Dendritic cells mature after they phagocytose M. tuberculosis. A. Human monocytes were separated from buffy coats by plastic adherence and cultured for 6 days in the presence of recombinant human IL-4 (40 ng/ml) and GM-CSF (50 ng/ml) to allow differentiation to DCs. Cells were analysed for CD14 and DC-SIGN expression by flow cytometry. DCs were CD14- and DC-SIGN+ (typically > 85% of gated cells; both before and after infection with Mtb). Plots show uninfected, immature DCs after 6 days of cytokine treatment from 1 representative donor of 3.. B. DCs were infected with live H37Ra at MOI 1 for 24 h and visualised by light microscopy. C. DCs were infected with live Mtb H37Rv at MOI 10 overnight. Bacteria were stained with auramine and nuclei with Hoechst and were visualised by confocal microscopy. Similar results were obtained with iH37Rv, live H37Ra and streptomycin-killed H37Ra (data not shown). D. DCs were infected with live Mtb H37Ra or streptomycin-killed H37Ra at MOI 1 for 48 h. Surface expression of CD83 and CD86 was assessed by flow cytometry. The histograms show 1 representative donor of 3.
Ryan et al. BMC Microbiology 2011 11:237 doi:10.1186/1471-2180-11-237