Population depletion leads to induction of CD40L transcripts in L3055 cells.

CD40L expression was assessed in L3055 (wild-type) cells cultured at various densities by using reverse transcriptase-PCR. Cells (2 × 10⁶) were plated out in increasing volumes of culture medium (10, 20, 40, 80, 160, 320, and one of 200 mL), in the absence or presence of sCD40L as indicated and cultured for 48 hours. Complementary DNAs were synthesized from total RNA isolated from these cultures and subjected to PCR by using primers specific for CD40L (upper panel) or glyceraldehyde phosphate dehydrogenase (lower panel). To increase the sensitivity of the detection, the CD40L PCR products were transferred onto nylon membrane and hybridized with a³²P-labeled CD40L-specific oligo-probe. As a positive control for CD40L expression, RNA from CD40L-transfected mouse L cells was used (upper panel, pos). Data shown are representative of 4 similar experiments.