Figure 3
The uptake of DNA-LL37 complexes by pDCs. (a-f) Representative flow cytograms showing the internalization of Alexa488 DNA-LL37 complexes by pDCs isolated from healthy controls at different time points (5, 15 and 30 minutes). (a) CD123 + BDCA-2+ pDCs, (b) Unstimulated cells, (c) Negative control (PFA treated cells) showing minimal uptake (6.5 ± 0.12%), (d) Uptake at 5 minutes (92.6 ± 0.85%), (e) Uptake at 15 minutes (90.1 ± 0.61%), (f) Uptake at 30 minutes (98.8 ± 0.52%). Uptake of complexes by pDCs occurred at a very fast rate as shown by their saturation just in 30 minutes of treatment with the complexes. Data is presented as mean ± SEM from 3 independent experiments. (g,h) Representative flow cytograms showing the internalization of Alexa488 DNA alone and Alexa488 DNA-LL37 complexes by pDCs of healthy controls. (g) BDCA-2 PE and ILT7 were used to gate pDCs, (h) uptake of Alexa488 DNA alone (92.96 ± 0.87%) and the uptake of Alexa488 DNA-LL37 complexes (97.09 ± 0.86%) (p = 0.01). Data is presented as mean+SEM from 3 independent experiments. (i-l) The representative confocal images showing uptake of Alexa488 DNA-LL37 complexes by pDCs of healthy controls. (i) Alexa488 DNA-LL37 complex (green), (j) Cell membrane marked by BDCA-2 PE (red) and (k) Nucleus (blue). (l) Merged overlay images showing the cytoplasmic location of complexes. All images were acquired at 60X magnification with 3X zoom on selected cells with a bar representing 10 µm on the scale.
