Catalogue Search | MBRL
Are you sure you want to remove the book from the shelf?
{{itemTitle}}
62
result(s) for
"Barnabé H"
Sort by:
Sperm cryodamage occurs after rapid freezing phase： flow cytometry approach and antioxidant enzymes activity at different stages of cryopreservation
Background： In order to improve the efficiency of bovine sperm cryopreservation process, it is important to understand how spermatozoa respond to differences in temperature as well as the ability to recover its own metabolism. The combination between flow cytometry approach and antioxidant enzymes activity allows a more sensible evaluation of sperm cell during cryopreservation. The aim of this study was to evaluate sperm attributes and antioxidant enzymes activity during different stages of cryopreservation process. Semen samples from Holstein bulls （n = 4） were separated in 3 treatments： fresh （37 ℃）; cooled （5 ℃）; and thawed. Evaluation occurred at 0 h and 2 h after incubation. Membrane integrity, mitochondrial membrane potential （MMP） and DNA damages were evaluated by flow cytometry; activities of antioxidant enzymes such as catalase, superoxide dismutase and gluthatione peroxidase were measured by spectrofotometry. Results： There was an increase in the percentage of sperm with DNA damage in the thawed group, compared to fresh and cooled, and for 2 hs of incubation when compared to 0 h. Considering MMP, there was an increase in the percentage of cells with medium potential in thawed group when compared to fresh and cooled groups. Opposingly, a decrease was observed in the thawed group considering high mitochondrial potential. Also in the thawed group, there was an increase on cells with damaged acrosome and membrane when compared to fresh and cooled groups. Significant correlations were found between antioxidant enzymes activity and membrane or mitochondrial parameters. Conclusion： Based on our results, we conclude that cryopreservation affects cellular and DNA integrity and that the critical moment is when sperm cells are exposed to freezing temperature. Also, our study indicates that intracellular antioxidant machinery （SOD and GPX enzymes） is not enough to control cryodamage.
Journal Article
The Stimulated Glycolytic Pathway Is Able to Maintain ATP Levels and Kinetic Patterns of Bovine Epididymal Sperm Subjected to Mitochondrial Uncoupling
Studies have reported the importance of mitochondria in sperm functionality. However, for some species, the glycolytic pathway appears to be as important as oxidative phosphorylation in ATP synthesis and sperm kinetics. These mechanisms have not been fully elucidated for bovine spermatozoa. Therefore, the aim of this study was to evaluate the role of mitochondria and the glycolytic pathway in ATP synthesis, sperm movement patterns, and oxidative homeostasis of epididymal spermatozoa in bovine specimens. We observed that mitochondrial uncoupling with protonophores significantly reduced ATP levels. However, these levels were reestablished after stimulation of the glycolytic pathway. We verified the same pattern of results for sperm kinetic variables and the production of reactive oxygen species (ROS). Thus, we suggest that, after its appropriate stimulation, the glycolytic pathway is capable of maintaining ATP levels, sperm kinetic patterns, and oxidative balance of bovine epididymal spermatozoa submitted to mitochondrial uncoupling.
Journal Article
Insights into soy lecithin and egg yolk-based extenders for chilling canine spermatozoa
The aim of this study was to compare different concentrations of soy lecithin (LEC0.01%, LEC0.05% and LEC0.1%) with egg yolk (Control) in cooling extenders during the storage of semen at 5ºC for 5 days. Twelve dogs (n = 12) were selected, and semen was cooled and assessed after 2, 24, 48, 72, 96 or 120 h. At each time point, sperm were analyzed for kinetic patterns (using computer-assisted sperm analysis), mitochondrial activity (3′3- diaminobenzidine assay), lipid peroxidation (TBARS assay), DNA fragmentation (SCSA®) and plasma and acrosome membrane integrity (eosin/nigrosin and fast green/rose Bengal stains, respectively). The Control group (1814.4 ± 197.2) presented the highest rates of lipid peroxidation at 120 h. Conversely, progressive motility (42.8 ± 4%), linearity (45.4 ± 1%), and VAP (88 ± 3%) were higher in the Control group. In addition, there was lower mitochondrial activity in the Control group at 72 h. Therefore, our data show that lecithin used at these concentrations was not able to maintain sperm viability at as high qualities as would egg yolk. Moreover, the decrease in high mitochondrial activity and the persistence of sperm motility may indicate a compensatory mechanism in canine spermatozoa (i.e., glycolytic pathway). Furthermore, these higher lipid peroxidation indexes could indicate the necessity for future therapy using extenders and antioxidants over a long cooling time for dog sperm.
Journal Article
The addition of docosahexaenoic acid (DHA) and antioxidants (glutathione peroxidase and superoxide dismutase) in extenders to epididymal sperm cryopreservation in bulls
The cryopreservation of epididymal sperm is an important technique that allows genetic material to be preserved, even post mortem. However, cryopreservation leads to increased oxidative stress and impaired sperm viability. Polyunsaturated fatty acid (PUFA) supplementation may improve certain sperm characteristics, but it also makes sperm more susceptible to oxidative stress, therefore adding antioxidants that counteract oxidative stress has become an option. In this context, this study aimed to evaluate the effect of the interaction between docosahexaenoic acid (DHA) and antioxidants on the quality after the cryopreservation of epididymal bull sperm. Twenty epididymides were collected after slaughter, and epididymal sperm was cryopreserved with bovine extender supplemented with docosahexaenoic acid (DHA), glutathione peroxidase (GPx) and superoxide dismutase (SOD). We verified an improvement in motility in the group that was treated only with DHA 5 µM and a concentration-dependent effect on susceptibility to lipid peroxidation that was associated with DHA concentration (1 µM, 5 µM or 10 µM). Moreover, treatment with DHA (5 µM) and SOD (20 IU/ml) resulted in higher sperm motility. Thus, the association between DHA (5 µM) and SOD (20 IU/ml) appears to be an option for increased epididymal sperm features in bulls.
Journal Article
The background in the0ν β β experiment Gerda
The GERmanium Detector Array (Gerda) experiment at the Gran Sasso underground laboratory (LNGS) of INFN is searching for neutrinoless double beta (0νββ) decay of 76 Ge. The signature of the signal is a monoenergetic peak at 2039 keV, the Q ββ value of the decay. To avoid bias in the signal search, the present analysis does not consider all those events, that fall in a 40 keV wide region centered around Q ββ. The main parameters needed for the 0νββ analysis are described. A background model was developed to describe the observed energy spectrum. The model contains several contributions, that are expected on the basis of material screening or that are established by the observation of characteristic structures in the energy spectrum. The model predicts a flat energy spectrum for the blinding window around Qββ with a background index ranging from 17.6 to 23.8 × 10 -3 cts/(keV kg yr). A part of the data not considered before has been used to test if the predictions of the background model are consistent. The observed number of events in this energy region is consistent with the background model. The background at Q ββ is dominated by close sources, mainly due to 42 K, 214 Bi, 228
Journal Article
Utilisation of sperm-binding assay combined with computer-assisted sperm analysis to evaluate frozen-thawed bull semen
Summary Due to homologies between the chicken egg perivitelline membrane with mammalian zona pellucida proteins, spermatozoa of several species are able to bind to this membrane. However, adequate standardisation is required to attest possible applications of this technique for semen evaluation of a given species. Therefore, we thawed and divided cryopreserved semen samples into two aliquotes, one kept in water bath at 37 °C (thawed) and the other submitted to snap‐freezing to damage sperm cells (dead spermatozoa). Aliquotes were mixed into different ratios of thawed:dead cells and analysed for motility, membrane and acrosomal integrity, and mitochondrial activity. In parallel, chicken egg perivitelline membranes were inseminated with these ratios, and the number of spermatozoa bound per mm2 of membrane was assessed by conventional microscopy (CM) and computer‐assisted sperm analysis (CASA). Linear regression showed high correlation between thawed:dead sperm ratio and number of spermatozoa bound to the membrane (CM: r2 = 0.91 and CASA: r2 = 0.92 respectively). Additionally, positive correlations were found between the number of spermatozoa bound to the membrane and acrosomal integrity, membrane integrity, mitochondrial activity and motility. These findings indicate that sperm‐egg‐binding assay associated with CASA is a reliable, practical and inexpensive method for examining the fertilising capacity of cryopreserved bull semen.
Journal Article
Identification of α6, β1, and β3 on Bovine Sperm
Sperm-egg interaction is a complex molecular process leading to gamete fusion mediated by a series of molecular interactions. Some integrin subunits, which are adhesion molecules, are expressed on human and mouse sperm, but major questions about integrins' roles in sperm-oocyte fusion remain unsolved. To investigate the potential role of integrins in fertilization, we determined the presence of integrin subunits on catlle (Bos indicus and Bos taurus) sperm by confocal microscopy. Frozen thawed sperm, donated by ABS Pecplan, were centrifuged at 700 x g for 10 min and washed once in warm PBS. Sperm were resuspended in PBS and treated with an equal volume of cold 4% paraformaldehyde (prepared fresh from formaldehyde) for 15 min on ice. All subsequent steps were conducted at RT. Fixed sperm were washed twice with PBS (10 000 rpm, 5 min) in an IEC Micromax (Needham, MA), and the pellet was resuspended in PBS containing 5% BSA. Samples of 1 ml, each containing 5 x 106 spermatozoa, were incubated overnight at 4°C with a monoclonal anti-integrin α6 IgG, monoclonal anti-integrin β1 IgG, and monoclonal anti-integrin β3 IgG (Chemicon;1:500 ). The next day, after being washed 3 times in PBS-BSA, sperm were resuspended in Alexa Fluor 488 goat anti-mouse IgG (Invitrogen). After being washed 3 times in PBS-BSA as before, a drop of sperm suspension was smeared on slide, air-dried, mounted with antifade reagent with DAPI (Invitrogen) and covered with a coverslip for analysis. Sperm were viewed by a confocal microscope. Immunoreactive α6, β1, and β3 were concentrated at the apical segment of the acrosome in all bulls tested. There was no detectable fluorescence in any of the controls, and not all the sperm in an ejaculate were immunoreactive. The presence of α6, β1, and β3 integrins on bovine sperm indicate a possible role in sperm-oocyte adhesion and/or fusion.
Journal Article
Evidence for Cardiomyocyte Renewal in Humans
It has been difficult to establish whether we are limited to the heart muscle cells we are born with or if cardiomyocytes are generated also later in life. We have taken advantage of the integration of carbon-14, generated by nuclear bomb tests during the Cold War, into DNA to establish the age of cardiomyocytes in humans. We report that cardiomyocytes renew, with a gradual decrease from 1% turning over annually at the age of 25 to 0.45% at the age of 75. Fewer than 50% of cardiomyocytes are exchanged during a normal life span. The capacity to generate cardiomyocytes in the adult human heart suggests that it may be rational to work toward the development of therapeutic strategies aimed at stimulating this process in cardiac pathologies.
Journal Article
Effects of Lidocaine Infusion during Experimental Endotoxemia in Horses
Background: The clinical efficacy of IV infusion of lidocaine for treatment of equine endotoxemia has not been studied. Hypothesis: Lidocaine infusion after exposure to lipopolysaccharide (LPS) will inhibit the inflammatory response and have inhibitory effects on the hemodynamic and cytokine responses to endotoxemia. Animals: Twelve horses. Methods: Two equal groups (n = 6): saline (GI) and lidocaine (GII). In all animals, endotoxin (500 ng/kg body weight [BW]) was injected intraperitoneally over 5 minutes. Twenty minutes later, animals received a bolus of GI or GII (1.3 mg/kg BW) over 5 minutes, followed by a 6-hour continuous rate infusion of GI or GII (0.05 mg/kg BW/min). Treatment efficacy was judged from change in arterial blood pressure, peripheral blood and peritoneal fluid (PF) variables (total and differential cell counts, enzyme activities, and cytokine concentrations), and clinical scores (CS) for behavioral evidence of abdominal pain or discomfort during the study. Results: Compared with the control group, horses treated with lidocaine had significantly lower CS and serum and PF tumor necrosis factor- (TNF-) activity. At several time points in both groups, total and differential cell counts, glucose, total protein and fibrinogen concentrations, and alkaline phosphatase, creatine kinase, and TNF- activities were significantly different from baseline values both in peripheral blood and in PF. Conclusions and Clinical Importance: Lidocaine significantly decreased severity of CS and inhibited TNF- activity in PF.
Journal Article