Reproductive function of the male obese Zucker rats: alteration in sperm production and sperm DNA damage.

Obesity has been considered a public health issue in many countries and is of increasing concern for authorities over the past 6 years. The Zucker rat is a good experimental model for obesity and diabetes studies due to its metabolic characteristics that are similar to those developed by humans. A total of 12 obese Zucker rats and their lean littermates were killed in pubertal and young adult phases for assessing organ weights (testis and epididymis), testicular histomorphometric and stereological analyses, daily sperm production, and transit time in the epididymis. Sperm integrity was also investigated in the adult animals using the Comet assay. Alterations in organ weights, seminiferous epithelium architecture, sperm production, and transit time were noticed in the pubertal fatty rats. The volume density of the lymphatic space was decreased in both the ages. Adult animals had a significant increase in the extent of damage found in sperm DNA. Our results show for the first time that leptin receptor deficiency compromises sperm production during puberty and that genetic obese Zucker rats have increased sperm DNA fragmentation.

Effects of pentoxifylline treatment before freezing on motility, viability and acrosome status of poor quality human spermatozoa cryopreserved by the liquid nitrogen vapor method.

The objective of the present study was to investigate the effects of the direct addition of pentoxifylline (PF) to the ejaculates of men with poor sperm quality before freezing on post-thaw sperm motility, viability, acrosome integrity, and agonist-induced acrosome reaction. Semen specimens from 16 infertile men with impaired sperm count and motility (oligoasthenozoospermia) were divided into two equal aliquots: one received no treatment (control) while the other was incubated with 5 mM PF (treated). Both aliquots were cryopreserved by the liquid nitrogen vapor method. Motility was assessed according to WHO criteria. Acrosome integrity and spontaneous and calcium ionophore-induced acrosome reactions were assessed with fluorescein isothiocyanate-conjugated peanut agglutinin combined with a supra-vital dye (Hoechst-33258). Cryopreservation impaired sperm motility (percentage reduction: 87.4 (interquartile range, IQ: 70.3-92.9) vs 89.1 (IQ: 72.7-96.0%)), viability (25.9 (IQ: 22.2-29.7) vs 25.6 (IQ: 19.7-40.3%)) and acrosome integrity (18.9 (IQ: 5.4-38.9) vs 26.8 (IQ: 0.0-45.2%)) to the same extent in both treated and control aliquots. However, PF treatment before freezing improved the acrosome reaction to ionophore challenge test scores in cryopreserved spermatozoa (9.7 (IQ: 6.6-19.7) vs 4.8 (IQ: 0.5-6.8%); P = 0.002). These data show that pre-freeze treatment of poor quality human sperm with pentoxifylline did not improve post-thaw motility or viability nor did it prevent acrosomal loss during the freeze-thaw process. However, PF, as used, improved the ability of thawed spermatozoa to undergo the acrosome reaction in response to calcium ionophore. The present data indicate that treatment of poor quality human sperm with PF may enhance post-thaw sperm fertilizing ability.

Effects of pentoxifylline treatment before freezing on motility, viability and acrosome status of poor quality human spermatozoa cryopreserved by the liquid nitrogen vapor method

The objective of the present study was to investigate the effects of the direct addition of pentoxifylline (PF) to the ejaculates of men with poor sperm quality before freezing on post-thaw sperm motility, viability, acrosome integrity, and agonist-induced acrosome reaction. Semen specimens from 16 infertile men with impaired sperm count and motility (oligoasthenozoospermia) were divided into two equal aliquots: one received no treatment (control) while the other was incubated with 5 mM PF (treated). Both aliquots were cryopreserved by the liquid nitrogen vapor method. Motility was assessed according to WHO criteria. Acrosome integrity and spontaneous and calcium ionophore-induced acrosome reactions were assessed with fluorescein isothiocyanate-conjugated peanut agglutinin combined with a supra-vital dye (Hoechst-33258). Cryopreservation impaired sperm motility (percentage reduction: 87.4 (interquartile range, IQ: 70.3-92.9) vs 89.1 (IQ: 72.7-96.0 percent)), viability (25.9 (IQ: 22.2-29.7) vs 25.6 (IQ: 19.7-40.3 percent)) and acrosome integrity (18.9 (IQ: 5.4-38.9) vs 26.8 (IQ: 0.0-45.2 percent)) to the same extent in both treated and control aliquots. However, PF treatment before freezing improved the acrosome reaction to ionophore challenge test scores in cryopreserved spermatozoa (9.7 (IQ: 6.6-19.7) vs 4.8 (IQ: 0.5-6.8 percent); P = 0.002). These data show that pre-freeze treatment of poor quality human sperm with pentoxifylline did not improve post-thaw motility or viability nor did it prevent acrosomal loss during the freeze-thaw process. However, PF, as used, improved the ability of thawed spermatozoa to undergo the acrosome reaction in response to calcium ionophore. The present data indicate that treatment of poor quality human sperm with PF may enhance post-thaw sperm fertilizing ability.

Oligozoospermia and heat-shock protein expression in ejaculated spermatozoa.

BACKGROUND: Heat-shock protein A2 (HspA2) is correlated with sperm maturity, function and fertility, and a dysfunctional expression of such a gene results in abnormal spermatogenesis. The purpose of this study was to compare HspA2 gene expression in spermatozoa from oligozoospermic men and normozoospermic controls. METHODS: Semen was obtained and analysed according to World Health Organization (World Health Organization, 1999) guidelines, morphology by Kruger's strict criteria. Seventeen patients with oligozoospermia and 21 fertile controls were studied. Total RNA was extracted from ejaculated and Percoll density-gradient-separated spermatozoa followed by semiquantitative RT-PCR analysis. The relative expression level of HspA2 was analysed according to the expression level of the housekeeping beta-actin gene. Serum hormonal profiles (FSH, LH and testosterone) and a peripheral karyotype were also performed. RESULTS: All patients possessed normal karyotype, and no significant hormonal differences were found between the two groups. The study group had significantly lower sperm concentration and normal morphology than the controls. Semiquantitative RT-PCR analysis of HspA2 showed significantly lower expression levels in the oligoteratozoospermic men when compared to controls (P=0.0021). CONCLUSIONS: The HspA2 gene was down-regulated in sperm from infertile men with idiopathic oligoteratozoospermia, suggesting that such anomalies of gene expression might be associated with pathogenesis in some subtypes of male infertility.

Prevalence of Y chromosome deletions in a Brazilian population of nonobstructive azoospermic and severely oligozoospermic men.

We determined the prevalence of Y chromosome deletions in a population of 60 Brazilian nonobstructive azoospermic and severely oligozoospermic men. PCR-based screening of microdeletions was performed on lymphocyte DNA for the presence of 14 sequence-tagged sites (STS) located in the azoospermic factor (AZF) on the Yq chromosome. All STS were amplified efficiently in samples from 12 fertile men tested, but failed to be amplified in samples from fertile women, indicating the specificity of PCR conditions for Yq screening. Overall, 4 of the 60 infertile patients tested (6.7%) exhibited deletion of the Y chromosome, 2 of them being severely oligozoospermic patients (P10 and P32) and 2 azoospermic men (patients P47 and P57). Patients P47 and P57 presented larger deletions in the AZFa, AZFb and AZFc subregions, with apparent loss of Yq material evidenced by karyotype analysis. Patients P10 and P32 presented deletions confined to the AZFc region, involving the DAZ locus. Male relatives of patients P10 and P32 had no Y chromosome deletions and presented a normal karyotype, suggesting a de novo status of the deletions found. Our data add to the growing literature showing that microdeletions of the Y chromosome can be the cause of male idiopathic infertility.

Prevalence of Y chromosome deletions in a Brazilian population of nonobstructive azoospermic and severely oligozoospermic men

We determined the prevalence of Y chromosome deletions in a population of 60 Brazilian nonobstructive azoospermic and severely oligozoospermic men. PCR-based screening of microdeletions was performed on lymphocyte DNA for the presence of 14 sequence-tagged sites (STS) located in the azoospermic factor (AZF) on the Yq chromosome. All STS were amplified efficiently in samples from 12 fertile men tested, but failed to be amplified in samples from fertile women, indicating the specificity of PCR conditions for Yq screening. Overall, 4 of the 60 infertile patients tested (6.7 percent) exhibited deletion of the Y chromosome, 2 of them being severely oligozoospermic patients (P10 and P32) and 2 azoospermic men (patients P47 and P57). Patients P47 and P57 presented larger deletions in the AZFa, AZFb and AZFc subregions, with apparent loss of Yq material evidenced by karyotype analysis. Patients P10 and P32 presented deletions confined to the AZFc region, involving the DAZ locus. Male relatives of patients P10 and P32 had no Y chromosome deletions and presented a normal karyotype, suggesting a de novo status of the deletions found. Our data add to the growing literature showing that microdeletions of the Y chromosome can be the cause of male idiopathic infertility

Microbiologic aerobic studies on normal male urethra.

OBJECTIVES: To carefully collect samples from the external urethral orifice, navicular fossa, and penile urethra and perform a semiquantitative evaluation and identification of gram-positive and gram-negative bacteria present in the normal male urethra. METHODS: Thirty uncircumcised male patients 18 to 40 years old without any inflammatory and/or infectious urethral processes were enrolled in this study. Samples were collected from the external urethral orifice, navicular fossa, and penile urethra with sterile alginate swabs that were immediately transferred to tubes containing buffered phosphate solution. Inoculation was done by spreading 0.01 mL of the buffered solutions on sheep blood agar plates and MacConkey agar plates; the plates were then incubated at 36.5 degrees C for 24 hours. After this period, the quantification and identification of each type of colony was performed. RESULTS: Among the 30 patients studied, 12 (40%) had bacteria isolated from the three segments, 10 (33.3%) had bacterial colonization in two segments, and 8 (26.7%) had colonization in only one segment (external urethral orifice). Staphylococcus coagulase-negative species, group viridans alpha-hemolytic streptococci, Corynebacterium species, and Enterococcus species were the bacteria more frequently isolated from these three segments. CONCLUSIONS: From the findings in this study, it was clear that the bacterial urethral flora was abundant, not evenly distributed, concentrated in the external urethral orifice and navicular fossa, and basically consisted of gram-positive aerobic bacteria.