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United States Department of Agriculture

Agricultural Research Service

Research Project: EFFICIENCY OF NUTRIENT USE IN CATTLE:IDENTIFICATION OF CRITICAL PHYSIOLOGIC AND GENOMIC REGULATORY PATHWAYS Title: Specific cell cycle synchronization with butyrate and cell cycle analysis

Author
item Li, Congjun

Submitted to: Methods in Molecular Biology
Publication Type: Book / Chapter
Publication Acceptance Date: June 10, 2010
Publication Date: July 31, 2011
Citation: Li, C. 2011. Specific cell cycle synchronization with butyrate and cell cycle analysis. In: Banfalvi, G., editor. Methods in Molecular Biology, Cell Cycle Synchronization: Methods and Protocols. New York NY: Humana Press. p. 125-136.

Interpretive Summary: Cultured cells usually do not grow at the same pace. However, study of cell growth regulation requires cultured cells proliferating at the same pace (i.e.,synchronization). Butyrate can block cells cycle, therefore can stop cell at specific stage of cell cycle. In this study, we investigated the property of the cell cycle arrest induced by butyrate and compared the differences of cell synchronization induced by butyrate, serum deprivation, and 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 the overlapping between different cells populations can be eliminates, but also the frequency and nature of individual cell that have synthesized DNA can be determined. We concluded that butyrate can be used as a novel method to synchronize the cell cycle.

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 the differences of cell synchronization induced by butyrate, serum deprivation, and 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 of 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 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 G1-synchronized cells induced by butyrate to start progression into S phase. One cycle of cell division for MDBK cells is about 20 h.

Last Modified: 10/24/2014
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