Diamine oxidase activity and biochemical markers in human seminal plasma.

Diamine oxidase (DAO), an enzyme which degrades polyamines, is present at a very high level in human seminal plasma and is assumed to come mainly from the prostate. The possible relationships between DAO activity and biochemical markers of accessory sex glands were evaluated in 139 men in barren marriages. Four groups were formed: normozoospermic (n = 41), asthenozoospermic (n = 29), oligoasthenozoospermic (n = 35) and azoospermic (n = 34). DAO activity was the highest in the asthenozoospermic group and was significantly different from that in the azoospermic one. For all specimens, a positive correlation was demonstrated between DAO activity and the prostatic markers citric acid and acid phosphatase. However, DAO activity was correlated with citric acid only in the oligoasthenozoospermic and the azoospermic groups. Acid phosphatase and citric acid were linked in all groups. These results implicate the DAO enzyme in changes in sperm metabolism leading to a loss of motility and suggest that DAO comes partly from the upper part of the genital tract (testis and/or epididymis), in addition to the prostatic gland secretion, accounting for the absence of correlation with prostatic markers in normozoospermic and asthenozoospermic groups.

Ultrastructural and immunocytochemical study of P1 protamine localization in human testis.

Two monoclonal antibodies (MAbs) directed against human protamine P1 were realized. Anti-P1 specificity was assessed by western-blot and confirmed by ELISA. Monoclonal antibody 97-3 was selected. Protamine P1 was specifically demonstrated in human testis by immunoelectron microscopy, using 97-3 MAb and an indirect post-embedding immunogold technique. Our results clearly demonstrated the precise time of appearance of P1 protamine in the nuclei of human spermatids. P1 first appeared in the nucleus of step 5 spermatids and its concentration was increased in steps 6-8 spermatids, cytoplasm was not labelled.

Preparation of spermatogenic cell populations at specific stages of differentiation in the human.

Studying biochemical events in human spermatogenesis requires separated populations of spermatogenic cells. Dissociation of these cells was performed by a Trypsin-DNAse method adapted from the technique used for rodents. Cell separation was performed by centrifugal elutriation. Seven populations were collected, one further purified by Percoll gradient centrifugation, giving nine different cell populations. The efficiency of the cell separation was evaluated by phase contrast microscopy, flow cytometric DNA analysis, and electron microscopy. Five populations were enriched in spermatids: two in round spermatids (87% and 73%), another in round (52%) and elongating (44%) spermatids, another constituted by 80% elongating spermatids, and the last by 90% elongated spermatids. Two of the four remaining populations were enriched in primary spermatocytes (74% and 54%); another population was the upper part of the Percoll gradient and constituted cytoplasmic lobes and residual bodies (89%); the last population was made up of various cells, with no specific enrichment. Electron microscopic observations revealed good preservation of the separated cells; only the flagella from elongated spermatids were lost. Furthermore, an unusual pattern of nucleoplasm distribution during stages 2-4 of spermatid differentiation was observed and its signification is discussed with regard to the shape of the human spermatozoon.