Use of primers derived from a new sequence of the bovine Y chromosome for sexing Bos taurus and Bos indicus embryos

A bovine male-specific marker was identified in our laboratory through random amplified polymorphic DNA (RAPD) analysis. This fragment of 3216 bp was cloned, sequenced and mapped by fluorescent in situ hybridization (FISH) on the taurine Yq. Primers derived from this sequence were initially screened by polymerase chain reaction (PCR) for their ability to detect Y-specific segments in zebu and taurine genomic DNA. Two of these primers amplified a 655 bp Y-specific sequence present in taurine and zebu male genomic DNA. These primers were then used for detecting the 655 bp male sequence in DNA from 173 zebu and 30 taurine embryos, which had been previously sexed using primers for the sequence BC 1.2. The results revealed an accuracy of 100%. (C) 2002 Elsevier B.V. All rights reserved.

Fracture strength of bovine incisors after intra-radicular treatment with MTA in an experimental immature tooth model

Aim To evaluate, using an experimental immature tooth model, the fracture resistance of bovine incisors submitted to different reinforcement treatments with mineral trioxide aggregate (MTA).Methodology An immature tooth model was created by sectioning the coronal and apical portions of 40 bovine incisors 8 mm above and 12 mm below the cementoenamel junction. The root canals were irrigated with 1.0% sodium hypochlorite. They were enlarged both coronally and apically using number 703 carbide burs (ISO: 500-104-168-007-021) and their internal diameter was standardized to 2.1 mm. The specimens were assigned to four groups (n = 10): GI-control (without filling); GII-apical MTA plug + filling with gutta-percha and endodontic sealer; GIII-filling with MTA; GIV-apical MTA plug + filling with MTA + metallic post (Reforpost I). A polyether impression material was used to simulate the periodontal ligament. The specimens were submitted to a compressive load at a crosshead speed of 0.5 mm min(-1) in a servo-hydraulic universal testing machine (MTS 810) applied at 45 degrees to the long axis of the tooth until failure. Data were submitted to statistical analysis by the Kruskal-Wallis test at 5% significance level.Results GIV presented the highest fracture resistance (32.7N) and differed significantly from the other groups (P < 0.05). No statistically difference was found between GII (16.6N) and GIII (23.4N) (P > 0.05). GIII had a significantly higher fracture resistance than GI (P < 0.05).Conclusions the use of MTA + metallic post as an intra-radicular reinforcement treatment increased the resistance to fracture of weakened bovine teeth in an experimental immature tooth model.

Cholinesterase measurements with an automated kit

Background the Test-mate kit determines acetylcholinesterase (AChE, EC 3.1.1.7) and hemoglobin content of a drop of blood, displaying enzyme activities normalized to 25degreesC. Previous models produced inconsistent results at different temperatures. This report focuses on the current model, ChE 400, and two instruments of a previous OP model.Methods AChE activities were determined by the Ellman assay, using the three kits and a 96-well microplate reader Temperatures ranged from 10 to 37degreesC. Fetal bovine serum was the source of AChE.Results Normalized activities decreased below 20degreesC in the ChE model and below 25 C in the OP models. Activities of the same serum sample differed between the three Test-mate kits, ranging from 1.03 to 1.49 mumoles/min/ml. Percent errors were greater than with the microplate reader at all temperatures.Conclusions Neither we nor the manufacturer recommend the current Test-mate model for fieldwork. Nevertheless, there have been field measurements with Test-Mate kits, and we recommend that an enzyme activity standard be run in parallel with their use. (C) 2002 Wiley-Liss, Inc.

Constituintes sangüíneos de vacas das raças Nelore (Bos indicus) e Holandesa (Bos taurus) e de bubalinas (Bubalus bubalis) da raça Murrah durante a gestação, no dia do parto e no puerpério

Blood constituents of Nelore, Hosltein and buffalo cows were determined during pregnancy, at the time of calving and at post-partum. It was verified that. 1. The pregnancy and post-partum periods had no influence in the erythrocyte values, but at calving the red blood cell counts and hemoglobin levels of Holstein and buffalo cows were lower than for Nelore cattle; 2. leukocyte counts were similar among groups; 3. total protein levels of Nelore cows were lower than Holstein and buffalo. The albumin levels were rite lowest at rite time of calving compared to late pregnancy; 4. glicose levels were lower ill buffalo cows during pregnancy as compared with Holstein cows. The glicemia of Nelore cows was lower as compared to Holstein cow's irt late pregnancy; 5. urea and creatinine levels were higher in buffalo cows than cattle. The urea and creatinine levels were greater in buffalo cows with maximum values at the post-partum and at the parturition, respectively; 6. bilirrubin levels were higher in bovine than buffalo cows; 7. aspartate aminotransferase activity was greater in buffalo cows and increased at the time of calving; 8. alkaline phosphatase activity was increased during pregnancy and decreased after the time of calving; 9. gammnglutamyltransferase activity,vas the highest for buffalo cows after calving; IO. calcium levels were the highest for Holstein cows at the post-partum and the phosphorus levels were higher in buffalo cows, which had the highest magnesium levels at the parturation.

Use of strontium in the activation of bovine oocytes reconstructed by somatic cell nuclear transfer

As an important step in the nuclear transfer (NT) procedure, we evaluated the effect of three different treatments for oocyte activation on the in vitro and in vivo developmental capacity of bovine reconstructed embryos: (1) strontium, which has been successfully used in mice but not yet tested in cattle; (2) ionomycin and 6-dimethylammopurine (6-DMAP), a standard treatment used in cattle; (3) ionomycin and strontium, in place of 6-DMAP. As regards NT blastocyst development, no difference was observed when strontium (20.1%) or ionomycin/6-DMAP (14.4%) were used. However, when 6-DMAP was substituted by strontium (3), the blastocyst rate (34.8%) was superior to that in the other activation groups (p < 0.05). Results of in vivo development showed the possibility of pregnancies when NT embryos activated in strontium were transferred to recipient cows (16.6%). A live female calf was obtained when ionomycin/strontium were used, but it died 30 days after birth. Our findings show that strontium can be used as an activation agent in bovine cloning procedures and that activation with a combination of strontium and ionomycin increased the in vitro developmental capacity of reconstructed embryos. This is the first report of a calf produced by adult somatic cell NT in Latin America.

Identification, content, and distribution of type VI collagen in bovine tendons

Tendon composition changes according to differentiation, mechanical load, and aging. In this study, we attempted to identify, localize, and quantify type VI collagen in bovine tendons. Type VI collagen was identified by the electrophoretic behavior of the alpha chains and Western blotting, and by rotary shadowing. Type VI collagen was extracted from powdered tendon with three sequential 24-h extractions with 4 M guanidine-HCl. The amount of type VI collagen was determined by enzyme-linked immunosorbent assay for purely tensional areas and for the compressive fibrocartilage regions of the deep flexor tendon of the digits, for the corresponding fetal and calf tendons, and for the extensor digital tendon. The distal fibrocartilaginous region of the adult tendon was richer in type VI collagen than the tensional area, reaching as much as 3.3 mg/g (0.33%) of the wet weight. Calf tendons showed an accumulation of type VI at the fibrocartilage site. Immunocytochemistry demonstrated that type VI collagen was evenly distributed in the tensional areas of tendons but was highly concentrated around the fibrochondrocytes in the fibrocartilages. The results demonstrate that tendons are variable with regard to the presence and distribution of type VI collagen. The early accumulation of type VI collagen in the region of calf tendon that will become fibrocartilage in the adult suggests that it is a good marker of fibrocartilage differentiation. Furthermore, the distribution of type VI collagen in tendon fibrocartilage indicates that it organizes the pericellular environment and may represent a survival factor for these cells.

Effect of post length on the fatigue resistance of bovine teeth restored with bonded fiber posts: A pilot study

This study evaluated the influence of the cementation length of glass fiber-reinforced composite (FRC) on the fatigue resistance of bovine teeth restored with an adhesively cemented FRC. Thirty roots of single-rooted bovine teeth were allocated to 3 groups (n = 10), according to the ratio of crown length/root length (post cementation length): group 1 = 2/3, group 2 = 1/2, and group 3 = 1/1. The roots were prepared, the fiber posts (FRC Postec Plus) were cemented, and the specimens were submitted to 2 million mechanical cycles. After fatigue testing, a score was given based on the number of fatigue cycles until fracture, and data were submitted to statistical analysis. All specimens were resistant to fatigue. Taking into account the methodology and results of this study, the evaluated fiber posts can be cemented based on the ratio of crown/root at 1/1. Further clinical studies must be conducted to verify this ratio.

Role of roscovitine and IBMX on kinetics of nuclear and cytoplasmic maturation of bovine oocytes in vitro

The 3-isobutyl-1-methylxanthine (IBMX) is able to prevent resumption of meiosis by maintaining elevated cyclic AMP (cAMP) concentrations in the oocyte, and roscovitine, a purine known to specifically inhibit MPF kinase activity, maintains bovine oocytes at the germinal vesicle (GV) stage. The present study was conducted to analyze whether cytoplasmic maturation (examined by the pattern of cortical granule (CG) distribution) of bovine oocytes is improved during meiotic arrest with IBMX and roscovitine. Oocytes were matured in vitro in a 10% Knockout(SR) supplemented TCM-199 medium (Control) with either 0.5 mM IBMX or 25 mu M roscovitine (ROSC). Oocytes were stained with fluorescein isothiocyanate conjugated Lens culinaris agglutinin (FITC-LCA) for CG evaluation and with Hoechst 33342 for nuclear stage assessment. At 16 h of culture, the percentage of oocytes remaining in the GV stage was higher (P < 0.05) in the ROSC group (32.41%) compared with the Control and IBMX groups (8.61% and 9.73%, respectively). At 24h of culture, progression of meiosis to M II stage was retarded (P < 0.05) in the ROSC group (24.05%) compared to the Control (60.20%), whereas the IBMX group (33.88%) showed no significant difference to the other two groups. At 16h of maturation, the proportion of oocytes with CG in clusters (immature cytoplasm) was similar between the groups, as was the percentage of peripheral CG (mature) at 24h of maturation. The results of the present study demonstrated that the meiotic inhibitors IBMX and roscovitine delay the progression of nuclear maturation without affecting cytoplasmic maturation, assessed by the analysis of CG repositioning. (c) 2006 Elsevier B.V. All rights reserved.

Penetration of 38% hydrogen peroxide into the pulp chamber in bovine and human teeth submitted to office bleach technique

This study evaluated the pulp chamber penetration of peroxide bleaching agent in human and bovine teeth after office bleach technique. All the teeth were sectioned 3 mm apical of the cement-enamel junction and were divided into 2 groups, A (70 third human molars) and B (70 bovine lateral incisors), that were subdivided into A1 and B1 restored by using composite resin, A2 and B2 by using glass ionomer cement, and A3 and B3 by using resin-modified glass ionomer cement; A4, A5, B4, and B5 were not restored. Acetate buffer was placed in the pulp chamber, and the bleaching agent was applied for 40 minutes as follows: A1-A4 and B1-B4, 38% hydrogen peroxide exposure and A5 and B5, immersion into distilled water. The buffer solution was transferred to a glass tube in which leuco crystal violet and horseradish peroxidase were added, producing a blue solution. The optical density of the blue solution was determined by spectrophotometer and converted into microgram equivalents of hydrogen peroxide. Data were submitted to analysis of variance and Dunnett, Kruskal-Wallis, and Tukey tests (5%). A higher level of hydrogen peroxide penetrated into the pulp chamber in resin-modified glass ionomer cements in bovine (0.79 +/- 0.61 mu g) and human (2.27 +/- 0.41 mu g) groups. The bleaching agent penetration into the pulp chamber was higher in human teeth for any experimental situation. The penetration of the hydrogen peroxide depends on restorative materials, and under the conditions of this study human teeth are more susceptible to penetration of bleaching agent into the pulp chamber than bovine teeth.