Fig. 5: Intercellular transfer of neutrophilic myeloperoxidase (MPO) is responsible for neutrophil-mediated tumor cell cytotoxicity.
From: Neutrophil-induced ferroptosis promotes tumor necrosis in glioblastoma progression
a MPO immunofluorescent staining of LN229TAZ(4SA) cells cultured alone or with indicated PKH26-labeled neutrophils. Non-diff undifferentiated, diff differentiated, Neu neutrophils, B.M. bone marrow. b, c PKH26 (b) and MPO (c) puncta per cell in a. Each data point represents an average of 10–15 cells in one image field. In b, nalone = 10, nnon-diff = 6, ndiff = 6, nw/TANs = 5, nNeu.Spl = 11, and nNeu.B.M. = 6. In c, nalone = 16, nnon-diff = 30, ndiff = 11, nw/TANs = 13, nNeu.Spl = 27, and nNeu.B.M. = 18. n indicates number of image fields from three independent experiments or animals. Ordinary one-way ANOVA. d Images of colony-formation assays of ZsGreen-expressing LN229TAZ(4SA) cells with many (PKHhigh) or few (PKHlow/-) PKH26 puncta when cocultured with PKH26-labeled differentiated 32Dcl3 cells. e Percent survival of conditions in d normalized to PKHlow/- LN229TAZ(4SA) cells. n = 3. Unpaired two-tailed t-test. f Chromogenic immunodetection of MPO in an endpoint LN229TAZ(4SA) tumor section. The outlined areas are shown on the bottom. g Immunofluorescent staining on an endpoint LN229TAZ(4SA) tumor section. For f and g, three animals were examined independently with similar observations. h Viability (luminescence) of LN229TAZ(4SA) cells cultured alone (n = 5) or with indicated neutrophils, treated with either of two MPO inhibitors, 4-ABAH (2 µM; nTANs = 5 and n32Dcl3 = 5) or PF06282999 (2 µM; nTANs = 3 and n32Dcl3 = 3), or with DMSO (nTANs = 5 and n32Dcl3 = 5). Percent survival normalized to LN229TAZ(4SA) cells cultured alone and treated with the same compound. Ordinary one-way ANOVA. i Viability (evaluated by Sytox-Blue) of LN229TAZ(4SA) cells cultured alone or with indicated neutrophils non-transduced (no-shRNA) or transduced with scrambled shRNA (sh-control) or shRNAs targeting MPO. n = 3. For e, h, and i, n indicates number of independent experiments. Each data point represents an animal or average of replicates from an independent experiment. Ordinary one-way ANOVA. a–i Numerical data presented as mean ± s.e.m. Scale bars in μm. Source data provided as a Source Data file.
