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United States Department of Agriculture

Agricultural Research Service

Title: Specific Cell Cycle Synchronization with Butyrate and Cell Cycle Analysis by Flow Cytometry for Madin Darby Bovine Kidney (Mdbk) Cell Line

Authors
item Li, Congjun
item Elsasser, Theodore

Submitted to: Journal of Animal and Veterinary Advances
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 25, 2006
Publication Date: November 15, 2006
Citation: Li, C., Elsasser, T.H. 2006. Specific cell cycle synchronization with butyrate and cell cycle analysis by flow cytometry for Madin Darby Bovine Kidney (MDBK) cell line. Journal of Animal and Veterinary Advances. 5(11):916-923.

Interpretive Summary: Synchronized cells have been invaluable for many kinds of cell cycle and cell proliferation studies. To explore the possibility of using butyrate-blocked cells to obtain synchronized cells, we investigated the property of the cell cycle arrest induced by butyrate and compared cell synchronization induced by butyrate, serum deprivation, and the combination of serum deprivation and aphidicolin. We also utilized the specific flow cytometric analysis to determine the cell cycle progression of the cells treated with butyrate. By combining BrdU incorporation and DNA content analysis, not only can the overlapping between different cell populations be eliminated, but also the frequency and nature of individual cells that have synthesized DNA can be determined. Exposure of MDBK cells to 10 mM butyrate caused inhibition of growth and cell cycle arrest in a reversible manner. Our data proved that butyrate affected the cell cycle at a specific point 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. The results showed that most serum deprivation treated cells were arrested at the G1 phase and aphidicolin treated cells were arrested at the early S phase. Using butyrate-synchronized cells, we determined that it takes about 8 h for G1 synchronized cells, induced by butyrate, to progress into the S phase and 8 h for the completion of the S phase. One cycle of cell division for MDBK cells is about 20 h. The results of this study are very important because MDBK cells and synchronized MDBK cells will provide us an important model for our study on animal growth regulation.

Technical Abstract: Synchronized cells have been invaluable for many kinds of cell cycle and cell proliferation studies. Butyrate induces cell cycle arrest and apoptosis in MDBK cells. To explore the possibility of using butyrate-blocked cells to obtain synchronized cells, we investigated the property of the cell cycle arrest induced by butyrate and compared cell synchronization induced by butyrate, serum deprivation, and the combination of serum deprivation and aphidicolin. The site of growth inhibition and cell cycle arrest was studied by BrdU incorporation and flow cytometry analyses. By combining BrdU incorporation and DNA content analysis, not only can the overlapping between different cell populations be eliminated, but also the frequency and nature of individual cells that have synthesized DNA can be determined. Exposure of MDBK cells to 10 mM butyrate caused inhibition of growth and cell cycle arrest in a reversible manner. Evidence is presented that butyrate affected the cell cycle at a specific point 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. The results showed that most serum deprivation treated cells were arrested at the G1 phase and aphidicolin treated cells were arrested at the early S phase. The effects of the three treatments were reversible. Using butyrate-synchronized cells, we determined that it takes about 8 h for G1 synchronized cells, induced by butyrate, to progress into the S phase and 8 h for the completion of the S phase. One cycle of cell division for MDBK cells is about 20 h.

Last Modified: 9/2/2014
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