Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Cellular and Molecular Biology

Cancer-associated fibroblasts mediate cancer progression and remodel the tumouroid stroma

Abstract

Background

Cancer-associated fibroblasts (CAFs) are highly differentiated and heterogeneous cancer-stromal cells that promote tumour growth, angiogenesis and matrix remodelling.

Methods

We utilised an adapted version of a previously developed 3D in vitro model of colorectal cancer, composed of a cancer mass and the surrounding stromal compartment. We compared cancer invasion with an acellular stromal surround, a “healthy” or normal cellular stroma and a cancerous stroma. For the cancerous stroma, we incorporated six patient-derived CAF samples to study their differential effects on cancer growth, vascular network formation and remodelling.

Results

CAFs enhanced the distance and surface area of the invasive cancer mass whilst inhibiting vascular-like network formation. These processes correlated with the upregulation of hepatocyte growth factor (HGF), metallopeptidase inhibitor 1 (TIMP1) and fibulin-5 (FBLN5). Vascular remodelling of previously formed endothelial structures occurred through the disruption of complex networks, and was associated with the upregulation of vascular endothelial growth factor (VEGFA) and downregulation in vascular endothelial cadherin (VE-Cadherin).

Conclusions

These results support, within a biomimetic 3D, in vitro framework, the direct role of CAFs in promoting cancer invasion, and their key function in driving vasculogenesis and angiogenesis.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Experimental setups.
Fig. 2: Patient-specific CAF tissue sample characterisation.
Fig. 3: Average invasion into an acellular or healthy cellular stroma.
Fig. 4: Invasion into the cancerous CAF stroma and subsequent gene upregulation.
Fig. 5: Endothelial structures formed within the cancerous CAF stroma.
Fig. 6: Disruption of a mature endothelial network caused by the addition of CAFs.

References

  1. 1.

    Chang, C. H., Qiu, J., O’Sullivan, D., Buck, M. D., Noguchi, T., Curtis, J. D. et al. Metabolic competition in the tumor microenvironment is a driver of cancer progression. Cell 162, 1229–1241 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Shiga, K., Hara, M., Nagasaki, T., Sato, T. & Takahashi, H. Cancer-associated fibroblasts: their characteristics and their roles in tumor growth. Cancers 7, 2443–2458 (2015).

    PubMed  PubMed Central  Google Scholar 

  3. 3.

    Augsten, M. Cancer-associated fibroblasts as another polarized cell type of the tumor microenvironment. Front. Oncol. 4, 1–34 (2014).

    Google Scholar 

  4. 4.

    Tommelein, J., Verset, L., Boterberg, T., Demetter, P., Bracke, M. & De Wever, O. Cancer-associated fibroblasts connect metastasis-promoting communication in colorectal cancer. Front. Oncol. 5, 1–11 (2015).

    Google Scholar 

  5. 5.

    Attieh, Y. & Vignjevic, D. M. The hallmarks of CAFs in cancer invasion. Eur. J. Cell Biol. 95, 493–502 (2016).

    CAS  PubMed  Google Scholar 

  6. 6.

    Kalluri, R. The biology and function of fibroblasts in cancer. Nat. Publ. Gr. 16, 582–598 (2016).

    CAS  Google Scholar 

  7. 7.

    Reid, S. E., Kay, E. J., Neilson, L. J., Henze, A., Serneels, J., McGhee, E. J. et al. Tumor matrix stiffness promotes metastatic cancer cell interaction with the endothelium. EMBO J. 36, 2373–2389 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Mantovani, A., Marchesi, F., Malesci, A., Laghi, L. & Allavena, P. Tumour-associated macrophages as treatment targets in oncology. Nat. Rev. Clin. Oncol. 14, 399–416 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Cirri, P. & Chiarugi, P. Cancer-associated-fibroblasts and tumour cells: a diabolic liaison driving cancer progression. Cancer Metastasis Rev. 31, 195–208 (2012).

    PubMed  Google Scholar 

  10. 10.

    Darby, I. A., Laverdet, B., Bonté, F. & Desmoulière, A. Fibroblasts and myofibroblasts in wound healing. Clin. Cosmet. Investig. Dermatol. 7, 301–311 (2014).

    PubMed  PubMed Central  Google Scholar 

  11. 11.

    Cirri, P. & Chiarugi, P. Cancer associated fibroblasts: the dark side of the coin. Am. J. 1 Cancer Res. 1, 482–497 (2011).

    CAS  Google Scholar 

  12. 12.

    Madsen, C. D., Pedersen, J. T., Venning, F. A., Singh, L. B., Charras, G., Cox, T. R. et al. Hypoxia and loss of PHD 2 inactivate stromal fibroblasts to decrease tumour stiffness and metastasis. EMBO Rep. 16, 1394–1408 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Glentis, A., Oertle, P., Mariani, P., Chikina, A., Marjou, F. El, Attieh, Y. et al. Cancer-associated fibroblasts induce metalloprotease-independent cancer cell invasion of the basement membrane. Nat. Commun. 8, 1–13 (2017).

  14. 14.

    Maller, O., Dufort, C. C. & Weaver, V. M. YAP forces fibroblasts to feel the tension. Nat. Cell Biol. 15, 570–572 (2013).

    CAS  PubMed  Google Scholar 

  15. 15.

    Calvo, F., Ege, N., Grande-Garcia, A., Hooper, S., Jenkins, R. P., Chaudhry, S. I. et al. Mechanotransduction and YAP-dependent matrix remodelling is required for the generation and maintenance of cancer-associated fibroblasts. Nat. Cell Biol. 15, 637–646 (2013).

    CAS  Article  Google Scholar 

  16. 16.

    Cao, H., Xu, E., Liu, H., Wan, L. & Lai, M. Epithelial-mesenchymal transition in colorectal cancer metastasis: a system review. Pathol. Res. Pr. 211, 557–569 (2015).

    CAS  Google Scholar 

  17. 17.

    Kakarla, S., Song, X.-T. & Gottschalk, S. Cancer-associated fibroblasts as targets for immunotherapy. Immunotherapy 4, 1129–1138 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Liu, T., Lin, B. & Qin, J. Carcinoma-associated fibroblasts promoted tumor spheroid invasion on a microfluidic 3D co-culture device. Lab. Chip 10, 1671–1677 (2010).

    CAS  PubMed  Google Scholar 

  19. 19.

    Pape, J., Magdeldin, T., Ali, M., Walsh, C., Lythgoe, M., Emberton, M. et al. Cancer invasion regulates vascular complexity in a three-dimensional biomimetic model. Eur. J. Cancer 119, 179–193 (2019).

    CAS  PubMed  Google Scholar 

  20. 20.

    Magdeldin, T., López-Dávila, V., Pape, J., Cameron, G. W. W., Emberton, M., Loizidou, M. et al. Engineering a vascularised 3D in vitro model of cancer progression. Sci. Rep. 7, 1–9 (2017).

    Google Scholar 

  21. 21.

    Stamati, K., Priestley, J. V., Mudera, V. & Cheema, U. Laminin promotes vascular network formation in 3D in vitro collagen scaffolds by regulating VEGF uptake. Exp. Cell Res. 327, 68–77 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Schindelin, J., Arganda-Carreras, I., Frise, E., Kaynig, V., Longair, M., Pietzsch, T. et al. Fiji: an open-source platform for biological-image analysis. Nat. Methods 9, 676–682 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. 23.

    Rio, D. C., Ares, M., Hannon, G. J. & Nilsen, T. W. Purification of RNA using TRIzol (TRI reagent). Cold Spring Harb. Protoc. 5, 1–4 (2010).

    Google Scholar 

  24. 24.

    Schmittgen, T. D. & Livak, K. J. Analyzing real-time PCR data by the comparative CTmethod. Nat. Protoc. 3, 1101–1108 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Bolander, J., Chai, Y. C., Geris, L., Schrooten, J., Lambrechts, D., Roberts, S. J. et al. Early BMP, Wnt and Ca2+/PKC pathway activation predicts the bone forming capacity of periosteal cells in combination with calcium phosphates. Biomaterials 86, 106–118 (2016).

    CAS  PubMed  Google Scholar 

  26. 26.

    Choi, S. Y., Sung, R., Lee, S. J., Lee, T. G., Kim, N., Yoon, S. M. et al. Podoplanin, α-smooth muscle actin or S100A4 expressing cancer-associated fibroblasts are associated with different prognosis in colorectal cancers. J. Korean Med. Sci. 28, 1293–1301 (2013).

    PubMed  PubMed Central  Google Scholar 

  27. 27.

    Madar, S., Goldstein, I. & Rotter, V. “Cancer associated fibroblasts”—more than meets the eye. Trends Mol. Med. 19, 447–453 (2013).

    CAS  PubMed  Google Scholar 

  28. 28.

    Knüpfer, H. & Preiss, R. Serum interleukin-6 levels in colorectal cancer patients-a summary of published results. Int J. Colorectal Dis. 25, 135–140 (2010).

    PubMed  Google Scholar 

  29. 29.

    Talele, N. P., Fradette, J., Davies, J. E., Kapus, A. & Hinz, B. Expression of α-smooth muscle actin determines the fate of mesenchymal stromal cells. Stem Cell Rep. 4, 1016–1030 (2015).

    CAS  Google Scholar 

  30. 30.

    Hamada, K., Monnai, M., Kawai, K., Nishime, C., Kito, C., Miyazaki, N. et al. Liver metastasis models of colon cancer for evaluation of drug efficacy using NOD/Shi-scid IL2Rgammanull (NOG) mice. Int. J. Oncol. 32, 153–159 (2008).

    CAS  PubMed  Google Scholar 

  31. 31.

    Roudsari, L. C. & West, J. L. Studying the influence of angiogenesis in in vitro cancer model systems. Adv. Drug Deliv. Rev. 97, 250–259 (2016).

    CAS  PubMed  Google Scholar 

  32. 32.

    Vestweber, D. VE-cadherin: The major endothelial adhesion molecule controlling cellular junctions and blood vessel formation. Arterioscler Thromb. Vasc. Biol. 28, 223–232 (2008).

    CAS  PubMed  Google Scholar 

  33. 33.

    Liao, Y., Zhao, H., Liu, Q. & Peng, R. Fibulin-5 inhibits the cell proliferation, migration and angiogenesis in glioma. Int. J. Clin. Exp. Pathol. 9, 8943–8952 (2016).

    CAS  Google Scholar 

  34. 34.

    Sullivan, K. M., Bissonnette, R., Yanagisawa, H., Hussain, S. N. & Davis, E. C. Fibulin-5 functions as an endogenous angiogenesis inhibitor. Lab. Investig. 87, 818–827 (2007).

    CAS  PubMed  Google Scholar 

  35. 35.

    Albig, A. R. & Schiemann, W. P. Fibulin-5 antagonizes vascular endothelial growth factor (VEGF) signaling and angiogenic sprouting by endothelial cells. DNA Cell Biol. 23, 367–379 (2004).

    CAS  PubMed  Google Scholar 

  36. 36.

    Jeanes, A., Gottardi, C. J. & Yap, A. S. Cadherins and cancer: how does cadherin dysfunction promote tumor progression? Oncogene 27, 6920–6929 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Grugan, K. D., Miller, C. G., Yao, Y., Michaylira, C. Z., Ohashi, S., Klein-Szanto, A. J. et al. Fibroblast-secreted hepatocyte growth factor plays a functional role in esophageal squamous cell carcinoma invasion. Proc. Natl Acad. Sci. USA 107, 11026–11031 (2010).

    CAS  PubMed  Google Scholar 

  38. 38.

    Zhang, Y., Tang, H., Cai, J., Zhang, T., Guo, J., Feng, D. et al. Ovarian cancer-associated fibroblasts contribute to epithelial ovarian carcinoma metastasis by promoting angiogenesis, lymphangiogenesis and tumor cell invasion. Cancer Lett. 303, 47–55 (2011).

    CAS  PubMed  Google Scholar 

  39. 39.

    Hwang, R. F., Moore, T., Arumugam, T., Ramachandran, V., Amos, K. D., Rivera, A. et al. Cancer-associated stromal fibroblasts promote pancreatic tumor progression. Cancer Res. 68, 918–926 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Barcus, C. E., Keely, P. J., Eliceiri, K. W. & Schuler, L. A. Stiff collagen matrices increase tumorigenic prolactin signaling in breast cancer cells. J. Biol. Chem. 288, 12722–12732 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Wei, S. C. & Yang, J. Forcing through tumor metastasis: the interplay between tissue rigidity and epithelial-mesenchymal transition. Trends Cell Biol. 26, 111–120 (2016).

    CAS  PubMed  Google Scholar 

  42. 42.

    Tang, D., Gao, J., Wang, S., Ye, N., Chong, Y., Huang, Y. et al. Cancer-associated fibroblasts promote angiogenesis in gastric cancer through galectin-1 expression. Tumor Biol. 37, 1889–1899 (2016).

    CAS  Google Scholar 

  43. 43.

    Hallaq, H. A null mutation of Hhex results in abnormal cardiac development, defective vasculogenesis and elevated Vegfa levels. Development 131, 5197–5209 (2004).

    CAS  PubMed  Google Scholar 

  44. 44.

    Kioi, M., Vogel, H., Schultz, G., Hoffman, R. M., Harsh, G. R. & Brown, J. M. Inhibition of vasculogenesis, but not angiogenesis, prevents the recurrence of glioblastoma after irradiation in mice. J. Clin. Invest. 120, 694–705 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. 45.

    Weis, S., Cui, J., Barnes, L. & Cheresh, D. Endothelial barrier disruption by VEGF-mediated Src activity potentiates tumor cell extravasation and metastasis. J. Cell Biol. 167, 223–229 (2004).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Li, W.-W., Wang, H., Nie, X., Liu, Y., Han, M. & Li, B.-H. Human colorectal cancer cells induce vascular smooth muscle cell apoptosis in an exocrine manner. Oncotarget 8, 62049–62056 (2017).

    PubMed  PubMed Central  Google Scholar 

  47. 47.

    De Francesco, E. M., Lappano, R., Santolla, M. F., Marsico, S., Caruso, A. & Maggiolini, M. HIF-1α/GPER signaling mediates the expression of VEGF induced by hypoxia in breast cancer associated fibroblasts (CAFs). Breast Cancer Res. 15, 1–18 (2013).

    Google Scholar 

  48. 48.

    Alarcón, T., Owen, M. R., Byrne, H. M. & Maini, P. K. Multiscale modelling of tumour growth and therapy: the influence of vessel normalisation on chemotherapy. Comput. Math. Methods Med. 7, 85–119 (2006).

    Google Scholar 

  49. 49.

    Torres, S., Bartolomé, R. A., Mendes, M., Barderas, R., Fernandez-Aceñero, M. J., Peláez-García, A. et al. Proteome profiling of cancer-associated fibroblasts identifies novel proinflammatory signatures and prognostic markers for colorectal cancer. Clin. Cancer Res. 21, 6006–6019 (2013).

    Google Scholar 

  50. 50.

    Ueno, H., Murphy, J., Jass, J. R., Mochizuki, H. & Talbot, I. C. Tumour “budding” as an index to estimate the potential of aggressiveness in rectal cancer. Histopathology 40, 127–132 (2002).

    CAS  PubMed  Google Scholar 

  51. 51.

    Pampaloni, F., Reynaud, E. G. & Stelzer, E. H. K. The third dimension bridges the gap between cell culture and live tissue. Nat. Rev. Mol. Cell Biol. 8, 839–845 (2007).

    CAS  PubMed  Google Scholar 

  52. 52.

    You, W. K. & McDonald, D. M. The hepatocyte growth factor/c-met signaling pathway as a therapeutic target to inhibit angiogenesis. J. Biochem. Mol. Biol. 41, 833–839 (2008).

    CAS  Google Scholar 

Download references

Author information

Affiliations

Authors

Contributions

J.P. planned the work and carried out experiments and subsequent analyses. T.M. conducted preceding optimisation and advised on the interpretation of the data. K.S. and A.N. received the tissue samples, processed and cultivated them in the first instance. M.E., M.L. and U.C. delivered design and guidance for the project. All authors read, provided edits and approved the paper.

Corresponding author

Correspondence to Umber Cheema.

Ethics declarations

Ethics approval and consent to participate

Primary human colorectal cancer-associated fibroblasts were obtained in accordance with the Declaration of Helsinki. Samples were isolated from tumour tissues acquired from surgeries at the Royal Free Hospital. Patients provided informed consent for tissue donation for research under ethics reference 11/WA/0077 obtained through the North Wales Research Ethics Committee (Central and East) through TAPb biobank.

Consent to publish

Not applicable.

Data availability

The data that support the findings of this study are available from the corresponding author (U.C.) on reasonable request.

Competing interests

The authors declare no competing interests.

Funding information

Judith Pape receives a stipend and EU fee funding from the EPSRC as part of the doctoral training programme (DTP). Mark Emberton receives research support from the United Kingdom’s National Institute of Health Research (NIHR) UCLH/UCL Biomedical Research Centre and became an NIHR Senior Investigator in 2015. This work was funded by the NIHR Invention for Innovation (i4i) programme. The views expressed are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health.

Additional information

Note This work is published under the standard license to publish agreement. After 12 months the work will become freely available and the license terms will switch to a Creative Commons Attribution 4.0 International (CC BY 4.0).

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Pape, J., Magdeldin, T., Stamati, K. et al. Cancer-associated fibroblasts mediate cancer progression and remodel the tumouroid stroma. Br J Cancer 123, 1178–1190 (2020). https://doi.org/10.1038/s41416-020-0973-9

Download citation

Further reading

Search

Quick links