Relief of tumor hypoxia unleashes the tumoricidal potential of neutrophils
Karim Mahiddine, Adam Blaisdell, Stephany Ma, Amandine Créquer-Grandhomme, Clifford A. Lowell, Adrian Erlebacher
Karim Mahiddine, Adam Blaisdell, Stephany Ma, Amandine Créquer-Grandhomme, Clifford A. Lowell, Adrian Erlebacher
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Research Article Immunology Oncology

Relief of tumor hypoxia unleashes the tumoricidal potential of neutrophils

Abstract

Polymorphonuclear neutrophils (PMNs) are increasingly recognized to influence solid tumor development, but why their effects are so context dependent and even frequently divergent remains poorly understood. Using an autochthonous mouse model of uterine cancer and the administration of respiratory hyperoxia as a means to improve tumor oxygenation, we provide in vivo evidence that hypoxia is a potent determinant of tumor-associated PMN phenotypes and direct PMN–tumor cell interactions. Upon relief of tumor hypoxia, PMNs were recruited less intensely to the tumor-bearing uterus, but the recruited cells much more effectively killed tumor cells, an activity our data moreover suggested was mediated via their production of NADPH oxidase–derived reactive oxygen species and MMP-9. Simultaneously, their ability to promote tumor cell proliferation, which appeared to be mediated via their production of neutrophil elastase, was rendered less effective. Relieving tumor hypoxia thus greatly improved net PMN-dependent tumor control, leading to a massive reduction in tumor burden. Remarkably, this outcome was T cell independent. Together, these findings identify key hypoxia-regulated molecular mechanisms through which PMNs directly induce tumor cell death and proliferation in vivo and suggest that the contrasting properties of PMNs in different tumor settings may in part reflect the effects of hypoxia on direct PMN–tumor cell interactions.

Authors

Karim Mahiddine, Adam Blaisdell, Stephany Ma, Amandine Créquer-Grandhomme, Clifford A. Lowell, Adrian Erlebacher

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Figure 1

Improved tumor oxygenation reduces PMN recruitment to PRPL tumors.

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Improved tumor oxygenation reduces PMN recruitment to PRPL tumors. Mice ...
Mice were housed in either ambient O2 (normoxia conditions) or 60% O2 (hyperoxia conditions) for the last 10 days prior to sacrifice on P28. (A and B) Uterine PMN densities and blood PMN concentrations in PRPL mice, as determined by flow cytometry. Uteri were enzymatically disaggregated prior to analysis. PMNs were identified as CD45+Ly6Ghi cells (see Supplemental Figure 1D for gating). (C) Quantification of CXCL5 expression by tumor cells. The cross-sectional area of tumor cells expressing CXCL5 was determined by immunofluorescence staining of uterine tissue sections and normalized to the cross-sectional area of all tumor cells, as identified by cytokeratin 8 (CK8) costaining. Supplemental Figure 2, A–D, shows representative images. (D) Quantification of nuclear HIF-1α, phospho-STAT3 (p-STAT3), and NF-κB p65 expression by tumor cells in PRPL-Csf3r–/– mice, as determined by immunofluorescence staining. The area of positive staining overlying tumor cell nuclei was normalized to the total tumor cell nuclear area per section. (E and F) Representative HIF-1α/CK8–stained sections of PPRL-Csf3r–/– mice, with closeups (lower panels) (n = 6 mice/group; DAPI counterstain). The staining in the myometrium (m) appeared artifactual as it was not cell associated. Graphs also show the mean ± SEM. *P < 0.05; **P < 0.01 by 2-tailed Mann-Whitney U test. NS, not significant.