Flow Cytometry Analysis of Cell Cycle and Specific Cell Synchronization with Butyrate.
Synchronized cells have been invaluable in many kinds of cell cycle and cell proliferation studies. Butyrate induces cell cycle arrest and apoptosis in MDBK cells. We explore the possibility of using butyrate-blocked cells to obtain synchronized cells and we characterize the properties of butyrate-induced cell cycle arrest. The site of growth inhibition and cell cycle arrest was analyzed using 5-bromo-2'-deoxyuridine (BrdU) incorporation and flow cytometry analyses. Exposure of MDBK cells to 10 mM butyrate caused growth inhibition and cell cycle arrest in a reversible manner. Butyrate affected the cell cycle at a specific point both immediately after mitosis and at a very early stage of the G1 phase. After release from butyrate arrest, MDBK cells underwent synchronous cycles of DNA synthesis and transited through the S phase. It takes at least 8 h for butyrate-induced G1-synchronized cells to begin the progression into the S phase. One cycle of cell division for MDBK cells is about 20 h. By combining BrdU incorporation and DNA content analysis, not only can the overlapping of different cell populations be eliminated, but the frequency and nature of individual cells that have synthesized DNA can be determined.
Animal Genomics and Improvement Laboratory, Beltsville Agricultural Research Center, ARS, USDA, 10300 Baltimore Avenue, Barc East, Bldg 306, Beltsville, MD, 20705, USA. congjun.li@ars.usda.gov.
Cong-Jun
CJ
Li
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D001973:Bromodeoxyuridine / Q000378:metabolism; D002087:Butyrates / Q000494:pharmacology*; D002417:Cattle; D002453:Cell Cycle / Q000187:drug effects*; D059447:Cell Cycle Checkpoints / Q000187:drug effects / Q000235:genetics; D004261:DNA Replication / Q000187:drug effects / Q000235:genetics; D004847:Epithelial Cells / Q000166:cytology* / Q000187:drug effects*; D005434:Flow Cytometry / Q000379:methods*; D007668:Kidney / Q000166:cytology; D008938:Mitosis / Q000187:drug effects / Q000235:genetics
D002087:Butyrates; D001973:Bromodeoxyuridine
Bovine kidney epithelial cell; BrdU incorporation; Butyrate; Cell cycle synchronization; Flow cytometry