Ανοσοϊστοχημική μελέτη γονιδίων σε επιμύες και σχετιζόμενοι με αυτά μηχανισμοί σταδιακής ογκογένεσης στη στοματική κοιλότητα

Abstract

Oral squamous cell carcinomas (OSCC) comprise the vast majority of cancers developing in the oral cavity. Oral oncogenesis is a multi-step procedure that requires the accumulation of several genetic alterations in oncogenes and tumour suppressor genes that result in destabilization of several growth control systems. The progression to malignancy includes sequential pathological alterations ranging from hyperplasia through dysplasia to carcinoma in situ and invasive carcinoma. Taking into consideration the high incidence of OSCC worldwide, the fact that prognosis remains unfavourable despite the substantial developments in both diagnostic and therapeutic possibilities and that OSCC can be cured in 80-90% of the cases when detected early, it would be of paramount importance to advance our understanding of molecular mechanisms involved in the induction and progression of this type of cancer in order to determine early and more accurate diagnostic and prognostic markers and achieve a more effective treatment. Objective: In this disquisition, the expression of the oncogene H-ras, the tumour suppressor gene p53 and the proliferation marker Ki-67, their role and probable correlation in sequential histological grades of oral carcinoma formation was studied in an experimental model in Syrian golden hamsters. The selection of these genes relied on the critical role they play in cellular functions such as cell cycle, DNA repair and apoptosis and their involvement in human carcinogenesis in general. Materials and Methods: Thirty-seven male Syrian golden hamsters were divided into three experimental groups (A,B,C;10 animals each) and one control group (7 animals). The left buccal pouches of animals in groups A, B and C were painted three times per week for 14 weeks with carcinogen 0.5% 9,10-dimethyl-1,2-benzanthracene (DMBA) and were excised at 10, 14, 19 weeks respectively. The left buccal pouch of control animals was excised following a ten-week period without carcinogen application. The biopsies were classified pathologically. There was an evaluation of the entire section and of the histological grades in each section. The presence of proteins H-ras, p53 and Ki-67 was evaluated immunohistochemically in all biopsies. Statistical analysis was performed per group and in each histological grade. Results: A progression towards OSCC formation in correlation to increased time of carcinogen application was evident either macroscopically or microscopically. The time of 14 weeks for carcinogen application and the latency time of 5 weeks until the animals were sacrificed (19 weeks totally) proved to be ample for the development of moderately differentiated OSCC, that has not been observed in other studies with the same experimental model. There were no samples with poorly differentiated carcinoma. A series of 122 lesions from 37 hamsters (15 normal mucosa, 5 hyperkeratosis, 16 hyperplasias, 21 dysplasias, 18 early invasions, 23 well differentiated carcinomas and 24 moderately differentiated carcinomas) was studied (methodology originality). H-ras expression decreased from dysplasia to well-differentiated carcinomas and from group A to B. This reduction was not persistent further on, as in moderately-differentiated carcinomas and in group C, H-ras expression increased again. The expression of mutant p53 increased significantly in oral mucosal hyperplasia and dysplasia in comparison to normal mucosa and in group A compared to control group, and then reached a plateau. Ki-67 expression increased sharply in initial stages of oral carcinogenesis, significantly decreased in well differentiated carcinoma, but elevated again in moderately differentiated carcinoma. There was a linear correlation between p53 and Ki- 67 expression, especially in precancerous lesions. In the later stages of oral oncogenesis there was a correlation 123 between Ki-67 and H-ras expression. No correlation between the expression of H-ras and mutant p53 was observed. Conclusions: ?his experimental model seems valid concerning the gradual development of OSCC and could largely contribute to the better understanding of the mechanisms involved in the several stages of oral carcinogenesis. It is demonstrated that the induction of poorly differentiated oral carcinoma requires a longer period of carcinogen application than 14 weeks or a later excision of the buccal pouches than 19 weeks. H-ras overexpression seems to be associated with later stages of OSCC development, since other genetic alterations have preceded and accumulated. A different signalling pathway, not involving H-ras, may be implicated in the pathogenesis of OSCC. The expression of p53 signals for an oral lesion prior to invasion. Cell proliferation is more intense in precancerous stages of oral cancer, while it reverts in the later stages of invasive carcinoma playing a role in its progression. The mutant p53 protein directly affects cell proliferation, inducing its increase. P53 and Ki-67 expression could be used in assays as markers of early recognition of precancerous oral lesions