Fig. 5. Inhibition of PLY in native and S.pn.-infected human alveolar epithelial cells.

PLY neutralized by PB-3 in human alveolar epithelial cells (A549 cells). Cells were grown in Ham’s F12 culture medium with inclusion of a red mitochondrial dye (TMRE: tetramethyl-rhodamine, ethyl ester) to indicate healthy cells and a fluorogenic peptide substrate (DEVD) with green fluorescence to highlight activity of caspase 3 and 7 activity in apoptotic cells (data here represent one of n = 3, distinct samples). a (I) Healthy cells as a negative control, 24 h; (II) cells treated with PLY 15 nM, 24 h; (III) cells treated with PLY (15 nM) + PB-3 (6 µM), 24 h; (IV) cells with PB-3 as a control (24 h). b PB-3 blocked cellular deterioration in A549 cells infected with S.pn. D39 strain (data shown is one of n = 3, distinct samples). (I) healthy control cells, (II) cells infected with D39Δcps induced cellular injury (16 h), (III) cells infected with D39Δcps treated with PB-3 (6 µM) 4-times after regular intervals of 4 h (16 h), and (IV) D39ΔcpsΔply control, and it represents that only PLY provokes the cellular injury. All images were recorded by laser confocal microscopy after the mentioned time intervals. Scale bars correspond to 50 µm.