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. 2009 Dec;175(6):2528–2539. doi: 10.2353/ajpath.2009.090147

Figure 5.

Figure 5

Increased microglial phagocytosis after stimulation of SIRPβ1. A: Increased phagocytosis of microsphere beads after stimulation of flag-tagged SIRPβ1 (fSIRPβ1). Primary microglial cells were lentivirally transduced with the flag-tagged SIRPβ1 (fSIRP) or the control GFP vector (GFP) and were cultured for 24 hours on dishes coated with flag-specific antibody (anti-flag) or control antibody (cont. Ab). The percentage of microglia having phagocytosed beads was quantified by flow cytometry. Stimulation of fSIRPβ1 on transduced primary microglia by the flag-specific antibody (anti-flag) increased the percentage of microglia having taken up microsphere beads compared with GFP-transduced microglia (GFP) and the isotype control antibody (Cont. Ab). Treatment of microglia with the actin polymerization inhibitor cytochalasin D (Cyt.) or the mitogen-activated protein kinase inhibitor PD98059 (ERK) prevented the increased phagocytosis induced by stimulation of fSIRPβ1 (no: no treatment). Data are presented as mean ± SEM of n = 3 independent experiments. P < 0.05 analysis of variance, followed by Bonferroni’s multiple comparison test. B: Phagocytosis of Aβ42 after stimulation of fSIRPβ1. Primary microglial cells were transduced with the fSIRPβ1 vector. Cells were cultured on dishes coated with flag-specific antibody and then Aβ42 was added for 6 hours before fixation. Phagocytosis of Aβ42 was analyzed by confocal microscopy. Serial sections along the z-axis were acquired and composed to images. Aβ42 was localized within the microglial cell. Scale bar: 10 μm. C: Increased phagocytosis of Aβ42 and neural debris after fSIRPβ1 stimulation. Primary microglial cells were transduced with the fSIRPβ1 vector and were cultured for 24 hours on dishes coated with flag-specific antibody (anti-flag) or isotype control antibody (Cont. Ab). Biotinylated Aβ42 or apoptotic neuronal material was added. Percentage of primary microglia having phagocytosed Aβ42 or neuronal debris was quantified by confocal microscopy. Stimulation of fSIRPβ1 increased the uptake of Aβ42 and apoptotic neuronal material. Data are presented as mean ± SEM of n = 3 independent experiments. P < 0.05, analysis of variance.

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