Tonacchera Massimo, Banco Maria Elena, Montanelli Lucia, Di Cosmo Caterina, Agretti Patrizia, De Marco Giuseppina, Ferrarini Eleonora, Ordookhani Arash, Perri Anna, Chiovato Luca, Santini Ferruccio, Vitti Paolo, Pinchera Aldo
Dipartimento di Endocrinologia e Metabolismo, Centro di Eccellenza AmbiSEN, Università di Pisa, Pisa, Italy.
Clin Endocrinol (Oxf). 2007 Jul;67(1):34-40. doi: 10.1111/j.1365-2265.2007.02831.x. Epub 2007 Apr 15.
To analyse the coding region of PAX8 in individuals with congenital (CH) or post neonatal hypothyroidism due to dysgenetic (TD) or eutopic thyroid glands.
Forty-three children with CH and TD (13 agenesis, 23 ectopia, and seven hypoplasia), one subject with post neonatal onset of hypothyroidism and thyroid ectopia, 15 children with CH and eutopic thyroid glands and six euthyroid adults with thyroid hemiagenesis were enrolled as cases, along with 120 healthy individuals as controls.
Exons 2-8 of the PAX8 were directly sequenced. HeLa and HEK293 cells were transfected with PAX8 wild-type (PAX8-WT), mutant PAX8, p300, thyroid transcription factor 1 (TTF-1) and thyroglobulin promoter pGL3 (TG prom-pGL3). Synergism of TTF-1 with PAX8-WT vs. mutant and activity of PAX8-WT vs. mutant in accompaniment with p300 on TG prom-pGL3 were also assessed. The luminescence produced by PAX8-WT and mutant PAX8 was measured.
Among patients and controls only a 15-year-old girl with thyroid ectopia showed a heterozygous transition of cytosine to thymine at position 674 in exon 6, which changed a conserved threonine at position 225 to methionine (PAX8-T225M). Her father and sister harboured PAX8-T225M without abnormal thyroid phenotypes. PAX8-T225M and PAX8-WT similarly increased luciferase activity and had a similar synergistic effect with TTF-1. At 500 ng p300, however, PAX8-T225M could not significantly increase TG promoter activity when compared to PAX8-T225M alone, while PAX8-WT +500 ng p300 induction was significantly higher than PAX8-WT alone (P < 0.001). Cotransfection of TTF-1 together with PAX8-T225M resulted in rescuing of the lack of synergism with p300.
PAX8 mutations in congenital hypothyroidism due to dysgenetic or orthotopic thyroid glands are rare. PAX8-T225M is probably a rare variant.
分析先天性（CH）或新生儿期后因甲状腺发育不全（TD）或甲状腺位置正常所致甲状腺功能减退患者中PAX8基因的编码区。
纳入43例CH和TD患儿（13例甲状腺缺如、23例异位甲状腺和7例甲状腺发育不全）、1例新生儿期后发生甲状腺功能减退且甲状腺异位的患者、15例CH且甲状腺位置正常的患儿以及6例甲状腺半侧发育不全的甲状腺功能正常成人作为病例组，同时纳入120例健康个体作为对照组。
对PAX8基因的第2至8外显子进行直接测序。将PAX8野生型（PAX8-WT）、突变型PAX8、p300、甲状腺转录因子1（TTF-1）和甲状腺球蛋白启动子pGL3（TG prom-pGL3）转染至HeLa和HEK293细胞。还评估了TTF-1与PAX8-WT相比与突变型PAX8的协同作用，以及PAX8-WT与突变型PAX8在与p300共同作用下对TG prom-pGL3的活性。测量PAX8-WT和突变型PAX8产生的荧光。
在患者和对照组中，仅1例15岁甲状腺异位女孩在外显子6的674位发生了胞嘧啶向胸腺嘧啶的杂合转变，该转变将225位保守的苏氨酸变为甲硫氨酸（PAX8-T225M）。她的父亲和姐姐携带PAX8-T225M，但无异常甲状腺表型。PAX8-T225M和PAX8-WT同样增加荧光素酶活性，且与TTF-1具有相似的协同作用。然而，在加入500 ng p300时，与单独的PAX8-T225M相比，PAX8-T225M不能显著增加TG启动子活性，而PAX8-WT + 500 ng p300诱导的活性显著高于单独的PAX8-WT（P < 0.001）。TTF-1与PAX8-T225M共转染可挽救与p300协同作用的缺失。
因甲状腺发育不全或位置正常所致先天性甲状腺功能减退中的PAX8突变罕见。PAX8-T225M可能是一种罕见变异。
