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Title: Karyotypic analysis of cervical neoplasia : chromosomal aberrations and human papillomavirus infection
Author: Southern, Shirley Anne
ISNI:       0000 0001 3473 1133
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2000
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Human papillomavirus (HPV) infection has been widely implicated in cervical carcinogenesis, but it appears to be an early event, with other genetic abnormalities required for biological transformation. The initial aim of this thesis was to examine the relationship between HPV presence and type, and numerical chromosome abnormalities in 60 low and 30 high grade squamous intraepithelial lesions (SIL) and 25 invasive squamous carcinomas. This involved the technical development of; i) a sensitive HPV in situ hybridisation technique; ii) a PCR system for accurate determination of HPV type; and iii) interphase cytogenetic methodology using pericentromeric probes to chromosomes 1, 3, 11, 17, 18 and X. Eleven of 33 low grade lesions infected with high risk HPVs (HPV 16, 18, 30, 31, 33, 35, 39, 45, 51, 52, 56, 58 and 66) but none of 24 lesions infected with low risk HPVs (HPV 6, 11, 42, 43 and 44) showed basal cell tetrasomy of all six chromosomes in the HPVinfected areas. Of the high grade lesions, all of which were infected with high risk HPV, 8 showed basal cell tetrasomy and 6 were aneusomic. Most of the invasive carcinomas contained high risk HPV and were also aneusomic. Two distinct patterns were observed; i) mainly disomic with additional loss/gain of individual chromosomes; and ii) mainly tetrasomic with additional loss/gain of chromosomes. This suggested that two different pathways/mechanisms are involved in neoplastic transformation and that tetrasomy may in some lesions, be an intermediate state. Cell cycle progression and genome maintenance are regulated by cyclins and cyclin-dependent kinases. These not only provide controlled passage through the cell cycle but also provide 'checkpoints' or restraints to prevent replication of damaged DNA. In vitro studies have shown that E6 and E7 oncoproteins can bind cell cycle regulatory proteins and disrupt cell cycle control. This not only accommodates viral replication but may also contribute to the accumulation of chromosome abnormalities that lead to immortalisation and transformation. In the second part of this thesis, the relationship between the expression of cyclins D1, E, A and B1 and both the HPV type and the presence of numerical chromosome abnormalities was examined. The expression of cyclins E, A and B was elevated in lesions infected with both low and high risk viruses. Cyclin D1 expression was absent in the majority of lesions infected with high risk HPVs, in contrast with overexpression observed in the low risk infected lesions. Taken together with the chromosome abnormalities, the data provided evidence that the G;S and G2M cell cycle checkpoints are abnormal in lesions infected with high risk HPVs but not in those infected with low risk HPVs. Finally, raft culture systems of human keratinocytes that were; i) transfected with whole HPV 18 viral genomes; and ii) acutely infected with recombinant retroviruses expressing E6, E7 or E6/E7 genes of HPV 18, were analysed for karyotype and cyclin expression to investigate which HPV genes induced tetrasomy, and under what cell cycle conditions. The data indicated that tetrasomy occurs in association with E7 gene expression.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: Carcinogenesis; HPV