The experiments to be described were designed to approach the following type of question: Will the differentiated state of a tissue cell survive multiple divisions in vitro? Do the cultured progeny of differentiated cells inherit the cellular mechanisms determining the unique somatic traits of their parental cells in a manner analogous to the way in which (1) Paramecia transmit Kappa particles to their daughter cells (Sonneborn,33 34 Beale,2 Preer3), (2) bacteria inherit genes for constitutive or inducible enzymes in the absence of substrate (Novick and Weiner,28 Spiegel-man35) or (3) temperate phage particles give rise to lysogenic progeny (Jacob and Wollman20)? Information of this kind is essential if the roles of the nucleus (i.e., genes) and the cytoplasm in cell differentiation are to be defined. An unambiguous approach to these problems requires that the following five conditions be fulfilled: (1) The initial population of differentiated cells must be homogeneous. (2) The cells must divide many times during the experiment. (3) The cells must not back-differentiate to the parental type immediately after each division. (4) The initial population must consist of differentiated cells and not dividing precursor cells whose progeny differentiate and thereafter cease dividing (e.g., presumptive muscle cells, presumptive blood cells, presumptive pigment cells, etc.). (5) The somatic traits used as markers must be specific to the parent cell and readily characterized. Given this list of conditions, it is clear that many kinds of somatic cells are excluded as test material. For example, both nerve cells and keratinizing skin cells do not divide. Liver and kidney cells offer difficulties of another kind; since it is difficult to secure a homogeneous population of either type of epithelial cell, it is possible that in long-term cultures the epithelial cells are selected against and the surviving population is derived from one of the "contaminating" cell types. Cartilage cells from vertebrae of 10-day chick embryos, on the other hand, satisfy the five conditions outlined. Pieces of cartilage may be obtained which consist * This investigation was supported, in part, by Research Grants B-493 and B-1629 from the

No abstract

No abstract

Observations were made on the behavior of chondrocytes grown under various conditions in vitro. The chondrocytes in 10-day embryonic chick vertebrae were grown as cultures of intact vertebrae, as pellets of chondrocytes liberated from their matrix, and as monodispersed cells plated out on plasma clots. Cartilage matrix was stained metachromatically with toluidine blue. Radioautographs were made of incorporated H3-thymidine, H3-proline, and S3~-sulfate to determine the extent of DNA synthesis, collagen synthesis, and chondroitin sulfate synthesis, respectively. Chondrocytes in intact vertebrae or in pellets are rounded and actively synthesizing chondroitin sulfate and collagen. There is little DNA synthesis by cells in either vertebrae or pellets. Chondrocytes grown as monodisperse cells rapidly cease synthesizing cytologically detectable chondroitin sulfate and are induced to synthesize DNA and divide. There is a change in the shape of these cbondrocytes from a rounded to a more stellate condition which accompanies the shift in metabolic activity: Conversely, when the cells attain a certain cell density, they reacquire a rounded shape, cease dividing, and again synthesize chondroitin sulfate. Clusters of chondrocytes synthesize more chondroitin sulfate than isolated chondrocytes. It is concluded that most chondrocytes synthesizing chondroitin ~ulfate do not concurrently synthesize DNA. Interaction between associated chondrocytes is important in inducing and maintaining chondroitin sulfate synthesis in genetically determined chondrocytes. Failure of interaction between chondrocytes leads to DNA synthesis and cell multiplication.

No abstract