A molecular analysis and long-term follow-up of two siblings with severe congenital hypothyroidism carrying the IVS30+1G>T intronic thyroglobulin mutation / Análise molecular e acompanhamento a longo prazo de dois irmãos com hipotireoidismo congênito portadores da mutação intrônica IVS30+1G>T no gene da tireoglobulina

OBJECTIVE: To extend the molecular analysis of the IVS30+1G>T intronic thyroglobulin (TG) mutation, and to report the eleven year follow-up of the affected patients. METHOSD: Two siblings with severe congenital hypothyroidism with fetal and neonatal goiter, harboring the IVS30+1G>T mutation were included. Nodular and non-nodular thyroid tissue specimens were collected. Specific thyroid genes expression was evaluated by real-timePCR and by immunohistochemistry. RESULTS: In non-nodular tissue specific thyroid genes mRNA were reduced when compared to normal thyroid sample. In the nodule, TPO and NIS expression was very low. Microscopic examinations showed very large follicular-lumina and swollen vesicles of endoplasmatic-reticulum. Strong cytoplasmatic and low follicular-lumen TG immunostaining were detected. Intracellular NIS, membrane TPO and TSHR immunostaining had higher positivity in non-nodular sample. Both patients had a long-term adequate developmental outcome, besides one patient have been lately-treated. CONCLUSIONS: IVS30+1G>T mutation not only lead to very enlarge endoplasmatic-reticulum, but also to alterations of specific thyroid genes expression. The clinical evolution of patients harboring these mutations strengthen the concept of the influence of environment, like iodine nutrition, to determine the final phenotypic appearance.

OBJETIVO: Aprofundar a análise molecular da mutação intrônica IVS30+1G>T do gene tireoglobulina (TG) e relatar a clínica de pacientes portadores da mutação, acompanhados por 11 anos. MÉTODOS: Foram estudados dois irmãos com hipotireoidismo congênito grave com bócio fetal e bócio neonatal, portadores da mutação IVS30+1G>T. Foram coletadas amostras de tecido nodular e não-nodular. Avaliou-se a expressão de genes específicos da tireóide por PCR em tempo real e imunohistoquímica. RESULTADOS: A expressão de genes específicos da tireóide foi menor no tecido não-nodular que no tecido normal controle. Expressões de TPO e NIS foram extremamente baixas no tecido nodular. Verificou-se lúmen folicular aumentado com grandes vesículas de retículo endoplasmático, e detectou-se forte marcação de TG no citoplasma e fraca no lúmen folicular. No tecido não-nodular observou-se forte positividade de NIS intracelular e, TPO e TSHR na membrana plasmática. O acompanhamento em longo prazo dos pacientes mostrou adequado desenvolvimento, apesar de um deles ter recebido tratamento tardio. CONCLUSÕES: A mutação IVS30+1G>T não só promove alterações no retículo endoplasmático, como alterações na expressão de genes específicos da tireóide. A evolução clínica destes pacientes reforça o conceito da influência do meio ambiente, como o aporte nutricional de iodo, no fenótipo final.

Morphometric analysis of the fate of secretory epithelial cells in the ventral lobe of the rat prostate during massive apoptosis

Orchiectomy causes marked, rapid involution of the prostatic secretory epithelium. Concurrently, macrophages, which in normal glands are small and rarely occur at the base of the secretory epithelium, increase in size and number. Apoptotic cells are engulfed by companion epithelial cells and also by macrophages. In secretory cells and macrophages, dense bodies progressively increase in number and store membranes derived from dead cells of the secretory epithelium. In this work, we examined the contributions of the various routes of disposal of demised secretory epithelial cells of the rat prostate, induced to enter in apoptosis by retrieval of androgen. Specifi cally, we sought to determine how much membrane surface area derived from apoptotic cells of the secretory epithelium could be stored in dense bodies, and how these data compared with the disposal of dead cells via the glandular lumen. Glands from unoperated controls (day 0) and from rats examined 12 h and 1, 2, 3, 4, 5, 6, 7, 8, and 9 days after orchiectomy were studied morphometrically. The total membrane surface area of rough and smooth endoplasmic reticulum, Golgi apparatus, mitochondria and vesicles declined from 6.75 x 103 ìm2 in non-castrated rats to 1.12 x 103 ìm2 nine days after castration. Similarly, the total surface area of the secretory epithelium decreased from 10.6 x 1011 ìm2 in non-castrated rats to 0.204 x 1011 ìm2 nine days after castration. Geometrical models revealed that 1 ìm3 of dense body accommodated at least 142 ìm2 of myelin-like membrane surface area. Three to four days after castration, the total volume of intramacrophage dense bodies peaked (~5 x 106 ìm3) and represented 1-2% of the volume of intraepithelial dense bodies (~4 x 108 ìm3). The minimum membrane surface area that could be stored in dense bodies of the secretory epithelium on post-castration days 0, 1, 2, 3, 4 and 9 was 1.4%, 9%, 16%, 23%, 28% and 44%, respectively, of the total membrane surface area of the...