Fig. 2

Drug-MPs increase ROS-dependent lysosomal pH. a IL-4 conditioned BMDMs were incubated with PKH26-labeled MTX-MPs and analyzed with early-endosomes, late-endosomes or lysosomes under a two-photon confocal microscope. b IL-4 conditioned BMDMs were treated with PKH26-labeled MTX-MPs in the presence or absence of amiloride hydrochloride (Ami, 75 μM) and the uptake of MTX-MPs was determined by flow cytometry. c IL-4 conditioned BMDMs were treated with Dox or Dox-MPs for 8 h and then the red-fluorescent Dox and lysosomes (Green) were observed under a two-photon confocal microscope. d, e IL-4 conditioned BMDMs were treated with T-MPs, MTX or MTX-MPs for 8 h. LysoSensor Green labeled acidic lysosomes were observed under a fluorescence microscope (d), pH value of lysosomes was detected by a microplate reader (e). f IL-4 conditioned BMDMs were treated with MTX or MTX-MPs and the MFI of LysoSensor Green was detected at different time points by flow cytometry. g IL-4 conditioned BMDMs were treated with MTX or MTX-MPs for 8 h and ROS level was detected by flow cytometry. h-i IL-4 conditioned BMDMs were pretreated with NAC (20 mM) for 1 h and treated with MTX or MTX-MPs. The MFI of LysoSensor was detected by flow cytometry (h), Nos2, Tnf and Il6 expression was determined by real-time PCR (i). All scale bars, 10 μm. Unless otherwise specified, n = 3 biologically independent experiments were performed. Data are presented as mean ± SEM. P values were calculated using one-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001