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Mutations in the TSH receptor gene of autonomous nodules, were searched
for by Nogueira and her colleagues from Sào Paulo,
Brazil. It is well known that constitutively activating thyrotropin receptor
mutations (TSHR) have been identified as the molecular cause of solitary
thyroid adenomas. It was also recently reported that a somatic point mutation
of the stimulatory guanine nucleotide binding protein (Gs-alpha) may cause
constitutive activation of the adenylate cyclase (AC) cascade in thyroid
adenomas. Tissues from five patients with hyperfunctioning adenomas were
screened for TSHR and for Gs-alpha gene mutations. DNA was extracted from
autonomous adenomas and normal peri-adenomatous thyroid tissue. The exons
9 and 10 of the TSH receptor gene were amplified by PCR and direct sequencing
was performed with fluorescently labeled primers and analysis on automated
sequencer. Four of five adenomas harbored a mutation. In three cases substitution
of alanine 623 (GCC) by valin (GTC) in the third intracellular loop of
the receptor was identified. In one case a substitution was observed of
aspartic acid 633 (GAC) by tyrosine (TAC) in the 6th transmembrane domain
of the receptor. This patient also had a polymorphism at position 727 where
aspartic acid (GAC) was replaced by glutamic acid (GAC). No mutations were
found in the GS-alpha gene. These results demonstrate a high incidence
of TSH receptor mutations in toxic adenomas from Brazil (80%). This is
in line with reports from Belgium (mildly iodine deficient area) but are
in contrast with reports from other areas (Japan, Italy, USA) where these
mutations are found only infrequently.
By contrast Carvalho and co-workers, also from Sào Paulo, Brazil, have not found mutations in the "hot spot" regions of the TSH receptor of autonomous nodules. They analyzed the mRNA encoding TSHR obtained by fine needle aspiration biopsy (FNAB) extracting total RNA from nodular and paranodular tissues using acid guanidinium-thiocyanate-phenol-chloroform. cDNA was obtained by reverse transcriptase. TSHR was amplified by PCR from the third cytoplasmic loop to the sixth transmembrane segment. The amplified fragments were cooked for mutations in aminoacids 619, 631, 632 and 633. The authors did not find any mutation in the 11 patients, which indicates that the presence of constitutively activating TSHR mutations in these aminoacids in AFTNs appears to be low in Sào Paulo, a result comparable with a Japanese study (1/45 nodules) and different from several European studies (7/11 in Naples and 18/22 in Brussels). This discrepancy can be related to the presence of mutations other than 619, 631, 632 and 633 in our population or to mutations in other steps of the stimulatory chain of the thyroid cell, like in GS-alpha, ras or other oncogenes. The study by Carvalho et al. (Sào Paulo, Brazil) has methodological limitations due to the use of samples obtained from needle aspiration as well as the use of Southern blot analysis in lieu of sequencing the cDNA of the TSH receptor. Thyroid hypoplasia is a common cause of congenital hypothyroidism and Nogueira and co-authors (Sào Paulo, Brazil) performed molecular analysis of the TSH receptor in these patients. Screening of neonatal hypothyroidism has indicated that, congenital hypothyroid neonates commonly have thyroid agenesis or an ectopic thyroid (about 60% of all cases). Another cause of neonatal hypothyroidism is the presence of hypoplastic thyroid tissue, normally present in the anterior portion of the neck. Lack of response of thyroid tissue to elevated levels of endogenous TSH may be the cause of this anomaly. The authors have hypothesized that patients with congenital hypothyroidism and normally localized hypoplastic thyroid glands may have inactivating mutations in the TSH receptor. Therefore they studied such 6 patients with congenital hypothyroidism and their direct relatives. Genomic DNA was extracted and all exons (1-10) encoding the TSH receptor cDNA with intronic primers (exons 1-9) and exonic primers (exon 10) were amplified by PCR and direct sequencing with fluorescently labeled primers and analysis on automated sequencer. No mutations were found in the coding region of the TSH receptor of these patients and their direct relatives. The polymorphisms of the TSH receptor found in previous studies were detected in some of these patients and probably has no direct relationship with the pathophysiologic conditions leading to this anomaly. In conclusion, somatic mutations (not germline) in the TSH receptor may be postulated as cause of sporadic congenital hypothyroidism due to thyroid hypoplasia. |
