TPO gene mutations are the main causes of thyroid dyshormonogenesis [6
]. In the present study, the whole gene scanning of
TPO gene by SSCP and sequencing was performed in 41 patients with permanent congenital hypothyroidism due to dyshormonogenesis. Only one mutation was detected in this group of patients. The reported mutation in this study c.2669G > A (NM_000547.5) is located in exon 15 of TPO gene and results in the hydrophobic glycin (G) to the positively charged arginine amino acid substitution in the TPO transmembrane region. This change is effected on insertion of the TPO enzyme into the plasma membrane of thyroid follicular cells, and results decrease protein activity in the patient [17
]. Both parents were heterozygous for this change so the mutation transmits as autosomal recessive traits in the affected family. Seven different single nucleotide polymorphisms (SNPs) in exons 1, 7, 8, 11, and 15 of the TPO gene were detected too.
As mentioned, different mutations of TPO gene have been reported previously, commonly in exons 8, 9, and 10. The mutation of c.2669G > A (NM_000547.5) has been reported by Avbelj et al. in Slovakia, and thereafter it was not reported in other studies [17
]. Mutation of exon 15 has been reported by Neves et al. in Brazil, but it was c.2630T > C mutation [32
The patient with mentioned mutation in Slovenia had nodular goiter according to the sonographic findings at the age of 16 years [17
]. In this study, the patients had not goiter according to both radiologic and clinical findings at the age of 3 years. Though it may be due to early initiation of treatment, it needs further studies in this field.
In our study, the frequency of TPO gene mutations was lower than Slovene and Portuguese population studies that had similar inclusion criteria, without doing perchlorate discharge test [17
In a population-based study in Japan, Narumi et al. have analyzed the prevalence of TPO gene mutation in fourteen CH patients with dyshormonogenesis and detected two biallelic mutations among them [33
In this study, total iodine organification defect (TIOD) or partial iodine organification defect (PIOD) as defined by the perchlorate discharge test was not determined in studied patients. Some studies have reported that homozygous and compound heterozygous TPO gene mutations are more frequently seen among dyshormonogenic CH patients with TIOD [32
]. It is suggested that low frequency of TPO gene mutations in our studied population may be due to the fact that most of them were dyshormonogenic CH patients with PIOD, which should be investigated in future studies.
In our study, number of patients and determination of permanent CH were similar to previous studies, so low sample size or transient disease cannot be the cause of low frequency of TPO gene mutations.
It is possible that mutations in intronic sequences or in the promoter region and unexamined regulatory regions of TPO gene are the cases of thyroid dyshormonogenesis in these patients. In addition, other genetic disorders may be more effective than TPO mutations in CH patients with dyshormonogenesis including the sodium symporter (NIS) gene, the pendrin gene (PDS), the thyroid oxidase gene 2 (THOX2 or DUOX2), and thyroglobulin gene [35
The technique used in this study, SSCP, is a cheap, simple, and suitable method with a good sensitivity. It has 70–90% sensitivity for the detection of single base substitutions [36
]. In addition, small deletions and insertions in the genome could be identified by this method [37
]. High prevalence of previously reported point mutations in TPO gene and high application of SSCP in detection of this type of mutations with another benefits of this method that is mentioned are reasons to choose SSCP for this study. Approximately 90% of the potential base exchanges are detectable by SSCP under optimal conditions [38
]. In present study, detection of several single nucleotide polymorphisms in different regions of
TPO gene with SSCP and similarly results of direct sequence analysis in four patients showed high sensitivity of SSCP in our study. But SSCP like another mutations screening methods may have percentage of error rate. The limitations of SSCP method are high depen. So in the present study, there is probability of existing mutations that are unidentified with SSCP.
We know, long homozygous deletions in the gene are caused of autosomal recessive disorders. In this group of patients, deletion regions cannot amplify with PCR method and they are detectable with this way, but patients with long heterozygous deletions because of having one normal copy of gene are not detectable with PCR-SSCP method and sequencing analysis. So far, there is no report about long deletions related with TPO gene but the probability of existence of these type of mutationsshould be investigated in studied population in our future studies.
In conclusion, because of low prevalence of TPO gene mutation in this study, it is necessary to investigate more studies with large sample by using another screening method besides SSCP and screening of intronic and regulatory TPO gene mutations and mutation detection of other genes that had effect on thyroid dyshormonogenesis.