The growth energy of a colony at a given instant is a function of several independent variables : its growth energy at the pre ceding instant, and the concentration in the pericellular fluid of the substances that increase or decrease cell activity. It could be ascertained by the activity displayed during an instant by the cells in a medium containing neither activating nor retarding sub stances, if such a measurement were possible. But the residual growth energy, which was found to vary at the same time as the inherent energy, may be used as a measurement of the latter. The residual energy is expressed by the duration of life and the pro liferative activity of the cells in Tyrode solution.
The morphology of the tissue cells growing in vitro is studied while they are living. Fragments of the coagulum containing cells are taken from the flasks and put on thin glass slides. They may also be readily cultivated in glass chambers especially con structed for cytological studies. The living unstained cells are photographed or cinematographed. When stained with neutral red, trypan blue, and Janus green, the cytoplasmic structures may be observed while in full activity. Cinematographic records can be taken at such a magnification that the details of the mitochondria, segregation apparatus, and surface membrane of the cells are clearly shown.
Mode and Rate of Growth of Tissues.—This method has already supplied some important information regarding the physi ological properties of tissue cells, the role of blood serum toward the tissues, and the nature of the chemical substances that deter mine the unlimited multiplication of fixed cells. The rate of growth of connective tissue and of epithelial cells in a nutrient medium of unvarying composition remains uniform indefinitely. Such a law appears to be the expression of one of the more funda mental properties of tissue cells. Since their rate of growth re mains unchanged as long as the medium does not vary, the cell colonies may be considered to be immortal. A strain is able to transform an unlimited amount of chemical energy. The energy spent by the cells must arise entirely from the substances con tained in the culture medium, and will never be exhausted as long as the same medium is supplied to them. It appears that the potentialities of tissues are much greater than those they display within the normal organism, and that the manifestation of their activity is determined by the physicochemical conditions of the pericellular fluid. The dependence of cells on the composition of the medium was further emphasized by a study of the rate of growth of colonies of fibroblasts or epithelial cells in media of varying composition. It was found that the activity of the col
onies is a function of the concentration of nutrient substances within the medium, when waste products and other inhibiting sub stances are constantly removed. If cell colonies in different condi tions of activity are placed in similar media, their rates of multi plication soon become identical. Fragments of connective tissue of chick embryos and adult chickens cultivated in a nutrient medium grew at first at varying rates, but after a few weeks no difference was observed in the activity of both colonies.
The residual energy of embryo tissues, which is always very large, varies in inverse ratio with the age of the animal from which the tissues have been extirpated. The duration of their sur vival in vitro may extend over a period of three months. On the other hand, the residual energy of the tissues of an adult mal is very small. The connective tissue cells of an old are in a dormant condition. If cultivated in a nutrient medium, a long time elapses before they begin to proliferate. Should their resting state be attributed to a factor preventing them from spend ing their energy, or to a complete lack of energy? An investigation of the behaviour of these tissues in a medium completely lacking in nutrient substances showed that they possess very little residual energy. This means that they have no reserves in store. How ever, they have not lost the power of accumulating the substances which supply them with energy. When they are placed in a nutrient medium, they begin to multiply and to store up reserves. Adult fibroblasts may be compared to a motor which possesses no energy of its own, but runs as soon as it is supplied with fuel.
The persistence of the essential characteristics of cells culti vated in vitro has opened a new field to histology. When the rate of growth and the constitution of the medium of a pure strain have been ascertained, the relations existing between the structure of the cells and their physiological state become apparent imme diately. Blood monocytes, tissue macrophages and fibroblasts have been studied according to these principles. The appearance of the cells varies greatly with their nutritional condition. The anatomy of a cell type can be modified at will by simple changes in the nature and the concentration of the substances contained in its medium.