D-Aspartic acid stimulates steroidogenesis through the delay of LH receptor internalization in a mammalian Leydig cell line.

PURPOSE: Recent experimental evidence on non-mammalian animal models showed that D-Aspartic acid (d-Asp) administration increases testosterone levels through upregulation of StAR in Leydig cells. In this study, we aimed to investigate in vitro the signaling pathway associated with d-Asp stimulation in MA-10 murine Leydig cells. METHODS: MA-10 cells were stimulated with different concentrations of d-Asp, in presence or absence of hCG. Then total testosterone (T) levels in the culture medium were evaluated by electrochemiluminescence immunoassay, and StAR and LHR protein expressions were quantified by the means of Western blotting. LHR cellular localization after hormonal stimulation was assessed by immunofluorescence. RESULTS: Stimulation with the sole d-Asp did not induce any relevant increase of T release from cultured cells. On the other hand, stimulation with hCG induced significant increase of T (P = 0.045). Concomitant stimulation with hCG and d-Asp, at the concentration of 0.1 and 1 nM, induced additional and significant increase of released T (P = 0.03 and P = 0.04, respectively). StAR protein levels increased after concomitant stimulation with hCG and d-Asp 0.1 nM, compared with stimulation with the sole hCG (P = 0.02), whereas no variation in LHR protein expression was observed. Finally, d-Asp attenuated displacement of LHR staining, from cell membrane to cytoplasm, subsequent to hCG stimulation. CONCLUSIONS: In this study, we confirmed a steroidogenic role for d-Asp, in concert with hCG, on murine Leydig cells, which is mediated by an increase in StAR protein levels. In addition, we showed that the possible mechanism subtending the effect of d-Asp could rely on the modulation of LHR exposure on the cell membrane.

Testicular fine needle aspiration as a diagnostic tool in non-obstructive azoospermia.

AIM: To report the fine needle aspiration cytology (FNAC) of the testes used as a diagnostic tool in non-obstructive azoospermic patients. METHODS: One hundred and twenty-five non-obstructive azoospermic male candidates to intracytoplasmic sperm injetion (ICSI) were analysed for follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone and inhibin B plasma levels. They were classified into three groups on the basis of FNAC: 1) Sertoli cell-only syndrome (SCOS) (70); 2) severe hypospermatogenesis (42); and 3) maturation arrest (13). Then, all men underwent testicular sperm extraction (TESE) for sperm recovery for ICSI. RESULTS: Mature spermatozoa were detected by FNAC in 24 of 42 men with severe hypospermatogenesis and nine of 13 men with maturation arrest; while they were retrieved by TESE in 29 of 70 men with SCOS, 35 of 42 men with severe hypospermatogenesis (including the 24 by FNAC) and 10 of 13 men with maturation arrest (including the nine by FNAC). The sensitivity and specificity of FNAC were 44.6 % and 100 %, respectively. There was no difference on testicular volume and hormonal parameters in men with and without sperm retrieved. CONCLUSION: These findings suggest that FNAC may be a simple and valid diagnostic parameter in non-obstructive azoospermic men and it may represent a valid positive prognostic parameter for sperm recovery at TESE.

PDE-5 inhibitor, Vardenafil, increases circulating progenitor cells in humans.

Bone marrow-derived endothelial progenitor cells (EPCs) originate from haematopoietic stem cells in bone marrow and migrate into the peripheral circulation to promote endothelial repair and neovascularization. The number of circulating progenitor cells is reduced in patients with cardiovascular risk factor. The aim of our study was to determine the number of these cells in healthy patients and to evaluate the effect of Vardenfil, a phosphodiesterases-5 (PDE-5) inhibitor, in the number of circulating EPCs. In our study, we found a significant increase in the number of these cells after the drug administration.

Human sperm express cannabinoid receptor Cb1, the activation of which inhibits motility, acrosome reaction, and mitochondrial function.

Cannabinoids and endocannabinoids negatively influence sperm functions. These substances have been demonstrated in many mammalian tissues, including male and female reproductive tracts, and previous studies have shown the presence of functional receptors for cannabinoids in human sperm. The present study, by means of RT-PCR and Western blot techniques, demonstrates that human sperm express the CB(1), but not CB(2), cannabinoid receptor (CB-R) subtype located in the head and middle piece of the sperm. The activation of this receptor by anandamide reduces sperm motility and inhibits capacitation-induced acrosome reaction. Activation of the CB(1)-R did not induce any variation in sperm intracellular calcium concentrations, but produced a rapid plasma membrane hyperpolarization that was reduced by the K(+) channel blocker tetraethylammonium. The effects of anandamide on human sperm motility were dependent on the reduction of sperm mitochondrial activity as determined by rhodamine 123 fluorescence. The specificity of anandamide effects in human sperm were confirmed by the effects of the CB(1)-R antagonist SR141716. These findings provide additional evidence that human sperm express functional CB(1)-R, the activation of which negatively influences important sperm functions, and suggest a possible role for the cannabinoid system in the pathogenesis of some forms of male infertility.

Antisperm antibodies modify plasma membrane functional integrity and inhibit osmosensitive calcium influx in human sperm.

BACKGROUND: The hypo-osmotic swelling (HOS) test evaluates the ability of the functional sperm plasma membrane to stretch following cell swelling when exposed to hypo-osmotic solutions. Sperm samples with low HOS scores show low fertilization and pregnancy rates during assisted reproductive techniques, though data are controversial. The aim of this study was to compare the results of the HOS test in a group of normozoospermic men with those in a group of subjects affected by autoimmune infertility due to the presence of antisperm antibodies (ASA) bound to the sperm surface. METHODS: Sperm from normozoospermic and from infertile subjects affected by autoimmune infertility were exposed to hypo-osmolar conditions to verify the effects on intracellular calcium concentrations and acrosome reaction. RESULTS: Sperm samples from infertile men with ASA showed HOS test scores that were significantly lower than those of normozoospermic subjects despite similar sperm viability percentages. Sperm with ASA bound to their plasma membrane showed a reduced rise in intracellular calcium concentrations and acrosome reaction after hypo-osmotic challenge with respect to sperm from normozoospermic subjects without ASA. CONCLUSIONS: Infertile subjects with ASA have a reduced sperm plasma membrane functional integrity that could explain, at least in part, the low fertilization and pregnancy rates observed in these subjects during assisted reproductive procedures. Evaluation for the presence of ASA in all sperm samples showing low HOS test scores in the presence of normal sperm viability percentages is suggested.