title: Case report of apoptosis in testis of four AZFc-deleted patients: increased DNA fragmentation during meiosis, but decreased apoptotic markers in post-meiotic germ cells.
abstract: AZFc deletions of the Y chromosome are the major known genetic cause of spermatogenetic failure. Meiotic studies have shown a prevalence of synaptonemal complex fragmentation and an excess of early-stage sperm cells, suggesting that the maturation block could involve apoptosis. We present a prospective and observational study of apoptotic markers in the sperm of four AZFc-deleted patients and two non-obstructive azoospermic controls without an AZFc deletion. Polycaspases assays and terminal deoxynucleotidyl transferase dUDP nick-end labelling (TUNEL) assays were combined to evaluate the incidence of apoptosis in pre-meiotic, meiotic and post-meiotic germs cells identified, respectively, using anti-melanoma-associated antigen A4 (MAGE-A4), anti-synaptonemal complex protein 3 (SCP3) and anti-sperm acrosome membrane-associated protein 1 (SPACA1) antibodies. We detected apoptosis at all stages of AZFc-deletion spermatogenesis. Using the caspase assay, the incidence of positive cells was found to be heterogeneous for pre-meiotic (from 4.8 to 84.5%) and meiotic stages (from 7.9 to 57.6%), while for post-meiotic cells, the mean incidence was 6% in AZFc-deleted patients compared with 26.5% in controls (P < 0.05). Using the TUNEL assay, the mean percentage with DNA fragmentation for meiotic cells was 54.0% in AZFc-deleted patients compared with 20.3% in controls (P < 0.05), while the percentage of TUNEL-positive post-meiotic cells ranged from 5.3 to 44.7%. Spermatocyte loss in AZFc-deleted patients occurs via the apoptotic pathway. In post-meiotic cells, the lower incidence of apoptosis in testis from three of the four AZFc-deleted patients, compared with controls, is consistent with AZFc deletions having little negative impact on sperm quality.
authors.affiliation: Inserm UMR 910, Génétique Médicale et Génomique Fonctionnelle, Aix-Marseille Université, Marseille cedex 5 13385, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Streichemberger
authors.forename: Jeanne
authors.initials: J
authors.lastname: Perrin
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Saias-Magnan
authors.forename: Gilles
authors.initials: G
authors.lastname: Karsenty
authors.forename: Perrine
authors.initials: P
authors.lastname: Malzac
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Grillo
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Mitchell
authors.forename: Catherine
authors.initials: C
authors.lastname: Metzler-Guillemain
mesh_terms: D000328:Adult; D017209:Apoptosis; D053713:Azoospermia / Q000235:genetics*; D020169:Caspases / Q000378:metabolism; D041322:Chromosomes, Human, Y / Q000235:genetics*; D053938:DNA Fragmentation; D004789:Enzyme Activation; D017353:Gene Deletion; D005854:Germ Cells / Q000166:cytology*; D006801:Humans; D020287:In Situ Nick-End Labeling; D007621:Karyotyping; D008297:Male; D008540:Meiosis; D008856:Microscopy, Fluorescence / Q000379:methods; D010641:Phenotype; D013090:Spermatocytes / Q000166:cytology; D013737:Testis / Q000473:pathology*
chemical_list: D020169:Caspases