Enrichment of cell cycle phases of synchronized estrogen receptor-positive cell models derived from breast adenocarcinomas

Date

2024-8-27

Author

Toker, Pelin

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17β‐estradiol (E2), the main circulating estrogen hormone, is a critical signaling factor for the growth, differentiation, and function of the breast tissue. The effects of E2 on the breast tissue are primarily mediated by the estrogen receptor α (ERα) and deregulation of the E2-ERα signaling contributes to the initiation/progression of breast cancer and resistance to treatments. Breast cancer cell lines as in vitro model systems provide invaluable insight into cellular events, drug discovery, and drug resistance. Among ERα-synthesizing cell lines, MCF7 and T47D cells are widely used to elucidate cell cycle phase-specific molecular events that coordinate cellular proliferation mediated by E2-ERα. Due to variable results in generating phase-enriched populations with various approaches, we wanted to reassess cell cycle synchronization-coupled phase enrichment with charcoal dextran-treated fetal bovine serum, CD-FBS, as an effective hormone withdrawal approach, alone or in combination with excess thymidine, as a DNA replication inhibitor, and/or nocodazole, a microtubule poison, in MCF7 and T47D cells. We find that hormone withdrawal synchronizes both MCF7 and T47D cells at the G0/G1 phase. Supplementation of CD-FBS with E2 enriches the S phase population and together with nocodazole and nocodazole-coupled mitotic shake-off augments the G2/M and M phase populations, respectively, of MCF7 cells. However, the double thymidine block approach with nocodazole or nocodazole-coupled mitotic shake-off is more effective in enriching S, G2/M, or M phase populations of T47D cells.

Subject Keywords

Breast cancer, Cell cycle, Estrogen, Estrogen receptor, Synchronization

URI

https://hdl.handle.net/11511/111027

Collections

Graduate School of Natural and Applied Sciences, Thesis