Fig. 7: LPS stimulation increases membrane ruffling and macropinocytosis.

Macrophages were treated with 100 ng ml⁻¹ LPS for 24 h prior to imaging. a Surface rendering of Mem-mNG on an LPS-stimulated macrophage provides a surface-level understanding of the membrane, exploration, ruffling, and PM structure (region 68 × 69 × 16 µm). b Dual-color volumetric intensity projections of Mem-mNG and AktPH-mSc for an LPS-stimulated cell provided the intensity activity during increased macrophage activity and shows the highly AktPH-rich regions of membrane ruffling (region 68 × 69 × 16 µm). c Untreated macrophage isosurface displaying less exploratory behavior (region 68 × 64 × 16 µm). d Dual-color volumetric intensity rendering of the untreated macrophage gives insight on the AktPH activity inside of the cell during macropinocytosis and allows for the quantitative comparison of macropinosomes formed between the stimulated and unstimulated cells (Region 68 × 64 × 16 µm). e) Differences in macropinocytic activity between untreated and LPS-treated macrophages. Box plot showing median (green line), IQR (blue box), range (dashed line) without outlier (asterisk), n = 10 biologically independent control cells examined over 7 independent coverslip experiments (blue circles), and n = 10 biologically independent LPS-stimulated cells examined over 5 independent coverslip experiments (red circles). All macropinosomes >1 µm were manually counted using a z-projection MIP in Fiji and were distinguished by the post closure spike in AktPH-mSc intensity.