Figure 7

Differential response to Y. pestis infection in human dendritic cells correlates with naturally-expressed c-KIT levels. (A) Differential expression of c-KIT in human dendritic cells. NHDCs (20,000) from seven different donors (D1-7) were cultured in LGM-3 for 4 days. Both adherent and suspension cells were collected, fixed, labeled with (PE)-conjugated c-KIT (Ab81) antibody, and subjected to flow cytometry analysis. 10,000 cells were acquired to generate histograms and a bar graph (B) that depict fluorescence intensity distribution and mean channel fluorescence intensity. The control sample (C) was generated from a pool of unlabeled NHDC from the seven donors. (C) NHDCs that express high levels of c-KIT exhibit increased inhibition of TNF-α release upon Y. pestis infection. NHDCs from seven donors were cultured in LGM-3 for 4 days prior to treatment. Cells from a single donor were plated in 6 replicates (in a 24-well cluster dish): 2 wells were treated with LPS (E. coli 055:B5, 5 μg/ml) and 4 wells received Y. pestis Ind195 at MOI 20. The inhibition of TNF-α production by Y. pestis-infected cells was determined relative to LPS-treated cells for each donor. The data presented was generated from an average of four replicates of Y. pestis-infected cells versus the average of two replicates treated with LPS. The ELISA for each experimental sample was performed in triplicate.