Cellular apoptosis in DHL6-eGFP, DHL6-PDE4B2-WT, and DHL6-PDE4B2-PI cells treated with forskolin. The indicated cells were incubated with 40 μM forskolin for 48 hours and analyzed by propidium iodide staining. ^*P < .05 when comparing rate of apoptosis following forskolin treatment in all 3 cell lines. Data in panels B-D represent the mean and SD of 3 independent experiments.