Efferocytosis produces a prometastatic landscape during postpartum mammary gland involution
Jamie C. Stanford, Christian Young, Donna Hicks, Philip Owens, Andrew Williams, David B. Vaught, Meghan M. Morrison, Jiyeon Lim, Michelle Williams, Dana M. Brantley-Sieders, Justin M. Balko, Debra Tonetti, H. Shelton Earp III, Rebecca S. Cook
Jamie C. Stanford, Christian Young, Donna Hicks, Philip Owens, Andrew Williams, David B. Vaught, Meghan M. Morrison, Jiyeon Lim, Michelle Williams, Dana M. Brantley-Sieders, Justin M. Balko, Debra Tonetti, H. Shelton Earp III, Rebecca S. Cook
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Research Article Oncology

Efferocytosis produces a prometastatic landscape during postpartum mammary gland involution

Abstract

Breast cancers that occur in women 2–5 years postpartum are more frequently diagnosed at metastatic stages and correlate with poorer outcomes compared with breast cancers diagnosed in young, premenopausal women. The molecular mechanisms underlying the malignant severity associated with postpartum breast cancers (ppBCs) are unclear but relate to stromal wound-healing events during postpartum involution, a dynamic process characterized by widespread cell death in milk-producing mammary epithelial cells (MECs). Using both spontaneous and allografted mammary tumors in fully immune–competent mice, we discovered that postpartum involution increases mammary tumor metastasis. Cell death was widespread, not only occurring in MECs but also in tumor epithelium. Dying tumor cells were cleared through receptor tyrosine kinase MerTK–dependent efferocytosis, which robustly induced the transcription of genes encoding wound-healing cytokines, including IL-4, IL-10, IL-13, and TGF-β. Animals lacking MerTK and animals treated with a MerTK inhibitor exhibited impaired efferocytosis in postpartum tumors, a reduction of M2-like macrophages but no change in total macrophage levels, decreased TGF-β expression, and a reduction of postpartum tumor metastasis that was similar to the metastasis frequencies observed in nulliparous mice. Moreover, TGF-β blockade reduced postpartum tumor metastasis. These data suggest that widespread cell death during postpartum involution triggers efferocytosis-induced wound-healing cytokines in the tumor microenvironment that promote metastatic tumor progression.

Authors

Jamie C. Stanford, Christian Young, Donna Hicks, Philip Owens, Andrew Williams, David B. Vaught, Meghan M. Morrison, Jiyeon Lim, Michelle Williams, Dana M. Brantley-Sieders, Justin M. Balko, Debra Tonetti, H. Shelton Earp III, Rebecca S. Cook

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Figure 4

Increased expression of Th2-like cytokines in postpartum tumors requires MerTK.

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Increased expression of Th2-like cytokines in postpartum tumors requires...
(A) A publicly available qRT-PCR dataset of RNA expression levels in normal breast biopsies harvested from premenopausal nulliparous women and premenopausal women 0–2 years after a full-term pregnancy (27) was queried for relative expression levels of the indicated transcripts, which were normalized to dataset housekeeping genes. P values were calculated by Student’s t test. n = 4–20 per group. (B) Cytokine antibody array was performed using lysates from spontaneous MMTV PyVmT tumors harvested from 71-day-old mice (n = 4 tumors per group). Individual cytokine spots are shown from identical exposures. (C) Whole-tumor RNA from 71-day-old virgin and postpartum (Inv d7) mice was assessed by qRT-PCR for Il10, Il4, and Tgfb1. Values are the average relative transcripts levels ± SD. n = 4 tumors analyzed in 6 replicate experiments. *P < 0.05 by Student’s t test. (D) Cytokine antibody array was performed using lysates from MerTK+/+ PyVmT and MerTK–/– PyVmT postpartum tumors (Inv d7). n = 4. (E) Densitometry-based quantitation of relative cytokine levels in whole-tumor lysates. Selected cytokines of interest are shown. Values are the average ± SD. n = 4. P values were calculated by Student’s t test. (F) RNA isolated from whole tumors collected from 71-day-old parous MerTK+/+ PyVmT and MerTK–/– PyVmT mice at Inv d7 was assessed by qRT-PCR to quantify Il10, Il4, and Tgfb1. Values shown are the average ± SD. n = 4. *P < 0.05; ****P < 0.0001 by Student’s t test. (G) RNA isolated from whole tumors from parous mice treated from Inv d0 to Inv d7 with or without BMS-777607 was assessed by qRT-PCR to quantify Il10, Il4, and Tgfb1. Values shown are the average ± SD. n = 4. *P < 0.05; **P < 0.01 by Student’s t test.