Fig 1.
A drawing of the electrotactic chip assembly.
The electrotactic chip had connecting holes for the medium inlet and outlet and the agar salt bridges. Cells were cultured in the cell culture region. The width, length, and thickness of the cell culture region were 3 mm, 45 mm, and 70 μm, respectively.
Fig 2.
Experimental scheme showing the electrical stimulation during differentiation.
(A) The configuration of the entire system for the electrical field stimulation study. (B) Experimental timeline of the electrical stimulation. D3, D7, and D14 denote 3, 7, and 14 d, respectively, of in vitro (DIV) culturing after seeding.
Fig 3.
Morphologic characteristics of mouse neural stem and progenitor cells (mNPCs).
Morphology of mNPCs without EF stimulation (A and C) and after stimulation (B and D) by DC pulses for 48 h. (A and B) Low-magnification view (×4) and (C and D) high-magnification view (×20). Scale bars: 200 μm (A and B) and 30 μm (C and D).
Fig 4.
Cell morphology in the control and electrical field (EF) groups after 3, 7, and 14 d in vitro (DIV).
Morphology of mNPCs without EF stimulation (A–C) and after stimulation (D–F) by the DC pulses. The images were taken at 3 DIV (A and D), 7 DIV (B and E), and 14 DIV (C and F). Low-magnification view (×4). Scale bars: 120 μm.
Fig 5.
Effect of electrical fields (EFs) on mouse neural stem and progenitor cell (mNPC) viability.
(A) The cells were labeled with SYTOX® (green) and Hoechst 33342 (blue). (a–f) No EF stimulation. (g–l) With DC pulses. Scale bars: 50 μm. (B) Cell viability of the control and EF groups after 3, 7, and 14 d in vitro.
Fig 6.
Immunofluorescence characterization of mouse neural stem and progenitor cells (mNPCs).
The cells were labeled with nestin (red), Sox2 (green), and Hoechst 33342 (blue). (A–I) Without EF stimulation. (J–R) With square-wave DC pulses. Scale bars: 75 μm.
Fig 7.
The expression of nestin and Sox2 without and with DC pulse stimulation.
The percentage of immunostained cells with (A) nestin and (B) Sox2 for mouse neural stem and progenitor cells (mNPCs) cultured with (electrical field [EF]) and without (control [CTL]) square-wave DC pulse stimulation. The P values of Sox2 expression at 3 and 14 DIV for the CTL and EF groups were 0.080 and 0.071, respectively.
Fig 8.
Fluorescence microscopy images of immunostained mouse neural stem and progenitor cells (mNPCs) labeled with glial fibrillary acidic protein (red), Tuj1 (green), and Hoechst 33342 (blue).
(A–I) mNPCs without electrical field (EF) stimulation. (J–R) mNPCs subjected to square-wave DC pulses. Scale bars: 75 μm.
Fig 9.
Fluorescence microscopy images of immunostained cells labeled with O4 (red), Tuj-1 (green), and Hoechst 33342 (blue).
(A–I) mouse neural stem and progenitor cells (mNPCs) without electrical field stimulation. (J–R) mNPCs subjected to square-wave DC pulses. Scale bars: 75 μm.
Fig 10.
Percentage of immunostained cells with Tuj1 (neurons), glial fibrillary acidic protein (astrocytes), and O4 (oligodendrocytes).
Differentiation of mouse neural stem and progenitor cells (A–C) in control conditions and (D–F) with electrical field stimulation after 3 (A and D), 7 (B and E), and 14 (C and F) d in vitro, respectively.