Hypercholesterolemia Impaired Sperm Functionality in Rabbits

Hypercholesterolemia represents a high risk factor for frequent diseases and it has also been associated with poor semen quality that may lead to male infertility. The aim of this study was to analyze semen and sperm function in diet-induced hypercholesterolemic rabbits. Twelve adult White New Zealand male rabbits were fed ad libitum a control diet or a diet supplemented with 0.05% cholesterol. Rabbits under cholesterol-enriched diet significantly increased total cholesterol level in the serum. Semen examination revealed a significant reduction in semen volume and sperm motility in hypercholesterolemic rabbits (HCR). Sperm cell morphology was seriously affected, displaying primarily a “folded head”-head fold along the major axe-, and the presence of cytoplasmic droplet on sperm flagellum. Cholesterol was particularly increased in acrosomal region when detected by filipin probe. The rise in cholesterol concentration in sperm cells was determined quantitatively by Gas chromatographic-mass spectrometric analyses. We also found a reduction of protein tyrosine phosphorylation in sperm incubated under capacitating conditions from HCR. Interestingly, the addition of Protein Kinase A pathway activators -dibutyryl-cyclic AMP and iso-butylmethylxanthine- to the medium restored sperm capacitation. Finally, it was also reported a significant decrease in the percentage of reacted sperm in the presence of progesterone. In conclusion, our data showed that diet-induced hypercholesterolemia adversely affects semen quality and sperm motility, capacitation and acrosomal reaction in rabbits; probably due to an increase in cellular cholesterol content that alters membrane related events

Nitrosative stress in human spermatozoa causes cell death characterized by induction of mitochondrial permeability transition-driven necrosis

Peroxynitrite is a highly reactive nitrogen species and a potent inducer of apoptosis and necrosis in somatic cells. Peroxynitrite-induced nitrosative stress has emerged as a major cause of impaired sperm function; however, its ability to trigger cell death has not been described in human spermatozoa. The objective here was to characterize biochemical and morphological features of cell death induced by peroxynitrite-mediated nitrosative stress in human spermatozoa. For this, spermatozoa were incubated with and without (untreated control) 3-morpholinosydnonimine (SIN-1), in order to generate peroxynitrite. Sperm viability, mitochondrial permeability transition (MPT), externalization of phosphatidylserine, DNA oxidation and fragmentation, caspase activation, tyrosine nitration, and sperm ultrastructure were analyzed. The results showed that at 24 h of incubation with SIN-1, the sperm viability was significantly reduced compared to untreated control (P<0.001). Furthermore, the MPT was induced (P<0.01) and increment in DNA oxidation (P<0.01), DNA fragmentation (P<0.01), tyrosine nitration (P<0.0001) and ultrastructural damage were observed when compared to untreated control. Caspase activation was not evidenced, and although phosphatidylserine externalization increased compared to untreated control (P<0.001), this process was observed in <10% of the cells and the gradual loss of viability was not characterized by an important increase in this parameter. In conclusion, peroxynitrite-mediated nitrosative stress induces the regulated variant of cell death known as MPT-driven necrosis in human spermatozoa. This study provides a new insight into the pathophysiology of nitrosative stress in human spermatozoa and opens up a new focus for developing specific therapeutic strategies to better preserve sperm viability or to avoid cell death

Ultrastructural changes.

<p>Transmission<b>-</b>electron micrographs of rabbit sperm heads from NCR (A) and HCR (B to D). Notice the small vesicles in the acrosome region in B and the long side fold of sperm head in D. Some sperm cells show the remaining residual body (white empty arrow, C). A and C, X 12,000; B and D, X 20,000.</p

Rabbits fed with fat-rich diet increased plasma cholesterol.

<p>Plasma cholesterol concentration from NCR (•) and HCR (<b>□</b>) during the 11 experimental months. Values are expressed as mean ± SEM. Arrowhead indicates fat intake start in HCR, and arrow indicates the moment from which HCR weight resulted significantly different from NCR, (p<0.001).</p

General characteristics of fresh rabbit semen samples (mean ± SEM).

<p>*p<0.001, <i>n</i> = 25.</p

Saturated-fat consumption damaged sperm plasma membrane in rabbits.

<p>Photographs (X 250) represent sperm cell morphology after hypo-osmotic stress. A (NCR): normal coiled tails and B (HCR): straight tails. C: Bars represent the percentage (means ± SEM) of spermatozoa swollen from NCR (black bar) and HCR (white bar) rabbits. **  =  significantly different from control (p<0.01).</p