title: Real time RT-PCR analysis of thyroglobulin mRNA in peripheral blood in patients with congenital athyreosis and with differentiated thyroid carcinoma after stimulation with recombinant human thyrotropin.
abstract: OBJECTIVE
  Thyroglobulin (Tg), measured by immunometric assay, is the most sensitive and widely used clinical marker for thyroid cancer progression and relapse. However, these Tg determinations are of limited sensitivity and susceptible to interference by Tg autoantibodies. As a possible diagnostic alternative, we tested a real time RT-PCR protocol to determine Tg mRNA levels in peripheral blood.
  
  
  METHODS
  Tg mRNA was determined by real-time RT-PCR using total RNA from peripheral blood. Tg mRNA blood levels were calibrated to the mRNA encoding the housekeeping enzyme glyceraldehyde phosphate dehydrogenase (GAPDH); pooled blood from ten healthy subjects served as a RT-PCR positive control.
  
  
  RESULTS
  Tg mRNA and serum Tg were detected in twelve patients with differentiated thyroid cancer (DTC) after thyroidectomy and radioiodine therapy, however, there was no correlation with the clinical stage. An increase in Tg mRNA and protein was observed after application of recombinant human thyrotropin (rhTSH) in four patients with DTC stimulated with rhTSH for postoperative follow up. Tg mRNA and protein were also detected in four congenital athyreotic patients. Analysis of Tg mRNA levels using a commercial multiple tissue Northern blot revealed Tg hybridization signals in several extrathyroidal tissues (salivary gland, trachea, kidney, pancreas, adrenal gland, etc.).
  
  
  CONCLUSIONS
  Our data suggests that RT-PCR detects Tg mRNA of extrathyroidal origin, from leukocytes or from metastasizing carcinoma cells under basal conditions or after TSH stimulation. However, considering the marked and highly variable individual Tg mRNA backgrounds, interpretation of real time PCR results requires caution. This limits the clinical use of Tg mRNA determination by real time PCR to an individual tumor progression marker in follow-up.
authors.affiliation: Institut für Experimentelle Endokrinologie, Charité, Universtätsmedizin Berlin, 10117 Berlin, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Kaufmann
authors.forename: Cornelia
authors.initials: C
authors.lastname: Schmutzler
authors.forename: Lutz
authors.initials: L
authors.lastname: Schomburg
authors.forename: Carsten
authors.initials: C
authors.lastname: Körber
authors.forename: Markus
authors.initials: M
authors.lastname: Luster
authors.forename: Johann
authors.initials: J
authors.lastname: Rendl
authors.forename: Christoph
authors.initials: C
authors.lastname: Reiners
authors.forename: Josef
authors.initials: J
authors.lastname: Köhrle
mesh_terms: D015152:Blotting, Northern; D018450:Disease Progression; D005500:Follow-Up Studies; D006801:Humans; D025202:Molecular Diagnostic Techniques; D012333:RNA, Messenger / Q000097:blood*; D011863:Radioimmunoassay; D011994:Recombinant Proteins / Q000627:therapeutic use; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013954:Thyroglobulin / Q000097:blood / Q000235:genetics*; D013961:Thyroid Gland / Q000002:abnormalities*; D013964:Thyroid Neoplasms / Q000097:blood*; D013965:Thyroidectomy; D013972:Thyrotropin / Q000627:therapeutic use*
chemical_list: D012333:RNA, Messenger; D011994:Recombinant Proteins; D013972:Thyrotropin; D013954:Thyroglobulin