Acrosome Biogenesis and Perinuclear Theca Defects in Globozoospermia

ObjectiveTo clarify the physiopathological mechanisms of the acrosome and perinuclear theca development (PT) in Globozoospermia.DesignDescriptive.Materials and methodsSemen samples (5 Globozoospermia and 6 fertile) and germ cells from testicular biopsies (1 Globozoospermia and 3 with obstructive azoospermia with complete spermatogenesis) were included. Samples were evaluated by transmission electron microscopy (diaminobenzidine pre-embedding and immunogold labelling), immunofluorescence and western blot. For immunodetection, acrosin (C5F10), tubulin αβ and perinuclear theca (PT427) were detected by monoclonal antibodies. Six semen samples (3 fertile and 3 Globo) were used for western blot procedure; equal sperm proteins amounts (from 1.5 sperm million) were loaded onto a 10% SDS-PAGE. Statistical analysis was performed by T-test (P<0.05).ResultsIn normal spermiogenesis, acrosomal development depends on proper proacrosomal vesicle formation and simultaneous spermatid nuclear envelope modifications. TP was consistently immunolocalized in proacrosomal membrane vesicles and below the acrosome during its formation and expansion over the nucleus. In fertile men the TP is composed of six proteins (16-22-29-34-50-68 kDa). In globozoospermia, abnormal proacrosomal vesicles and paranuclear multivesicular and multilamellar structures were observed that resulted in acrosomes insufficiently developed or detached from the nuclear envelope. PT proteins, dissociated from the acrosomes, were ectopically localized in the cytoplasm. Proteomic analysis showed a significant decrease in all six PT proteins.ConclusionThe alterations observed during early acrosome biogenesis in Globozoospermia are due to anomalous development of Golgi-derived proacrosomic vesicles and a significant deficiency of PT proteins. These findings demonstrate a key role of PT for proper acrosome formation, implatation and expansion over the spermatid nucleus. ObjectiveTo clarify the physiopathological mechanisms of the acrosome and perinuclear theca development (PT) in Globozoospermia. To clarify the physiopathological mechanisms of the acrosome and perinuclear theca development (PT) in Globozoospermia. DesignDescriptive. Descriptive. Materials and methodsSemen samples (5 Globozoospermia and 6 fertile) and germ cells from testicular biopsies (1 Globozoospermia and 3 with obstructive azoospermia with complete spermatogenesis) were included. Samples were evaluated by transmission electron microscopy (diaminobenzidine pre-embedding and immunogold labelling), immunofluorescence and western blot. For immunodetection, acrosin (C5F10), tubulin αβ and perinuclear theca (PT427) were detected by monoclonal antibodies. Six semen samples (3 fertile and 3 Globo) were used for western blot procedure; equal sperm proteins amounts (from 1.5 sperm million) were loaded onto a 10% SDS-PAGE. Statistical analysis was performed by T-test (P<0.05). Semen samples (5 Globozoospermia and 6 fertile) and germ cells from testicular biopsies (1 Globozoospermia and 3 with obstructive azoospermia with complete spermatogenesis) were included. Samples were evaluated by transmission electron microscopy (diaminobenzidine pre-embedding and immunogold labelling), immunofluorescence and western blot. For immunodetection, acrosin (C5F10), tubulin αβ and perinuclear theca (PT427) were detected by monoclonal antibodies. Six semen samples (3 fertile and 3 Globo) were used for western blot procedure; equal sperm proteins amounts (from 1.5 sperm million) were loaded onto a 10% SDS-PAGE. Statistical analysis was performed by T-test (P<0.05). ResultsIn normal spermiogenesis, acrosomal development depends on proper proacrosomal vesicle formation and simultaneous spermatid nuclear envelope modifications. TP was consistently immunolocalized in proacrosomal membrane vesicles and below the acrosome during its formation and expansion over the nucleus. In fertile men the TP is composed of six proteins (16-22-29-34-50-68 kDa). In globozoospermia, abnormal proacrosomal vesicles and paranuclear multivesicular and multilamellar structures were observed that resulted in acrosomes insufficiently developed or detached from the nuclear envelope. PT proteins, dissociated from the acrosomes, were ectopically localized in the cytoplasm. Proteomic analysis showed a significant decrease in all six PT proteins. In normal spermiogenesis, acrosomal development depends on proper proacrosomal vesicle formation and simultaneous spermatid nuclear envelope modifications. TP was consistently immunolocalized in proacrosomal membrane vesicles and below the acrosome during its formation and expansion over the nucleus. In fertile men the TP is composed of six proteins (16-22-29-34-50-68 kDa). In globozoospermia, abnormal proacrosomal vesicles and paranuclear multivesicular and multilamellar structures were observed that resulted in acrosomes insufficiently developed or detached from the nuclear envelope. PT proteins, dissociated from the acrosomes, were ectopically localized in the cytoplasm. Proteomic analysis showed a significant decrease in all six PT proteins. ConclusionThe alterations observed during early acrosome biogenesis in Globozoospermia are due to anomalous development of Golgi-derived proacrosomic vesicles and a significant deficiency of PT proteins. These findings demonstrate a key role of PT for proper acrosome formation, implatation and expansion over the spermatid nucleus. The alterations observed during early acrosome biogenesis in Globozoospermia are due to anomalous development of Golgi-derived proacrosomic vesicles and a significant deficiency of PT proteins. These findings demonstrate a key role of PT for proper acrosome formation, implatation and expansion over the spermatid nucleus.