Download PDF
Horm Metab Res 2013; 45(06): 401-407
DOI: 10.1055/s-0032-1333224
Original Basic
© Georg Thieme Verlag KG Stuttgart · New York

Examination of Orbital Tissues in Murine Models of Graves’ Disease Reveals Expression of UCP-1 and the TSHR in Retrobulbar Adipose Tissues

K.T. M. Johnson
1   Department for Ophthalmology, University of Duisburg-Essen, Essen, Germany
,
B. Wiesweg
1   Department for Ophthalmology, University of Duisburg-Essen, Essen, Germany
,
M. Schott
2   Department of Endocrinology and Diabetology, University of Düsseldorf, Düsseldorf, Germany
,
M. Ehlers
2   Department of Endocrinology and Diabetology, University of Düsseldorf, Düsseldorf, Germany
,
M. Müller
1   Department for Ophthalmology, University of Duisburg-Essen, Essen, Germany
,
W. B. Minich
3   Institute for Experimental Endocrinology – EnForCé, Charité, Berlin, ­Germany
,
Y. Nagayama
4   Department of Molecular Medicine, Atomic Bomb Disease Institute, Japan University Graduate School of Biomedical Sciences, Nagasaki, Japan
,
E. Gulbins
5   Department of Molecular Biology, University of Duisburg-Essen, Essen, Germany
,
A. K. Eckstein
1   Department for Ophthalmology, University of Duisburg-Essen, Essen, Germany
,
U. Berchner-Pfannschmidt
1   Department for Ophthalmology, University of Duisburg-Essen, Essen, Germany
› Author Affiliations
Further Information

Publication History

received 04 June 2012

accepted 10 December 2012

Publication Date:
05 February 2013 (online)

Also available at
   Permissions and Reprints

Abstract

Over the past decade a number of murine models of Graves’ disease (GD) have been described. The full symptom complex, including typical orbital changes, however, could not yet be induced. In this report, we examined the influence of modified immunization protocols on orbital pathology. C57BL/6 and BALB/c mice were immunized against the human TSH receptor (TSHR), using either a TSHR encoding plasmid or a TSHR A-subunit adenovirus. Prior to immunization with the TSHR plasmid, regulatory T cells were depleted in one group of each strain. TSHR-stimulating antibodies (TSAbs) were evaluated and orbits were stained immunohistochemically for F4/80, uncoupling protein-1 (UCP-1) and the TSHR. We found that after depletion of regulatory T cells, incidence of TSAb was increased in TSHR plasmid immunized C57BL/6 mice. Examination of early immunized mice showed no antibody production. However, a TSHR epitope-specific cellular immune response could be detected by tetramer-analyses. Adenoviral immunization lead to TSAb production in all but one animal. Analysis of F4/80 positive cells in retrobulbar fat revealed no significant macrophage infiltration in the orbits of immunized mice. Immunohistochemical staining shows co-localization of F4/80 positive cells, UCP-1 and the TSHR in retrobulbar fat. Though targets for TSHR autoimmunity could clearly be shown, immunization methods were not efficient enough to cause clear signs of orbital inflammation.

Key words

Graves’ orbitopathy - animal model - brown adipose tissue - macrophages
 

  • References

  • 1 Eckstein AK, Johnson KT, Thanos M, Esser J, Ludgate M. Current insights into the pathogenesis of Graves’ orbitopathy. Horm Metab Res 2009; 41: 456-464
    Thieme ConnectPubMedSearch in Google Scholar
  • 2 Prabhakar BS, Bahn RS, Smith TJ. Current perspective on the pathogenesis of Graves’ disease and ophthalmopathy. Endocr Rev 2003; 24: 802-835
    CrossrefPubMedSearch in Google Scholar
  • 3 Bahn RS. Clinical review 157: Pathophysiology of Graves’ ophthalmopathy: the cycle of disease. J Clin Endocrinol Metab 2003; 88: 1939-1946
    CrossrefPubMedSearch in Google Scholar
  • 4 Khoo DH, Ho SC, Seah LL, Fong KS, Tai ES, Chee SP, Eng PH, Aw SE, Fok AC. The combination of absent thyroid peroxidase antibodies and high thyroid-stimulating immunoglobulin levels in Graves’ disease identifies a group at markedly increased risk of ophthalmopathy. Thyroid 1999; 9: 1175-1180
    CrossrefPubMedSearch in Google Scholar
  • 5 Gerding MN, van der Meer JW, Broenink M, Bakker O, Wiersinga WM, Prummel MF. Association of thyrotrophin receptor antibodies with the clinical features of Graves’ ophthalmopathy. Clin Endocrinol (Oxf) 2000; 52: 267-271
    Search in Google Scholar
  • 6 Noh JY, Hamada N, Inoue Y, Abe Y, Ito K. Thyroid-stimulating antibody is related to Graves’ ophthalmopathy, but thyrotropin-binding inhibitor immunoglobulin is related to hyperthyroidism in patients with Graves’ disease. Thyroid 2000; 10: 809-813
    CrossrefPubMedSearch in Google Scholar
  • 7 Eckstein AK, Plicht M, Lax H, Neuhauser M, Mann K, Lederbogen S, Heckmann C, Esser J, Morgenthaler NG. TSH-receptor autoantibodies are independent risk factors for Graves’ ophthalmopathy and help to predict severity and outcome of the disease. J Clin Endocrinol Metab 2006; 91: 3464-3470
    Search in Google Scholar
  • 8 Kaithamana S, Fan J, Osuga Y, Liang SG, Prabhakar BS. Induction of experimental autoimmune Graves’ disease in BALB/c mice. J Immunol 1999; 163: 5157-5164
    Search in Google Scholar
  • 9 Shimojo N, Kohno Y, Yamaguchi K, Kikuoka S, Hoshioka A, Niimi H, Hirai A, Tamura Y, Saito Y, Kohn LD, Tahara K. Induction of Graves-like disease in mice by immunization with fibroblasts transfected with the thyrotropin receptor and a class II molecule. Proc Natl Acad Sci USA 1996; 93: 11074-11079
    CrossrefPubMedSearch in Google Scholar
  • 10 Costagliola S, Alcalde L, Tonacchera M, Ruf J, Vassart G, Ludgate M. Induction of thyrotropin receptor (TSH-R) autoantibodies and thyroiditis in mice immunised with the recombinant TSH-R. Biochem Biophys Res Commun 1994; 199: 1027-1034
    CrossrefPubMedSearch in Google Scholar
  • 11 Costagliola S, Rodien P, Many MC, Ludgate M, Vassart G. Genetic immunization against the human thyrotropin receptor causes thyroiditis and allows production of monoclonal antibodies recognizing the native receptor. J Immunol 1998; 160: 1458-1465
    Search in Google Scholar
  • 12 Nagayama Y. Animal models of Graves’ hyperthyroidism. Endocr J 2005; 52: 385-394
    CrossrefPubMedSearch in Google Scholar
  • 13 Nagayama Y. Graves’ animal models of Graves’ hyperthyroidism. Thyroid 2007; 17: 981-988
    CrossrefPubMedSearch in Google Scholar
  • 14 Kita-Furuyama M, Nagayama Y, Pichurin P, McLachlan SM, Rapoport B, Eguchi K. Dendritic cells infected with adenovirus expressing the thyrotrophin receptor induce Graves’ hyperthyroidism in BALB/c mice. Clin Exp Immunol 2003; 131: 234-240
    CrossrefPubMedSearch in Google Scholar
  • 15 Yamaguchi K, Shimojo N, Kikuoka S, Hoshioka A, Hirai A, Tahara K, Kohn LD, Kohno Y, Niimi H. Genetic control of anti-thyrotropin receptor antibody generation in H-2K mice immunized with thyrotropin receptor-transfected fibroblasts. J Clin Endocrinol Metab 1997; 82: 4266-4269
    CrossrefPubMedSearch in Google Scholar
  • 16 Nagayama Y, Kita-Furuyama M, Ando T, Nakao K, Mizuguchi H, Hayakawa T, Eguchi K, Niwa M. A novel murine model of Graves’ hyperthyroidism with intramuscular injection of adenovirus expressing the thyrotropin receptor. J Immunol 2002; 168: 2789-2794
    Search in Google Scholar
  • 17 Saitoh O, Nagayama Y. Regulation of Graves’ hyperthyroidism with naturally occurring CD4+CD25+ regulatory T cells in a mouse model. Endocrinology 2006; 147: 2417-2422
    CrossrefPubMedSearch in Google Scholar
  • 18 Patibandla SA, Fan JL, Prabhakar BS, Seetharamaiah GS. Comparison of immune responses to extracellular domains of mouse and human thyrotropin receptor. J Autoimmun 1999; 13: 205-213
    CrossrefPubMedSearch in Google Scholar
  • 19 Land KJ, Gudapati P, Kaplan MH, Seetharamaiah GS. Differential requirement of signal transducer and activator of transcription-4 (Stat4) and Stat6 in a thyrotropin receptor-289-adenovirus-induced model of Graves’ hyperthyroidism. Endocrinology 2006; 147: 111-119
    CrossrefPubMedSearch in Google Scholar
  • 20 Seetharamaiah GS, Land KJ. Differential Susceptibility of BALB/c and BALB/cBy mice to Graves’ hyperthyroidism. Thyroid 2006; 16: 651-658
    CrossrefPubMedSearch in Google Scholar
  • 21 Kaneda T, Honda A, Hakozaki A, Fuse T, Muto A, Yoshida T. An improved Graves’ disease model established by using in vivo electroporation exhibited long-term immunity to hyperthyroidism in BALB/c mice. Endocrinology 2007; 148: 2335-2344
    CrossrefPubMedSearch in Google Scholar
  • 22 Many MC, Costagliola S, Detrait M, Denef F, Vassart G, Ludgate MC. Development of an animal model of autoimmune thyroid eye disease. J Immunol 1999; 162: 4966-4974
    Search in Google Scholar
  • 23 Ludgate M, Baker G. Inducing Graves’ ophthalmopathy. J Endocrinol Invest 2004; 27: 211-215
    PubMedSearch in Google Scholar
  • 24 Ludgate ME. Animal models of thyroid-associated ophthalmopathy. Thyroid 2002; 12: 205-208
    CrossrefPubMedSearch in Google Scholar
  • 25 Ho SC, Goh SS, Kee IH, Chow PK, Yeo CP, Khoo DH. Effects of genetic immunization of Swiss outbred mice with human thyroid stimulating hormone receptor cDNA plasmids harboring gain-of-function mutations. J Mol Endocrinol 2007; 38: 277-288
    CrossrefPubMedSearch in Google Scholar
  • 26 Ulmer JB, DeWitt CM, Chastain M, Friedman A, Donnelly JJ, McClements WL, Caulfield MJ, Bohannon KE, Volkin DB, Evans RK. Enhancement of DNA vaccine potency using conventional aluminum adjuvants. Vaccine 1999; 18: 18-28
    CrossrefPubMedSearch in Google Scholar
  • 27 Andrew SM, Titus JA. Purification of immunoglobulin G. Curr Protoc Immunol 2001; Chapter 2: Unit 27
    PubMedSearch in Google Scholar
  • 28 Rammensee H, Bachmann J, Emmerich NP, Bachor OA, Stevanovic S. SYFPEITHI: database for MHC ligands and peptide motifs. Immunogenetics 1999; 50: 213-219
    CrossrefPubMedSearch in Google Scholar
  • 29 Morgenthaler NG, Ho SC, Minich WB. Stimulating and blocking thyroid-stimulating hormone (TSH) receptor autoantibodies from patients with Graves’ disease and autoimmune hypothyroidism have very similar concentration, TSH receptor affinity, and binding sites. J Clin Endocrinol Metab 2007; 92: 1058-1065
    CrossrefPubMedSearch in Google Scholar
  • 30 Mark M, Teletin M, Antal C, Wendling O, Auwerx J, Heikkinen S, Khetchoumian K, Argmann CA, Dgheem M. Histopathology in mouse metabolic investigations. In: Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K. (eds.). Current Protocols in Molecular Biology. Hoboken, N. J.: Wiley; 2007. Chapter 29: Unit 29B 24
    Search in Google Scholar
  • 31 Payne AP. The harderian gland: a tercentennial review. Journal of anatomy 1994; 185 (Pt 1) 1-49
    PubMedSearch in Google Scholar
  • 32 Baker G, Mazziotti G, von Ruhland C, Ludgate M. Reevaluating thyrotropin receptor-induced mouse models of Graves’ disease and ophthalmopathy. Endocrinology 2005; 146: 835-844
    CrossrefPubMedSearch in Google Scholar
  • 33 Endo T, Kobayashi T. Thyroid-stimulating hormone receptor in brown adipose tissue is involved in the regulation of thermogenesis. American journal of physiology Endocrinology and metabolism 2008; 295: E514-E518
    CrossrefPubMedSearch in Google Scholar
  • 34 Zhang L, Baker G, Janus D, Paddon CA, Fuhrer D, Ludgate M. Biological effects of thyrotropin receptor activation on human orbital preadipocytes. Invest Ophthalmol Vis Sci 2006; 47: 5197-5203
    CrossrefPubMedSearch in Google Scholar
  • 35 Zhao SX, Tsui S, Cheung A, Douglas RS, Smith TJ, Banga JP. Orbital fibrosis in a mouse model of Graves’ disease induced by genetic immunization of thyrotropin receptor cDNA. J Endocrinol 2011; 210: 369-377
    Search in Google Scholar
  • 36 Ho SC, Goh SS, Su Q, Khoo DH. Cysteine 390 mutation of the TSH receptor modulates its ectodomain as an inverse agonist on the serpentine domain with decrease in basal constitutive activity. Mol Cell Endocrinol 2005; 245: 158-168
    CrossrefPubMedSearch in Google Scholar
  • 37 Nakahara M, Johnson K, Eckstein A, Taguchi R, Yamada M, Abiru N, Nagayama Y. Adoptive transfer of antithyrotropin receptor (TSHR) autoimmunity from TSHR knockout mice to athymic nude mice. Endocrinology 2012; 153: 2034-2042
    CrossrefPubMedSearch in Google Scholar