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. 2023 Jul 24;13(12):4197–4216. doi: 10.7150/thno.84059

Figure 1.

Figure 1

Identification of primary cortical microglia and the impact of hypoxia on microglial polarization. Primary microglia were extracted from neonatal C57BL/6 mice and plated for 24 h before use. Immunofluorescence staining found microglia positive for the expression of CD68 (A), CD11b (B), Iba1 (C), and CX3CR1 (D). DAPI was used as a counterstain for the cell nuclei. (E-F) Light microscopy of p3 passaged primary microglia in culture. Microglia appeared healthy and displayed processes and a ramified morphology. (G) MTT (Thiazolyl Blue Tetrazolium Bromide) was used to test the microglia viability exposed to 2, 4, 6, and 8 h of OGD followed by 24-h reoxygenation. Cells incubated under standard cell culture conditions ('Normoxia') were defined as 100% cell survival (n = 6). (H-I) RT-qPCR was employed to detect the M2 signature genes IL-10 and CD206 on the mRNA levels (n = 4). (J) MTT was used to test the cell viability exposed to 4 h of OGD followed by 24-, 48-, and 72-h reoxygenation. Cells incubated under standard cell culture conditions ('Normoxia') were defined as 100 % cell survival (n = 6). *p < 0.05; **p < 0.01; ****p < 0.0001; P-microglia, primary microglia; NS, not statistically significant; IL, Interleukin; OGD, oxygen-glucose deprivation; RT-qPCR, quantitative real-time PCR analysis; RO, reoxygenation.