title: Serum starvation and thymidine double blocking achieved efficient cell cycle synchronization and altered the expression of p27, p53, bcl-2 in canine breast cancer cells.
abstract: Cell synchronization is an approach to obtain cell populations of the same stage, which is a prerequisite to studying the regulation of cell cycle progression in vivo. Serum starvation and thymidine double blocking (TdR) are two important practices in studying cell cycle synchronization. However, their effects on canine cancer cells as well as the regulatory mechanisms by these two methods are poorly understood. In this study, we determined the optimum conditions of serum starvation and TdR and their effects on cell cycle synchronization. We further explored the involvement of PI3K/Akt signaling pathway in the cell cycle synchronization by investigating the expression of three key genes (p27, p53 and bcl-2). Serum starvation resulted in a reversible cell cycle arrest and synchronously progress through G0/G1. The highest percentage of CHMm cells (87.47%) in G0/G1 stage was obtained after 42 h incubation with 0.5% fetal bovine serum (FBS). TdR double blocking could arrest 98.9% of CHMm cells in G1/S phase (0 h of release), and could arrest 93.74% of CHMm cells in S phase after 4h of release. We also found that the p27, p53, bcl-2 genes were most highly expressed in G0/G1 phase. Our current work revealed that serum starvation and TdR methods could achieve sufficient synchronization of CHMm cells. Moreover, the expression of p27, p53 and bcl-2 genes was related to cyclical movements and apoptosis. Our results will provide a new insight into cell cycle regulation and reprogramming of canine cancer cells induced by serum starvation and TdR blocking.
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Jinjin
authors.initials: J
authors.lastname: Tong
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Dongdong
authors.initials: D
authors.lastname: Sun
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Yang
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Yingxue
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Sichao
authors.initials: S
authors.lastname: Sun
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Li
authors.affiliation: College of Animal Science and Technology, Northeast Agricultural University, Harbin, China; Synergetic Innovation Center of Food Safety and Nutrition, Northeast Agricultural University, Harbin, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Bao
authors.affiliation: Department of Veterinary Surgery, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China. Electronic address: abliuyun@yeah.net.
authors.forename: Yun
authors.initials: Y
authors.lastname: Liu
mesh_terms: D000818:Animals; D002453:Cell Cycle; D002455:Cell Division; D045744:Cell Line, Tumor; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics* / Q000378:metabolism; D004285:Dogs; D005260:Female; D015674:Mammary Neoplasms, Animal / Q000235:genetics* / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics* / Q000378:metabolism; D044967:Serum / Q000502:physiology*; D015398:Signal Transduction; D013936:Thymidine / Q000031:analogs & derivatives*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism
chemical_list: D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D050760:Cyclin-Dependent Kinase Inhibitor p27; D013936:Thymidine
keywords: Bcl-2; Canine breast cancer cell; Cell cycle synchronization; P27; P53