INVESTIGATIONS ON HISTOCHEMICAL LOCALIZATION OF LEUCINE- AND CYSTINEAMINOPEPTIDASE (OXYTOCINASE) IN PLACENTA

The histochemical demonstration of oxytocinase in human placenta described by Semm and Waidl (1962) is based on the use of L-cystine-di-β-naphthylamide (CBNA) as a substrate for enzymatic cleavage, on diazotising the enzymically hydrolysed β-naphthalamine, and of the development of a blue color by coupling the diazocompound with N-1 (naphthyl)-ethylenediamine. Yet the method was not realisable chemically, because of the high pH during diazotising β-naphthylamine. After lowering the pH under 2,0 we succeeded in obtaining the results described by Semm and Waidl. They have to be regarded however as an artefact for tissue staining was of secondary nature. Diffusion of oxytocinase during incubation period lead to an enzymatic reaction within the solvent and not within the tissue as should be supposed. Following the method of Nachlas, Crawford and Seligman (1957) with L-leucine-β-naphthylamide (LBNA) as substrate we succeded in the real histochemical demonstration of oxytocinase. The cleavage of LBNA by oxytocinase proceeds twenty times faster than the cleavage of the more specific CBNA. Inhibition of Oxytocinase with EDTA was followed by a nearly complete loss of enzyme specific staining, inhibition of leucineaminopeptidase with D,L-methionine displayed no significant loss in enzyme activities. We can conclude therefore that these aminopeptidase activities being located in the trophoblast and in the X-cells of the human placenta are depending chiefly on oxytocinase. © 1969 Springer-Verlag.