Cleavage of negatively charged sialic acids impairs macrophage galvanotaxis.

(A) Representative confocal images of surface MAL-2 (green) of BMDMs incubated with 0 (Ctrl) or 100 mU ml−1 Neu’se for 30 min. Nuclei were counterstained with DAPI (blue). Bar, 20 μm. (B) Representative flow cytograms and (C) independent data of standardized MAL-2 fluorescence (green) intensity of BMDMs incubated with 0 or 100 mU ml−1 Neu’se for 30 min. *P t test. (D) Zeta potential of BMDMs incubated with 0 or 100 mU ml−1 Neu’se for 30 min. Data were quantified from a representative of 3 independent experiments. *P t test. (E) Directedness, (F) rose plots, and (G) polarity of BMDMs treated with or without Neu’se, followed by 3 h exposure to an EF of 4 V cm−1. Data were quantified from a representative of 4 independent experiments. ***P t test (Panel E). ***P 2 test (Panel G) (See S1 Data). (H) Proposed model of Salmonella-infection–dependent directional switch in macrophage galvanotaxis. Upon exposure to an IGEF-like EF (marked gradient), negatively charged (sign with small green circles) macrophages undergo robust directional migration to the anode. Phagocytosing Salmonella reduces surface negativity of the macrophage through catalytic activities of certain glycosidases, as exemplified by the cleavage of sialic acids with Neu’se (scissors). Consequently, activated macrophages can either undergo defective directional migration or switch direction to the cathode. Macrophages that phagocytosed beads through binding of nonsialylated surface components (brown squares) still migrate to the anode. Arrows indicate both direction and strength of macrophage galvanotaxis. BMDM, bone marrow-derived macrophage; cosθ, cosine theta; Ctrl, control; EF, electric field; IGEF, infection-generated electric field; MAL-2, Maackia Amurensis lectin II; Neu’se, neuraminidase; nMFI, normalized mean fluorescence intensity; ns, nonsignificant.