Selenium is an essential chemopreventive antioxidant element to oxidative stress, although high concentrations of selenium induce toxic and oxidative effects on the human body. However, the mechanisms behind these effects remain elusive. We investigated toxic effects of different selenium concentrations in human promyelocytic leukemia HL-60 cells by evaluating Ca(2+) mobilization, cell viability and caspase-3 and -9 activities at different sample times. We found the toxic concentration and toxic time of H(2)O(2) as 100 microM: and 10 h on cell viability in the cells using four different concentrations of H(2)O(2) (1 microM: -1 mM: ) and six different incubation times (30 min, 1, 2, 5, 10, 24 h). Then, we found the therapeutic concentration of selenium to be 200 nM: by cells incubated in eight different concentrations of selenium (10 nM: -1 mM: ) for 1 h. We measured Ca(2+) release, cell viability and caspase-3 and -9 activities in cells incubated with high and low selenium concentrations at 30 min and 1, 2, 5, 10 and 24 h. Selenium (200 nM: ) elicited mild endoplasmic reticulum stress and mediated cell survival by modulating Ca(2+) release, the caspases and cell apoptosis, whereas selenium concentrations as high as 1 mM: induced severe endoplasmic reticulum stress and caused cell death by activating modulating Ca(2+) release, the caspases and cell apoptosis. In conclusion, these results explained the molecular mechanisms of the chemoprotective effect of different concentrations of selenium on oxidative stress-induced apoptosis.

The aim of this study was to determine the effect of aerobic exercise on uric acid (UA), total antioxidant activity (TAA), lipid hydroperoxides, and nitric oxide (NO) metabolites in human saliva. Twenty-four healthy male and female subjects were studied during a 10,000-m race. Saliva samples were collected 1 h before and immediately after exercise. The NO concentration was determined by the Griess reaction, UA by enzymatic method, TAA by the ABTS method, and lipid hydroperoxide by the ferrous iron/xylenol orange (FOX) method. A repeated measures ANOVA was used to examine the effect of aerobic exercise on salivary UA, TAA, lipid hydroperoxides, and NO metabolites. Aerobic exercise caused an increase in both salivary UA and TAA, and a decrease in salivary lipid hydroperoxide. There was no, however, change in nitrite concentration. These results suggested that aerobic exercise-induced increment in both UA and TAA seems to inhibit lipid hydroperoxide generation, a marker of oxidative stress in human saliva.

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Abstract. Apoptosis plays an essential role in normal spermatogenesis, but deregulations of this biological process, which is closely associated with male infertility, have been found. Whereas calcium homeostasis is a key regulator of cell survival, sustained elevation of intracellular calcium plays a role in apoptosis. The aim of this research was to determine the role of two different calcium mobilizing agents, hydrogen peroxide (H2O2) and the physiological agonist progesterone, on the apoptosis process of human ejaculated spermatozoa. Translocation of membrane phosphatidylserine was examined with an annexin V binding assay, DNA damage was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL assay) and caspase-3 activity was assessed using a fluorometric assay. After incubation of spermatozoa for 1 h with either 10 μM H2O2 or 20 μM of progesterone, there was a significant increase in both caspase-3 activity and the percentage of annexin V-positive cells. Similarly, the TUNEL results were significantly higher 1 h after incubation with either 10 μM H2O2 or 20 μM of progesterone. In fact, progesterone-treated cells showed a three-fold increase (from 17.6 to 52.9%) of TUNEL-positive cells compared to untreated cells, while H2O2-treated cells exhibited a two-fold increase (from 17.6 to 37.9%). In sum, our results suggest that spermatozoa treated with calcium mobilizing agents, such as H2O2 and progesterone, seem to undergo an apoptosis process that is dependent on caspase-3 activation. Key words: Apoptosis, Caspases, Phosphatidylserine exposure, Spermatozoa, TUNEL (J. Reprod. Dev. 55: [615][616][617][618][619][620][621] 2009) poptosis is a distinctive form of cell death characterized by a series of morphological and biochemical changes that plays a critical role in many physiological processes such as development, tissue homeostasis, inflammatory responses and disease [1]. It also plays an essential role in gamete maturation and embryogenesis, contributing to appropriate formation of various organs and structures. In fact, many studies in animal models have demonstrated that apoptosis is the underlying mechanism of germ cell death during normal spermatogenesis [2], which has also been reported in humans [3].On the other hand, deregulations of this biological process, which is closely associated with male infertility, have been found, and both men with azoospermia and men with severe oligozoospermia have an increased frequency of apoptotic germ cells in their testicular tissues compared with patients having normal spermatogenesis [2,4,5]. In addition, a certain degree of spontaneous apoptosis and relatively high rates of apoptosis have been detected using an in situ end-labelling (TUNEL) assay in testicular biopsies of infertile patients with testicular insufficiency [6,7]. Similarly, disturbance of cell membrane symmetry with translocation of phospatidylserine residues to the outer layer of the plasma membrane has been observed in germ cells from patients with complete spermi...

Phenylketonuria (PKU), the most common inborn error of amino acid metabolism, is caused by mutations in the phenylalanine-4-hydroxylase (PAH) gene. This study aimed to assess the genotype-phenotype correlation in the PKU Spanish population and the usefulness in establishing genotype-based predictions of BH4 responsiveness in our population. It involved the molecular characterization of 411 Spanish PKU patients: mild hyperphenylalaninemia non-treated (mild HPA-NT) (34%), mild HPA (8.8%), mild-moderate (20.7%) and classic (36.5%) PKU. BH4 responsiveness was evaluated using a 6R-BH4 loading test. We assessed genotype-phenotype associations and genotype-BH4 responsiveness in our population according to literature and classification of the mutations. The mutational spectrum analysis showed 116 distinct mutations, most missense (70.7%) and located in the catalytic domain (62.9%). The most prevalent mutations were c.1066-11G>A (9.7%), p.Val388Met (6.6%) and p.Arg261Gln (6.3%). Three novel mutations (c.61-13del9, p.Ile283Val and p.Gly148Val) were reported. Although good genotype-phenotype correlation was observed, there was no exact correlation for some genotypes. Among the patients monitored for the 6R-BH4 loading test: 102 were responders (87, carried either one or two BH4-responsive alleles) and 194 non-responders (50, had two non-responsive mutations). More discrepancies were observed in non-responders. Our data reveal a great genetic heterogeneity in our population. Genotype is quite a good predictor of phenotype and BH4 responsiveness, which is relevant for patient management, treatment and follow-up.

Assisted sperm morphometry analysis (ASMA) was used in this study to determine the effects of cryopreservation on bull spermatozoa distribution in morphometrically distinct subpopulations. Ejaculates were collected from five bulls and were divided. One portion was diluted at 30 degrees C in a skim milk-egg yolk medium, containing glycerol. A microscope slide was prepared from single extended sperm samples prior to freezing. The remainder of each sample was frozen in nitrogen vapours. After thawing, sperm smears were prepared as described above. All slides were air dried and stained with Hemacolor. The sperm-head dimensions for a minimum of 200 sperm heads were analysed from each sample by means of the Sperm-Class Analyser (SCA), and the mean measurements recorded. Our results showed that applying the ASMA technology and multivariate cluster analyses, it was possible to determine that three separate subpopulations of spermatozoa with different morphometric characteristics coexist in bull ejaculates (large, average and small spermatozoa). The mean values of each sperm head dimension among the three subpopulations of spermatozoa were significantly different (p < 0.001). Besides, there were significant (p < 0.001) differences in the distribution of these three sperm subpopulations between fresh and thawed samples. Thus, the percentage of representation of the subpopulation that includes those spermatozoa whose dimensions are the biggest, decreased from 52.06% in extended fresh samples to 15.51% in the thawed ones. Contrarily, the percent of representation of the subpopulation containing the smallest spermatozoa, increased from 8.70% in extended fresh samples to 34.04% in the thawed ones. In conclusion, the present study confirms the heterogeneity of sperm head dimensions in bull semen, heterogeneity that vary through the cryopreservation procedure.

Rapid weight gain in childhood is associated with increased risk of chronic diseases in adults. C-reactive protein (CRP) is a mediator of atherosclerosis and chronically elevated levels predict cardiovascular outcomes. The effects of life course weight gain on CRP levels are not clear. The 1982 Pelotas (Brazil) birth cohort study (n = 5,914) has prospectively collected weight and health data at several follow-ups since birth. The most recent was in 2004-05, when 77.4% of the cohort was traced and CRP levels were measured in 89% of those interviewed (n = 3827). Geometric mean (SE) C-reactive protein levels were 0.89 mg/l (0.03) and 1.66 mg/l (0.04) in men and women, respectively. In analyses adjusted for confounding variables, weight gain in infancy showed a weak negative association among males, but from the second year onwards, weight gain was positively associated with CRP levels. In females, weight gain was associated with higher CRP at every period tested. The strongest associations were observed in the most recent (18-23 years) period; CRP ratios (95% CI) per z score increase in weight gain were 1.78 (1.57-2.00) and 1.52 (1.30-1.78) for men and women, respectively. Males who were stunted at 2 years and centrally obese at 23 years had the highest CRP levels (P = 0.002 for interaction). In summary, rapid weight gain throughout life predicted higher CRP levels. Public health efforts need to tackle chronic under-nutrition in infancy, together with rapid weight gain in later childhood and adolescence, especially in countries undergoing the nutritional transition.

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