Prednisolone as preservation additive prevents from ischemia reperfusion injury in a rat model of orthotopic lung transplantation

The lung is, more than other solid organs, susceptible for ischemia reperfusion injury after orthotopic transplantation. Corticosteroids are known to potently suppress pro-inflammatory processes when given in the post-operative setting or during rejection episodes. Whereas their use has been approved for these clinical indications, there is no study investigating its potential as a preservation additive in preventing vascular damage already in the phase of ischemia. To investigate these effects we performed orthotopic lung transplantations (LTX) in the rat. Prednisolone was either added to the perfusion solution for lung preservation or omitted and rats were followed for 48 hours after LTX. Prednisolone preconditioning significantly increased survival and diminished reperfusion edema. Hypoxia induced vasoactive cytokines such as VEGF were reduced. Markers of leukocyte invasiveness like matrix metalloprotease (MMP)-2, or common pro-inflammatory molecules like the CXCR4 receptor or the chemokine (C-C motif) ligand (CCL)-2 were downregulated by prednisolone. Neutrophil recruitment to the grafts was only increased in Perfadex treated lungs. Together with this, prednisolone treated animals displayed significantly reduced lung protein levels of neutrophil chemoattractants like CINC-1, CINC-2α/β and LIX and upregulated tissue inhibitor of matrix metalloproteinase (TIMP)-1. Interestingly, lung macrophage invasion was increased in both, Perfadex and prednisolone treated grafts, as measured by MMP-12 or RM4. Markers of anti-inflammatory macrophage transdifferentiation like MRC-1, IL-13, IL-4 and CD163, significantly correlated with prednisolone treatment. These observations lead to the conclusion that prednisolone as an additive to the perfusion solution protects from hypoxia triggered danger signals already in the phase of ischemia and thus reduces graft edema in the phase of reperfusion. Additionally, prednisolone preconditioning might also lead to macrophage polarization as a beneficial long-term effect

Recent advances in understanding liver fibrosis: bridging basic science and individualized treatment concepts [version 1; referees: 2 approved]

Hepatic fibrosis is characterized by the formation and deposition of excess fibrous connective tissue, leading to progressive architectural tissue remodeling. Irrespective of the underlying noxious trigger, tissue damage induces an inflammatory response involving the local vascular system and the immune system and a systemic mobilization of endocrine and neurological mediators, ultimately leading to the activation of matrix-producing cell populations. Genetic disorders, chronic viral infection, alcohol abuse, autoimmune attacks, metabolic disorders, cholestasis, alterations in bile acid composition or concentration, venous obstruction, and parasite infections are well-established factors that predispose one to hepatic fibrosis. In addition, excess fat and other lipotoxic mediators provoking endoplasmic reticulum stress, alteration of mitochondrial function, oxidative stress, and modifications in the microbiota are associated with non-alcoholic fatty liver disease and, subsequently, the initiation and progression of hepatic fibrosis. Multidisciplinary panels of experts have developed practice guidelines, including recommendations of preferred therapeutic approaches to a specific cause of hepatic disease, stage of fibrosis, or occurring co-morbidities associated with ongoing loss of hepatic function. Here, we summarize the factors leading to liver fibrosis and the current concepts in anti-fibrotic therapies

Antinociceptive effects of epidural magnesium sulphate alone and in combination with morphine in dogs

OBJECTIVE : To compare the antinociceptive effects of magnesium sulphate (MgSO4) when administered epidurally alone and in combination with morphine. STUDY DESIGN : Experimental, randomized, ‘blinded’, crossover study. ANIMALS : Six healthy adult Beagle dogs. METHODS : Evaluated treatments were MgSO4 (2.5 mg kg 1) alone (Mg), morphine (0.1 mg kg 1) alone (Mo), MgSO4 in combination with morphine (Mm), and sterile water (0.115 mL kg 1; Co) that were injected in the lumbosacral epidural space using an epidural catheter. Antinociception was measured using the von Frey mechanical threshold device applied to the carpal pads, both sides of the thorax and metatarsi. Measurements were obtained at time points: before treatment (baseline) and 0.5, 1, 2, 4, 6, 12, 18 and 24 hours after the epidural injection. Sedation, behaviour score and presence of motor deficits were assessed. Data were analyzed using a linear mixed model and Bonferroni adjustments, with significance set at p < 0.05. RESULTS : There were significant effects of treatment and time in all regions. Overall threshold values in grammes force [median (interquartile range)] when stimulation regions were combined were significantly higher in Mg [164 (135–200)], Mo 156 (129–195)] and Mm [158 (131–192)] compared to Co [145 (120–179)]. Thresholds were significantly higher compared to Co in Mg, Mo and Mm at the thorax and metatarsi, but only in Mg and Mo at the carpal pads. No motor deficits were observed at any time point. Thresholds (combined regions) were increased from baseline at one or more time points with all treatments, including control. CONCLUSION AND CLINICAL RELEVANCE : Epidural MgSO4 produced an antinociceptive effect characterised by an increase in the mechanical thresholds of similar magnitude to that produced by epidural morphine, compared with the control group, without causing any motor deficits. No potentiation of morphine antinociception was observed. The onset and offset times of antinociception could not be clearly established. To what extent these results can be extrapolated to clinical cases requires further investigation.Department of Companion Animal Clinical Studies of the University of Pretoria and the University of Pretoria Research Fund.http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1467-29952016-05-31hb201

Aspartoacylase-LacZ Knockin Mice: An Engineered Model of Canavan Disease

Canavan Disease (CD) is a recessive leukodystrophy caused by loss of function mutations in the gene encoding aspartoacylase (ASPA), an oligodendrocyte-enriched enzyme that hydrolyses N-acetylaspartate (NAA) to acetate and aspartate. The neurological phenotypes of different rodent models of CD vary considerably. Here we report on a novel targeted aspa mouse mutant expressing the bacterial β-Galactosidase (lacZ) gene under the control of the aspa regulatory elements. X-Gal staining in known ASPA expression domains confirms the integrity of the modified locus in heterozygous aspa lacZ-knockin (aspalacZ/+) mice. In addition, abundant ASPA expression was detected in Schwann cells. Homozygous (aspalacZ/lacZ) mutants are ASPA-deficient, show CD-like histopathology and moderate neurological impairment with behavioural deficits that are more pronounced in aspalacZ/lacZ males than females. Non-invasive ultrahigh field proton magnetic resonance spectroscopy revealed increased levels of NAA, myo-inositol and taurine in the aspalacZ/lacZ brain. Spongy degeneration was prominent in hippocampus, thalamus, brain stem, and cerebellum, whereas white matter of optic nerve and corpus callosum was spared. Intracellular vacuolisation in astrocytes coincides with axonal swellings in cerebellum and brain stem of aspalacZ/lacZ mutants indicating that astroglia may act as an osmolyte buffer in the aspa-deficient CNS. In summary, the aspalacZ mouse is an accurate model of CD and an important tool to identify novel aspects of its complex pathology

Deguelin Attenuates Reperfusion Injury and Improves Outcome after Orthotopic Lung Transplantation in the Rat

The main goal of adequate organ preservation is to avoid further cellular metabolism during the phase of ischemia. However, modern preservation solutions do rarely achieve this target. In donor organs hypoxia and ischemia induce a broad spectrum of pathologic molecular mechanisms favoring primary graft dysfunction (PGD) after transplantation. Increased hypoxia-induced transcriptional activity leads to increased vascular permeability which in turn is the soil of a reperfusion edema and the enhancement of a pro-inflammatory response in the graft after reperfusion. We hypothesize that inhibition of the respiration chain in mitochondria and thus inhibition of the hypoxia induced mechanisms might reduce reperfusion edema and consecutively improve survival in vivo. In this study we demonstrate that the rotenoid Deguelin reduces the expression of hypoxia induced target genes, and especially VEGF-A, dose-dependently in hypoxic human lung derived cells. Furthermore, Deguelin significantly suppresses the mRNA expression of the HIF target genes VEGF-A, the pro-inflammatory CXCR4 and ICAM-1 in ischemic lungs vs. control lungs. After lung transplantation, the VEGF-A induced reperfusion-edema is significantly lower in Deguelin-treated animals than in controls. Deguelin-treated rats exhibit a significantly increased survival-rate after transplantation. Additionally, a downregulation of the pro-inflammatory molecules ICAM-1 and CXCR4 and an increase in the recruitment of immunomodulatory monocytes (CD163+ and CD68+) to the transplanted organ involving the IL4 pathway was observed. Therefore, we conclude that ischemic periods preceding reperfusion are mainly responsible for the increased vascular permeability via upregulation of VEGF. Together with this, the resulting endothelial dysfunction also enhances inflammation and consequently lung dysfunction. Deguelin significantly decreases a VEGF-A induced reperfusion edema, induces the recruitment of immunomodulatory monocytes and thus improves organ function and survival after lung transplantation by interfering with hypoxia induced signaling

Repeated anaesthesia with isoflurane and medetomidine-midazolam-fentanyl in guinea pigs and its influence on physiological parameters.

Repeated anaesthesia may be required in experimental protocols and in daily veterinary practice, but anaesthesia is known to alter physiological parameters in GPs (Cavia porcellus, GPs). This study investigated the effects of repeated anaesthesia with either medetomidine-midazolam-fentanyl (MMF) or isoflurane (Iso) on physiological parameters in the GP. Twelve GPs were repeatedly administered with MMF or Iso in two anaesthesia sets. One set consisted of six 40-min anaesthesias, performed over 3 weeks (2 per week); the anaesthetic used first was randomized. Prior to Iso anaesthesia, atropine was injected. MMF anaesthesia was antagonized with AFN (atipamezole-flumazenil-naloxone). Abdominally implanted radio-telemetry devices recorded the mean arterial blood pressure (MAP), heart rate (HR) and core body temperature continuously. Additionally, respiratory rate, blood glucose and body weight were assessed. An operable state could be achieved and maintained for 40 min in all GPs. During the surgical tolerance with MMF, the GPs showed a large MAP range between the individuals. In the MMF wake- up phase, the time was shortened until the righting reflex (RR) returned and that occurred at lower MAP and HR values. Repeated Iso anaesthesia led to an increasing HR during induction (anaesthesias 2-6), non-surgical tolerance (anaesthesias 3-6) and surgical tolerance (anaesthesias 4, 6). Both anaesthetics may be used repeatedly, as repeating the anaesthesias resulted in only slightly different physiological parameters, compared to those seen with single anaesthesias. The regular atropine premedication induced HR increases and repeated MMF anaesthesia resulted in a metabolism increase which led to the faster return of RR. Nevertheless, Iso's anaesthesia effects of strong respiratory depression and severe hypotension remained. Based on this increased anaesthesia risk with Iso, MMF anaesthesia is preferable for repeated use in GPs