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

Abstract

OvBH-1 cells from a patient with ovarian clear cell carcinoma were established and their biochemical status was analyzed. Cells grown at 37°C exhibited normal cell cycle distribution, whereas the cells shifted to 31°C were arrested in the G 2 /M phase of the cell cycle. Immunochemical analysis using anti-p53 antibodies (DO-1, PAb240, PAb421, and PAb1620) revealed that only the DO-1 antibody reacted with p53 with a high and similar percentage at both temperatures. PAb240 reacted with a low percentage of cells at 37°C and no reaction was observed at 31°C. PAb421 antibody stained a significantly lower percentage of cells at 37°C than at 31°C. Cells were not stained with PAb1620 antibody and were negative for antibodies against p21 WAF1 and MDM2 proteins independently of the temperature. Sequencing of all coding exons of the p53 gene demonstrated only a neutral genetic polymorphism, i.e. a G-to-A substitution (GAG to GAA) at nucleotide position 13 432. Thus, the observed temperature sensitivity of OvBH-1 cells cannot be ascribed to a p53 primary structure mutation. Based upon immunochemical analyses, we consider, however, that p53 in nuclei of OvBH-1 cells is in a highly unstable conformation. Furthermore, the N-terminal portion of the p53 protein at Ser20 has not been modified, and Lys373 and/or Ser378 of the C-terminus is acetylated and/or phosphorylated. The nuclear location signal of p53 is preserved. Induction of MDM2 protein is uncoupled from the cell regulatory machinery and the induction of p21 WAF1 by p53 is impaired in OvBH-1 cells.

Comparative Analysis of Primary Ovarian Cancer Cells and Established Cell Lines as a New Tool for Studies on Ovarian Cancer Cell Complexity

“…Based on published data, it can be assumed that only the cell clones with unique stemness features were selected during the immortalization procedure [26,27]. This suggestion might be confirmed by the high nuclear accumulation of the p53 protein found in the majority of immortalized cells, which indicated that only malignant cells were able to survive and proliferate [7]. Strong p53 immunopositivity in immortalized cells found in the current study indicates that the selection of cells carried the TP53 gene mutation, similar to the published data on the correlation between p53 overexpression and gene mutation [28][29][30].…”

“…One such procedure is spontaneous immortalization, which occurs during the numerous passages of primary cells (hundreds of cell passages). An example of spontaneous immortalization is the ovarian cancer OvBH-1 cell line derived from the ascitic fluid of a patient with an ovarian clear-cell adenocarcinoma [7] and a high-grade serous ovarian cancer cell line OVPA8 [8]. Another way to obtain a cell line is to immortalize the cells using vectors carrying telomerase or Simian virus 40 (SV40) genes.…”

Growth inhibition and apoptosis induction in ovarian cancer cells

“…We found that both cell lines expressed high levels of p53 indicating a stabilization and inactivation of the protein. In the case of OvBH-1 cells this stabilization is not due to a mutation in the p53 gene because sequence analysis revealed only a neutral genetic polymorphism (17,39). OV-MZ-32 cells express a p53 mutant with a deletion between amino acids 314 and 318 (15).…”

“…OV-MZ-32 cells carry a 13-bp deletion from codon 314 to codon 318 of the p53 gene. OvBH-1 is a temperature-sensitive ovarian clear cell carcinoma cell line, characterized in detail elsewhere (16,17). Polish Patent Nr.…”

Mesenchymal Stem Cell Microvesicles from Adipose Tissue: Unraveling Their Impact on Primary Ovarian Cancer Cells and Their Therapeutic Opportunities

“…The OptiMEM GlutaMax medium was supplemented with 3% FBS and a 1% penicillin/streptomycin solution. Tumor cells from the ascitic fluid were isolated according to the protocol described by [57]. Briefly, the ascitic fluid was centrifuged at 1000× g for 10 min.…”