Analytical The Kinetics of Granulopoiesis in Normal Man

I N 1955, when studies were initiated in our laboratory to determine the suitability of radioactive diisopropylfluorophosphate ( DFP32) as a label for granulocytes, information concerning the rate of production and movement of these cells was very meager as compared with knowledge of erythrokinetics. Since that time, with the development and application of the DFP32 cytoplasmic granulocyte labeling technic11’ and with DNA nuclear labeling by means of radiophosphorus1216 and tritiated thymidine,’722 knowledge of the kinetics of granulocytes has grown. The purpose of this report is to summarize our own studies with DFP32, to review the limitations of this method, to compare the results obtained with those obtained by others by other means, and to formulate a concept of granulopoiesis in normal man based on information obtained with the DFP32 label. Alkyl phosphates such as DFP are potent irreversible inhibitors of a number of proteases and esterases such as chymotrypsin,23 trypsin,24 thrombin25 and the various cholinesterases.26’27 The general reaction of DFP with one of these enzymes ( E ) , is illustrated in figure 1. One molecule of DFP binds to the hydroxyl group of a serine residue in the active site of the enzyme and one molecule of HF is liberated. The inactive and stable diisopropylphosphateenzyme ( DIP-E ) derivative remains intact until the protein is degraded. In the case of erythrocytes at least, the DIP-cholinesterase does not turn over since the enzyme remains inactive during the normal life-span of the erythrocyte.28’29 When the DIP-enzyme is degraded enzymatically, DIP-serine is formed ( reaction 2, fig. 2) . The fate of DIP-serine within the body has not been investigated. Presumably, the phosphate-serine bond is cleaved and DIP is released ( reaction 3, fig. 2 ) . However, it is conceivable30 that the alkyl groups are hydrolyzed and that 0-serine phosphate is formed (reaction 4, fig. 2) . If this reaction occurs, serine would be incorporated into phospholipids ( reaction 5, fig. 2) through the phosphatidyl serine and phosphatidyl ethanolamine pathways. 31 Such a possibility was suggested by Mizuno et al.32 Any phosphoserine not utilized directly for phospholipid synthesis would undergo enzymic