Contrasting effects of heat shock during in vitro maturation on development of in vitro‐fertilized and parthenogenetic bovine embryos

This study investigated the influence of heat shock during in vitro maturation on embryo development following in vitro fertilization (IVF) or parthenogenesis (Part). Immature bovine cumulus–oocyte complexes were exposed to heat shock (41.0°C) during the first 12 hr of in vitro maturation (IVM), followed by 12 hr at 38.5°C. Control group consisted of in vitro maturation for 24 hr at 38.5°C. Oocytes were in vitro‐fertilized or activated with ionomycin and cultured in vitro for 192 hr post‐in vitro insemination or parthenogenetic activation (hpia). There was an interaction (p < .01) between temperature of IVM and method of oocyte activation (IVF or Part) for cleavage at 48 hpia. Heat shock had a negative impact (p < .01) on cleavage of IVF embryos, whereas no (p > .05) effect was found in the Part embryos. Embryo development towards blastocyst stage at 168 and 192 hpia decreased in both IVF and Part embryos derived from heat‐shocked oocytes. Heat shock increased (p < .05) the apoptotic index in Part blastocysts, but no effect (p > .05) was found in IVF counterparts. Heat shock also down‐regulated the expression of AQP3 (p < .01) and up‐regulated the expression of HSP70.1 (p < .01) in Part blastocysts, whereas it down‐regulated the expression of ATP1A1 (p < .05) in IVF blastocysts. In conclusion, the effects of heat shock during IVM on early embryo cleavage and blastocyst apoptosis are influenced by the method of oocyte activation and expression of some genes can be disturbed in embryos derived from heat‐shocked oocytes.     

Exogenous paraoxonase‐1 during oocyte maturation improves bovine embryo development in vitro

Paraoxonase‐1 (PON1) is an enzyme found in serum and follicular fluid that protects cell membrane and circulating lipids against oxidative damage. The aims of this study were to measure the direct effects of recombinant PON1 (rPON1) on bovine oocyte maturation at the molecular level (gene expression) and to measure the carry‐over effects of PON1 on pre‐implantation embryo development in vitro. COCs were submitted to IVM with the addition of 0.0, 0.02, 0.04 and 0.08 mg ml^?1 of rPON1, corresponding to an average PON1 arylesterase enzyme activity of 2.2 ± 0.4, 15.5 ± 1.5, 30.2 ± 3.0 and 57.9 ± 5.0 U ml^?1, respectively. The results indicated that addition of rPON1 during IVM improved embryo development in a dose‐dependent manner as D7 embryo development was 22.2%, 29.4%, 32.2% and 37.0% for the treatment groups, respectively (p = 0.02). In conclusion, addition of PON1 enzyme during IVM exerted dose‐related positive effects on embryo development rates to blastocysts.     

Agarose gel electrophoresis of cerebrospinal fluid proteins of dogs after sample concentration using a membrane microconcentrator technique

BACKGROUND: Cerebrospinal fluid (CSF) is produced in the cerebral ventricles through ultrafiltration of plasma and active transport mechanisms. Evaluation of proteins in CSF may provide important information about the production of immunoglobulins within the central nervous system as well as possible disturbances in the blood-brain barrier. OBJECTIVE: The objective of this study was to measure the concentration and fractions of protein in CSF samples using a membrane microconcentrator technique followed by electrophoresis, and to compare the protein fractions obtained with those in serum. METHODS: CSF samples from 3 healthy dogs and 3 dogs with canine distemper virus infection were concentrated using a membrane microconcentrator having a 0.5 to 30,000 d nominal molecular weight limit (Ultrafree, Millipore, Billerica, MA, USA). Protein concentration was determined before and after concentration. Agarose gel electrophoresis was done on concentrated CSF samples, serum, and serial dilutions of one of the CSF samples. RESULTS: Electrophoretic bands were clearly identified in densitometer tracings in CSF samples with protein concentrations as low as 1.3 g/dL. The higher CSF protein concentration in dogs with distemper was mainly the result of increased albumin concentration. CONCLUSION: The microconcentrating method used in this study enables characterization of the main protein fractions in CSF by routine electrophoresis and may be useful for interpreting the underlying cause of changes in CSF protein concentrations.     

The importance of the proportion of heme/nonheme iron in the diet to minimize the interference with calcium, phosphorus, and magnesium metabolism on recovery from nutritional ferropenic anemia.

The digestive utilization of Fe and its nutritive interaction with Ca, P, and Mg were studied in rats with nutritional ferropenic anemia. The diet contained 80% ferric citrate and 20% heme iron (80/20 diet). The weight gain, digestive utilization of Fe, and regeneration efficiency of hemoglobin and seric Fe were higher in iron-deficient rats (ID) fed the 80/20 diet than in iron-deficient rats fed the 50/50 diet (Campos et al., 1996). The phospho-calcic metabolism, which is adversely affected in ferropenic anemia, returned to normal values when iron was added to the diet. The digestive utilization of Mg, which fell with the 50/50 diet (Campos et al., 1996), returned to normal values when the ferropenic anemia was reversed with the 80/20 diet. In a state of iron deficiency, certain parameters related to the glucose and lipid metabolism are affected; the glucose and triglycerides values return to a normal range with the 80/20 diet.     

Self-oriented regular arrays of carbon nanotubes and their field emission properties

The synthesis of massive arrays of monodispersed carbon nanotubes that are self-oriented on patterned porous silicon and plain silicon substrates is reported. The approach involves chemical vapor deposition, catalytic particle size control by substrate design, nanotube positioning by patterning, and nanotube self-assembly for orientation. The mechanisms of nanotube growth and self-orientation are elucidated. The well-ordered nanotubes can be used as electron field emission arrays. Scaling up of the synthesis process should be entirely compatible with the existing semiconductor processes, and should allow the development of nanotube devices integrated into silicon technology.     

Pathogenesis of porcine Actinobacillus pleuropneumonia, part II: roles of proinflammatory cytokines.

The in vitro production of proinflammatory cytokines after stimulation with Actinobacillus pleuropneumoniae and the relation of these cytokines in vivo with the disease caused by A. pleuropneumoniae were investigated. Within 24 h, in vitro stimulation by A. pleuropneumoniae (serotype 1) preparations, including killed bacteria, bacterial culture supernatant, lipopolysaccharide, and bacterial extracts, porcine pulmonary alveolar macrophages (PAM) produced significant (P < 0.05) amounts of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1) as measured by bioassays. The supernatants containing interleukin-8 from PAM after stimulation by bacterial preparations showed significant neutrophil chemotaxis, while bacterial preparations alone did not. After in vivo infection with A. pleuropneumoniae, the mean levels of TNF-alpha and IL-1 in serum, as measured by bioassays, were elevated 37- to 27836-fold for TNF-alpha and 11- to 5941-fold higher for IL-1 within 4 d post-infection, depending on the treatments, and remained elevated up to Day 7. Both cytokines were also detected in porcine lungs by bioassays and immunocytochemistry. The results indicated that both secreted and surface components of A. pleuropneumoniae can stimulate PAM to produce proinflammatory mediators. Neutrophil chemoattractants rather than bacterial components are the major factor causing acute lung inflammation. The elevation of TNF-alpha and IL-1 in pigs occurred coincident with the onset of acute clinical disease.     

Chemical composition of Lippia spp. essential oil and antimicrobial activity against Aeromonas hydrophila

Phytotherapy can replace antibiotic administration as an alternative to control Aeromonas hydrophila, one of the main bacteria involved in the aetiology of farmed fish diseases. Given that plants of the Lippia spp. genus show biological potential for antimicrobial activity, this study evaluated the chemical composition of essential oils extracted from Lippia alba, Lippia origanoides and Lippia sidoides and their activity against A. hydrophila. The oils were obtained by steam distillation in a Clevenger‐type apparatus and their composition determined by gas chromatography and mass spectrometry (CG/MS). Antibacterial activity was assessed by calculating the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) using broth microdilution method. The main compounds identified were geranial (25.4%) and neral (16.6%) in L. alba oil, carvacrol (40.4%) and p‐cymene (11.4%) in L. origanoides oil and thymol (76.6%) and ortho‐cymene (6.3%) in L. sidoides oil. The three Lippia species showed bacteriostatic and bactericidal action against A. hydrophila, with MICs and MBCs ranging from 1250 to 5000 μg mL^?1. Of the species tested, the best performance was obtained with essential oil of L. sidoides.     

Effect of the addition of Chaetoceros calcitrans,Navicula sp. and Phaeodactylum tricornutum (diatoms) on phytoplankton composition and growth of Litopenaeus vannamei (Boone) postlarvae reared in a biofloc system

The aim of this study was to evaluate the effect of the addition of Chaetoceros calcitrans, Navicula sp. and Phaeodactylum tricornutum (diatoms) on phytoplankton composition and the growth of Litopenaeus vannamei postlarvae reared in a biofloc system . Four treatments were used: BFT (biofloc system without feed and no addition of diatoms); BFT‐F (biofloc system with feed and no addition of diatoms); BFT‐D (biofloc system with the addition of diatoms and no feed); and BFT‐FD (biofloc system with the addition of feed and diatoms), all in triplicate. The shrimp (16 ± 0.02 mg) were stocked at 2500 postlarvae m^?3 and fed a commercial feed. Diatoms were added on the 1st, 5th, 10th and 15th day at a density of 5 × 10⁴ cells mL^?1 for each species. No significant differences (P > 0.05) between treatments were observed for gross primary production, net ecosystem production and water column respiration rate. However, significant differences (P < 0.05) were observed for nitrite, orthophosphate, alkalinity, final weight, weight gain, yield, feed conversion ratio (FCR), phytoplankton and cyanobacteria composition. The BFT‐FD treatment had better performance parameters for final weight (270 mg), weight gain (254 mg), yield (0.67 Kg m^?3) and FCR (0.61), indicating the benefits of the diatoms C. calcitrans, Navicula sp. and P. tricornutum for decreasing cyanobacteria and improving growth of L. vannamei postlarvae reared in biofloc systems.