208 CELLS, TISSUES, AND ORGANISMS 



of micromolecules, addition of horse serum at a concentration of 5 to 

 10 per cent was required. The aggregation-supporting activity of whole 

 serum could not be replaced by serum dialysates or by various non- 

 proteinaceous macromolecular additives. Therefore a preliminary survey 

 was made to determine the ability of various serum fractions to support 

 histogenetic cohesion of dissociated cells in synthetic culture media in 

 rotating cultures. It was found that in the presence of 0.5 mg/cc of 

 fraction IV-1 of adult bovine serum, containing alpha-1-globulin and 

 lipid protein ("Alpha Lipoprotein" of Nutritional Biochemicals Corp., 

 Cleveland) as the only. macromolecular additive to the medium, there 

 was rapid cohesion of cells and typical histogenesis of the ensuing ag- 

 gregates. It cannot yet be stated conclusively whether this is the only 

 serum fraction capable of eflPectively supporting histogenetic bonding 

 of cells; nor is it clear whether only one or both of the major constitu- 

 ents are involved, or whether the eflFect might or might not be due to 

 an "impurity." It is of interest that "fetuin," the alpha-1-globulin 

 fraction of fetal calf serum which promotes adhesion of adult cells to 

 glass ( Fisher, et al., 1958 ) , has been found by us to enable mutual co- 

 hesion of embryonic cells in rotating cultures, albeit at concentrations 

 higher than the alpha-lipoprotein preparation. Both of these serum 

 derivatives are known to be physically heterogeneous (Lieberman et 

 al., 1959; Fisher et al., 1959), and therefore a comparison of their spe- 

 cific functions on cells must await further analysis. Similarly, the nature 

 of their aggregation-promoting function remains to be determined: 

 whether they act as cell-surface stabilizers, or assist in transport of 

 molecules, or supply essential metabolites. It is, however, of some in- 

 terest that lipid and globular proteins, which have long been thought 

 to contribute to the constitution and activities of cell surfaces ( Danielli, 

 1958; Willmer, 1961), may also be involved in supporting the establish- 

 ment of functional contacts between cells. 



Further light on this area is being shed by collateral information. 

 Thus it has been found that in cultures maintained at 15° C. in optimal 

 concentrations of these proteins the cells do not cohere, though brought 

 together by rotation. It is also known that in the absence of calcium, at 

 otherwise optimal conditions of temperature and protein concentration, 

 dissociated cells do not cohere. Glucose is another essential requirement 

 for morphogenetically effective aggregation of embryonic cells— pre- 

 sumably as a source of energy for cell activities bearing on this process. 

 All these indications point to a missing link in this series of require- 

 ments and unavoidably draw us to the already postulated notion that 

 this pivotal prerequisite for cell cohesion and aggregation may be a 

 cellular product. As a matter of fact, an extracellular material ( ECM ) 

 has been found to accumulate between cells in early phases of aggrega- 

 tion and is chiefly responsible for holding together the newly bunched 



