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Ferritin stimulates breast cancer cells through an iron-independent mechanism and is localized within tumor-associated macrophages

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Abstract

Tumor-associated macrophages play a critical role in breast tumor progression; however, it is still unclear what effector molecular mechanisms they employ to impact tumorigenesis. Ferritin is the primary intracellular iron storage protein and is also abundant in circulation. In breast cancer patients, ferritin is detected at higher levels in both serum and tumor lysates, and its increase correlates with poor clinical outcome. In this study, we comprehensively examined the distribution of ferritin in normal and malignant breast tissue at different stages in tumor development. Decreased ferritin expression in cancer cells but increased infiltration of ferritin-rich CD68-positive macrophages was observed with increased tumor histological grade. Interestingly, ferritin stained within the stroma surrounding tumors suggesting local release within the breast. In cell culture, macrophages, but not breast cancer cells, were capable of ferritin secretion, and this secretion was further increased in response to pro-inflammatory cytokines. We next examined the possible functional significance of extracellular ferritin in a breast cancer cell culture model. Ferritin stimulated the proliferation of the epithelial breast cancer cell lines MCF7 and T47D. Moreover, this proliferative effect was independent of the iron content of ferritin and did not increase intracellular iron levels in cancer cells indicating a novel iron-independent function for this protein. Together, these findings suggest that the release of ferritin by infiltrating macrophages in breast tumors may represent an inflammatory effector mechanism by which ferritin directly stimulates tumorigenesis.

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Ethical Standards

The experiments presented in this manuscript comply with the current laws and standards of the USA and have been approved by the institutional review board at Penn State Hershey Medical Center.

Conflict of Interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Neurosurgery, The Pennsylvania State University Hershey Medical Center, Hershey, PA, USA

    Ahmed A. Alkhateeb & James R. Connor

  2. Department of Pathology, The Pennsylvania State University Hershey Medical Center, Hershey, PA, USA

    Bing Han

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  1. Ahmed A. Alkhateeb
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  2. Bing Han
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Correspondence to James R. Connor.

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10549_2012_2405_MOESM1_ESM.tif

Supplementary Fig. 1. a The L-ferritin antibody used in immunohistochemistry specifically recognized recombinant L-ferritin (rLFr) but not recombinant H-ferritin (rHFR). The L-subunit-rich human liver ferritin was used as a positive control. The appearance of the 17 kDa band in liver ferritin has been shown to be due to lysosomal cleavage of the L-ferritin protein [24]. Each lane was loaded with 0.25 μg of protein b Western blot analysis showing no phosphorylation of IKKα/β following exposure to 25nM ferritin extracted from the human liver. c Western blot analysis of total protein lysates from breast cancer cell lines shows that both responsive (i.e. MCF7) and unresponsive (i.e. MDA-MB-231) cell lines express Scara5. The expected molecular weight of Scara5 based on amino acid sequence is 53 kDa. (TIFF 2260 kb)

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Alkhateeb, A.A., Han, B. & Connor, J.R. Ferritin stimulates breast cancer cells through an iron-independent mechanism and is localized within tumor-associated macrophages. Breast Cancer Res Treat 137, 733–744 (2013). https://doi.org/10.1007/s10549-012-2405-x

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