Adult tissue is characterized by a lag period of several days preceding the onset of growth in vitro.
Treatment of fresh adult tissues with trypsin before planting them in culture flasks stimulated the tissues to grow sooner and more rapidly.
Best stimulation was obtained by slow digestion at low temperature. The tissues lost nitrogen during the digestion. Lowering the temperature from 22°C. to 5°C. reduced the digestion of aorta tissue much less than it reduced the digestion of casein.
Washing the tissue after trypsin treatment resulted in better stimulation.
Trypsin solutions of different degrees of purity, when diluted to equal activity toward casein, gave equal stimulation to the tissue growth. These included solutions of Northrop's crystalline trypsin and chymo-trypsin.
Papain also stimulated growth in a similar manner.
The results indicate that this stimulation of tissue growth is due entirely to proteolytic action.
Cultures of adult fibroblasts (and some tumor cultures) having reached a state of retarded growth have been treated with trypsin to digest away most of the plasma clot (used as a medium). Fresh plasma has been added to renew the clot. This treatment has resulted in an immediate renewal of growth.
Reasons are given for supposing that the cells produce an inhibitor in vitro which they deposit in the surrounding clot, and which is removed by the action of trypsin.