Effects of Partial Replacement of Corn with Glycerin on Ruminal Fermentation in a Dual-Flow Continuous Culture System

The objective of this study was to evaluate the effects of partially replacing dry ground corn with glycerin on ruminal fermentation using a dual-flow continuous culture system. Six fermenters (1,223 ± 21 ml) were used in a replicated 3x3 Latin square arrangement with three periods of 10 d each, with 7 d for diet adaptation and 3 d for sample collections. All diets contained 75% concentrate and three dietary glycerin levels (0, 15, and 30% on dry matter basis), totaling six replicates per treatment. Fermenters were fed 72 g of dry matter/d equally divided in two meals/d, at 0800 and 2000 h. Solid and liquid dilution rates were adjusted daily to 5.5 and 11%/h, respectively. On d 8, 9, and 10, samples of 500 ml of solid and liquid digesta effluent were mixed, homogenized, and stored at -20°C. Subsamples of 10 ml were collected and preserved with 0.2 mL of a 50% H₂SO₄ solution for later determination of NH₃-N and volatile fatty acids. Microbial biomass was isolated from fermenters for chemical analysis at the end of each experimental period. Data were analyzed using the MIXED procedure in SAS with α = 0.05. Glycerin levels did not affect apparent digestibility of DM (P _Lin. = 0.13; P _Quad. = 0.40), OM (P _Lin. = 0.72; P _Quad. = 0.15), NDF (P _Lin. = 0.38; P _Quad. = 0.50) and ADF (P _Lin. = 0.91; P _Quad. = 0.18). Also, glycerin inclusion did not affect true digestibility of DM (P _Lin. = 0.35; P _Quad. = 0.48), and OM (P _Lin. = 0.08; P _Quad. = 0.19). Concentrations of propionate (P < 0.01) and total volatile fatty acids (P < 0.01) increased linearly and concentrations of acetate (P < 0.01), butyrate (P = 0.01), iso-valerate (P < 0.01), and total branched-chain volatile fatty acids, as well as the acetate: propionate ratio (P < 0.01) decreased with glycerin inclusion. Linear increases on NH₃-N concentration in digesta effluent (P < 0.01) and on NH₃-N flow (P < 0.01) were observed due to glycerin inclusion in the diets. Crude protein digestibility (P = 0.04) and microbial N flow (P = 0.04) were greater in the control treatment compared with the other treatments and responded quadratically with glycerin inclusion. Furthermore, the inclusion of glycerin linearly decreased (P = 0.02) non-ammonia N flow. Glycerin levels did not affect the flows of total N (P _Lin. = 0.79; P _Quad. = 0.35), and dietary N (P _Lin. = 0.99; P _Quad. = 0.07), as well as microbial efficiency (P _Lin. = 0.09; P _Quad. = 0.07). These results suggest that partially replacing dry ground corn with glycerin may change ruminal fermentation, by increasing total volatile fatty acids, and propionate concentration without affecting microbial efficiency, which may improve glucogenic potential of beef cattle diets.     

Issues in using whole slide imaging for diagnostic pathology: “routine” stains,immunohistochemistry and predictive markers

The traditional microscope, together with the “routine” hematoxylin and eosin (H & E) stain, remains the “gold standard” for diagnosis of cancer and other diseases; remarkably, it and the majority of associated biological stains are more than 150 years old. Immunohistochemistry has added to the repertoire of “stains” available. Because of the need for specific identification and even measurement of “biomarkers,” immunohistochemistry has increased the demand for consistency of performance and interpretation of staining results. Rapid advances in the capabilities of digital imaging hardware and software now offer a realistic route to improved reproducibility, accuracy and quantification by utilizing whole slide digital images for diagnosis, education and research. There also are potential efficiencies in work flow and the promise of powerful new analytical methods; however, there also are challenges with respect to validation of the quality and fidelity of digital images, including the standard H & E stain, so that diagnostic performance by pathologists is not compromised when they rely on whole slide images instead of traditional stained tissues on glass slides.     

Identification of DNA sequence variation in Campylobacter jejuni strains associated with the Guillain-Barré syndrome by high-throughput AFLP analysis

Background  

Campylobacter jejuni is the predominant cause of antecedent infection in post-infectious neuropathies such as the Guillain-Barré (GBS) and Miller Fisher syndromes (MFS). GBS and MFS are probably induced by molecular mimicry between human gangliosides and bacterial lipo-oligosaccharides (LOS). This study describes a new C. jejuni-specific high-throughput AFLP (htAFLP) approach for detection and identification of DNA polymorphism, in general, and of putative GBS/MFS-markers, in particular.     

Selective decrease in paracellular conductance of tight junctions: role of the first extracellular domain of claudin-5

Claudin-5 is a protein component of many endothelial tight junctions, including those at the blood-brain barrier, a barrier that limits molecular exchanges between the central nervous system and the circulatory system. To test the contribution of claudin-5 to this barrier function of tight junctions, we expressed murine claudin-5 in Madin-Darby canine kidney II cells. The result was a fivefold increase in transepithelial resistance in claudin-5 transductants and a reduction in conductance of monovalent cations. However, the paracellular flux of neither neutral nor charged monosaccharides was significantly changed in claudin-5 transductants compared to controls. Therefore, expression of claudin-5 selectively decreased the permeability to ions. Additionally, site-directed mutations of particular amino acid residues in the first extracellular domain of claudin-5 altered the properties of the tight junctions formed in response to claudin-5 expression. In particular, the conserved cysteines were crucial: mutation of either cysteine abolishted the ability of claudin-5 to increase transepithelial resistance, and mutation of Cys(64) strikingly increased the paracellular flux of monosaccharides. These new insights into the functions of claudin-5 at the molecular level in tight junctions may account for some aspects of the blood-brain barrier's selective permeability.     

Pharmacological evidence for beta2-adrenoceptor in right atria from stressed female rats.

The purpose of the present study was to demonstrate a physiological response to TA2005, a potent m-adrenoceptor (beta2-AR) selective agonist, in right atria isolated from stressed female rats under the influence of the estrous cycle. We obtained concentration-response curves to the agonist in the presence and in the absence of selective antagonists in right atria isolated from female rats submitted to three daily foot-shock sessions (30 min duration, 120 pulses of 1.0 mA, 1.0 s, applied at random intervals of 5-25 s) and sacrificed at estrus or diestrus. Our results showed that the pD2 values of TA2005 were not influenced by estrous cycle phase or foot-shock stress. However, in right atria from stressed rats sacrificed during diestrus, the concentration-response curve to TA2005 was biphasic, with a response being obtained at concentrations of 0.1 nM, whereas during estrus no response was observed at doses lower than 3 nM. ICI 1118,551, a beta2-AR antagonist, abolished the response to nanomolar concentrations of TA2005 in right atria from stressed rats at diestrus, with no changes in agonist pD2 values in right atria from control rats (7.47 +/- 0.09, p > 0.05) but a 3-fold decrease in pD2 values of TA2005 in right atria from foot shock stressed rats (7.90 +/- 0.07, p < or = 0.05). Concentration-response curves to TA2005 in the presence of ICI118,551 were best fitted by a one-site model equation. The beta1-AR antagonist, CGP20712A, shifted to the right only the second part of the concentration-response curves to the agonist, unmasking the putative beta2-AR-mediated response to the agonist in tissues isolated from stressed rats at diestrus. Under this condition, concentration-response curves to the agonist were best fitted by a two-site model equation. pD2 and maximum response of TA2005 interaction with beta1- and putative beta2-adrenoceptor components were calculated. Schild analyses gave a pK(B) value for CGP20712A that was typical for the interaction with beta1-AR in each experimental group. pK(B) values for ICI118,551 could not be obtained in stressed rats sacrificed at diestrus since Schild plot slopes were lower than 1.0. In right atria from control rats, ICI118,551 pK(B) values were similar to reported values for the interaction of the antagonist with beta1-AR. These results confirm that a heterogenous beta-AR population mediating the chronotropic response to catecholamines can be demonstrated in right atria from foot shock stressed female rats sacrificed at diestrus. The stress-induced response seems to be mediated by the beta2-AR subtype. Right atria from rats sacrificed during estrus are protected against stress-induced alterations on the homogeneity of beta-AR population.     

Morphological and molecular characterization of populations of plains bristlegrass (<i>Setaria macrostachya</i> Kunth) in Chihuahua,México

Plains bristlegrass (Setaria macrostachya Kunth) is a native grass with forage value. However, due to the lack of grazing management practices, populations and thus genetic diversity, have been reduced. Morphological and genetic variability were analyzed on 44 populations of plains bristlegrass in the State of Chihuahua. Plants were transplanted in a common area under natural conditions. Two years later, morphological characterization was evaluated measuring nine variables, and genetic variability using AFLP molecular markers. The principal components analysis (PC) showed that the three first principal components explained 73.74% of the variation. The variables with the greatest contribution to the variance in PC1 were plant height and inflorescence length; in CP2, tiller number and leaf width; and in PC3, tiller thickness. Application of four pairs of primers, presented 186 total bands, from which 87.10% showed polymorphism and 12.90% monomorphism. The combination of EcoRI-AGG MseI-CAG primers detected the highest percentage (93%) of polymorphism with 40 polymorphic bands. The cluster analysis and Dice coefficient indicated that populations clump into two groups. The wide genetic variability and morphological characteristics detected among populations represent the basis for the selection of populations that could be used with different purposes in the rehabilitation of ecosystems. In addition, this study will allow establishment of in situ conservation strategies.     

Aerobic training prevents cardiometabolic changes triggered by myocardial infarction in ovariectomized rats

This study aimed to evaluate the impact of aerobic training (AT) on autonomic, cardiometabolic, ubiquitin‐proteasome activity, and inflammatory changes evoked by myocardial infarction (MI) in ovariectomized rats. Female Wistar rats were ovariectomized and divided into four groups: sedentary + sham (SS), sedentary + MI (SI), AT + sham surgery (TS), AT + MI (TI). AT was performed on a treadmill for 8 weeks before MI. Infarcted rats previously subjected to AT presented improved physical capacity, increased interleukin‐10, and decreased pro‐inflammatory cytokines. Metabolomic analysis identified and quantified 62 metabolites, 9 were considered significant by the Vip Score. SS, SI, and TS groups presented distinct metabolic profiles; however, TI could not be distinguished from the SS group. MI dramatically increased levels of dimethylamine, and AT prevented this response. Our findings suggest that AT may be useful in preventing the negative changes in functional, inflammatory, and metabolic parameters related to MI in ovariectomized rats.     

Immunohistochemistry in the evaluation of neovascularization in tumor xenografts

Angiogenesis, or neovascularization, is known to play an important role in the neoplastic progression leading to metastasis. CD31 or Factor VIII-related antigen (F VIII RAg) immunohistochemistry is widely used in experimental studies for quantifying tumor neovascularization in immunocompromised animal models implanted with transformed human cell lines. Quantification, however, can be affected by variations in the methodology used to measure vascularization including antibody selection, antigen retrieval (AR) pretreatment, and evaluation techniques. To examine this further, we investigated the microvessel density (MVD) and the intensity of microvascular staining among five different human tumor xenografts and a mouse syngeneic tumor using anti-CD31 and F VIII RAg immunohistochemical staining. Different AR methods also were evaluated. Maximal retrieval of CD31 was achieved using 0.5 M Tris (pH 10) buffer, while maximum retrieval of F VIII RAg was achieved using 0.05% pepsin treatment of tissue sections. For each optimized retrieval condition, anti-CD31 highlighted small vessels better than F VIII RAg. Furthermore, the MVD of CD31 was significantly greater than that of F VIII RAg decorated vessels (p<0.001). The choice of antibody and AR method has a significant affect on immunohistochemical findings when studying angiogenesis. One also must use caution when comparing studies in the literature that use different techniques and reagents.     

A quarter of a world away: female humpback whale moves 10,000 km between breeding areas

Fidelity of individual animals to breeding sites is a primary determinant of population structure. The degree and scale of philopatry in a population reflect the fitness effects of social facilitation, ecological adaptation and optimal inbreeding. Patterns of breeding-site movement and fidelity are functions of social structure and are frequently sex biased. We report on a female humpback whale (Megaptera novaeangliae) first identified by natural markings off Brazil that subsequently was photographed off Madagascar. The minimum travel distance between these locations is greater than 9800 km, approximately 4000 km longer than any previously reported movement between breeding grounds, more than twice the species' typical seasonal migratory distance and the longest documented movement by a mammal. It is unexpected to find this exceptional long-distance movement between breeding groups by a female, as models of philopatry suggest that male mammals move more frequently or over longer distances in search of mating opportunities. While such movement may be advantageous, especially in changeable or unpredictable circumstances, it is not possible to unambiguously ascribe causality to this rare observation. This finding illustrates the behavioural flexibility in movement patterns that may be demonstrated within a typically philopatric species.     

Influence of estradiol and progesterone on the sensitivity of rat thoracic aorta to noradrenaline

The aim of this study was to investigate the effects of low and high doses of estradiol, and of progesterone on the response to noradrenaline in rat thoracic aorta. Two weeks after bilateral ovariectomy, female rats received a s.c. injection of vehicle (corn oil, 0.1 mL/day), estradiol (10 microg/kg/day or 4 mg/kg/day) and/or progesterone (20 mg/kg/day), for eight days. On the ninth day, the rats were sacrificed and aortic rings, with or without endothelium, were used to generate concentration-response curves to noradrenaline. Aortic rings with intact endothelium from the high-dose (4 mg/kg/day) estradiol group were supersensitive to noradrenaline compared to the vehicle or low-dose (10 microg/kg/day) estradiol groups (pD2 values = 7.86+/-0.09, 7.30+/-0.11 and 7.35+/-0.04, respectively). Endothelium-intact aortic rings from high-estradiol rats were supersensitive to noradrenaline when compared to vehicle-, progesterone- and progesterone + high-estradiol-treated rats (pD2 values = 7.77+/-0.12, 7.21+/-0.13, 6.93+/-0.04 and 7.22+/-0.18, respectively). There were no significant differences among the pD2 values for noradrenaline in aortic rings without endothelium. In conclusion, at high but not low doses, estradiol increased the sensitivity to noradrenaline and this was prevented by progesterone. Both of these effects were endothelium-dependent.