5-Bromodeoxyuridine inhibition of differentiation. Kinetics of inhibition and reversal in myoblasts

This paper describes experiments on the kinetics of inhibition of muscle differentiation in vitro in the presence of 5-bromodeoxyuridine (BrdUrd) and the recovery phenomena that occur when such inhibited cells are permitted growth in normal medium. The studies consist of a quantitation of cell fusion in the presence of the analog and during recovery in its absence coupled with simultaneous studies on changes in buoyant density of cellular DNA. We find that if myoblasts are exposed to BrdUrd during the last doubling before cell fusion would normally occur, most cells do not differentiate, but as many as 18% of the cells can fuse in spite of the incorporation of BrdUrd into their nuclei. These nuclei contain approximately the amount of BrdUrd expected for a full round of DNA synthesis. Studies on the rate of recovery of inhibition of cell fusion following one generation in BrdUrd reveal that after one doubling of inhibited cells in the presence of normal medium. fusion reaches about 50% of the control value; after two doublings it reaches 75% of control value; and after 2.5 doublings of reversal, recovery is essentially complete. We find that both the degree of inhibition after approximately one round of BrdUrd incorporation and the rate of cell differentiation after two generations of reversal are consistent with a model which assumes that BrdUrd “sensitivity” resides on single pair of chromosomes and that inhibition occurs in a dominant fashion if approximately 30% or more of the thymidine is replaced by BrdUrd in the readout strand of either chromosome.