Masters Theses

Date of Award

8-1987

Degree Type

Thesis

Degree Name

Master of Science

Major Professor

R. L. Ullrich

Committee Members

R. Julian Preston, Margaret Terzaghi-Howe

Abstract

Considerable evidence indicates that many cellular and molecular alterations can accompany tumor progression. We have developed and characterized a model system to study the multistage neoplastic progression of nonviral carcinogen altered murine mammary epithelial cells. The cell line, EF-42, was derived from mammary tissue of a virgin BALB/c mouse that was exposed to gamma irradiation four weeks prior to tissue removal. Cellular studies reveal that this is a heterogeneous cell line that undergoes in vitro alterations that are associated with neoplastic progression as shown by tumor induction in vivo. At low passage EF-42 cells contain a near 2C DNA content and form normal mammary ductal outgrowths in vivo. With increase in passage, several characteristic changes in cell growth are noted, including; loss of contact inhibition; increase in growth rate; increase in DNA content; and the ability to form tumors when injected into animals. Several morphologically distinct subpopulations have been isolated from EF-42 as single cell clones via a limiting dilution assay. These clonal lines are heterogeneous for many phenotypic characteristics, including: culture morphology, tissue histology, karyotype, growth rate, growth factor response, tumor potential, invasiveness and metastasis. Further, these clonal lines undergo several sequential in vitro alterations, including: decrease in cell size, loss of contact inhibition, dome formation, increase in growth rate, and loss of serum dependence, which correspond with an increase in tumorigenicity in vivo. Chromosomal analysis suggests the presence of cell heterogeneity and karyotypic instability both interclonally as well as intraclonally. These studies also suggest genetic mechanism(s) involving gene amplification and/or chromosome translocations that may play a role in the neoplastic progression of these cells. Karyotypic instability was indicated by changes in chromosome modal number and range, as well as by the presence of metacentrics, dicentrics, and acentric fragments. Double minutes (DMs) and homogeneously staining regions (HSRs), signifying gene amplification units, and metacentric indicator chromosomes (MICs) appear to be associated with the late stages of neoplastic progression. The type and frequency of these chromosomal structures is variable interclonally and intraclonally and there are differences with increase in passage. Comparative studies of cells from tumors obtained from injection of the in vitro clonal lines into animals indicate that in vivo cell selection is involved in tumor formation. Tumor metaphases from five of the six clonal lines studied contain a near diploid chromosome number. A high frequency of all tumor cells contain an MIC as well as DMs and/or HSRs. Three of the clonal lines contain amplified amounts of DMs in tumor cells relative to the in vitro cells. The possible role of specific gene amplification and chromosomal aberrations in the neoplastic progression are currently being studied using this model system.

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