Cytogenetic, morphologic and oncogene analysis of a cell line derived from a heterologous mixed mullerian tumor of the ovary

Abstract

A cell line was established from a mixed mullerian tumor of the ovary and designated LN1. Histopathologic analysis of the fresh tumor specimen demonstrated a highly aneuploid heterologous tumor comprised of undifferentiated mesodermal components with carcinomatous cells present as a smaller population. Long-term in vitro culture resulted in the establishment of a cell line that exhibits an epithelial-like morphology and expresses epithelial antigens cytokeratin, epithelial membrane antigen, and carcinoma antigen TAG-72. These cells also express mesenchymal intermediate filaments, vimentin, and desmin. Karyotypic analysis revealed a basic triploid pattern with multiple chromosomal abnormalities, most notably an isochromosome of the short arm of five present in three copies. Analysis of oncogene expression revealed that LN1 cells constitutively express mRNA for c-ras, c-erbB2, and p53. The expression of mRNA for cellular oncogenes correlated with the presence of corresponding oncoproteins, p21H-ras, p21K-ras, and p185erB2 and mutant p53 protein. In summary, coexpression of epithelial and mesenchymal antigens by LN1 cells lends support to the hypothesis that epithelial and mesenchymal elements comprising mixed mullerian tumors of the ovary are derived from a common stem cell precursor. Furthermore, this cell line represents a functional in vitro model to evaluate the biologic activities of these unusual and highly aggressive ovarian malignancies.

Three-dimensional culture of a mixed mullerian tumor of the ovary: Expression of in vivo characteristics

“…The expression of mRNA for HER-2/ neu, H-ras, K-ras, and p53 was determined by reverse transcription (RT) polymerase chain reaction (PCR) amplification (2). Briefly, cells grown for 32 d in the RWV were removed from microcarrier beads by digestion for 30-45 rain at 37 ° C in a solution of 2.4 U/ml dispase (Boehringer Mannheim) containing 20 mg bovine serum albumin (BSA) fraction V (Sigma).…”

“…Briefly, cells grown for 32 d in the RWV were removed from microcarrier beads by digestion for 30-45 rain at 37 ° C in a solution of 2.4 U/ml dispase (Boehringer Mannheim) containing 20 mg bovine serum albumin (BSA) fraction V (Sigma). Cells were washed by centrifugation four times with PBS (GIBCO, Grand Island, NY), after which total RNA was extracfed with TriReagent (MRC, Inc., Cincinnati, OH), reverse transcribed to cDNA, and amplified by PCR as described (2). All reagents for reverse transcription were obtained from Promega Corp. (Madison, WI).…”

“…Intermediate filament proteins (cytokeratin, vimentin), epithelial membrane antigen, TAG-72, and oncoproteins (p21H-r% p21K-r% and mutant p53) present within LN1 cells were determined flow cytometrically utilizing a Becton Dickinson FACScan (Mountain View, CA) as described (2). Monoclonal antibodies to p21H-r~*, p21K-~, and p53 mutant protein were obtained from Oncogene Science (San Diego, CA).…”

Characterization of Primary Breast Carcinomas Grown in Three-Dimensional Cultures

“…DNA ploidy and cell cycle were assessed using a modified Krishan procedure, as previously described [19,20]. Data were analyzed with FACScan Cellfit software (Becton-Dickinson) for doublet discrimination and with Lysys II software (Becton-Dickinson) for multiparameter analysis.…”

Temperature‐sensitive Ovarian Carcinoma Cell Line (OvBH‐1)

“…12) Reports on alterations in the p53 gene in ovarian cell lines are scarce. 2,[13][14][15][16][17] Some authors reported point mutations in the highly conserved DNA binding domain encoded by exons 5-8 of the gene, resulting in single amino acid changes. 2,13,15,18) However, no data are available concerning the p53 status in ovarian clear cell carcinoma cell lines.…”