Fig. 4: TIPE2-deficient macrophages dictate ASC ferroptosis in vitro. | Nature Communications

Fig. 4: TIPE2-deficient macrophages dictate ASC ferroptosis in vitro.

From: Obesity-associated macrophages dictate adipose stem cell ferroptosis and visceral fat dysfunction by propagating mitochondrial fragmentation

Fig. 4

a Schematic of coculture assay for ASCs (wild-type) and macrophages (Tipe2+/+ or Tipe2-/-) in the presence of FA for 24 h. b, c Flow cytometry frequency (b) of lipid ROS in ASCs (CD45-) with MFI quantification (c; n = 4) after coculture with Tipe2+/+ or Tipe2-/- macrophages. d–g Flow cytometry frequency (d, e; n = 3) of ASC death after coculture with Tipe2+/+ or Tipe2-/- macrophages, or in the presence of 100 μM DFO (f, g; n = 3). h–j Representative confocal imaging (h) of intracellular Fe2+ with MFI quantification by ImageJ (i; n ≥ 51 cells examined over two independent experiments) and immunoblots for FTH1 protein (j) in ASCs that were incubated with condition medium (CM) from Tipe2+/+ or Tipe2-/- macrophages for 24 h. Scale bars, 50 μm. k, l Representative immunoblots for FTH1 protein in Tipe2+/+ or Tipe2-/- macrophages (k) and their exosomes (l). m–o Representative immunoblots for FTH1 protein (m) and confocal imaging (n) of intracellular Fe2+ with MFI quantification by ImageJ (o; n ≥ 42 cells examined over two independent experiments) in ASCs that were incubated with 50 μg exosomes from Tipe2+/+ or Tipe2-/- macrophages for 24 h. Scale bars, 50 μm. Data are presented as mean ± s.e.m. **P < 0.01, ***P < 0.001 determined by two-tailed student’s t-test (c, e, i, o) or one-way ANOVA (g). Data are representative of two or three independent experiments.

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