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. 2019 Jan 7;38(4):e99430. doi: 10.15252/embj.201899430

Figure EV5. LC3‐NLRP3 colocalization is present in adult microglia as well, but not in neonatal astrocytes.

Figure EV5

  • A
    Adult microglia were isolated from the brains of 8‐month‐old wild‐type and APPPS1 mice with magnetic beads sorting, stained 4 h after plating on coverslips for NLRP3 (green) and LC3 (red) and observed by confocal microscopy. NLRP3 aggregates (arrows) as well as LC3‐NLRP3 colocalization (arrowheads) were found in microglia from both genotypes, confirming that the described in vitro phenotype occurs also in vivo; scale bar: 10 μm.
  • B, C
    Neonatal astrocytes of wild‐type and Becn1 +/− mice do not present NLRP3 upon stimulation with LPS/ATP in Western blot (B), nor in immunocytochemistry (C); scale bar: 25 μm.
  • D, E
    Microglia from Becn1 +/− mice were transfected with a scrambled siRNA or a siRNA for CALCOCO2. 6 d after transfection, cells were stimulated with LPS/ATP. The amount of TNFalpha (D) and IL‐6 (E) in the supernatant was determined by ELISA. Microglia with a CALCOCO2 knockdown show no changes in their TNFalpha or IL‐6 signals compared to cells transfected with a scrambled siRNA; mean ± SEM, n = 3, two‐tailed t‐test ns.