Abstract
Purpose
As one form of tumor invasion, cancer cells can invade the extracellular matrix (ECM) through tracks that have been physically remodeled by cancer-associated fibroblasts (CAFs). However, CAFs are a heterogeneous population with diverse matrix-remodeling capacities. The purpose of this study was to investigate how CAFs with various matrix-remodeling capacities influence cancer cell invasion.
Methods
We established single-cell-derived clones from three primary cultures of CAFs from lung adenocarcinoma patients (Case 1, 5 clones; Case 2, 5 clones; and Case 3, 7 clones). Using a co-culture model, we evaluated the correlations between the number of invaded cancer cells and the remodeling areas generated by CAF clones in each case.
Results
When A549 lung adenocarcinoma cells and CAF clones were co-cultured, both the numbers of invaded cancer cells and the remodeling areas generated by the CAF clones varied greatly. The number of invaded cancer cells was moderately and strongly correlated with the remodeling areas generated by each CAF clone originating from Cases 1 and 2 (R 2 value = 0.53 and 0.68, respectively), suggesting that the remodeling areas in the ECM may determine the number of invaded cancer cells. In contrast, the number of invaded cancer cells was not correlated with the remodeling areas generated by CAF clones originating from Case 3, suggesting that factors other than the remodeling areas might determine the number of invading cancer cells.
Conclusions
These findings showed two types of fibroblast-dependent cancer cell invasion that are dependent on and independent of the remodeling areas generated by CAFs.
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Acknowledgments
S. Neri is a recipient of the Research Resident Fellowship from the Research for Promotion of Cancer Control Programmes (Japan).
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The authors have no conflicts of interest to disclose.
Funding
This work was supported by the National Cancer Center Research and Development Fund (23-A-12 and 23-K-18), the Foundation for the Promotion of Cancer Research, the Third-Term Comprehensive 10-Year Strategy for Cancer Control, the Advanced Research for Medical Products Mining Programme of the National Institute of Biomedical Innovation (NIBIO), and JSPS KAKENHI (24659185).
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All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.
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Informed consent was obtained from all individual participants included in the study.
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432_2015_2046_MOESM1_ESM.pptx
Supplementary Fig. 1. Examples of image analysis using CL-Quant software. Detection of the invasion region using the first frame of the phase-contrast images (upper panel). Detection of the remodeling areas generated by the invaded CAFs in the invasion region using phase-contrast images (middle panel, pink region). Detection of the number of cancer cells in the invasion region using fluorescence images (lower panel, green region). (PPTX 624 kb)
432_2015_2046_MOESM2_ESM.pptx
Supplementary Fig. 2. Relationship between the relative mRNA expressions (/GAPDH) of the CAF clones and the invaded cancer cells in Case 3. R2 = coefficient of determination. (PPTX 107 kb)
432_2015_2046_MOESM3_ESM.wmv
Supplementary Movie 1. Time-lapse imaging of single-cell cloning of CAFs. Representative time-lapse images for 1 week of the single-cell cloning of CAFs expressing Venus in their cytoplasm (green). (WMV 6466 kb)
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Neri, S., Hashimoto, H., Kii, H. et al. Cancer cell invasion driven by extracellular matrix remodeling is dependent on the properties of cancer-associated fibroblasts. J Cancer Res Clin Oncol 142, 437–446 (2016). https://doi.org/10.1007/s00432-015-2046-7
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DOI: https://doi.org/10.1007/s00432-015-2046-7