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Hölzlwimmer, Gabriele (2007): Histologische und molekulargenetische Untersuchungen von strahleninduzierten Schilddrüsentumoren im Mausmodell. Dissertation, LMU München: Tierärztliche Fakultät



Thyroid cancer derived from follicular epithelial cells is the most common endocrine malignancy in man. An increased incidence of predominantly papillary thyroid carcinomas (PTC) was found in children exposed to radiation after the Chernobyl nuclear accident in 1986. Therefore, in this study, the goal was to establish a mouse model of thyroid carcinogenesis, based on a standardized histological classification scheme for the murine thyroid tumors, and complemented by molecular genetic analyses. In previous studies, radioiodine (I131, 111 kBq) was injected into iodine deficient fed mothers of various mouse strains (F1-hybrids and backcrosses of C57/BL6, C3H, BALB/c, and JF1). The first injection was applied during gestation and the second during lactation. The necropsy tissue was submitted for the analysis in this study. A set of 365 thyroid glands (203 irradiated and 162 control mice) was histological examined following the current WHO classification of human thyroid tumors (2004) for comparative purposes. The irradiated mice showed 24 % of cases with simple hyperplasia (SH), 20 % with nodular hyperplasia (NH), 7 % with follicular thyroid adenoma (FTA), and 5 % with follicular thyroid carcinoma (FTC) whereas in the control group only 3 % SH, 3 % of NH, 1 % of FTA, and 1 % of FTC were observed. Interestingly, no PTC was diagnosed in the mice, which is the most frequent irradiation-related type of thyroid cancer in human. Therefore, the histological type of the radiation-associated thyroid tumors in mice differs from that in human. However, some cases of murine FTC presented PTC-like biological behavior. In addition to the significant increase of hyperplasias in irradiated mice, most of the FTC (82 %) arose amongst a background of hyperplastic nodules. Therefore, a progression from NH to FTC, based on genetic instability, cannot be ruled out. The following molecular methods were used: PCR- (polymerase chain reaction-) based loss of heterozygosity (LOH), comparative genomic hybridization (CGH), and fluorescence-in-situ-hybridization (FISH). Since the CGH-study in mice using formalin-fixed paraffin-embedded tissue (FFPE) is not yet established, an important part of the study was dedicated to evaluate this methodology. The LOH-study was performed with thyroid gland tissue from 40 mice (seven normal thyroid glands, 12 SH, 10 NH, 10 FTA, and one FTC) using 36 microsatellites for nine different loci. With the exception of an LOH with a single microsatellite on chromosome 14 in 40 % of NH, LOH was found in 75 % of the irradiated male mice with H6F1-background on chromosomes 4, 5, 6, 11, 14, and / or 19. This suggests the existence of a mouse strain specific genetic predisposition, which influence on the genetic stability. One of the FTA (an atypical FTA) was highly suspicious for a deletion of the tumorsuppressorgene Rb1 (supported by intragenic FISH-analysis), which could play an important role in the thyroid carcinogenesis. For the CGH-study, thyroid tissue derived from 21 different mice (F2-hybrids) was analyzed (two normal thyroid glands, one SH, 12 NH, two FTA, and eight FTC). In 46 % of the hyperplasias, small chromosomal gains and losses located on different chromosomes were observed; suggesting that there exists a genetic instability, which may lead eventually to malignant progression. Regional polyploidies on chromosomes 4 and 5 were demonstrated in one of the FTAs, which could be a hint for the location of oncogenes. Taken together, in FTA development there is a broad spectrum of genetic alteration, and by inference mechanisms. In contrast, the FTC exhibited a significant increase of specific aneuploidies, mainly deletions of the chromosomes 4 (88 %), 9 (50 %), and 14 (38 %). Identical alterations of chromosomes 4 and 9 were also observed in the one case of an FTC from a non-irradiated mouse. These data indicate that irradiation, most probably, increases the frequency of genetic changes, but does not change the type of genetic alterations, which play a crucial role in thyroid carcinogenesis in mice. A better understanding of molecular genetics involved in thyroid tumorigenesis in standardized mouse models may give insight into the pathogenesis of the various tumor types. Together with the results from human pathology and in vitro studies, this may lead to a better knowledge about the molecular pathways with diagnostic, prognostic, and therapeutic relevance. The results of this study demonstrate a morphological and genetical difference between human (PTC) and murine (FTC) radiation-associated thyroid tumors, but a strong similarity to the human follicular tumors. Therefore, this mouse model serves as a good model of carcinogenetic mechanisms, tumor induction, and progression in the human follicular tumors FTA and FTC, resulting from the cooperative effect of radioiodine exposition and iodine deficiency.