title: Biological effects of thyrotropin receptor activation on human orbital preadipocytes.
abstract: OBJECTIVE
  Thyrotropin receptor (TSHR) expression is upregulated in the orbits of patients with Graves ophthalmopathy (GO), most of whom have TSHR-stimulating antibodies. The authors investigated the biological effects of TSHR activation in vitro in adipose tissue, the site of orbital TSHR expression.
  
  
  METHODS
  Activating mutant TSHR (TSHR*) or wild-type (WT) was introduced into human orbital preadipocytes using retroviral vectors. Their proliferation (Coulter counting), basal cAMP accumulation (radioimmunoassay), and spontaneous and peroxisome proliferator-activated receptor (PPARgamma)-induced adipogenesis (quantitative oil red O staining) were assessed and compared with those of nonmodified cells. QRT-PCR was used to measure transcripts of CCAT/enhancer binding protein (C/EBP)beta, PPARgamma, and lipoprotein lipase (LPL; early, intermediate, and late markers of adipogenesis) and for uncoupling protein (UCP)-1 (brown adipose tissue [BAT]).
  
  
  RESULTS
  Expression of TSHR* significantly inhibited the proliferation of preadipocytes and produced an increase in unstimulated cAMP of 200% to 600%. Basal lipid levels were significantly increased in TSHR* (127%-275%) compared with nonmodified (100%) or WT-expressing (104%-187%) cells. This was accompanied by 2- to 10-fold increases in early-intermediate markers and UCP-1 transcripts (2- to 8-fold); LPL was at the limit of detection. In nonmodified cells, adipogenesis produced significant increases in transcripts of all markers, including LPL (approximately 30-fold). This was not the case in TSHR*-expressing cells, which also displayed 67% to 84% reductions in lipid levels.
  
  
  CONCLUSIONS
  TSHR activation stimulates early differentiation (favoring BAT formation?) but renders preadipocytes refractory to PPARgamma-induced adipogenesis. In neither case did lipid-containing vacuoles accumulate, suggesting that terminal stages of differentiation were inhibited.
authors.affiliation: Centre for Endocrine and Diabetes Sciences, School of Medicine, Cardiff University, United Kingdom.
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.forename: Glynn
authors.initials: G
authors.lastname: Baker
authors.forename: Dominika
authors.initials: D
authors.lastname: Janus
authors.forename: Carol A
authors.initials: CA
authors.lastname: Paddon
authors.forename: Dagmar
authors.initials: D
authors.lastname: Fuhrer
authors.forename: Marian
authors.initials: M
authors.lastname: Ludgate
mesh_terms: D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D050156:Adipogenesis / Q000187:drug effects / Q000502:physiology*; D002001:Adipose Tissue, Brown / Q000378:metabolism; D049109:Cell Proliferation; D000242:Cyclic AMP / Q000378:metabolism; D005822:Genetic Vectors; D049970:Graves Ophthalmopathy / Q000378:metabolism*; D006801:Humans; D007473:Ion Channels / Q000235:genetics; D050356:Lipid Metabolism; D008071:Lipoprotein Lipase / Q000235:genetics; D024101:Mitochondrial Proteins / Q000235:genetics; D009915:Orbit / Q000378:metabolism; D047495:PPAR gamma / Q000235:genetics / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D011863:Radioimmunoassay; D011989:Receptors, Thyrotropin / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050656:Transcription Factor AP-2 / Q000235:genetics; D000071256:Uncoupling Protein 1
chemical_list: D007473:Ion Channels; D024101:Mitochondrial Proteins; D047495:PPAR gamma; D012333:RNA, Messenger; D011989:Receptors, Thyrotropin; D050656:Transcription Factor AP-2; C000604371:UCP1 protein, human; D000071256:Uncoupling Protein 1; D000242:Cyclic AMP; D008071:Lipoprotein Lipase