A 3D Biometric Platform to Study Breast Cancer Metabolism and the Tumor Microenvironment
Yoshiko Toyoda, B.A., John P. Morgan, Ph.D., Julia Jin, B.S., Jaime L. Bernstein, B.S., Jason A. Spector, M.D. F.A.C.S..
Weill Cornell Medical College, New York, NY, USA.
Introduction: Obesity is a known risk factor for the development and prognosis of breast cancer (BC), and metastasis to breast adipose tissue results in its transformation from soft tissue, primarily composed of adipocytes, to a solid tumor histologically devoid of adipocytes. Although many recent BC studies have focused on tumor-microenvironment interactions, they have emphasized adipose stromal cells (ASC), with scant attention devoted to the mature adipocytes—despite the fact that in BCs, early local tumor invasion results in immediate proximity of BC cells to adipocytes. Indeed, adipocytes have been identified as a source for exogenous lipids in various other cancer cell types and may similarly provide energy to fuel malignant survival and growth in BC. This relationship is of particular relevance to plastic surgery, as many autologous reconstructions and most prosthetic based reconstructions after oncologic mastectomy achieve optimal aesthetics and maximum durability using autogenous fat transfer (AFT). Despite the increasing ubiquity and promise of this straightforward procedure, many unanswered questions remain, including the safety of AFT in the setting of BC. Although clinical studies to examine this question are underway, an in vitro system is critical for elucidating the complex interplay between the cells that normally reside at the surgical recipient site. In order to study these interactions and characterize possible lipid transfer between breast adipocytes to BC cells, we designed a three dimensional in vitro model using primary patient tissues.
Methods: Breast adipose tissue was acquired from patients undergoing breast reduction surgery. The tissue was enzymatically digested and sorted by differential centrifugation to retrieve adipocytes and ASCs. Polydimethylsiloxane wells were filled with type I 0.3% w/v collagen and seeded with varying concentrations of adipocytes labeled with the fluorescent lipid dye boron-dipyrromethene (BODIPY), ASCs and fluorescently-labeled BC cells in the bulk and on the surface. Cultures of BC cells in non-adipocyte containing collagen matrices served as controls. Lipid transfer and BC cell invasion into the collagen-adipocyte bulk were analyzed using laser scanning confocal microscopy and image analysis.
Results: Fluorescent and confocal microscopy revealed a dense culture of native adipocytes containing fluorescent lipid droplets in the 3D collagen culture platform. Lipid transfer from adipocytes to BC cells was evaluated by characterizing the presence of BODIPY positive lipid droplets within RFP-labeled BC cells. Metastatic potential was evaluated by comparing BC cell invasion into adipocyte-containing hydrogels versus those lacking adipocytes.
Conclusion: We have established a 3D platform to study BC-adipose tissue interactions, including lipid transfer from primary human breast adipocytes to BC cells and tumor invasion potential in the presence of adipocytes and ASCs. Transfer of fluorescently-labeled lipids directly from adipocytes to BC cells can indicate aberrant metabolism to fuel malignant growth and adaptive survival, while the presence or absence of ASCs and adipocytes enables analysis of their effect on metastatic progression. Our novel, 3D platform can untangle the complex interplay within the entire breast cancer tumor microenvironment for high-throughput analysis and can elucidate the safety of adipose tissue transfer in breast reconstruction in post-oncologic mastectomy.
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