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. 2019 Feb 11;120(4):453–465. doi: 10.1038/s41416-019-0382-0

Fig. 4.

Fig. 4

Correlation between EDP-stimulated Hsp90 secretion and EDP-induced membrane blebbing. a Different cell types were treated with EDPs and extracellular Hsp90 was analysed by western blot. Actin level attested the equal amount of protein loaded in the control and EDP. Data from one experiment, representative of three independent experiments, are shown. b Quantitative evaluation of Hsp90 secretion by different cell types. c Quantification of the blebbing cell number was evaluated by counting 10 fields per well under a phase contrast optical microscope after EDP stimulation for 40 min. Data from one experiment, representative of three independent experiments, are shown. d Correlation between Δblebbing and ΔHsp90 secretion in different cell types. Δblebbing = EDP-dependent blebbing – unstimulated blebbing. ΔHsp90 = EDP-dependent Hsp90 secretion – unstimulated Hsp90 secretion. Linear regression test was performed for each analysis and the R and p values are indicated below the graph. e Blebbing inhibition in HT-1080 cells in the presence of blebbistatin (50 µM), evaluated by counting 10 fields per well under a phase contrast optical microscope after EDP stimulation for 40 min. Data from one experiment, representative of three independent experiments, are shown. f HT-1080 cells were pretreated with blebbistatin (50 µM) for 1 h then stimulated by EDPs. Cell culture media were analysed for Hsp90, MMP-2, MMP-9 and uPA secretions (24 h of treatment) by western bot, gelatin zymography and gelatin-plasminogen zymography. g Blebbing inhibition in HT-1080 cells in the presence of Y27632 (25 µM), evaluated by counting 10 fields per well under a phase contrast optical microscope after EDP stimulation for 40 min. Data from one experiment, representative of three independent experiments, are shown. h HT-1080 cells were pretreated with Y27632 (25 µM) for 1 h then stimulated by EDPs. Cell culture media were analysed for Hsp90, MMP-2, MMP-9 and uPA secretions (24 h of treatment) by western bot, gelatin zymography and gelatin-plasminogen zymography