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Fabrication of Functional Cell Sheets with Human Thyrocytes from Non-Tumorous Thyroid Tissue

  • Yu Huang
  • Kosho YamanouchiEmail author
  • Yusuke Sakai
  • Sayaka Kuba
  • Chika Sakimura
  • Michi Morita
  • Kengo Kanetaka
  • Mitsuhisa Takatsuki
  • Susumu Eguchi
Original Article
  • 14 Downloads

Abstract

Background:

Engineered cell sheet transplantation has been considered an alternative physiological therapy for endocrine disorders. In this study, we attempted to fabricate functional human thyroid cell sheets using the engineering technology by culturing primary thyrocytes in free-feeder monolayers and assessed their proliferation and function in two different media.

Methods:

The non-tumorous tissues (approximately 2 g) were dissected during surgery. Primary human thyroid cells were isolated by mechanical dispersion and treatment with isolation solution. The cells were cultured on tissue culture dishes or temperature-responsive culture dishes to induce the formation of detached cell sheets.

Results:

Primary thyroid cells isolated from nine patients were positive for thyroid transcription factor 1, thyroglobulin (TG) and cytokeratin 7. Cell sheets with follicles were fabricated by cells incubated in both Dulbecco’s Modified Eagle Medium (DMEM) and hepatocyte-defined medium (HDM) culture medium. The diameter and thickness of sheets fabricated in HDM were larger and thicker than those fabricated from DMEM. Furthermore, the cells incubated in HDM secreted higher levels of fT3 and fT4 than those incubated in DMEM. The thyroid peroxidase and TG mRNA of cells maintained in HDM were higher than those in cells maintained in DMEM.

Conclusion:

HDM appears suitable as a culture medium for maintaining primary thyrocytes and fabricating functional cell sheets. These in vitro findings may contribute to the development of appropriate culture conditions for human thyrocytes as well as engineered functional cell sheets.

Keywords

Cell sheet Thyroid Free-feeder monolayer Hepatocyte-defined medium 

Notes

Acknowledgement

This work was supported by Grant-in-Aid for Scientific Research (No. 26461950).

Compliance with Ethical Standards

Conflict of interest

The authors have no conflict of interest to declare.

Ethical Statement

This study was approved by the institutional review board of the Nagasaki University Hospital (Approval Number 14052642) and we obtained informed consent by each patient.

References

  1. 1.
    Celi FS, Zemskova M, Linderman JD, Smith S, Drinkard B, Sachdev V, et al. Metabolic effects of liothyronine therapy in hypothyroidism: a randomized, double-blind, crossover trial of liothyronine versus levothyroxine. J Clin Endocrinol Metab. 2011;96:3466–74.CrossRefGoogle Scholar
  2. 2.
    Arauchi A, Shimizu T, Yamato M, Obara T, Okano T. Tissue-engineered thyroid cell sheet rescued hypothyroidism in rat models after receiving total thyroidectomy comparing with nontransplantation models. Tissue Eng Part A. 2009;15:3943–9.CrossRefGoogle Scholar
  3. 3.
    Memon IA, Sawa Y, Fukushima N, Matsumiya G, Miyagawa S, Taketani S, et al. Repair of impaired myocardium by means of implantation of engineered autologous myoblast sheets. J Thorac Cardiovasc Surg. 2005;130:1333–41.CrossRefGoogle Scholar
  4. 4.
    Ohashi K, Yokoyama T, Yamato M, Kuge H, Kanehiro H, Tsutsumi M, et al. Engineering functional two- and three-dimensional liver systems in vivo using hepatic tissue sheets. Nat Med. 2007;13:880–5.CrossRefGoogle Scholar
  5. 5.
    Perrod G, Rahmi G, Pidial L, Camilleri S, Bellucci A, Casanova A, et al. Cell sheet transplantation for esophageal stricture prevention after endoscopic submucosal dissection in a porcine model. PLoS One. 2016;11:e0148249.CrossRefGoogle Scholar
  6. 6.
    Sakai Y, Yamanouchi K, Ohashi K, Koike M, Utoh R, Hasegawa H, et al. Vascularized subcutaneous human liver tissue from engineered hepatocyte/fibroblast sheets in mice. Biomaterials. 2015;65:66–75.CrossRefGoogle Scholar
  7. 7.
    Bell E, Moore H, Mitchie C, Sher S, Coon H. Reconstruction of a thyroid gland equivalent from cells and matrix materials. J Exp Zool. 1984;232:277–85.CrossRefGoogle Scholar
  8. 8.
    Toda S, Koike N, Sugihara H. Thyrocyte integration, and thyroid folliculogenesis and tissue regeneration: perspective for thyroid tissue engineering. Pathol Int. 2001;51:403–17.CrossRefGoogle Scholar
  9. 9.
    Toda S, Sugihara H. Reconstruction of thyroid follicles from isolated porcine follicle cells in three-dimensional collagen gel culture. Endocrinology. 1990;126:2027–34.CrossRefGoogle Scholar
  10. 10.
    Bechtner G, Schopohl D, Rafferzeder M, Gärtner R, Welsch U. Stimulation of thyroid cell proliferation by epidermal growth factor is different from cell growth induced by thyrotropin or insulin-like growth factor I. Eur J Endocrinol. 1996;134:639–48.CrossRefGoogle Scholar
  11. 11.
    Fröhlich E, Wahl R, Reutter K. Basal lamina formation by porcine thyroid cells grown in collagen- and laminin-deficient medium. Histochem J. 1995;27:602–8.CrossRefGoogle Scholar
  12. 12.
    Takasu N, Ohno S, Komiya I, Yamada T. Requirements of follicle structure for thyroid hormone synthesis; cytoskeletons and iodine metabolism in polarized monolayer cells on collagen gel and in double layered, follicle-forming cells. Endocrinology. 1992;131:1143–8.CrossRefGoogle Scholar
  13. 13.
    Zeng L, Geng Y, Tretiakova M, Yu X, Sicinski P, Kroll TG. Peroxisome proliferator-activated receptor-delta induces cell proliferation by a cyclin E1-dependent mechanism and is up-regulated in thyroid tumors. Cancer Res. 2008;68:6578–86.CrossRefGoogle Scholar
  14. 14.
    Penna-Martinez M, Winten C, Fichtel T, Caspar-Bell G, Usadel KH, Schumm-Draeger PM. Isolation of thyroid cells obtained by fine-needle aspiration biopsy. Thyroid. 2005;15:989–95.CrossRefGoogle Scholar
  15. 15.
    Baimakhanov Z, Sakai Y, Yamanouchi K, Hidaka M, Soyama A, Takatsuki M, et al. Spontaneous hepatocyte migration towards an endothelial cell tube network. J Tissue Eng Regen Med. 2018;12:e1767–71.CrossRefGoogle Scholar
  16. 16.
    Toda S, Yonernitsu N, Hikichi Y, Sugihara H, Koike N. Differentiation of human thyroid follicle cells from normal subjects and Basedow’s disease in three-dimensional collagen gel culture. Pathol Res Pract. 1992;188:874–82.CrossRefGoogle Scholar
  17. 17.
    Goretzki PE, Frilling A, Simon D, Roeher HD. Growth regulation of normal thyroids and thyroid tumors in man. Recent Results Cancer Res. 1990;118:48–63.CrossRefGoogle Scholar
  18. 18.
    Hegyesi O, Földes A, Bori E, Németh Z, Barabás J, Steward MC, et al. Evidence for active electrolyte transport by two-dimensional monolayers of human salivary epithelial cells. Tissue Eng Part C Methods. 2015;21:1226–36.CrossRefGoogle Scholar
  19. 19.
    Tiwari S, Nair RM, Vamadevan P, Ali MJ, Naik MN, Honavar SG, et al. Establishing and characterizing lacrispheres from human lacrimal gland for potential clinical application. Graefes Arch Clin Exp Ophthalmol. 2018;256:717–27.CrossRefGoogle Scholar
  20. 20.
    Vassart G, Dumont JE. The thyrotropin receptor and the regulation of thyrocyte function and growth. Endocr Rev. 1992;13:596–611.Google Scholar
  21. 21.
    Yamazaki K, Yamada E, Kanaji Y, Shizume K, Wang DS, Maruo N, et al. Interleukin-6 (IL-6) inhibits thyroid function in the presence of soluble IL-6 receptor in cultured human thyroid follicles. Endocrinology. 1996;137:4857–63.CrossRefGoogle Scholar
  22. 22.
    Colletta G, Cirafici AM, Di Carlo A. Dual effect of transforming growth factor beta on rat thyroid cells: inhibition of thyrotropin-induced proliferation and reduction of thyroid-specific differentiation markers. Cancer Res. 1989;49:3457–62.Google Scholar
  23. 23.
    Asmis LM, Gerber H, Kaempf J, Studer H. Epidermal growth factor stimulates cell proliferation and inhibits iodide uptake of FRTL-5 cells in vitro. J Endocrinol. 1995;145:513–20.CrossRefGoogle Scholar
  24. 24.
    Westermark K, Karlsson FA, Westermark B. Epidermal growth factor modulates thyroid growth and function in culture. Endocrinology. 1983;112:1680–6.CrossRefGoogle Scholar
  25. 25.
    Bravo SB, Garcia-Rendueles ME, Garcia-Rendueles AR, Rodrigues JS, Perez-Romero S, Garcia-Lavandeira M, et al. Humanized medium (h7H) allows long-term primary follicular thyroid cultures from human normal thyroid, benign neoplasm, and cancer. J Clin Endocrinol Metab. 2013;98:2431–41.CrossRefGoogle Scholar
  26. 26.
    Nishida K, Yamato M, Hayashida Y, Watanabe K, Yamamoto K, Adachi E, et al. Corneal reconstruction with tissue-engineered cell sheets composed of autologous oral mucosal epithelium. N Engl J Med. 2004;351:1187–96.CrossRefGoogle Scholar
  27. 27.
    Sawa Y, Yoshikawa Y, Toda K, Fukushima S, Yamazaki K, Ono M, et al. Safety and efficacy of autologous skeletal myoblast sheets (TCD-51073) for the treatment of severe chronic heart failure due to ischemic heart disease. Circ J. 2015;79:991–9.CrossRefGoogle Scholar

Copyright information

© The Korean Tissue Engineering and Regenerative Medicine Society 2019

Authors and Affiliations

  1. 1.Department of SurgeryNagasaki University Graduate School of Biomedical SciencesNagasakiJapan

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