RESUMO
Ionizing radiation is a well known risk factor of thyroid cancer development, but the mechanism of radiation induced carcinogenesis is not clear. The RET/PTC oncogene, an activated form of the RET proto-oncogene, is frequently observed in papillary thyroid carcinoma (PTC); RET/PTC1, -2 and -3 are known to be the three major forms. High frequencies of RET/PTC rearrangements have been observed in radiation-associated PTC, such as those appearing post-Chernobyl or post-radiotherapy, but the rearrangement types differ between these two populations. We investigated whether a specific type of RET/PTC rearrangement was induced by X-rays in vivo and in vitro. In human normal thyroid tissues transplanted in scid mice, the RET/PTC1 rearrangement was predominantly detected throughout the observation period (up to 60 days) after X-ray exposure of 50 Gy. On the other hand, RET/PTC3 was detected only 7 days after X-irradiation, and no transcript of RET/PTC2 was detected. These results are supported by the results of an in vitro study. The RET/PTC1 rearrangement was preferentially induced in a dose-dependent manner by X-rays within a high dose range (10, 50 and 100 Gy) in four cell lines. On the other hand, RET/PTC3 was induced at a much lower frequency, and no induction of RET/PTC2 was observed. These results suggest that the preferential induction of the RET/PTC1 rearrangement may play an important role in the early steps of thyroid carcinogenesis induced by acute X-irradiation.
Assuntos
Carcinoma Papilar/genética , Proteínas de Drosophila , Rearranjo Gênico , Neoplasias Induzidas por Radiação/etiologia , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Neoplasias da Glândula Tireoide/genética , Animais , Carcinoma Papilar/etiologia , Proteínas de Fusão bcr-abl/genética , Humanos , Camundongos , Camundongos SCID , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-ret , Neoplasias da Glândula Tireoide/etiologia , Células Tumorais Cultivadas , Raios XRESUMO
The validity of molecular studies using DNA and RNA extracted from decades-old formalin-fixed and paraffin-embedded tissue blocks has been demonstrated. The quality and usability of DNA and RNA from archival tissues are modified by various factors, such as the fixative, the fixation time, and the postmortem time. However, in contrast to DNA, there are no comprehensive studies quantitatively addressing the feasibility of RNA from old (more than 10 years) archival samples. This study examined the integrity of RNA extracted from 738 autopsy liver and 63 autopsy thyroid cancer tissue blocks procured during a span of nearly four decades, beginning in 1952 and ending in 1989, from the atomic bomb survivors. The integrity of RNA was assessed by amplification of c-BCR messenger RNA (mRNA) between two sequential exons with an intervening intron by reverse-transcription polymerase chain reaction (RT-PCR). The integrity of RNA was influenced by the age of the samples and the postmortem time, but not by the formalin-fixation period. It was possible to amplify more than 60% of the samples. Using these RNAs, the HCV genome in liver cancers and the H4-RET gene in thyroid cancers were detectable. This study illustrates the possibility of molecular studies using RNA from routinely prepared paraffin blocks stored for long periods and provides the statistics and critical factors to consider in assessing the feasibility of such contemplated studies.
Assuntos
Proteínas de Drosophila , Hepacivirus/genética , Neoplasias Hepáticas/genética , Proteínas Tirosina Quinases , RNA Neoplásico/análise , RNA Viral/análise , Neoplasias da Glândula Tireoide/genética , Estudos de Viabilidade , Hepatite C/epidemiologia , Hepatite C/genética , Hepatite C/virologia , Humanos , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/virologia , Guerra Nuclear , Inclusão em Parafina , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcr , Proteínas Proto-Oncogênicas c-ret , RNA Mensageiro/biossíntese , RNA Neoplásico/isolamento & purificação , RNA Viral/isolamento & purificação , Receptores Proteína Tirosina Quinases/genética , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias da Glândula Tireoide/patologia , Bancos de TecidosRESUMO
Significance of the binding of hexokinase to mitochondria was examined with respect to stabilization of the enzyme by the binding. Stability during the incubation of the mitochondria-bound forms of hexokinases I and II, both prepared from Ehrlich-Lettre ascites hyperdiploid tumor cells (ELD cells), were compared with that of the corresponding free forms. During the incubation at pH 7.4 and 37 degrees C up to 60 min, hexokinase activities decreased gradually, and the decrease in the activity of the free form was much more marked than that of the bound form for both hexokinases. Hexokinase II was much less stable than I, and the activity of the free form of the former was almost lost by the incubation for 15 min. But, more than a half of the original activity of hexokinase II was retained even after 60 min of the incubation when the enzyme was bound to mitochondria. Addition of 50 mM glucose increased the stability of hexokinase II, but the stabilizing effect was less marked for hexokinase I. On the other hand, addition of 28 mg/ml of bovine serum albumin markedly stabilized hexokinase I to almost the same extent as was observed with mitochondria. On the contrary, the serum albumin had little stabilizing effect on hexokinase II. These findings indicate that the binding to mitochondria stabilizes the hexokinases of ELD cells, though the stability is different by nature between hexokinases I and II.
