SPERM, OVA, AND PREGNANCY 



In view of the above considerations, a 

 brief review of the topography and histo- 

 logic structure of the placentas of man and 

 rhesus monkey will be given early in this 

 chapter. Later a histologic description of 

 yolk sac placentation, as it occurs in the 

 rat, will be presented. These morphologic 

 descriptions will provide a background 

 against which the various features of the 

 localization of hormones, enzymes, and 

 other compounds in these placentas will be 

 more readily understood. 



11. Histoohemical Methods Utilized 

 in the Study of the Placenta 



The placenta is a most reactive organ 

 histochemically; some portion of it reacts 

 positively in most histochemical tests. This 

 is not surprising in view of its manifold 

 functions which surpass those of any other 

 organ, with the possible exception of the 

 adult liver. The cytologic methods which 

 have been applied to placentas, such as 

 those for the demonstration of mitochon- 

 dria, Golgi apparatus, brush borders, re- 

 ticular fibers, and nuclear morphology, are 

 standard and need not be specifically cited 

 here. On the other hand, histochemical 

 methods have been more recently developed, 

 and are also changing constantly in the light 

 of new advances. To aid the reader in assess- 

 ing the localizations illustrated in this re- 

 view, a brief outline of some of the histo- 

 chemical methods, as they were used in this 

 laboratory, will be presented. The reader is 

 referred to Pearse's Textbook of Histochem- 

 istry (1960) for fuller accounts of methods. 



A. BASOPHILIA AND ACIDOPHILIA 



Nucleoproteins and acid mucopolysaccha- 

 rides are conspicuously basophilic sub- 

 stances. Desoxyribonucleoprotein is respon- 

 sible for most of the basophilia of cell 

 nuclei, whereas much cytoplasmic baso- 

 philia, especially in cells which are rapidly 

 growing or synthesizing large amounts of 

 protein, is attributable to ribonucleoprotein. 

 The latter is also present in the nucleoli. 

 The strong affinity of nucleoproteins and 

 acid mucopolysaccharides for cationic or 

 basic dyes is dependent on the presence in 

 them of phosphate and sulfate groups which 

 bear strong negative charge under many 



routine conditions of staining. Under con- 

 trolled conditions of staining, in which the 

 pH of the staining solution is varied, it is 

 possible to distinguish basophilia caused by 

 nucleic acids and acid mucopolysaccharides 

 from that of proteins. Because the isoelec- 

 tric point of proteins differs, some stain 

 readily with basic dyes whereas others re- 

 act with acid dyes at a given pH. A discus- 

 sion of the factors influencing basophilia 

 and acidophilia of tissues is given in papers 

 by Singer and Morrison (1948) and Singer 

 (1952) . The controlled use of acid and basic 

 dyes on the human and rat placenta is illus- 

 trated in the papers by Singer and Wislocki 

 (1948) and Wislocki, Weiss, Burgos and 

 Ellis (1957 ». 



Many cell types of the growing, differen- 

 tiating ])lacenta are rich in cytoplasmic ri- 

 bonucleoprotein (Dempsey and Wislocki, 

 1946). It can be identified by com})aring 

 histologic sections stained by methylene 

 blue with control sections treated with ribo- 

 nuclease before staining (Brachet, 1953). 

 Deso.xyribonucleoprotein can be distin- 

 guished from ril)onucleoprotein by the Feul- 

 gen method. 



The acid mucopolysaccharides which con- 

 tain sulfuric acid, such as heparin, chon- 

 droitin sulfuric acid esters, and certain mu- 

 coid substances, are strikingly basophilic. 

 This class of mucopolysaccharides can be 

 identified in tissue sections which have been 

 stained with basic dyes in a series of solu- 

 tions of descending pH, inasmuch as the 

 sulfate-containing moiety continues to stain 

 at pH as low as 1 and 2, whereas the weaker 

 acid groupings of polysaccharides, such as 

 hyaluronic acid, cease to stain at pH 4.5 

 (Dempsey, Bunting, Singer and Wislocki, 

 1947). 



In addition to exhibiting basophilia, acid 

 mucopolysaccharides possess the property 

 of shifting the absorption spectrum of cer- 

 tain thiazine dyes, thereby inducing meta- 

 chromatic staining. Thus, toluidin blue and 

 thionin, which color most basophilic sub- 

 stances blue, will stain acid mucopolysac- 

 charides and, under some conditions, nu- 

 cleoproteins red (Wislocki, Bunting and 

 Dempsey, 1947). Michaelis (1947) attril)- 

 uted the metachromasia of dye molecules 

 to their polymerization. An excellent analy- 



