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

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Journal of Bone and Mineral Metabolism Aims and scope Submit manuscript

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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References

  1. Arguello F, Baggs RB, Frantz CN (1988) A murine model of experimental metastasis to bone and bone marrow. Cancer Res 48:6876–6881

    CAS  PubMed  Google Scholar 

  2. Hiraga T, Nakajima T, Ozawa H (1995) Bone resorption induced by a metastatic human melanoma cell line. Bone 16:349–356

    Article  CAS  Google Scholar 

  3. Shimamura T, Amizuka N, Li M, Luiz de Freitas PH, White JH, Henderson JE, Shingaki S, Nakajima T, Ozawa H (2005) Histological observations on the microenvironment of osteolytic bone metastasis by breast carcinoma cell line. Biomed Res 26:159–172

    Article  CAS  Google Scholar 

  4. Yoneda T, Sasaki A, Mundy GR (1994) Osteolytic bone metastasis in breast cancer. Breast Cancer Res Treat 32:73–84

    Article  CAS  Google Scholar 

  5. Hiraga T (2019) Bone metastasis: Interaction between cancer cells and bone microenvironment. J Oral Biosci 61:95–98

    Article  Google Scholar 

  6. Seccareccia D (2010) Cancer-related hypercalcemia. Can Fam Physician 56:244–246

    PubMed  PubMed Central  Google Scholar 

  7. Shimada T, Muto T, Urakawa I, Yoneya T, Yamazaki Y, Okawa K, Takeuchi Y, Fujita T, Fukumoto S, Yamashita T (2002) Mutant FGF-23 responsible for autosomal dominant hypophosphatemic rickets is resistant to proteolytic cleavage and causes hypophosphatemia in vivo. Endocrinology 143:3179–3182

    Article  CAS  Google Scholar 

  8. Yamazaki Y, Okazaki R, Shibata M, Hasegawa Y, Satoh K, Tajima T, Takeuchi Y, Fujita T, Nakahara K, Yamashita T, Fukumoto S (2002) Increased circulatory level of biologically active full-length FGF-23 in patients with Hypophosphatemic Rickets/Osteomalacia. J Clin Endocrinol Metab 87:4957–4960

    Article  CAS  Google Scholar 

  9. Folpe AL, Fanburg-Smith JC, Billings SD, Bisceglia M, Bertoni F, Cho JY, Econs MJ, Inwards CY, Jan de Beur SM, Mentzel T, Montgomery E, Michal M, Miettinen M, Mills SE, Reith JD, O’Connell JX, Rosenberg AE, Rubin BP, Sweet DE, Vinh TN, Wold LE, Wehrli BM, White KE, Zaino RJ, Weiss SW (2004) Most osteomalacia-associated mesenchymal tumors are a single histopathologic entity: an analysis of 32 cases and a comprehensive review of the literature. Am J Surg Pathol 28:1–30

    Article  Google Scholar 

  10. Nasu T, Kurisu S, Matsuno S, Tatsumi K, Kakimoto T, Kobayashi M, Nakano Y, Wakasaki H, Furuta H, Nishi M, Sasaki H, Suzuki H, Ito N, Fukumoto S, Nanjo K (2008) Tumor-induced hypophosphatemic osteomalacia diagnosed by the combinatory procedures of magnetic resonance imaging and venous sampling for FGF23. Intern Med 47:957–961

    Article  Google Scholar 

  11. Jiang Y, Xia WB, Xing XP, Silva BC, Li M, Wang O, Zhang HB, Li F, Jing HL, Zhong DR, Jin J, Gao P, Zhou L, Qi F, Yu W, Bilezikian JP, Meng XW (2012) Tumor-induced osteomalacia: an important cause of adult-onset hypophosphatemic osteomalacia in China: report of 39 cases and review of the literature. J Bone Miner Res 27:1967–1975

    Article  Google Scholar 

  12. Minisola S, Peacock M, Fukumoto S, Cipriani C, Pepe J, Tella SH, Collins MT (2017) Tumour-induced osteomalacia. Nat Rev Dis Primers 3:17044. https://doi.org/10.1038/nrdp.2017.44

    Article  PubMed  Google Scholar 

  13. Kinoshita Y, Fukumoto S (2018) X-Linked hypophosphatemia and FGF23-related hypophosphatemic diseases: prospect for new treatment. Endocr Rev 39:274–291

    Article  Google Scholar 

  14. Mansinho A, Ferreira AR, Casimiro S, Alho I, Vendrell I, Costa AL, Sousa R, Abreu C, Pulido C, Macedo D, Pacheco TR, Correia L, Costa L (2019) Levels of circulating fibroblast growth factor 23 (FGF23) and prognosis in cancer patients with bone metastases. Int J Mol Sci 20:pii: E695. https://doi.org/10.3390/ijms20030695

    Article  CAS  Google Scholar 

  15. Ubaidus S, Li M, Sultana S, Luiz de Freitas PH, Oda K, Maeda T, Takagi R, Amizuka N (2009) FGF23 is mainly synthesized by osteocytes in the regularly distributed osteocytic lacunar canalicular system established after physiological bone remodeling. J Electron Microsc (Tokyo) 58:381–392

    Article  CAS  Google Scholar 

  16. Sakurai A, Hasegawa T, Kudo A, Shen Z, Nagai T, Abe M, Yoshida T, Hongo H, Yamamoto T, Yamamoto T, Oda K, Freitas PHL, Li M, Sano H, Amizuka N (2017) Chronological immunolocalization of sclerostin and FGF23 in the mouse metaphyseal trabecular and cortical bone. Biomed Res 38:257–267

    Article  CAS  Google Scholar 

  17. Nabeshima Y (2006) Toward a better understanding of Klotho. Sci Aging Knowledge Environ 8:pe11. https://doi.org/10.1126/sageke.2006.8.pe11

    Article  Google Scholar 

  18. Urakawa I, Yamazaki Y, Shimada T, Iijima K, Hasegawa H, Okawa K, Fujita T, Fukumoto S, Yamashita T (2006) Klotho converts canonical FGF receptor into a specific receptor for FGF23. Nature 444:770–777

    Article  CAS  Google Scholar 

  19. Nakatani T, Sarraj B, Ohnishi M, Densmore MJ, Taguchi T, Goetz R, Mohammadi M, Lanske B, Razzaque MS (2009) In vivo genetic evidence for klotho-dependent, fibroblast growth factor 23 (Fgf23)-mediated regulation of systemic phosphate homeostasis. FASEB J 23:433–441

    Article  CAS  Google Scholar 

  20. Fukumoto S (2009) The role of bone in phosphate metabolism. Mol Cell Endocrinol 310:63–70

    Article  CAS  Google Scholar 

  21. Hasegawa T, Sasaki M, Yamada T, Ookido I, Yamamoto T, Hongo H, Yamamoto T, Oda K, Yokoyama K, Amizuka N (2013) Histochemical examination of vascular medial calcification of aorta in klotho-deficient mice. J Oral Biosci 55:10–15

    Article  Google Scholar 

  22. Hasegawa T, Yamamoto T, Sakai S, Miyamoto Y, Hongo H, Qiu Z, Abe M, Takeda S, Oda K, de Freitas PHL, Li M, Endo K, Amizuka N (2019) Histological effects of the combined administration of eldecalcitol and a parathyroid hormone in the Metaphyseal Trabeculae of Ovariectomized Rats. J Histochem Cytochem 67:169–184

    Article  CAS  Google Scholar 

  23. Hasegawa T, Endo T, Tsuchiya E, Kudo A, Zhao S, Moritani Y, Abe M, Yamamoto T, Hongo H, Tsuboi K, Yoshida T, Nagai T, Khadiza N, Yokoyama A, Luiz de Freitas PH, Li M, Amizuka N (2017) Biological application of focus ion beam-scanning electron microscopy (FIB-SEM) to the imaging of cartilaginous fibrils and osteoblastic cytoplasmic processes. J Oral Biosci 59:55–62

    Article  Google Scholar 

  24. Hu X, Du S, Yu J, Yang X, Yang C, Zhou D, Wang Q, Qin S, Yan X, He L, Han D, Wan C (2016) Common housekeeping proteins are upregulated in colorectal adenocarcinoma and hepatocellular carcinoma, making the total protein a better “housekeeper.” Oncotarget 7:66679–66688

    Article  Google Scholar 

  25. Nie X, Li C, Hu S, Xue F, Kang YJ, Zhang W (2017) An appropriate loading control for western blot analysis in animal models of myocardial ischemic infarction. Biochem Biophys Rep 12:108–113

    PubMed  PubMed Central  Google Scholar 

  26. Yamamoto T, Hasegawa T, Sasaki M, Hongo H, Tsuboi K, Shimizu T, Ota M, Haraguchi M, Takahata M, Oda K, Luiz de Freitas PH, Takakura A, Takao-Kawabata R, Isogai Y, Amizuka N (2016) Frequency of teriparatide administration affects the histological pattern of bone formation in young adult male mice. Endocrinology 157:2604–2620

    Article  CAS  Google Scholar 

  27. Imanishi Y, Hashimoto J, Ando W, Kobayashi K, Ueda T, Nagata Y, Miyauchi A, Koyano HM, Kaji H, Saito T, Oba K, Komatsu Y, Morioka T, Mori K, Miki T, Inaba M (2012) Matrix extracellular phosphoglycoprotein is expressed in causative tumors of oncogenic osteomalacia. J Bone Miner Metab 30:93–99

    Article  CAS  Google Scholar 

  28. Ito N, Wijenayaka AR, Prideaux M, Kogawa M, Ormsby RT, Evdokiou A, Bonewald LF, Findlay DM, Atkins GJ (2015) Regulation of FGF23 expression in IDG-SW3 osteocytes and human bone by pro-inflammatory stimuli. Mol Cell Endocrinol 399:208–218

    Article  CAS  Google Scholar 

  29. Bär L, Feger M, Fajol A, Klotz LO, Zeng S, Lang F, Hocher B, Föller M (2018) Insulin suppresses the production of fibroblast growth factor 23 (FGF23). Proc Natl Acad Sci USA 115:5804–5809

    Article  Google Scholar 

  30. Kamiya N, Yamaguchi R, Aruwajoye O, Kim AJ, Kuroyanagi G, Phipps M, Adapala NS, Feng JQ, Kim HK (2017) Targeted disruption of NF1 in osteocytes increases FGF23 and osteoid with osteomalacia-like bone phenotype. J Bone Miner Res 32:1716–1726

    Article  CAS  Google Scholar 

  31. Egli-Spichtig D, Imenez Silva PH, Glaudemans B, Gehring N, Bettoni C, Zhang MYH, Pastor-Arroyo EM, Schönenberger D, Rajski M, Hoogewijs D, Knauf F, Misselwitz B, Frey-Wagner I, Rogler G, Ackermann D, Ponte B, Pruijm M, Leichtle A, Fiedler GM, Bochud M, Ballotta V, Hofmann S, Perwad F, Föller M, Lang F, Wenger RH, Frew I, Wagner CA (2019) Tumor necrosis factor stimulates fibroblast growth factor 23 levels in chronic kidney disease and non-renal inflammation. Kidney Int 96:890–905

    Article  CAS  Google Scholar 

  32. Kaludjerovic J, Komaba H, Sato T, Rg E, Baron R, Olauson H, Larsson TE, Lanske B (2017) Klotho expression in long bones regulates FGF23 production during renal failure. FASEB J 31:2050–2064

    Article  CAS  Google Scholar 

  33. Xiao Z, King G, Mancarella S, Munkhsaikhan U, Cao L, Cai C, Quarle LD (2019) FGF23 expression is stimulated in transgenic α-Klotho longevity mouse model. JCI Insight 4:e132820

    Article  Google Scholar 

  34. Kinoshita Y, Takashi Y, Ito N, Ikegawa S, Mano H, Ushiku T, Fukayama M, Nangaku M, Fukumoto S (2018) Ectopic expression of Klotho in fibroblast growth factor 23 (FGF23)-producing tumors that cause tumor-induced rickets/osteomalacia (TIO). Bone Rep 10:100192. https://doi.org/10.1016/j.bonr.2018.100192

    Article  PubMed  PubMed Central  Google Scholar 

  35. Toyosawa S, Shintani S, Fujiwara T, Ooshima T, Sato A, Ijuhin N, Komori T (2001) Dentin matrix protein 1 is predominantly expressed in chicken and rat osteocytes but not in osteoblasts. J Bone Miner Res 16:2017–2026

    Article  CAS  Google Scholar 

  36. Kitai Y, Matsubara T, Yanagita M (2015) Onco-nephrology: current concepts and future perspectives. Jpn J Clin Oncol 45:617–628

    Article  Google Scholar 

  37. Ben-Dov IZ, Galitzer H, Lavi-Moshayoff V, Goetz R, Kuro-o M, Mohammadi M, Sirkis R, Naveh-Many T, Silver J (2007) The parathyroid is a target organ for FGF23 in rats. J Clin Invest 117:4003–4008

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Olauson H, Lindberg K, Amin R, Sato T, Jia T, Goetz R, Mohammadi M, Andersson G, Lanske B, Larsson TE (2013) Parathyroid-specific deletion of klotho unravels a novel calcineurin-dependent FGF23 signaling pathway that regulates PTH secretion. PLoS Genet 9:e1003975

    Article  Google Scholar 

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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).

Author information

Authors and Affiliations

  1. Developmental Biology of Hard Tissue, Graduate School of Dental Medicine and Faculty of Dental Medicine, Hokkaido University, Kita-13, Nishi-7, Kita-Ku, Sapporo, Japan

    Ayako Yokoyama, Tomoka Hasegawa, Hiromi Hongo, Tomomaya Yamamoto, Miki Abe, Taiji Yoshida & Norio Amizuka

  2. Gerodontology, Graduate School of Dental Medicine and Faculty of Dental Medicine, Hokkaido University, Sapporo, Japan

    Ayako Yokoyama & Yutaka Yamazaki

  3. Oral and Maxillofacial Surgery, Graduate School of Dental Medicine and Faculty of Dental Medicine, Hokkaido University, Sapporo, Japan

    Tamaki Yamada

  4. Department of Oral Anatomy, Matsumoto Dental University, Shiojiri, Japan

    Toru Hiraga

  5. Central Research Institute, Graduate School of Medicine, Hokkaido University, Sapporo, Japan

    Yimin

  6. Northern Army Medical Unit, Camp Makomanai, Japan Ground Self-Defense Forces,, Sapporo, Japan

    Tomomaya Yamamoto

  7. Department of Nephrology, Graduate School of Medicine, Osaka City University, Osaka, Japan

    Yasuo Imanishi

  8. Department of Applied Prosthodontics, Unit of Translational Medicine, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki, Japan

    Shinichiro Kuroshima & Muneteru Sasaki

  9. Department of Dentistry, Federal University of Sergipe at Lagarto, Sergipe, Brazil

    Paulo Henrique Luiz de Fraitas

  10. Division of Basic Science of Stomatology, The School of Stomatology, Shandong University, Jinan, China

    Minqi Li

Authors
  1. Ayako Yokoyama
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  2. Tomoka Hasegawa
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  4. Tamaki Yamada
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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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  • DOI: https://doi.org/10.1007/s00774-021-01220-7

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