Structure of the thyrotrophin-releasing hormone receptor in human pituitary adenomas.
OBJECTIVE
TRH acts on specific G-protein coupled receptors sited in cells of the anterior pituitary gland. Pituitary tumours expressing either TSH, PRL or GH may respond to TRH by enhanced, blunted or paradoxical hormone release. Non-functioning pituitary tumours may also show abnormal responses to TRH. Little is understood of the mechanisms regulating inappropriate hormone release in these tumours. Activating or inactivating mutations found in G-protein coupled receptors have been implicated in human pathological conditions. Mutations in the G-protein coupled TRH receptor might be involved in the aetiology of pituitary adenomas resulting in aberrant hormone secretion. We therefore screened samples of pituitary adenomas for the presence of somatic mutations in the TRH receptor gene.


METHODS
Pituitary adenoma tissue samples were obtained at surgery from 50 patients with pituitary adenoma (17 acromegaly, 15 prolactinoma, 11 TSH-secreting and 7 non-functioning adenoma) along with blood samples to provide lymphocyte DNA as control sequence.


METHODS
Genomic DNA was extracted from adenoma and lymphocyte samples and the entire coding region of the TRH receptor was amplified using 5 overlapping pairs of PCR primers. The PCR products were analysed for mutations by non-denaturing polyacrylamide gel electrophoresis which reveals single-strand conformational polymorphisms (SSCP) as a mobility shift in product migration. Wild-type and mutant TRH receptor cDNA were similarly analysed to confirm the sensitivity of the method. Additionally, PCR products were ligated into a PCR cloning vector and DNA sequencing carried out to confirm the findings of SSCP analysis.


RESULTS
The human TRH receptor retained normal wild-type sequence in the large group of TSH secreting, PRL secreting, GH secreting and non-functioning pituitary adenomas investigated in this study.


CONCLUSIONS
Our observations suggest that the TRH receptor structure is normal in TSH secreting, PRL secreting, GH secreting and non-functioning pituitary adenomas. It is therefore unlikely that the TRH receptor is involved in the pathology associated with the types of pituitary adenomas investigated in this study. It is possible that some other component of the pathway controlling TRH-signalling events may be implicated in pituitary tumorigenesis.
Faccenda
E
E
MRC Reproductive Biology Unit, Centre for Reproductive Biology, Edinburgh, UK.
Melmed
S
S
Bevan
J S
JS
Eidne
K A
KA
D000172:Acromegaly / Q000235:genetics; D000236:Adenoma / Q000235:genetics; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D005260:Female; D013006:Growth Hormone / Q000378:metabolism; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D010911:Pituitary Neoplasms / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D015175:Prolactinoma / Q000235:genetics; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013972:Thyrotropin / Q000378:metabolism
D017931:DNA Primers; D018025:Receptors, Thyrotropin-Releasing Hormone; D013972:Thyrotropin; D013006:Growth Hormone