ASB2α is expressed in immature DCs and is down-regulated on their activation. (A) Autoradiogram of ASB2 mRNA expression in mouse BMDCs, spleen, BM, and C2C12 myoblasts as a negative control. Northern blot was performed with 4 μg of total RNA. Arbp was used for assessment of RNA quantities in each lane. (B) Relative expression of ASB2 mRNAs in BM, spleen, and BMDCs. Quantitative real-time RT-PCR was carried out with primers specific to ASB2α, ASB2β, or common to both isoforms. (C) BMDCs and spleen-derived DCs were obtained as described in the Materials and methods section. Relative expression of ASB2α mRNAs in BM cells or in spleen cells induced to differentiate into DCs (left panel). Quantitative real-time RT-PCR was carried out with ASB2α-specific primers. Cells were stained with antibodies to CD11c for flow cytometry as assessment of differentiation (right panel). (D) Relative expression of ASB2α mRNAs during LPS-induced BMDC activation (left panel). Immature BMDCs were treated with LPS as indicated. Quantitative real-time RT-PCR was carried out with ASB2α-specific primers. BMDC activation was monitored by flow cytometry with antibodies directed against CD11c and CD86 (right panel). In C and D, data show means and standard error of the mean (SEM) of at least 3 independent experiments.