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Sparse and dense cultures of chick embryo cells were affected differently by pH. The rates of cell multiplication and of thymidine-(3)H incorporation into DNA of dense cultures were increased as the pH was increased from 6.6 to 7.6. At pH higher than 7.6 the rate of multiplication decreased slightly in the dense cultures, but the rate of thymidine-(3)H incorporation continued to increase. The discrepancy was due in part to cell death and detachment at very high pH, and in part to a more rapid uptake of thymidine-(3)H at very high pH. Sparse cultures were much less sensitive to pH reduction and, when a suitably conditioned medium was used to minimize cell damage, very sparse cultures grew almost as well at pH 6.7 as at higher pH. The rates of cell multiplication and thymidine-(3)H incorporation at low pH decreased in the initially sparse cultures before they reached confluent cell densities. There was no microscope evidence of direct contact between plasma membranes of cells at these densities although the parallel orientation indicated that the cells were influencing locally each other's behavior. Even at much higher cell densities, electron microscopy revealed large intercellular gaps partly filled with a fragmentary electron-opaque material suspected to be glycoprotein. Wounding experiments showed that pH affected cell migration in a manner similar to its effects on cell multiplication. Low pH inhibited cell migration, but those cells which migrated into the denuded region multiplied as rapidly at low pH as at high pH. The effects of pH on growth were correlated with effects on the uptake of 2-deoxyglucose-(3)H. Dense populations of cells inhibited by low pH were stimulated to incorporate thymidine-(3)H by the addition of small amounts of diethylaminoethyl-dextran. Rous sarcoma cells at high cell density were less sensitive to pH than were normal cells at the same density, but were more sensitive than sparse normal cultures. The results suggest that cell growth is inhibited through the combined effects of both lowered pH and high cell density on cell surface permeability.

pH AND POPULATION DENSITY IN THE REGULATION OF ANIMAL CELL MULTIPLICATION