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Expression Pattern of the Pro-apoptotic Gene PAR-4 During the Morphogenesis of MCF-10A Human Mammary Epithelial Cells

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Cancer Microenvironment

Abstract

The histological organization of the mammary gland involves a spatial interaction of epithelial and myoepithelial cells with the specialized basement membrane (BM), composed of extra-cellular matrix (ECM) proteins, which is disrupted during the tumorigenic process. The interactions between mammary epithelial cells and ECM components play a major role in mammary gland branching morphogenesis. Critical signals for mammary epithelial cell proliferation, differentiation, and survival are provided by the ECM proteins. Three-dimensional (3D) cell culture was developed to establish a system that simulates several features of the breast epithelium in vivo; 3D cell culture of the spontaneously immortalized cell line, MCF10A, is a well-established model system to study breast epithelial cell biology and morphogenesis. Mammary epithelial cells grown in 3D form spheroids, acquire apicobasal polarization, and form lumens that resemble acini structures, processes that involve cell death. Using this system, we evaluated the expression of the pro-apoptotic gene PAWR (PKC apoptosis WT1 regulator; also named PAR-4, prostate apoptosis response-4) by immunofluorescence and quantitative real time PCR (qPCR). A time-dependent increase in PAR-4 mRNA expression was found during the process of MCF10A acinar morphogenesis. Confocal microscopy analysis also showed that PAR-4 protein was highly expressed in the MCF10A cells inside the acini structure. During the morphogenesis of MCF10A cells in 3D cell culture, the cells within the lumen showed caspase-3 activation, indicating apoptotic activity. PAR-4 was only partially co-expressed with activated caspase-3 on these cells. Our results provide evidence, for the first time, that PAR-4 is differentially expressed during the process of MCF10A acinar morphogenesis.

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Abbreviations

BM:

basement membrane

ECM:

extra-cellular matrix

PAR-4:

prostate apoptosis response-4

qPCR:

quantitative real time PCR

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Acknowledgments

This work was supported by FAPESP—Fundação de Amparo a Pesquisa do Estado de São Paulo (grant number 06/01026-0). This work was also supported in part by CNPq (304949/2006-0). We thank Dr. Ana Lucia Garippo from Rede Multiusuários HC-FMUSP (FAPESP no. 04/08908-2) for the excellent technical assistance in confocal immunofluorescence image capture.

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  1. Disciplina de Oncologia, Departamento de Radiologia da Faculdade de Medicina da, Universidade de São Paulo, Av. Dr. Arnaldo 455, 4 andar, sala 4112, 01246-903, São Paulo, Brazil

    Simone A. de Bessa Garcia, Michelly C. Pereira & Maria A. Nagai

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  1. Simone A. de Bessa Garcia
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  2. Michelly C. Pereira
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  3. Maria A. Nagai
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Correspondence to Maria A. Nagai.

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de Bessa Garcia, S.A., Pereira, M.C. & Nagai, M.A. Expression Pattern of the Pro-apoptotic Gene PAR-4 During the Morphogenesis of MCF-10A Human Mammary Epithelial Cells. Cancer Microenvironment 4, 33–38 (2011). https://doi.org/10.1007/s12307-010-0059-y

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  • DOI: https://doi.org/10.1007/s12307-010-0059-y

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