Higher clusterin immunolabeling and sperm DNA damage levels in hypertensive men compared with controls.

BACKGROUND: Clusterin, a heterodimeric glycoprotein found at several sites in the human male reproductive tract, could be a marker of morphologically abnormal spermatozoa, while TUNEL positivity indicates DNA fragmentation. Metabolic disorders such as diabetes mellitus and obesity may compromise sperm quality and fertility of men; however, little evidence specifically links hypertension with the impairment of male reproductive function. METHODS: By flow cytometric, immunofluorescence (TUNEL assay and clusterin immunolabeling) and immunohistochemical (peroxidase-streptavidin method) analyses, we have compared both clusterin- and TUNEL labeling in ejaculated spermatozoa from healthy normotensive donors and hypertensive subjects with the purpose to reveal possible differences between the two conditions. RESULTS: Data analysis from the normotensive (n=25) and hypertensive subjects (n=25) demonstrate a significant correlation between high levels of clusterin immunolabeling and the presence of sperm DNA damage, which is often associated with abnormal morphology. In the normotensive subjects, a low percentage (15.3±4.5) of spermatozoa positive for high levels of clusterin was detected; however, this percentage significantly increased (30.9±13.0) (P<0.01) in hypertensive subjects. Standard semen evaluations does not reveal any significant differences between the two groups of subjects, except for a reduced forward motility and lower sperm vitality in the hypertensive subjects. CONCLUSIONS: This pilot study strongly suggests a relationship between hypertension and markers indicative of poor sperm quality. In hypertensive subjects, high levels of clusterin immunolabeling identified a consistent fraction of ejaculated spermatozoa carrying both DNA fragmentation and strong morphological alterations, which was not correlated with age or with sperm cell mortality. The alternative possibility that sperm damage observed is due to adverse effects of anti-hypertensive drugs does not find support in the literature nor in the drug data sheets. The relationship observed between hypertension and human semen represents a novel and possibly relevant information to be considered in the study of male fertility.

Expression of a truncated form of KIT tyrosine kinase in human spermatozoa correlates with sperm DNA integrity.

BACKGROUND: TR-KIT, a truncated form of KIT (the KITL receptor), corresponding to the c-terminal half of the intracellular split tyrosine kinase domain, is expressed during the haploid stages of mouse spermatogenesis, and is one of the candidate sperm factors possibly involved in egg activation at fertilization. METHODS: Immunocytochemistry of adult human testis, and studies of human semen samples from volunteer donors through immunofluorescence, confocal microscopy, flow cytometry, western blot and RT-PCR analyses were performed. RESULTS: We show that the TR-KIT is expressed during spermiogenesis in the human testis, and that it is maintained in human ejaculated spermatozoa. TR-KIT is localized both in the equatorial segment and in the sub-acrosomal region of the human sperm head. The equatorial localization of the TR-KIT persists after the spontaneous acrosome reaction. Cytometric analysis of several sperm samples from volunteer donors, showed variable degrees of the TR-KIT-specific immunolabeling, and a significant inverse correlation (Pearson's coefficient, r = -0.76, P < 0.0001, n = 23) of the TR-KIT positivity with markers of sperm damage, i.e. DNA fragmentation, as revealed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labeling (TUNEL) analysis and the intense clusterin positivity. We also found less significant inverse correlation with altered head morphology (r = -0.47, P < 0.05, n = 23) and direct correlation with sperm forward motility parameters (r = 0.59, P < 0.01, n = 23). CONCLUSIONS: The TR-KIT is present in the equatorial region of human spermatozoa, which is the first sperm component entering into the oocyte cytoplasm after fusion with the egg. This localization is consistent with the function previously proposed for this protein in mice. In addition, the TR-KIT represents a potential predictive parameter of human sperm quality.

Peculiar subcellular localization of Fas antigen in human and mouse spermatozoa.

The highly polarized structure and function of mammalian spermatozoa dictate that these cells compartmentalize specific metabolic and signaling pathways to regions where they are needed. Fas was initially identified as membrane receptor for pro-apoptotic signals, has been recently recognized as a molecule with pleiotropic functions. In this article, we provide evidence of a peculiar Fas localization: it is closely associated to the perinucleus, mainly at the level of the inner acrosomal membrane, as well as in the inner compartment of mitochondria. Immunoelectron microscopy and Western blot analysis indicated that intracellular Fas was associated with mitochondria in mouse epididymal spermatozoa. Accordingly, also in human ejaculated sperm, immunofluorescence analysis showed Fas localized in the middle piece of sperm flagellum where mitochondria are grouped. The potential functional implications of these findings are discussed.