Immunohistochemical identification and fiber type specific localization of protein kinase C isoforms in equine skeletal muscle

OBJECTIVE: To investigate whether protein kinase C (PKC) isoforms are expressed in equine skeletal muscle and determine their distribution in various types of fibers by use of immunofluorescence microscopy. ANIMALS: 5 healthy adult Dutch Warmblood horses. PROCEDURE: In each horse, 2 biopsy specimens were obtained from the vastus lateralis muscle. Cryosections of equine muscle were stained with PKC isoform (alpha, beta1, beta2, delta, epsilon, or zeta)-specific polyclonal antibodies and examined by use of a fluorescence microscope. Homogenized muscle samples were evaluated via western blot analysis. RESULTS: The PKC alpha, beta1, beta2, delta, epsilon, and zeta isoforms were localized within the fibers of equine skeletal muscle. In addition, PKC alpha and beta2 were detected near or in the plasma membrane of muscle cells. For some PKC isoforms, distribution was specific for fiber type. Staining of cell membranes for PKC alpha was observed predominantly in fibers that reacted positively with myosin heavy chain (MHC)-IIa; PKC delta and epsilon staining were more pronounced in MHC-I-positive fibers. In contrast, MHC-I negative fibers contained more PKC zeta than MHC-I-positive fibers. Distribution of PKC beta1 was equal among the different fiber types. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that PKC isoforms are expressed in equine skeletal muscle in a fiber type-specific manner. Therefore, the involvement of PKC isoforms in signal transduction in equine skeletal muscle might be dependent on fiber type.     

The effect of bpV(HOpic) on in vitro activation of primordial follicles in cultured swine ovarian cortical strips

The vanadate‐derivative dipotassium bisperoxo (5‐hydroxy‐pyridine‐2‐carboxylic) oxovanadate (V) (bpV(HOpic)), a pharmacological inhibitor of phosphatase and tensin homolog (PTEN), has been used in ovarian follicle culture systems for activation of follicular growth in vitro and suggested to be responsible for primordial follicle survival through indirect Akt activation. For pig ovarian tissue, it is still not clear which culture medium needs to be used, as well as which factors and hormones could influence follicular development; this also applies to bpV(HOpic) exposure. Therefore, ovarian cortical strips from pigs were cultured in 1 µM bpV(HOpic) (N = 24) or control medium (N = 24) for 48 hr. Media were then replaced with control medium and all tissue pieces incubated for additional 4 days. The strips were embedded in paraffin for histological determination of follicle proportions at the end of the culture period and compared to histological sections from tissue pieces without cultivation, which had been embedded right after preparation; comparison of healthy follicles for each developmental stage was performed to quantify follicle survival and activation. After 6‐day culture, follicle activation occurred in tissue samples from both cultured groups but significantly more follicles showed progression of follicular development in the presence of 1 µM bpV(HOpic). The amount of non‐vital follicles was not significantly increased during cultivation. BpV(HOpic) affects pig ovarian follicle development by promoting the initiation of follicle growth and development, similar as in rodent species and humans.     

Soil Sorption of Nickel in presence of Citrate or Arginine

The influence from 0.5 and 5 mmol L^-1 citrate andarginine on nickel sorption in sandy loamy topsoil ofa Mollic Hapludalf has been investigated in order toevaluate the effect of organic complexing agents onnickel retention in soils. Both ligands, which formstrong complexes with nickel, represent common soilsolutes but also serve as models for oxygen andnitrogen donor ligands in fulvic acids and otherorganic soil solutes. In the pH range 4.5–8,sorption edges showed a dramatic decrease in nickelsorption in presence of citrate, but not in thepresence of arginine. These differences were furtherquantified at pH 6.0 through sorption isothermscovering initial nickel concentrations in the range3.4–42.6 μmol L^-1. Again, the amounts ofnickel sorbed without and in the presence of 0.5 and5 mmol L^-1 arginine were not significantlydifferent, whereas 0.5 and 5 mmol L^-1 citratereduced sorption by 50 to 90%. The ligand effectscould be expressed through variation of the Freundlichaffinity constant K. The results indicate thattrivalent positively charged nickel-arginine complexes(NiH₃arg₂ ³⁺) are sorbed to cationexchange sites, while in the presence of citrate sitesfor the bonding of monovalent negatively chargednickel-citrate complexes (Nicit^-) are sparse.     

Bioactive benzoxazinoids in rye bread are absorbed and metabolized in pigs

Recently, bioactive benzoxazinoids were discovered in cereal grains and bakery products. In this study, we studied the uptake, distribution, and metabolism of these secondary metabolites using a pig model. Twelve benzoxazinoid compounds and their 4 transformation products were quantified in the pigs' diets and biofluids using high-performance liquid chromatography coupled to electrospray ionization triple quadrupole mass spectrometry. The 2-β-D-glucopyranosyloxy-4-hydroxy-1,4-benzoxazin-3-one (DIBOA-glc) was the most dominant benzoxazinoid (232 nmol/g DM) seconded by the double-hexose derivative of DIBOA (provisionally characterized here as DIBOA-glc-hex) in the rye-based diet. DIBOA-glc (derived from the diet and intestinal deglycosylation of DIBOA-glc-hex) was apparently reduced to 2-β-D-glucopyranosyloxy-1,4-benzoxazin-3-one (HBOA-glc), the most dominant benzoxazinoid in the blood (829 nmol/L). The benzoxazinoid compounds were excreted in the urine, with HBOA-glc (18 μmol/L) as a major metabolite. In this study, we determined for the first time the bioavailability of dietary benzoxazinoids that have high digestibility, distribution, and metabolism in mammals. These findings could be a milestone for the exploitation of healthful and pharmacological properties of benzoxazinoids.     

Initial and subsequent effects of hydrochar amendment on germination and nitrogen uptake of spring barley

The application of hydrochars as soil amendments could be an option to ameliorate soil quality by enhancing nutrient and water‐holding capacity of sandy soils. But when hydrochar application is directly followed by sowing, it can decrease the soil mineral‐N concentration and the germination rate of crops. We currently lack evidence, whether these effects are persistent or transient and thus can be avoided by adjusting the application strategy, e.g., by prolonging the period between application and sowing. A set of pot trials with spring barley (Hordeum vulgare L.) was performed to study the initial and subsequent effects of different hydrochar amendments on germination, biomass production, and plant N availability. To this end, two subsequent cultivations of barley were grown on hydrochar‐amended soil–peat substrates. The first barley cultivation was sown directly after hydrochar application. After the harvest, the same substrates were cultivated with barley again. Germination, biomass production, and N concentration in the biomass were reduced when sowing directly followed the application of hydrochars. Differences in germination rate and biomass production between hydrochar treatments depended on the hydrochar feedstock. A recultivation of the hydrochar‐amended substrates 9 weeks after hydrochar application showed no significant effects on germination and biomass production. The N concentration in plants was still lower in the hydrochar treatments despite additional fertilization. The N immobilized during the first cultivation period was not released in the following weeks. We conclude that the germination‐inhibiting substances in hydrochars were removed in the weeks after soil incorporation. This is probably due to microbial degradation of hydrochar components and a consecutive immobilization of mineral N. Hydrochar amendment had only short‐term effects on germination and biomass production of barley that can be avoided by applying hydrochar at least 4 weeks prior to sowing. The N concentration in plants was persistently low even 4 months after hydrochar application which can only be redeemed by adjusting the fertilization strategy.     

Radical surgical resection of locally invasive oro-cutaneous tumors in the buccal region of 2 horses

OBJECTIVE: To report treatment and wound healing after radical surgical mass excision in the equine buccal region. STUDY DESIGN: Clinical case reports. ANIMALS: An 11-year-old Warmblood gelding and a 9-year-old Norwegian Trotter gelding. METHODS: Large tumoral masses were removed from the buccal region by radical electrosurgery, creating large defects into the oral cavity. Wound ultimately healed by second intention. RESULTS: Buccal defects in both horses healed well by second intention without tumor recurrence. CONCLUSIONS: Locally invasive tumors involving the cheek region of the horse can be successfully treated by radical electroexcision alone, creating transmural defects up to 14 cm length x 8 cm height. CLINICAL RELEVANCE: Large transmural defects in the cheek region of the horse, formed by radical surgery can heal satisfactorily by second intention healing without undue discomfort to the patient or major complications.     

Sequence variation in the barley genes encoding sucrose synthase I and sucrose phosphate synthase II,and its association with variation in grain traits and malting quality

Sequence diversity in the two barley (Hordeum vulgare L.) genes encoding sucrose synthase I (SSI) and sucrose phosphate synthase II (SPSII), both of which are involved in sucrose accumulation and grain filling, was studied by partial resequencing of eight reference genotypes and SNP analysis by pyrosequencing in a panel of 94 spring and 96 winter European barley varieties. The resequencing was based on two adjacent SSI fragments of size 880 and 820 bp, and a 2,322 bp SPSII fragment. In the SSI gene, 26 SNPs were present in the larger fragment, and 25 in the smaller one, and 11 of these were exploited to develop high-throughput SNP assays used for haplotype analysis. An association analysis based on either a general or a mixed linear model suggested that the predominant three haplotypes influenced certain components of both kernel and malting quality. However, the level of phenotype/haplotype association shown with the SPSII gene was rather low. SNP variation of SSI was used to map the locus to chromosome 7H.     

Non-target effects of the microbial control agents Pseudomonas fluorescens DR54 and Clonostachys rosea IK726 in soils cropped with barley followed by sugar beet: a greenhouse assessment

Non-target effects of a bacterial (Pseudomonas fluorescens DR54) and a fungal (Clonostachys rosea IK726) microbial control agent (MCA), on the indigenous microbiota in bulk soil and rhizosphere of barley, and subsequent a sugar beet crop, were studied in a greenhouse experiment. MCAs were introduced by seed and soil inoculation. Bulk and rhizosphere soils were sampled regularly during the growth of barley and sugar beet. The soils were assayed for the fate of MCAs and various features of the indigenous soil microbiota. At the end of the experiment (193 d), DR54 and IK726 had declined by a factor of 10⁶ and 20, respectively, and DR54 showed a short-lasting growth increase in the sugar beet rhizosphere. In general, the non-target effects were small and transient. IK726 seemed to have general stimulating effects on soil enzyme activity and the soil microbiota, and resulted in a significant increase in plant dry weight. The plant growth-promoting effect of DR54 was less pronounced and the DR54 displaced indigenous pseudomonads. DR54 stimulated growth of protozoans with a tolerance for the anti-fungal compound viscosinamide produced by DR54. Treatment with the fungicide Fungazil had no effects on plant growth or soil microorganisms. Phospholipid fatty acid (PLFA) analysis detected the perturbations of the soil microbial community structure in the MCA treatments as well as the return to non- perturbed conditions reflecting the decline of inoculant populations. The PLFA technique appears to be suitable for in situ monitoring of MCA non-target effects on the soil microbiota, but should be combined with assays for MCA survival and soil enzyme activity.     

Hemostatic biomarkers in dogs with chronic congestive heart failure

BACKGROUND: Chronic congestive heart failure (CHF) in humans is associated with abnormal hemostasis, and abnormalities in hemostatic biomarkers carry a poor prognosis. Alterations in hemostatic pathways can be involved in the pathogenesis of CHF in dogs, and microthrombosis in the myocardium could contribute to increased mortality. HYPOTHESIS: That plasma concentration or activity of hemostatic biomarkers is altered in dogs with CHF and that these factors predict mortality. ANIMALS: Thirty-four dogs with CHF caused by either dilated cardiomyopathy (DCM, n=14) or degenerative valvular disease (CDVD, n=20) compared with 23 healthy age-matched control dogs were included in this study. Dogs with CHF were recruited from 2 referral cardiology clinics, and control dogs were owned by friends or colleagues of the investigators. METHODS: Clinical examination and echocardiography were performed in all dogs. Plasma fibrinogen and D-dimer concentrations, antithrombin and protein C activity, and thrombin-antithrombin complex (TAT) were measured in all dogs. RESULTS: Dogs with CHF had significantly higher fibrinogen (P = .04), D-dimer (P = .002), and TAT concentration (P < .0001), lower antithrombin (P < .0001) and protein C activity (P < .001) compared with control dogs. None of the hemostatic biomarkers were associated with risk of death. CONCLUSIONS AND CLINICAL IMPORTANCE: There is evidence of a procoagulant state in dogs with CHF. The lack of predictive value for survival might be due to the small number of dogs examined. Further studies are necessary to investigate the presence and importance of microthrombosis in dogs with CHF.