The changing cell number and composition of chick heart explants growing in vitro

Abstract 1. 1. Cell number and changing cell composition have been determined in chick heart expiants from 12 day embryos during growth in vitro . The determination of cell number has been based on the deoxyribonucleic acid (DNA) content of the cultures, since it has been established that there is a constant amount of DNA phosphorus (DNAP) per nucleus of about 2.35 × 10 −7 μg in all the somatic cells of the fowl. By reference to the DNAP of the cultures it has also been possible to express as amounts per cell the protein nitrogen (PN), ribonucleic acid phosphorus (RNAP), lipid phosphorus (LP), and acid-soluble phosphorus (ASP) of the cultures during the various phases of growth. 2. 2. There was an initial fall of two-thirds in the number of cells per 48 expiants when these were in contact with Tyrode-serum mixture during the 24 hours following planting. This initial loss in cell number could be reduced but not prevented by keeping the cells in growth-promoting medium from the time of planting. In both media there were similar alterations in cell composition, namely large increases in the amounts of PN and LP per cell. 3. 3. After this initial stage in Tyrode-serum the cultures received fresh amounts of growth-promoting medium every 24 hours. Cell number increased most rapidly between 48 and 144 hours when the numbers per 48 expiants doubled in each 48 hour interval. After 144 hours the increase was slower and linear until 240 hours. 4. 4. Within certain limits the number of cells in the initial explants determined the final number in the fully grown cultures. The largest increases in cell number recorded in these experiments were between six and seven-fold. 5. 5. Growth of the cultures was accompanied by a consistent pattern of changes in the chemical composition of the cells. The intensive cell division in the cultures was characterized by large increases in the PN and LP per cell in the first stage of culture growth, and by later increases in the RNAP and ASP per cell to maximal amounts during the period of rapid cell multiplication. The amounts of all four cell constituents (PN, RNAP, LP, and ASP) decreased to approximately their original amounts per cell in the chick heart cells when the culture growth slowed down after 144 hours. 6. 6. These results suggest that the synthesis of protein and phospholipid precede the synthesis of DNA in the chemical processes leading to cell division, and that RNA are in some way related to DNA synthesis. 7. 7. It was found that keeping 48-hour growing cultures in Tyrode-serum for a further 24 hours resulted in a small increase in cell number, but produced little or no change in cell composition; it did not impair their ability to grow normally when returned to growth-promoting medium.