Use of corticosteroid therapy on the modulation of uterine inflammatory response in mares after artificial insemination with frozen semen

In order to modulate uterine inflammatory response and evaluate the effect of corticosteroid therapy on fertility, 90 cycles of 45 mares were used for artificial insemination with frozen semen, using three different protocols: G1 - inseminated with frozen semen (800 x 10(6) viable spermatozoa pre-freezing) + 20 mL of seminal plasma; G2 - inseminated with frozen semen (800 x 10(6) viable spermatozoa pre-freezing) + corticosteroid therapy; G3 - inseminated with frozen semen (800 x 10(6) viable spermatozoa pre-freezing) + 20 mL of seminal plasma + corticosteroid therapy. Corticosteroid therapy consisted on one administration of prednisolone acetate (0.1 mg/Kg - Predef (R)) when mares presented 35mm follicles and uterine edema, concomitantly with the unique dose of hCG (human chorionic gonadotropin), then repeated each 12 hours until ovulation. on first fertility trial, with normal mares, there was no difference between control and treated groups (p>0.05), using seminal plasma associated with corticosteroid therapy (40 vs. 38%, respectively) or corticosteroid therapy alone (40 vs. 45% respectively). The second fertility trial, performed with mares with previous history of post-insemination endometritis, demonstrated a significant increase of pregnancy rate when mares were submitted to corticosteroid therapy (0.0 vs. 64.5%, respectively; p<0.05). Corticosteroid therapy was shown to be safe, with no physical or reproductive alterations on treated mares, demonstrating to be an adequate option to those animals with history of post-breeding or post-insemination endometritis. Further clinical research is necessary to confirm these results and contribute to the establishment of preventive therapy for cases of post-insemination endometritis.

Fertilizing Capacity of Frozen Epididymal Sperm Collected from Dogs

The collection of epididymal sperm may be a valuable tool for canine reproduction especially since it can enable collection of cells after death of a valuable dog. The aim of the present study was to evaluate the viability of epididymal sperm after freeze-thawing. Epididymides were obtained from four adult dogs by elective orchiectomy. The caudal portion of the epididymides and part of the deferential ducts were squeezed by means of an anatomic clamp into a Petri dish containing either 0.9% saline solution (Group 1) or Ringer solution without lactate (Group 2). Samples were centrifuged at 800 x g for 10 min, the supernatant was removed and the pellet was diluted in one step with a Tris/citric acid/OEP (Orvus Es Paste) extender containing 7% glycerol and subjected to semen freezing. Oocytes were obtained from canine ovaries, after ovariohysterectomy. Only oocytes that were approximately 100 mu m in diameter, with a dark ooplasm surrounded by three-or four-well formed cumulus cell layers were used for sperm testing. Frozen semen samples were thawed in a water bath at 70 degrees C for 8 s and analysed at room temperature for sperm motility and velocity. Oocytes were incubated with spermatozoa in humidified atmosphere containing 5% CO(2) at 38 degrees C for 18 h. Morphological and functional characteristics of spermatozoa were similar in both groups. However, the percentage of sperm cells bound to oocytes was significantly higher in Group 2 than in Group 1. This result suggests that the Ringer solution without lactate was a more suitable medium for collecting epididymal canine sperm than 0.9% saline.

Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen supplemented with catalase or Trolox

Semen manipulation and cryopreservation-thaw procedures may accelerate the generation of reactive oxygen species (ROS). Sperm exposure to large amounts of ROS has been shown to cause membrane lipid peroxidation and cellular injury to the sperm. The objective of this study was to overcome the ROS production in frozen-thawed ram semen by the addition of the antioxidants catalase or Trolox to semen following thawing. Frozen-thawed ram semen (100 x 10(6) sperm/straw) was supplemented with PBS (control group), 100 mu g/ml catalase, or 100 mu M Trolox/10(8) sperm (catalase and Trolox being dissolved in PBS) and incubated (37 degrees C) for 5 min. Under the experimental conditions used in this study, the catalase and Trolox antioxidants failed to protect the sperm from the spontaneous production of ROS. However, when lipid peroxidation was induced by iron (FeSO(4)), the addition of Trolox promoted a reduction (P < 0.05) in the formation of TBARS in the semen, compared to the control and catalase semen samples. The generation of TBARS and H(2)O(2) occurred in the extender alone, without the presence of sperm cells. In conclusion, the addition of Trolox to frozen-thawed ram semen could be beneficial as it decreases the production of TBARS when oxidative stress is induced. It is possible that a longer incubation period could lead to different results. The concentration of catalase also needs to be further evaluated. The extender could contribute to the oxidative stress of sperm, as it is a source of ROS during the cryopreservation of semen. (C) 2010 Elsevier B.V. All rights reserved.

Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen cryopreserved in extenders with antioxidants

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effect of refrigeration systems upon frozen bull sperm viability assessed by computer-assisted sperm analysis and fluorescent probes

Sperm cryopreservation success depends upon the maintenance of spermatozoa fertility potential. Sperm cells must preserve both integrity and functionality of several cell structures. The stabilization phase must allow the exit of water from the sperm cells via osmosis. This study aimed to compare the effect of refrigeration in the commercial refrigerator (CR) and the transport/refrigeration box (TRB) upon the viability of frozen bull sperm diluted in three different extenders (A, B and C). Ten Nellore bulls, Bos taurus indicus maintained in Artificial Insemination Center were used and the spermatozoa samples was assessed for Plasma Membrane Integrity and CASA evaluation. The stabilization phase (5 degrees C/4 hours) was performed in the CR as well as in the TRB, and then samples were exposed to nitrogen vapor during 20 minutes and then plunged into nitrogen. The statistical analysis was done using the variance analysis and the significance level was set at 5%. In the CR the post-thawing parameters for PM and ALH were higher (p < 0.05) in the extender A (glicine egg-yolk) and extender B (glicine egg-free) when compared with extender C (TRIS egg-yolk). As for BCF, STR and LIN, the parameters were higher (p < 0.05) in extender B than in C. Samples that were stabilized in the TRB presented higher post-thawing parameters (p < 0.05) for PM and LIN in extender A and extender B when compared with C. BCF and STR parameters were higher (p < 0.05) in extemder B when compared with C. Extender B samples had higher (p < 0.05) PMI when stabilized in CR. The findings in this experiment enable us to say that both CR and TRB were effective in keeping the viability of post-thawing bull semen.

Impact of proximal cytoplasmic droplets on quality traits and in-vitro embryo production efficiency of cryopreserved bull spermatozoa

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anterior Tooth Rehabilitation With Frozen Homogenous Bone Graft and Immediately Loaded Titanium Implant Using Computer-Guided Surgery

Computed tomographic scanning is a precise, noinvasive surveying technique that enables the professionals to improve the precision of implant placement by building a prototype that allows the confection of surgical guides. The authors present a clinical case of anterior tooth rehabilitation with frozen homogenous bone graft and immediately loaded titanium implant using computer-guided surgery. A multislice computed tomography was realized, and a prototype was built. All the procedures were previously realized in the prototype before started in the patient. This technique allows a better surgical planning, makes the procedures more accurate, and reduces surgery time.

Force system evaluation of symmetrical beta-titanium T-loop springs preactivated by curvature and concentrated bends

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Saliva substitute in combination with high-concentrated fluoride toothpaste: Effects on demineralised dentin in vitro

Objectives: The aim of this in vitro study was to assess the effects of saliva substitutes (modified with respect to calcium, phosphates, and fluorides) in combination with a high-concentrated fluoride toothpaste on demineralised dentin.Methods: Before and after demineralisation of bovine dentin specimens (subsurface lesions; 37 degrees C, pH 5.0, 5 d), one-quarter of each specimen's surface was covered with nail varnish (control of sound/demineralised tissue). Subsequently, specimens were exposed to original Saliva natura (saturation with respect to octacalciumphosphate [S(OCP)]: 0.03; SN 0), or to three lab-produced Saliva natura modifications (S(OCP): 1, 2, and 3; SN 1-3) for 2 and 5 weeks (37 degrees C). An aqueous solution (S(OCP): 2.5) served as positive control (PC). Two times daily (2 min each), Duraphat toothpaste (5000 ppm F(-); Colgate)/saliva substitute slurry (ratio 1:3) was applied gently. Differences in mineral losses (Delta Delta Z) and lesion depths (Delta LD) between values before and after exposure were microradiographically evaluated.Results: After both treatment periods specimens immersed in SN 0 revealed significantly higher mineral losses (lower Delta Delta Z values) and lesion depths (lower Delta LD) compared to PC (p < 0.05; ANOVA). After 5 weeks, specimens stored in SN 1 and 2 showed significantly higher mineral losses compared to PC (p < 0.05), while those stored in SN 3 showed similar results (p > 0.05). No differences in mineral loss could be observed between SN 2 and 3 (p > 0.05).Conclusions: Under the conditions of this limited protocol, the combination of Saliva natura solutions slightly saturated with respect to OCP in combination with a high-concentrated fluoride toothpaste enabled remineralisation of dentin in vitro. Crown Copyright (c) 2009 Published by Elsevier Ltd. All rights reserved.

Gas exchange rates at different vapor pressure deficits and water relations of 'Pera' sweet orange plants with citrus variegated chlorosis (CVC)

Net photosynthesis (A) and transpiration rates (E), stomatal conductance (g), water use efficiency (WUE), intrinsic water use efficiency (IWUE) and internal leaf CO2 concentration (C) in response to different vapor pressure deficit (1.2 and 2.5 kPa) were investigated in 'Pera' sweet orange plants affected by citrus variegated chlorosis (CVC), a disease caused by Xylella fastidiosa. All plants were well watered and leaf water potential (Pw) was also measured by the psychrometric technique. Results showed that healthy plants responded to higher vapor pressure deficit (VPD), lowering its net photosynthesis and transpiration rates, and stomatal conductance. However, diseased plants presented no clear response to VPD, showing lower A, E and g for both VPDs studied and very similar values to these variables in healthy plants at the highest VPD. Internal leaf CO2 concentration also decreased for healthy plants when under the highest VPD, and surprisingly, the same pattern of response was found in plants with CVC. These results, the lower Psi(w) and higher WUE values for diseased plants, indicated that this disease may cause stomatal dysfunction and affect the water resistance through xylem vessels, which ultimately may play some role in photosynthetic metabolism. (C) 2003 Elsevier B.V. B.V. All rights reserved.

Rooting of healthy and CVC-affected 'Valência' sweet orange stem cuttings, through the use of plant regulators

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Enzymes in juice processing: a review

P>The use of enzymes in juice industry has contributed in increasing the yield and production of various types of juices. The addition pectinases aims in particular to degrade the pectic substances, in the cell wall and middle lamella of the cells of plants, aiming to minimise the impacts of these compounds on the characteristics of the final product, such as colour, turbidity and viscosity. Enzymes able to remove bitterness of citrus juice, extract pigments, among other applications, have also had great interest in the juice industry.

Studies on productivity and characterization of polygalacturonase from Aspergillus giganteus submerged culture using citrus pectin and orange waste

Polygalacturonases are part of the group of enzymes involved in pectin degradation. The aim of this work was to investigate some of the factors affecting polygalacturonase production by an Aspergillus giganteus strain and to characterize this pectinolytic activity. Several carbon sources, both pure substances and natural substrates, were tested in standing cultures, and the best results were obtained with orange bagasse and purified citrus pectin. on citrus pectin as sole carbon source, the highest extracellular activity (9.5 U/ml and 40.6 U/mg protein) was obtained in 4.5-day-old cultures shaken at 120 rpm, pH 3.5 and 30 degrees C, while on orange bagasse, the highest extracellular activity (48.5 U/ml and 78.3 U/mg protein) was obtained in 3.5-day-old cultures shaken at 120 rpm, pH 6.0 and 30 degrees C. Optimal polygalacturonase activity was observed in assays conducted at pH 5.5-6.5 and 55-60 degrees C. The activity showed good thermal stability, with half-lives of 90 and 30 min when incubated at 55 and 60 degrees C, respectively. High stability was observed from pH 4.5 to 8.5; more than 90% of the activity remained after 24 h in this pH range.