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Bone morphogenetic protein 4 provides cancer-supportive phenotypes to liver fibroblasts in patients with hepatocellular carcinoma

  • Original Article—Liver, Pancreas, and Biliary Tract
  • Published:
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Abstract

Background

Cancer-associated fibroblasts (CAFs) are essential constituents of cancer-supportive microenvironments. The high incidence of hepatocellular carcinoma (HCC) in advanced fibrosis patients implies that fibroblasts have a promoting effect on HCC development. We aimed to explore the regulators of phenotypes and function of CAFs in the liver.

Methods

We established primary cancer-associated fibroblasts (CAFs) and non-cancerous liver fibroblasts (NFs) from 15 patients who underwent HCC resection. We compared phenotypes, capacity of cytokine/chemokine production and gene expression profiles between pairs of CAFs and NFs from the same donors. We examined resected tissue from additional 50 patients with HCC for immunohistochemical analyses.

Results

The CAFs expressed more ACTA2 and COL1A1 than the NFs, suggesting that CAFs are more activated phenotype. The CAFs produced larger amounts of IL-6, IL-8 and CCL2 than the NFs, which led to invasiveness of HuH7 in vitro. We found that Bone Morphogenetic Protein-4 (BMP4) is up-regulated in CAFs compared to NFs. The CAF phenotype and function were gained by BMP4 over-expression or recombinant BMP4 given to fibroblasts, all of which decreased with BMP4 knockdown. In tissues obtained from the patients, BMP4-positive cells are mainly observed in encapsulated fibrous lesions and HCC. Positive expression of BMP4 in HCC in resected tissues, not in fibroblasts, was associated with poorer postoperative overall survival in patients with HCC.

Conclusion

Endogenous and exogenous BMP4 activate liver fibroblasts to gain capacity of secreting cytokines and enhancing invasiveness of cancer cells in the liver. BMP4 is one of the regulatory factors of CAFs functioning in the microenvironment of HCC.

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Abbreviations

BMP4:

Bone morphogenetic protein-4

CAFs:

Cancer-associated fibroblasts

CM:

Conditioned media

DAAs:

Direct anti-viral agents

HCC:

Hepatocellular carcinoma

HSCs:

Hepatic stellate cells

NFs:

Non-cancerous liver fibroblasts

OS:

Overall survival

References

  1. Forner A, Llovet JM, Bruix J. Hepatocellular carcinoma. Lancet (London, England). 2012;379:1245–55.

    Article  Google Scholar 

  2. Webster DP, Klenerman P, Dusheiko GM. Hepatitis C. Lancet (London, England). 2015;385:1124–35.

    Article  Google Scholar 

  3. Younossi ZM, Otgonsuren M, Henry L, et al. Association of nonalcoholic fatty liver disease (NAFLD) with hepatocellular carcinoma (HCC) in the United States from 2004 to 2009. Hepatology (Baltimore, MD). 2015;62:1723–30.

    Article  CAS  Google Scholar 

  4. El-Serag HB, Kanwal F. Epidemiology of hepatocellular carcinoma in the United States: where are we? Where do we go? Hepatology (Baltimore, MD). 2014;60:1767–75.

    Article  Google Scholar 

  5. El-Serag HB, Kanwal F, Richardson P, et al. Risk of hepatocellular carcinoma after sustained virologic response in veterans with HCV-infection. Hepatology (Baltimore, Md) 2016;64:130–137.

    Article  CAS  Google Scholar 

  6. Iwaisako K, Jiang C, Zhang M, et al. Origin of myofibroblasts in the fibrotic liver in mice. Proc Natl Acad Sci USA. 2014;111:E3297–305.

    Article  CAS  Google Scholar 

  7. Holt AP, Salmon M, Buckley CD, et al. Immune interactions in hepatic fibrosis. Clin Liver Dis. 2008;12(861–82):x.

    Google Scholar 

  8. Franco OE, Shaw AK, Strand DW, et al. Cancer associated fibroblasts in cancer pathogenesis. Semin Cell Dev Biol. 2010;21:33–9.

    Article  CAS  Google Scholar 

  9. Sharon Y, Raz Y, Cohen N, et al. Tumor-derived osteopontin reprograms normal mammary fibroblasts to promote inflammation and tumor growth in breast cancer. Can Res. 2015;75:963–73.

    Article  CAS  Google Scholar 

  10. Tjomsland V, Spangeus A, Valila J, et al. Interleukin 1alpha sustains the expression of inflammatory factors in human pancreatic cancer microenvironment by targeting cancer-associated fibroblasts. Neoplasia (New York, NY). 2011;13:664–75.

    Article  CAS  Google Scholar 

  11. Rupp C, Scherzer M, Rudisch A, et al. IGFBP7, a novel tumor stroma marker, with growth-promoting effects in colon cancer through a paracrine tumor-stroma interaction. Oncogene. 2015;34:815–25.

    Article  CAS  Google Scholar 

  12. Lau EY, Lo J, Cheng BY, et al. Cancer-Associated fibroblasts regulate tumor-initiating cell plasticity in hepatocellular carcinoma through c-Met/FRA1/HEY1 signaling. Cell Rep. 2016;15:1175–89.

    Article  CAS  Google Scholar 

  13. Kuchnio A, Moens S, Bruning U, et al. The cancer cell oxygen sensor PHD2 promotes metastasis via activation of cancer-associated fibroblasts. Cell Rep. 2015;12:992–1005.

    Article  CAS  Google Scholar 

  14. Kouwaki T, Okamoto T, Ito A, et al. Hepatocyte factor JMJD5 regulates hepatitis B virus replication through interaction with HBx. J Virol. 2016;90:3530–42.

    Article  CAS  Google Scholar 

  15. Nwani NG, Deguiz ML, Jimenez B, et al. Melanoma cells block PEDF production in fibroblasts to induce the tumor-promoting phenotype of cancer-associated fibroblasts. Can Res. 2016;76:2265–76.

    Article  CAS  Google Scholar 

  16. Orimo A, Gupta PB, Sgroi DC, et al. Stromal fibroblasts present in invasive human breast carcinomas promote tumor growth and angiogenesis through elevated SDF-1/CXCL12 secretion. Cell. 2005;121:335–48.

    Article  CAS  Google Scholar 

  17. Ohlund D, Elyada E, Tuveson D. Fibroblast heterogeneity in the cancer wound. J Exp Med. 2014;211:1503–23.

    Article  CAS  Google Scholar 

  18. Madar S, Goldstein I, Rotter V. ‘Cancer associated fibroblasts’—more than meets the eye. Trends Mol Med. 2013;19:447–53.

    Article  CAS  Google Scholar 

  19. Salazar VS, Gamer LW, Rosen V. BMP signalling in skeletal development, disease and repair. Nat Rev Endocrinol. 2016;12:203–21.

    Article  CAS  Google Scholar 

  20. Lee KW, Yeo SY, Sung CO, et al. Twist1 is a key regulator of cancer-associated fibroblasts. Can Res. 2015;75:73–85.

    Article  CAS  Google Scholar 

  21. Nemer G, Nemer M. Transcriptional activation of BMP-4 and regulation of mammalian organogenesis by GATA-4 and -6. Dev Biol. 2003;254:131–48.

    Article  CAS  Google Scholar 

  22. Kim JS, Kurie JM, Ahn YH. BMP4 depletion by miR-200 inhibits tumorigenesis and metastasis of lung adenocarcinoma cells. Mol Cancer. 2015;14:173.

    Article  Google Scholar 

  23. Li Z, Fei T, Zhang J, et al. BMP4 Signaling Acts via dual-specificity phosphatase 9 to control ERK activity in mouse embryonic stem cells. Cell Stem Cell. 2012;10:171–82.

    Article  CAS  Google Scholar 

  24. Ma J, Zeng S, Zhang Y, et al. BMP4 promotes oxaliplatin resistance by an induction of epithelial-mesenchymal transition via MEK1/ERK/ELK1 signaling in hepatocellular carcinoma. Cancer Lett. 2017;411:117–29.

    Article  CAS  Google Scholar 

  25. Ma J, Zeng S, Zhang Y, et al. BMP4 enhances hepatocellular carcinoma proliferation by promoting cell cycle progression via ID2/CDKN1B signaling. Mol Carcinog. 2017;56:2279–89.

    Article  CAS  Google Scholar 

  26. Zeng S, Zhang Y, Ma J, et al. BMP4 promotes metastasis of hepatocellular carcinoma by an induction of epithelial-mesenchymal transition via upregulating ID2. Cancer Lett. 2017;390:67–76.

    Article  CAS  Google Scholar 

  27. Maegdefrau U, Amann T, Winklmeier A, et al. Bone morphogenetic protein 4 is induced in hepatocellular carcinoma by hypoxia and promotes tumour progression. J Pathol. 2009;218:520–9.

    Article  CAS  Google Scholar 

  28. Daher R, Kannengiesser C, Houamel D, et al. Heterozygous mutations in BMP6 pro-peptide lead to inappropriate hepcidin synthesis and moderate iron overload in humans. Gastroenterology. 2016;150(672–83):e4.

    Google Scholar 

  29. Latour C, Besson-Fournier C, Meynard D, et al. Differing impact of the deletion of hemochromatosis-associated molecules HFE and transferrin receptor-2 on the iron phenotype of mice lacking bone morphogenetic protein 6 or hemojuvelin. Hepatology. 2016;63:126–37.

    Article  CAS  Google Scholar 

  30. Sadlonova A, Novak Z, Johnson MR, et al. Breast fibroblasts modulate epithelial cell proliferation in three-dimensional in vitro co-culture. Breast Cancer Res. 2005;7:R46–59.

    Article  Google Scholar 

  31. Berdiel-Acer M, Bohem ME, Lopez-Doriga A, et al. Hepatic carcinoma-associated fibroblasts promote an adaptative response in colorectal cancer cells that inhibit proliferation and apoptosis: nonresistant cells die by nonapoptotic cell death. Neoplasia (New York, NY). 2011;13:931–46.

    Article  CAS  Google Scholar 

  32. Burton DG, Krizhanovsky V. Physiological and pathological consequences of cellular senescence. Cell Mol Life Sci. 2014;71:4373–86.

    Article  CAS  Google Scholar 

  33. Trautwein C, Friedman SL, Schuppan D, et al. Hepatic fibrosis: concept to treatment. J Hepatol. 2015;62:S15–24.

    Article  CAS  Google Scholar 

  34. Mederacke I, Hsu CC, Troeger JS, et al. Fate tracing reveals hepatic stellate cells as dominant contributors to liver fibrosis independent of its aetiology. Nat Commun. 2013;4:2823.

    Article  Google Scholar 

Download references

Acknowledgements

We thank Ms. Chizu Kanokoda for her technical assistance.

Funding

This study was supported by Grants-in-aid for Research from the National Center for Global Health and Medicine (26-shi-109 and 26A201) and by the Research Program on Hepatitis from Japan Agency for Medical Research and Development (17fk0210305h0003).

Author information

Author notes

  1. Yohei Mano and Sachiyo Yoshio equally contributed to this work.

Authors and Affiliations

  1. The Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, 1-7-1, Kohnodai, Ichikawa, Chiba, 272-8516, Japan

    Yohei Mano, Sachiyo Yoshio, Hirotaka Shoji, Shimagaki Tomonari, Yoshihiko Aoki, Yosuke Osawa & Tatsuya Kanto

  2. Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan

    Yohei Mano, Shimagaki Tomonari, Kyohei Yugawa, Huanlin Wang & Tomoharu Yoshizumi

  3. Department of Surgery, Kohnodai Hospital, National Center for Global Health and Medicine, Ichikawa, Chiba, Japan

    Nobuyoshi Aoyanagi

  4. Department of Molecular Virology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan

    Toru Okamoto & Yoshiharu Matsuura

  5. Department of Hepatology, Tokyo Metropolitan Cancer and Infectious Diseases Center Komagome Hospital, Tokyo, Japan

    Kiminori Kimura

  6. Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan

    Kyohei Yugawa, Huanlin Wang & Yoshinao Oda

  7. Department of Surgery, Kyushu Central Hospital, Fukuoka, Japan

    Yoshihiko Maehara

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  1. Yohei Mano
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Correspondence to Tatsuya Kanto.

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Mano, Y., Yoshio, S., Shoji, H. et al. Bone morphogenetic protein 4 provides cancer-supportive phenotypes to liver fibroblasts in patients with hepatocellular carcinoma. J Gastroenterol 54, 1007–1018 (2019). https://doi.org/10.1007/s00535-019-01579-5

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  • DOI: https://doi.org/10.1007/s00535-019-01579-5

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