7^ 

 BINUCLEATE CELLS IX TISSUE CULTURES. 81 



Considering next (c), we find that this also is improbable. It is easy to observe 

 tiie process of mitosis in vitro, and to follow the various changes. IMany such cases 

 have been observed, and in none has there been seen a failure of the cytoplasm to 

 divide following separation of the chromosomes. This process of cytoplasmic con- 

 striction is well shown in figures 68, 69, and 70, and in the living culture it is very 

 evident and easy to watch. In no case has it been observed, in following these cells 

 dividing by karyokinesis, that a binucleate cell was formed; always the end result 

 was two distinct daughter cells, often widely separated, connected by a thin strand 

 of protoplasm (fig. 1, /). If crowding of the cells occurs, separation of the daughter 

 cells may be interfered with to some extent, but it is doubtful if this interference ever 

 is so serious as to prevent cytoplasmic fission altogether and thus result in the forma- 

 tion of a single cell containing two nuclei. At least no evidence has been found 

 from observation of the cells of tissue cultures that this is ever the case. 



Upon this point my observations agree with those of Child (1911, p. 283). He 

 says: "In Moniezia nuclei which arise by mitosis are separated by an appreciable dis- 

 tance when they form." Again (p. 292), in describing a "double" nucleus, repre- 

 sented in his figure 1 1 , he says : "The two parts of the nucleus . . . are in immediate 

 contact and flattened against each other. It is difficult to understand how they could 

 attain such a position as the result of mitotic cleavage, like that of the earlier stages." 



It must be said that my observations upon living cells have principally been 

 made with cells of the connective-tissue type. In the case of membranes, however, 

 there is always a well-marked dividing line between the cells, which is made evident 

 by staining with iron hematoxylin or the use of silver; also this potential isolation 

 of the cells is made apparent by the fact, when cells do separate, that the cleavage 

 is along this line of partition, as is shown from the study of fixed preparations 

 (Lewis and Lewis, 1912c, figs. 14, 13, and 12). No such partition is ever found 

 between the nuclear parts of binucleate cells. 



In fixed preparations of connective-tissue cells there is no indication of an\' 

 failure of the cytoplasm to divide in the later stages of mitosis; that is to saj'-, we 

 find no telophases where separation of the cytoplasm is not evident (fig. 17). 



Again, these double nuclei almost always have only a single centrosj^here (fig. 7), 

 whereas nuclei arising by karyokinesis have each a centrosi^here. This finding as 

 to the centrosphere agrees entirely with that of Deineka (1912) for the Netzapparat 

 in the dividing epithelial cells of Descemet's membrane and connective-tissue cells 

 of the cornea. This author is of the opinion that the Netzapparat surrounds the 

 centrosome, and its changes ai)i)ear to follow the variations of the latter body. In 

 binucleate cells of these tissues, in which the nucleus divides by amitosis, the Netz- 

 apparat remains single, whereas if the nuclear division takes place by mitosis each 

 of the daughter nuclei obtains a separate Netzapparat. By reference to this dis- 

 position of the Netzapparat, Deineka is even able to tell the mann(>r of origin of such 

 double nucleus, whether by amitosis or mitosis in which cleavage of the cytoplasm 

 has been delayed. I liave never observed this cell organ in living tissue-cultures. 



The fact that the centrosphere in the binucleate cell is single seems to indicate 

 that the twin nucleus is single so far as its reproductive capacity is concerned. This 

 infcronre is borno out bv olisorvations. later to be referred to. 



