Chronic Active B Cell Receptor Signaling in Diffuse Large B Cell Lymphoma

To determine gene expression of DLBCL upon NF-kB inhibition or knockdown of BCR signaling components. Total RNA (Trizol reagent; Invitrogen, Carlsbad, CA) was prepared from 8 HBL-1 samples following incubation with 25 µmol/L MLN120B (Millennium Pharmaceuticals, Cambridge, MA) for 2, 3, 4, 6, 8, 12, 16, and 24 hours. In addition, 16 HBL-1 samples were infected with retroviral vectors expressing various shRNAs (CARD11, SYK, BTK, CD79A) in a doxycycline-inducible fashion, selected with puromycin, treated with doxycycline for 24 or 48 hr and then harvested for total RNA. Uninduced cultures were prepared in parallel as control samples. Biological replicates were performed for the CARD11, BTK, and CD79A samples.