Trimming and Re-shoeing Results in More Steps per Day and More Time Spent Lying per Day.

To examine the impact of trimming and re-shoeing on behavior, light horse geldings (3-21 year-old Quarter Horse, Dutch Warmblood, or Thoroughbred) were fitted with three-axis accelerometers (IceTag, Ice Robotics, Edinburgh, Scotland) on the left rear limb. Boots were placed under the accelerometer, and both were removed daily for approximately 1 hour while horses were stalled for morning feeding to examine the horses' limb. After a two-day adaptation period and five days of activity tracking, horses were treated by having shoes removed, feet trimmed, and new shoes fitted (re-shod; n = 3) or being handled but not trimmed or fitted with new shoes (sham; n = 4). Horse activity was monitored for five days after treatment. Steps per day, time spent lying per day, and the number of lying bouts were tested for effects of treatment, time (before or after treatment), and interaction of treatment by time using procedures for repeated measures with JMP Software (version 7, SAS Inst. Inc, Cary, NC). Means separation was performed using the Student's t-test. There was a treatment by time interaction on time spent lying per day (P = .0447) and steps per day (P = .0501). Re-shod spent more time lying after treatment than before (121.4 ± 22.8 vs 66.8 ± 22.8 minutes per day, respectively). After treatment, re-shod also took more steps than sham (3499 ± 527 vs 3056 ± 456 steps per day, respectively). These results may indicate increased mental and physical comfort following trimming and re-shoeing in horses.

Angiopoietin-2 Inhibition Rescues Arteriovenous Malformation in a Smad4 Hereditary Hemorrhagic Telangiectasia Mouse Model.

BACKGROUND: Hereditary hemorrhagic telangiectasia is an autosomal dominant vascular disorder caused by heterozygous, loss-of-function mutations in 4 transforming growth factor beta (TGFß) pathway members, including the central transcriptional mediator of the TGFß pathway, Smad4. Loss of Smad4 causes the formation of inappropriate, fragile connections between arteries and veins called arteriovenous malformations (AVMs), which can hemorrhage leading to stroke, aneurysm, or death. Unfortunately, the molecular mechanisms underlying AVM pathogenesis remain poorly understood, and the TGFß downstream effectors responsible for hereditary hemorrhagic telangiectasia-associated AVM formation are currently unknown. METHODS: To identify potential biological targets of the TGFß pathway involved in AVM formation, we performed RNA- and chromatin immunoprecipitation-sequencing experiments on BMP9 (bone morphogenetic protein 9)-stimulated endothelial cells (ECs) and isolated ECs from a Smad4-inducible, EC-specific knockout ( Smad4-iECKO) mouse model that develops retinal AVMs. These sequencing studies identified the angiopoietin-Tek signaling pathway as a downstream target of SMAD4. We used monoclonal blocking antibodies to target a specific component in this pathway and assess its effects on AVM development. RESULTS: Sequencing studies uncovered 212 potential biological targets involved in AVM formation, including the EC surface receptor, TEK (TEK receptor tyrosine kinase) and its antagonistic ligand, ANGPT2 (angiopoietin-2). In Smad4-iECKO mice, Angpt2 expression is robustly increased, whereas Tek levels are decreased, resulting in an overall reduction in angiopoietin-Tek signaling. We provide evidence that SMAD4 directly represses Angpt2 transcription in ECs. Inhibition of ANGPT2 function in Smad4-deficient mice, either before or after AVMs form, prevents and alleviates AVM formation and normalizes vessel diameters. These rescue effects are attributed to a reversion in EC morphological changes, such as cell size and shape that are altered in the absence of Smad4. CONCLUSIONS: Our studies provide a novel mechanism whereby the loss of Smad4 causes increased Angpt2 transcription in ECs leading to AVM formation, increased blood vessel calibers, and changes in EC morphology in the retina. Blockade of ANGPT2 function in an in vivo Smad4 model of hereditary hemorrhagic telangiectasia alleviated these vascular phenotypes, further implicating ANGPT2 as an important TGFß downstream mediator of AVM formation. Therefore, alternative approaches that target ANGPT2 function may have therapeutic value for the alleviation of hereditary hemorrhagic telangiectasia symptoms, such as AVMs.

Lying Time and Its Importance to the Dairy Cow: Impact of Stocking Density and Time Budget Stresses.

The stocking density of confinement dairy operations is one of the most important management factors in determining cows' ability to achieve the 12 hours per day of lying and 3 hours to 5 hours per day feeding that cows are highly motivated to engage in. Overstocking facilities consistently decreases lying time, alters feeding behaviors, decreases rumination times, and increases social stress. This article reviews the relevant literature to establish the recommended stocking density with freestall systems. Novel housing systems and the considerations of transition cows are also reviewed.

Effects of endophyte-infected tall fescue seed and red clover isoflavones on rumen microbial populations and physiological parameters of beef cattle.

Lolium arundinaceum [(Darbyshire) tall fescue] toxicosis is responsible for substantial beef production losses in the United States, due to its negative effects on reproduction, growth, and feed efficiency. These effects are consequences of toxic alkaloids within tall fescue. Interseeding legumes, such as Trifolium pratense (red clover), into pastures has been shown to mitigate a portion of these effects. Clovers contain isoflavones, which may play a role in tall fescue toxicosis mitigation. The present study utilized 36 Angus steers to determine the effects of daily supplementation with a red clover-isolated isoflavone feed additive on physiological symptoms of tall fescue toxicosis and the rumen microbial environment over a 21-d period. Angus steers were initially stratified based upon their single nucleotide polymorphism genotype at the DRD2 receptor. Treatments were then randomly assigned in a 2 × 2 factorial arrangement within a completely randomized design, where treatment factors consisted of tall fescue seed type (endophyte-infected tall fescue seed vs. endophyte-free tall fescue seed) supplemented with and without the isoflavone additive. Steers that consumed endophyte-infected tall fescue seed had lower serum prolactin concentrations (P = 0.0007), average daily gain (ADG; P = 0.003), final body weight (BW; P = 0.004), and feed efficiency (P = 0.018) when compared with steers that consumed endophyte-free tall fescue seed. Serum insulin-like growth factor-1 (IGF-1) tended to be reduced with supplementation of isoflavones (P = 0.06) but was unaffected by seed type (P ≥ 0.10) and seed by treatment interaction (P ≥ 0.10). Isoflavones reduced serum glucose levels (P = 0.023), but neither seed type, isoflavones, or their interaction affected serum urea nitrogen (SUN), nonesterified fatty acids (NEFA), or insulin (P ≥ 0.10). Volatile fatty acid concentrations, dry matter intake (DMI), ruminal pH, and overall feeding behaviors were also unaffected by seed type or isoflavone treatments (P ≥ 0.10). Twenty-eight ruminal bacteria taxa shifted as a result of seed type or isoflavone treatment (P < 0.05). In this experiment, feeding isoflavones to Angus cattle did not completely mitigate all symptoms of fescue toxicosis. However, dose-response trials may aid future research to determine if dietary supplementation with isoflavones alleviates fescue toxicosis symptoms and promotes livestock growth and performance.

Vascular deficiency of Smad4 causes arteriovenous malformations: a mouse model of Hereditary Hemorrhagic Telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular disorder that leads to abnormal connections between arteries and veins termed arteriovenous malformations (AVM). Mutations in TGFß pathway members ALK1, ENG and SMAD4 lead to HHT. However, a Smad4 mouse model of HHT does not currently exist. We aimed to create and characterize a Smad4 endothelial cell (EC)-specific, inducible knockout mouse (Smad4f/f;Cdh5-CreERT2) that could be used to study AVM development in HHT. We found that postnatal ablation of Smad4 caused various vascular defects, including the formation of distinct AVMs in the neonate retina. Our analyses demonstrated that increased EC proliferation and size, altered mural cell coverage and distorted artery-vein gene expression are associated with Smad4 deficiency in the vasculature. Furthermore, we show that depletion of Smad4 leads to decreased Vegfr2 expression, and concurrent loss of endothelial Smad4 and Vegfr2 in vivo leads to AVM enlargement. Our work provides a new model in which to study HHT-associated phenotypes and links the TGFß and VEGF signaling pathways in AVM pathogenesis.

Cellular Uptake and Delivery of Myeloperoxidase to Lysosomes Promote Lipofuscin Degradation and Lysosomal Stress in Retinal Cells.

Neutrophil myeloperoxidase (MPO) catalyzes the H2O2-dependent oxidation of chloride anion to generate hypochlorous acid, a potent antimicrobial agent. Besides its well defined role in innate immunity, aberrant degranulation of neutrophils in several inflammatory diseases leads to redistribution of MPO to the extracellular space, where it can mediate tissue damage by promoting the oxidation of several additional substrates. Here, we demonstrate that mannose 6-phosphate receptor-mediated cellular uptake and delivery of MPO to lysosomes of retinal pigmented epithelial (RPE) cells acts to clear this harmful enzyme from the extracellular space, with lysosomal-delivered MPO exhibiting a half-life of 10 h. Lysosomal-targeted MPO exerts both cell-protective and cytotoxic functions. From a therapeutic standpoint, MPO catalyzes the in vitro degradation of N-retinylidene-N-retinylethanolamine, a toxic form of retinal lipofuscin that accumulates in RPE lysosomes and drives the pathogenesis of Stargardt macular degeneration. Furthermore, chronic cellular uptake and accumulation of MPO in lysosomes coincides with N-retinylidene-N-retinylethanolamine elimination in a cell-based model of macular degeneration. However, lysosomal-delivered MPO also disrupts lysosomal acidification in RPE cells, which coincides with nuclear translocation of the lysosomal stress-sensing transcription factor EB and, eventually, cell death. Based on these findings we predict that under periods of acute exposure, cellular uptake and lysosomal degradation of MPO mediates elimination of this harmful enzyme, whereas chronic exposure results in progressive accumulation of MPO in lysosomes. Lysosomal-accumulated MPO can be both cell-protective, by promoting the degradation of toxic retinal lipofuscin deposits, and cytotoxic, by triggering lysosomal stress and cell death.