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Altered immunolocalization of FGF23 in murine femora metastasized with human breast carcinoma MDA-MB-231 cells

Abstract

Introduction

After the onset of bone metastasis, tumor cells appear to modify surrounding microenvironments for their benefit, and particularly, the levels of circulating fibroblast growth factor (FGF) 23 in patients with tumors have been highlighted.

Materials and methods

We have attempted to verify if human breast carcinoma MDA-MB-231 cells metastasized in the long bone of nu/nu mice would synthesize FGF23. Serum concentrations of calcium, phosphate (Pi) and FGF23 were measured in control nu/nu mice, bone-metastasized mice, and mice with mammary gland injected with MDA-MB-231 cells mimicking primary mammary tumors.

Results and conclusions

MDA-MB-231 cells revealed intense FGF23 reactivity in metastasized lesions, whereas MDA-MB-231 cells cultured in vitro or when injected into the mammary glands (without bone metastasis) showed weak FGF23 immunoreactivity. Although the bone-metastasized MDA-MB-231 cells abundantly synthesized FGF23, osteocytes adjacent to the FGF23-immunopositive tumors, unlike intact osteocytes, showed no FGF23. Despite significantly elevated serum FGF23 levels in bone-metastasized mice, there was no significant decrease in the serum Pi concentration when compared with the intact mice and mice with a mass of MDA-MB-231 cells in mammary glands. The metastasized femora showed increased expression and FGFR1 immunoreactivity in fibroblastic stromal cells, whereas femora of control mice showed no obvious FGFR1 immunoreactivity. Taken together, it seems likely that MDA-MB-231 cells synthesize FGF23 when metastasized to a bone, and thus affect FGFR1-positive stromal cells in the metastasized tumor nest in a paracrine manner.

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Acknowledgements

This study was partially supported by the Grants-in Aid for Scientific Research of Japan Society for the Promotion of Science (JSPS; Hasegawa T and Amizuka N) and Promoting International Joint Research (Bilateral Collaborations) of JSPS in Japan and NSFC in China (Amizuka N and Li M).

Funding

The research was supported by Japan Society for the Promotion of Science (Grants 19K10040, 18K19628).

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Authors

Contributions

All authors contributed to the study conception and design. AY is the main researcher who contributed to this work, including histochemical analyses. Hasegawa T, Yamada T, HH, and MA under the guidance by Hiraga T performed animal experiments, such as preparation of cultured MDA-MB-231 cells, their injection into nu/nu mice, and fixation of the metastasized mice. Y and Yoshida T performed western blotting, and YI and Yamamoto T conducted serum collection from mice, measurements of FGF23, and statistical analysis on serum concentration of calcium, Pi, and FGF23. SK, MS, PHLF, ML, NA, and YY participated in the discussion and preparation of the manuscript. Hasegawa T is the chief of this research project who organized collaborators and provided the whole outline of this experiment. The first draft of the manuscript was written by Hasegawa T and all authors have provided their inputs on the manuscript. All authors have read and approved the final manuscript.

Corresponding author

Correspondence to Tomoka Hasegawa.

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Yokoyama, A., Hasegawa, T., Hiraga, T. et al. Altered immunolocalization of FGF23 in murine femora metastasized with human breast carcinoma MDA-MB-231 cells. J Bone Miner Metab 39, 810–823 (2021). https://doi.org/10.1007/s00774-021-01220-7

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Keywords

  • Fibroblast growth factor 23
  • Osteocyte
  • Immunohistochemistry
  • Bone metastasis
  • MDA-MB-231