I EFFECTS OF ENVIRONMENT 55 1 



hypothesis provides an explanation for the radial pattern of outgrowth which is 

 typical in tissue fragments freshly explanted in a plasma clot. It should not be 

 thought, however, that mutual repulsion is a general phenomenon. Holtfreter 

 (1947) showed that cells from amphibian embryos adhere to and grow over each 

 other in preference to growing on a glass surface; and Abercrombie and Heays- 

 man (1954b) themselves have demonstrated that certain tumour cells (Sarcoma 

 37 and Crocker mouse sarcoma), grown together with neonatal mouse cells (skele- 

 tal muscle, bone, heart or liver), do not show "contact-inhibition" with respect 

 to the normal cells, but penetrate among them in a manner which, it is suggested, 

 is analogous to the invasiveness of malignant tumours in vivo. 



Almost from the earliest time that successful tissue culture has been practiced, 

 the idea of growing colonies from a single cell has been an attractive possibility. 

 Attempts to realize this possibility were made as early as 191 6 by Rous and Jones, 

 and repeatedly since that time. Important questions about cellular differentiation 

 and about cellular responses to different stimuli have been awaiting answers 

 which depend upon a knowledge of cell lineage, or answers which become more 

 meaningful when the cell population is more uniform than can be obtained from 

 a mixed population of cells. The long period of failure to derive clones from single 

 cells gave rise to many speculations on its cause. Burrows (1923-24; 1924; 1926) 

 was among the first to discuss the problem. He recognized that growth "may be 

 a property of any fixed tissue cell when placed in the proper environment", and 

 concluded that a certain density of cell population was a necessary part of this 

 environment. The situation in tissue cultures was compared with that in yeast, and 

 in some bacterial cultures, where it had been shown that a few cells in a large 

 volume of fluid would not divide. Burrows suggested that, as in these examples, 

 some material formed by the cells, which he named "archusia", had to accumulate 

 in a certain concentration before growth could occur. He suggested, but apparently 

 did not actually try, restricted volumes of medium. Fischer (1923) and Barnard 

 (1925) had unsuccessfully made attempts to produce cultures from isolated cells, 

 and in 1928 Harrison was still puzzled by the "inexplicable fact that single somatic 

 cells isolated in tissue culture media do not proliferate". 



A number of attempts was made to determine in a broader way the relationships 

 between size of explant, volume of medium and successful cultivation (Fischer, 1 923 ; 

 Earle and Thompson, 1930; Buchsbaum, 1932; Ephrussi and Teissier, 1932-33; 

 Olivo, 1932). Buchsbaum, using fresh explants from 6-day embryonic chick heart 

 in hanging drop cultures, examined the "growth" of cells in these cultures in the 

 light of the hypothesis that, by analogy with the phenomenon of "allelocatalysis" 

 applicable to protozoa, multiplication of one cell may be accelerated, in a restricted 

 volume of fluid, by contiguity with other cells. These experiments were made 

 with explants of 0.1-2.5 iimi^j and with volumes of media of 0.004-0.12 ml. 

 Within the limits of these variables, smaller explants increased in area relatively 

 more than larger ones, in a 48 h. period. Size of explant was of greater influence 

 than volume of medium, though equal explants "grew" (in the sense of area 

 increase) more in larger than in smaller volumes of nutrient. Buchsbaum con- 

 cluded that the hypothesis that cells are mutually beneficial or that cell crowding 

 was favorable to growth was not supported by these results. However, his explants 



Literature p. 581 



