Combined defects in cell proliferation and survival and aberrant expression of cell cycle inhibitor genes in Dicer-deficient GC B cells. (A) Flow cytometric analysis of BrdU incorporation and CaspGLOW stainings in splenic GC B cells after immunization with NP₃₈-CGG. A representative FACS profile and gating strategy for the detection of BrdU⁺ and CaspGLOW⁺ cells among CD19⁺Fas⁺ Dump⁻NIP⁻ and CD19⁺Fas⁺Dump⁻NIP⁺ cells from Dicer1^+/+Aicda^Cre/+ mice at day 10 after immunization are shown. Mice were pulsed with BrdU (2 mg) 4 hours before analysis. Numbers adjacent to the outlined area indicate the percentage of cells in the individual gate. Data are representative of more than 10 experiments. (B-C) The frequencies of splenic B cells that are BrdU⁺ (B) or CaspGLOW⁺ (C) among splenic CD19⁺Fas⁺Dump⁻NIP⁻ and CD19⁺Fas⁺Dump⁻NIP⁺ cells from Dicer1^+/+Aicda^Cre/+ (●) and Dicer1^fl/flAicda^Cre/+ (○) mice 10 days after immunization are quantified. Each symbol represents 1 mouse. (D) Real-time RT-PCR analyses of various CDK inhibitor gene expression in Dicer1^+/+Aicda^Cre/+ (filled) and Dicer1^fl/flAicda^Cre/+ (empty) GC B cells (CD19⁺Fas⁺PNA⁺CD38⁻). Bars represent relative expression of genes normalized to the expression of hprt (mean values of 3 independently sorted GC B-cell samples). **P < .01 (Student t test).