Hours Of Ischemia Research Articles

Effects of alprostadil and iloprost on renal, lung, and skeletal muscle injury following hindlimb ischemia-reperfusion injury in rats.

ObjectivesTo evaluate the effects of alprostadil (prostaglandin [PGE1] analog) and iloprost (prostacyclin [PGI2] analog) on renal, lung, and skeletal muscle tissues after ischemia reperfusion (I/R) injury in an experimental rat model.Materials and methodsWistar albino rats underwent 2 hours of ischemia via infrarenal aorta clamping with subsequent 2 hours of reperfusion. Alprostadil and iloprost were given starting simultaneously with the reperfusion period. Effects of agents on renal, lung, and skeletal muscle (gastrocnemius) tissue specimens were examined.ResultsRenal medullary congestion, cytoplasmic swelling, and mean tubular dilatation scores were significantly lower in the alprostadil-treated group than those found in the I/R-only group (P<0.0001, P=0.015, and P<0.01, respectively). Polymorphonuclear leukocyte infiltration, pulmonary partial destruction, consolidation, alveolar edema, and hemorrhage scores were significantly lower in alprostadil- and iloprost-treated groups (P=0.017 and P=0.001; P<0.01 and P<0.0001). Polymorphonuclear leukocyte infiltration scores in skeletal muscle tissue were significantly lower in the iloprost-treated group than the scores found in the nontreated I/R group (P<0.0001).ConclusionAlprostadil and iloprost significantly reduce lung tissue I/R injury. Alprostadil has more prominent protective effects against renal I/R injury, while iloprost is superior in terms of protecting the skeletal muscle tissue against I/R injury.

Absolute Cerebral Blood Flow Infarction Threshold for 3-Hour Ischemia Time Determined with CT Perfusion and 18F-FFMZ-PET Imaging in a Porcine Model of Cerebral Ischemia.

CT Perfusion (CTP) derived cerebral blood flow (CBF) thresholds have been proposed as the optimal parameter for distinguishing the infarct core prior to reperfusion. Previous threshold-derivation studies have been limited by uncertainties introduced by infarct expansion between the acute phase of stroke and follow-up imaging, or DWI lesion reversibility. In this study a model is proposed for determining infarction CBF thresholds at 3hr ischemia time by comparing contemporaneously acquired CTP derived CBF maps to 18F-FFMZ-PET imaging, with the objective of deriving a CBF threshold for infarction after 3 hours of ischemia. Endothelin-1 (ET-1) was injected into the brain of Duroc-Cross pigs (n = 11) through a burr hole in the skull. CTP images were acquired 10 and 30 minutes post ET-1 injection and then every 30 minutes for 150 minutes. 370 MBq of 18F-FFMZ was injected ~120 minutes post ET-1 injection and PET images were acquired for 25 minutes starting ~155–180 minutes post ET-1 injection. CBF maps from each CTP acquisition were co-registered and converted into a median CBF map. The median CBF map was co-registered to blood volume maps for vessel exclusion, an average CT image for grey/white matter segmentation, and 18F-FFMZ-PET images for infarct delineation. Logistic regression and ROC analysis were performed on infarcted and non-infarcted pixel CBF values for each animal that developed infarct. Six of the eleven animals developed infarction. The mean CBF value corresponding to the optimal operating point of the ROC curves for the 6 animals was 12.6 ± 2.8 mL·min-1·100g-1 for infarction after 3 hours of ischemia. The porcine ET-1 model of cerebral ischemia is easier to implement then other large animal models of stroke, and performs similarly as long as CBF is monitored using CTP to prevent reperfusion.

Prolonged and Intermittent Bilateral Common Carotid Artery Occlusion Induces Brain Lipidome Changes in a Rat Stroke Model

Background: Recent studies suggest that prolonged and intermittent bilateral common carotid artery occlusion (prolonged bilateral common carotid artery occlusion (PO) and intermittent bilateral common carotid artery occlusion, respectively) can lead to reduced ischemia brain injury in ischemic tolerance. Objectives: In this study, we aimed to investigate the changes in brain lipidomics following prolonged and intermittent ischemic preconditioning. Materials and Methods: Two groups underwent bilateral common carotid artery occlusion either intermittently (for 3 continuous periods within a timeframe of 20 minutes, releasing 3 consecutive times, yielding a total of 9 minutes (intermittent bilateral common carotid artery occlusion)) or for 9 minutes continuously (prolonged bilateral common carotid artery occlusion). The third and fourth groups were used as control and sham (without ischemia) groups, respectively. The first three groups were subdivided into middle cerebral artery occlusion-operated (middle cerebral artery occlusion MCAO), for 60 minutes of ischemia) and intact (without any surgery) subgroups. After 1 hour of ischemia and 24 hours of reperfusion, the neurologic deficit score (neurological deficit score (NDS)) and the infarct volume (infarct volume) were assessed in the middle cerebral artery occlusion-operated subgroup. Brain lipidomics were measured in the intact subgroup and the sham group. Results: Preconditioning with prolonged bilateral common carotid artery occlusion and intermittent bilateral common carotid artery occlusion significantly decreased the neurological deficit scores and infarct volume and increased the levels of phosphatidylethanolamine, sphingomyelin, cholesterol ester, cholesterol, phosphatidylcholine, and cerebroside in the brain compared with the control and sham groups. Prolonged bilateral common carotid artery occlusion and intermittent bilateral common carotid artery occlusion significantly decreased the brain ceramide and lyso-phosphatidylcholine levels. The triglyceride levels in both groups (intermittent bilateral common carotid artery occlusion and prolonged bilateral common carotid artery occlusion) did not change in comparison with the control and sham groups. Conclusions: Although it seems that further studies are needed to clarify the real mechanisms of ischemic tolerance, ischemic preconditioning could decrease brain ischemia injury via changes in brain lipidomics.

NOX2 Activation of Natural Killer T Cells Is Blocked by the Adenosine A2A Receptor to Inhibit Lung Ischemia-Reperfusion Injury.

Ischemia-reperfusion (IR) injury after lung transplantation, which affects both short- and long-term allograft survival, involves activation of NADPH oxidase 2 (NOX2) and activation of invariant natural killer T (iNKT) cells to produce IL-17. Adenosine A2A receptor (A2AR) agonists are known to potently attenuate lung IR injury and IL-17 production. However, mechanisms for iNKT cell activation after IR and A2AR agonist-mediated protection remain unclear. We tested the hypothesis that NOX2 mediates IL-17 production by iNKT cells after IR and that A2AR agonism prevents IR injury by blocking NOX2 activation in iNKT cells. An in vivo murine hilar ligation model of IR injury was used, in which left lungs underwent 1 hour of ischemia and 2 hours of reperfusion. Adoptive transfer of iNKT cells from p47(phox-/-) or NOX2(-/-) mice to Jα18(-/-) (iNKT cell-deficient) mice significantly attenuated lung IR injury and IL-17 production. Treatment with an A2AR agonist attenuated IR injury and IL-17 production in wild-type (WT) mice and in Jα18(-/-) mice reconstituted with WT, but not A2AR(-/-), iNKT cells. Furthermore, the A2AR agonist prevented IL-17 production by murine and human iNKT cells after acute hypoxia-reoxygenation by blocking p47(phox) phosphorylation, a critical step for NOX2 activation. NOX2 plays a key role in inducing iNKT cell-mediated IL-17 production and subsequent lung injury after IR. A primary mechanism for A2AR agonist-mediated protection entails inhibition of NOX2 in iNKT cells. Therefore, agonism of A2ARs on iNKT cells may be a novel therapeutic strategy to prevent primary graft dysfunction after lung transplantation.

Mechanism of vacuum sealing drainage therapy attenuating ischemia-reperfusion injury of skeletal muscle in rabbit

To investigate the mechanism of how vacuum sealing drainage (VSD) ameliorating ischemia reperfusion (I/R) injury in skeletal muscle I/R model. Thirty New Zealand white rabbits were divided into three groups: control (sham operation) group, I/R group, VSD+ I/R group.The ischemia of the left hind limb of the animal was induced by clamping the common femoral artery and vein. After 4 hours of ischemia, the clamp was removed and the hind limp underwent 6 hours reperfusion. VSD treated animals received the treatment at the beginning of reperfusion. The concentrations of myeloperoxidase (MPO), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) in muscular tissues were assayed. HE stained pathological section was used to evaluate the degree of edema of muscular tissues, and the immunohistochemistry was used to detect the percentage of positive cells expressing high mobility group protein B1 (HMGB1). Q-RT-PCR and Western Blot were used to detect the mRNA levels and protein expression of HMGB1 in myocyte respectively. The experimental data was tested using variance analysis. The levels of inflammatory factors and antioxidant factors in muscular tissues were significantly different in the I/R group compared to the VSD group and control group (the levels of MPO in I/R group, I/R+ VSD group and control group were 0.91±0.22, 0.53±0.08, 0.31±0.10, respectively, F=26.48, P=0.000; MDA were 2.04±0.92, 1.65±1.02, 1.01±0.12, F=4.250, P=0.040; SOD were 35.97±9.23, 55.99±18.97, 61.83±14.91, F=5.240, P=0.020; CAT were 31.42±16.27, 48.50±17.86, 75.95±13.09, F=9.720, P=0.002; GSH were 1.48±0.90, 3.54±1.88, 3.84±2.08, F=5.240, P=0.020). HE staining showed an increased intercellular space ratio in the I/R group (F=16.47, P<0.05). Immunohistochemistry staining showed that percentage of HMGB1 positive myocytes in control, I/R and I/R+ VSD group are 1.94%, 18.63% and 61.36%, respectively. There was significant difference among groups (F=853.886, P<0.01). A significantly inhibited HMGB1 expression by VSD therapy was also validated by the results of Q-RT-PCR (F=50.653, P<0.01) and Western blot (F=963.489, P<0.01). The results from the present research suggest that VSD may attenuate skeletal muscles I/R injury by increasing the cellular antioxidative stress reaction and inhibiting the reactive oxygen species as well as the inflammatory mediators.

MP28-09 A NOVEL MODEL OF PELVIC ORGAN ISCHEMIA: PELVIC FLOOR CONTRACTILE FUNCTION IS MORE RESISTANT TO ISCHEMIA AND POST-ISCHEMIA REPERFUSION THAN BLADDER DETRUSOR MUSCLE

You have accessJournal of UrologyBladder & Urethra: Anatomy, Physiology & Pharmacology I1 Apr 2016MP28-09 A NOVEL MODEL OF PELVIC ORGAN ISCHEMIA: PELVIC FLOOR CONTRACTILE FUNCTION IS MORE RESISTANT TO ISCHEMIA AND POST-ISCHEMIA REPERFUSION THAN BLADDER DETRUSOR MUSCLE Amy D. Dobberfuhl, Catherine Schuler, Robert E. Leggett, Elise J.B. De, and Robert M. Levin Amy D. DobberfuhlAmy D. Dobberfuhl More articles by this author , Catherine SchulerCatherine Schuler More articles by this author , Robert E. LeggettRobert E. Leggett More articles by this author , Elise J.B. DeElise J.B. De More articles by this author , and Robert M. LevinRobert M. Levin More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.1059AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Weakness of the pelvic floor musculature (PFM) is well described in stress urinary incontinence and pelvic organ prolapse. Periods of low blood flow, hypoxia and ischemia have been implicated as causes of pelvic floor and bladder dysfunction. We explored the contractile characteristics of PFM and bladder detrusor muscle (BDM) in a novel model of in-vitro ischemia-reperfusion. METHODS Female adult virgin rabbits were selected for their well developed pelvic floor. Five animals were euthanized. BDM, pubococcygeus (PC) and coccygeous (CC) were isolated into 150 mg strips. Tissues were equilibrated with oxygenated Tyrodes containing glucose, stimulated with field stimulation, and then stimulated under ischemic conditions for 1 hour in nitrogenated Tyrodes without glucose. Tissues were then incubated in oxygenated Tyrodes for 2 hours and stimulated. RESULTS PFM required 10 times the stimulus duration to generate contractions under baseline conditions. Both maximal contractile response and rate of tension generation were significantly greater for BDM than either PC or CC. However, PC and CC were both significantly less sensitive to the effects of ischemia than BDM and continued to have stable contractile responses to field stimulation during the entire hour of ischemia (Figure 1). There was progressive decline in BDM contractile strength after 1 hour of ischemia. Following the ischemic period, PFM contractile recovery was significantly superior to BDM (Figure 2) and consistent with markers of oxidative stress. CONCLUSIONS PFM contractions and recovery were significantly slower than BDM, yet not as sensitive to either in-vitro ischemia or the effects of post-ischemia reperfusion. © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e374 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Amy D. Dobberfuhl More articles by this author Catherine Schuler More articles by this author Robert E. Leggett More articles by this author Elise J.B. De More articles by this author Robert M. Levin More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Image Analysis of Oxidative and Glycolytic Muscle Fibers During Reperfusion Injury by Segmentation Based on Regions

Different situations cause ischemia and reperfusion injury, affecting tissues under the level of compression. In this research, abnormal characteristics in distribution of muscle fibers types in soleus and extensor carpi radialis longus, during short periods of ischemia and short and long periods of reperfusion, were determined. Fibers were classified by enzyme histochemistry techniques NADH-TR and myosin-ATPase. Measurements of areas were carried out through semiautomatic image processing by using segmentation based on regions, which evidenced significant changes in distribution during reperfusion followed to one and three hours of ischemia, as well as in comparisons of areas for all periods of reperfusion. This study strengthens the evidence about using practical procedures of image analysis in the diagnosis of tisular abnormalities.

Hypothermia Promotes Survival of Ischemic Retinal Ganglion Cells.

Ischemic stroke in retinal arteries leads to death of neural tissue and ultimately to blindness. The retina is known to die within 4 hours after onset of ischemia. It is debated whether hypothermia might increase the time window for medical treatment and thereby the chance of recovering sight. In order to characterize the time course of cell death during ischemia and potential beneficial effects of hypothermia in more detail, we investigated the survival of ganglion cells in ischemic pig and human retina as a function of time and temperature. Eyes were obtained from minipigs and from human donors post mortem. Enucleated minipig eyes were stored for defined durations at three different temperatures (37 °C, 21 °C, and 4 °C). In order to assess the viability of the tissue, we measured ganglion cell activity (spiking) with multielectrode arrays. Minipig retinal ganglion cell function was severely compromised after 2 hours of ischemia at body temperature. After 4 hours, ganglion cells did not fire action potentials anymore. However, at 21 °C, ganglion cell activity was maintained under ischemic conditions for up to 12 hours, and for at least 50 hours at 4 °C. In postmortem human retina, we recorded ganglion cell activity in retinas received up to 27 hours after death. Our results demonstrate that hypothermia greatly increases survival of retinal ganglion cells exposed to ischemia. These results might be relevant for the future treatment of retinal ischemia.

Platelet-derived Growth Factor-B Protects Rat Cardiac Allografts From Ischemia-reperfusion Injury.

Microvascular dysfunction and cardiomyocyte injury are hallmarks of ischemia-reperfusion injury (IRI) after heart transplantation. Platelet-derived growth factors (PDGF) have an ambiguous role in this deleterious cascade. On one hand, PDGF may exert vascular stabilizing and antiapoptotic actions through endothelial-pericyte and endothelial-cardiomyocyte crosstalk in the heart; and on the other hand, PDGF signaling mediates neointimal formation and exacerbates chronic rejection in cardiac allografts. The balance between these potentially harmful and beneficial actions determines the final outcome of cardiac allografts. We transplanted cardiac allografts from Dark Agouti rat and Balb mouse donors to fully major histocompatibility complex-mismatched Wistar Furth rat or C57 mouse recipients with a clinically relevant 2-hour cold ischemia and 1-hour warm ischemia. Ex vivo intracoronary delivery of adenovirus-mediated gene transfer of recombinant human PDGF-BB upregulated messenger RNA expression of anti-mesenchymal transition and survival factors BMP-7 and Bcl-2 and preserved capillary density in rat cardiac allografts at day 10. In mouse cardiac allografts PDGF receptor-β, but not -α intragraft messenger RNA levels were reduced and capillary protein localization was lost during IRI. The PDGF receptor tyrosine kinase inhibitor imatinib mesylate and a monoclonal antibody against PDGF receptor-α enhanced myocardial damage evidenced by serum cardiac troponin T release in the rat and mouse cardiac allografts 6 hours after reperfusion, respectively. Moreover, imatinib mesylate enhanced rat cardiac allograft vasculopathy, cardiac fibrosis, and late allograft loss at day 56. Our results suggest that PDGF-B signaling may play a role in endothelial and cardiomyocyte recovery from IRI after heart transplantation.

What is the relationship among penumbra volume, collaterals, and time since onset in the first 6 h after acute ischemic stroke?

The steep, time-dependent loss of benefit from reperfusion in clinical trials is consistent with loss of penumbra over the early hours of ischemia, as observed in animal models. Human imaging studies, however, show persistent penumbra for up to 48 h. We investigated core and penumbra volumes and collateral status in relation to time after stroke onset within the first 6 h. Using data from three multimodal computer tomography-based studies in acute ischemic stroke patients <6 h after onset, we measured core and penumbra volumes, collateral status, and target mismatch (defined as core volume < 50 ml, perfusion lesion volume > 15 ml, mismatch ratio > 1.8). Patients were grouped by onset to imaging time (<3, 3-4.5, 4.5-6 h). We explored correlates of penumbra proportion by multivariable linear regression. Analysis included 144 subjects. Across time epochs, neither proportions of penumbra (59%, 64%, 75% at <3, 3-4.5, >4. 5 h, respectively, p = 0.4) nor poor collaterals (15/56 (27%), 14/47 (30%), 4/15 (27%) at <3, 3-4.5, >4.5 h, p = 0.9) differed significantly. Penumbra proportion was not clearly related to time to imaging (R(2) = 0.003; p = 0.5) but a trend for divergent effects by collateral status was seen (slight increase in penumbra over time with good collaterals versus reduced with poor, interaction = 0.08). The proportion of patients with target mismatch did not vary by time (56%, 74%, and 67% at <3, 3-4.5, >4.5 h, p = 0.09). In a cross-sectional sample imaged within 6 h, neither the proportions of penumbral tissue nor "target mismatch" varied by time from onset. A trend for reducing penumbra proportion only among those with poor collaterals may have pathophysiological and therapeutic importance.

Effects of alpha lipoic acid on ishemia/reperfusion injury in rat hindlimb ischemia model

This study was performed to evaluate the effect of alpha lipoic acid (ALA) on prevention of ischemia-reperfusion (IR) injury in rat hindlimb ischemia model. Forty male Sprague Dawley rats weighing between 250 and 300 g were divided into 4 groups of 10 rats. Hindlimb composite island flaps were raised in all rats. Clamps were applied to femoral vessels of all subjects, but immediately released without causing ischemia in Group 1. In Group 2, after 4 hours of ischemia, 24 hours of reperfusion was performed. Following 4 hours of ischemia, saline was administered to rats in Group 3 and flaps were reperfused for 24 hours. In Group 4, ALA was administered intraperitoneally after 4 hours of ischemia and flaps were reperfused for 24 hours. In Group 4, there was a significant decrease of liver malondialdehyde compared to Group 2 and decrease of muscle tumor necrosis factor-alpha compared to Group 3. There was also increase in levels of glutathione in erythrocytes compared to Group 3 and increase of plasma vitamin C compared to all groups. ALA was found to be effective in prevention of ischemia-reperfusion injury. Further studies are needed before clinical application.

Ischemic postconditioning protects against ischemic brain injury by up-regulation of acid-sensing ion channel 2a.

Ischemic postconditioning renders brain tissue tolerant to brain ischemia, thereby alleviating ischemic brain injury. However, the exact mechanism of action is still unclear. In this study, a rat model of global brain ischemia was subjected to ischemic postconditioning treatment using the vessel occlusion method. After 2 hours of ischemia, the bilateral common carotid arteries were blocked immediately for 10 seconds and then perfused for 10 seconds. This procedure was repeated six times. Ischemic postconditioning was found to mitigate hippocampal CA1 neuronal damage in rats with brain ischemia, and up-regulate acid-sensing ion channel 2a expression at the mRNA and protein level. These findings suggest that ischemic postconditioning up-regulates acid-sensing ion channel 2a expression in the rat hippocampus after global brain ischemia, which promotes neuronal tolerance to ischemic brain injury.

Can intravoxel incoherent motion diffusion-weighted imaging characterize the cellular injury and microcirculation alteration in hepatic ischemia-reperfusion injury? An animal study.

To investigate whether intravoxel incoherent motion diffusion-weighted imaging (IVIM-DWI) can be used to quantitatively analyze the cellular injury and microcirculation alterations in hepatic ischemia-reperfusion injury (HIRI). Thirty-two New Zealand white rabbits were randomly and equally assigned to the sham group, 1-hour, 4-hour, and 12-hour groups according to the reperfusion time after 1 hour of ischemia using a 70% liver ischemia-reperfusion injury model. All the animals underwent IVIM-DWI with 12 b values at 1.5T. The imaging parameters (IVIM parameters and apparent diffusion coefficient [ADC]) among different groups were compared. The correlations between imaging parameters and histological scores, and the ratio of serum aspartate aminotransferase to serum alanine aminotransferase (serum AST/ALT) were analyzed. During the first hour of HIRI, true diffusion coefficient (D) and ADC significantly decreased (P < 0.05), while there was no significant decrease in perfusion fraction (f) (P = 0.708). There was fair to good correlation between histological scores and f (rs = -0.493 with the sham cases excluded, and -0.682 with all cases, both P < 0.05) and ADC (rs = -0.479 with the sham cases excluded, and -0.766 with all cases, both P < 0.05). There was no correlation between imaging parameters and serum AST/ALT with the sham cases excluded (P = 0.673 for f, 0.568 for D, 0.403 for ADC), and good correlation between D, ADC, and serum AST/ALT (r = 0.747 and 0.748, both P < 0.001) with all cases. IVIM-DWI can quantitatively characterize an animal model of HIRI, with D and ADC sensitive in early detection of cellular injury, as well as fair to good correlation between f, ADC, and microcirculation alteration. J. Magn. Reson. Imaging 2016;43:1327-1336.

Post-conditioning with Cyclosporine A after a 24-hour cold ischemia in ex vivo reperfused pig lungs

ABSTRACTObjective: To evaluate the effects of 1 and 5 μM of Cyclosporine A (CsA), administered 24 hours after a cold ischemic period, in an ex vivo reperfused pig lung model. Methods: The experiments were performed in 15 pigs. Each pair of lungs was surgically separated. Extracorporeal perfusion and mechanical ventilation were started after a cold ischemia of 2 hours for one lung and 24 hours for the contralateral. We constituted three groups (n = 5 each): two groups for which the lung underwent a 24-hour ischemia received either 1 or 5 μM of CsA at the time of reperfusion, and a control group without CsA. For each group, lungs undergoing a 2-hour ischemia did not receive CsA. Results: Reperfusion with either CsA increased the PO2 levels in a dose dependent manner, and reduced concentrations of the receptor for advanced glycation endproducts, compared to the control. The pulmonary arterial pressure, the capillary pressure, and the pulmonary vascular resistances were not increased, even with 5 μM of CsA. No significant change was shown on cytokines levels. Discussion: Postconditioning with CsA improves lung function, after a 24-hour cold ischemic period. Either 1 or 5 μM seemed to be safe regarding the pulmonary vascular pressures and resistances.

Matrix Metalloproteinase-12 Induces Blood-Brain Barrier Damage After Focal Cerebral Ischemia.

Matrix metalloproteinases (MMPs) have a central role in compromising the integrity of the blood-brain barrier (BBB). The role of MMP-12 in brain damage after ischemic stroke remains unknown. The main objective of the current study is to investigate the effect of MMP-12 suppression at an early time point before reperfusion on the BBB damage in rats. Sprague-Dawley rats were subjected to middle cerebral artery occlusion and reperfusion. MMP-12 shRNA-expressing plasmids formulated as nanoparticles were administered at a dose of 1 mg/kg body weight. The involvement of MMP-12 on BBB damage was assessed by performing various techniques, including Evans blue dye extravasation, 2,3,5-triphenyltetrazolium chloride staining, immunoblot, gelatin zymography, and immunofluorescence analysis. MMP-12 is upregulated ≈31-, 47-, and 66-fold in rats subjected 1-, 2-, or 4-hour ischemia, respectively, followed by 1-day reperfusion. MMP-12 suppression protected the BBB integrity by inhibiting the degradation of tight-junction proteins. Either intravenous or intra-arterial delivery of MMP-12 shRNA-expressing plasmid significantly reduced the percent Evans blue dye extravasation and infarct size. Furthermore, MMP-12 suppression reduced the endogenous levels of other proteases, such as tissue-type plasminogen activator and MMP-9, which are also known to be the key players involved in BBB damage. These results demonstrate the adverse role of MMP-12 in acute brain damage that occurs after ischemic stroke and, thereby, suggesting that MMP-12 suppression could be a promising therapeutic target for cerebral ischemia.

Early Growth Response-1 Plays an Important Role in Ischemia-Reperfusion Injury in Lung Transplants by Regulating Polymorphonuclear Neutrophil Infiltration.

Early growth response-1 (Egr-1) has been shown to be a trigger-switch transcription factor that is involved in lung ischemia-reperfusion injury (IRI). Mouse lung transplants were performed in wild-type (WT) C57BL/6 and Egr1-knockout (KO) mice in the following donor → recipient combinations: WT → WT, KO → WT, WT → KO, and KO → KO to determine whether the presence of Egr-1 in the donor or recipient is the most critical factor for IRI. Pulmonary grafts were retrieved after 18 hours of ischemia after 4 hours of reperfusion. We analyzed graft function by analyzing arterial blood gas and histology in each combination and assessed the effects of Egr1 depletion on inflammatory cytokines that are regulated by Egr-1 as well on polymorphonuclear neutrophil (PMN) infiltration. Deletion of Egr1 improved pulmonary graft function in the following order of donor → recipient combinations: WT → WT < WT → KO < KO → WT < KO → KO. Polymerase chain reaction assays for Il1B, Il6, Mcp1, Mip2, Icam1, and Cox2 showed significantly lower expression levels in the KO → KO group than in the other groups. Immunohistochemistry demonstrated clear Egr-1 expression in the nuclei of pulmonary artery endothelial cells and PMN cytoplasm in the WT grafts. Flow cytometry analysis showed that Egr1 deletion reduced PMN infiltration and that the extent of reduction correlated with graft function. Both graft and recipient Egr-1 played a role in lung IRI, but the graft side contributed more to this phenomenon through regulation of PMN infiltration. Donor Egr-1 expression in pulmonary artery endothelial cells may play an important role in PMN infiltration, which results in IRI after lung transplantation.

Progressively heterogeneous mismatch of regional oxygen delivery to consumption during graded coronary stenosis in pig left ventricle.

In normal hearts, myocardial perfusion is fairly well matched to regional metabolic demand, although both are distributed heterogeneously. Nonuniform regional metabolic vulnerability during coronary stenosis would help to explain nonuniform necrosis during myocardial infarction. In the present study, we investigated whether metabolism-perfusion correlation diminishes during coronary stenosis, indicating increasing mismatch of regional oxygen supply to demand. Thirty anesthetized male pigs were studied: controls without coronary stenosis (n = 11); group I, left anterior descending (LAD) coronary stenosis leading to coronary perfusion pressure reduction to 70 mmHg (n = 6); group II, stenosis with perfusion pressure of about 35 mmHg (n = 6); and group III, stenosis with perfusion pressure of 45 mmHg combined with adenosine infusion (n = 7). [2-(13)C]- and [1,2-(13)C]acetate infusion was used to calculate regional O2 consumption from glutamate NMR spectra measured for multiple tissue samples of about 100 mg dry mass in the LAD region. Blood flow was measured with microspheres in the same regions. In control hearts without stenosis, regional oxygen extraction did not correlate with basal blood flow. Average myocardial O2 delivery and consumption decreased during coronary stenosis, but vasodilation with adenosine counteracted this. Regional oxygen extraction was on average decreased during stenosis, suggesting adaptation of metabolism to lower oxygen supply after half an hour of ischemia. Whereas regional O2 delivery correlated with O2 consumption in controls, this relation was progressively lost with graded coronary hypotension but partially reestablished by adenosine infusion. Therefore, coronary stenosis leads to heterogeneous metabolic stress indicated by decreasing regional O2 supply to demand matching in myocardium during partial coronary obstruction.

Dictionary-Driven Ischemia Detection From Cardiac Phase-Resolved Myocardial BOLD MRI at Rest.

Cardiac Phase-resolved Blood-Oxygen-Level Dependent (CP-BOLD) MRI provides a unique opportunity to image an ongoing ischemia at rest. However, it requires post-processing to evaluate the extent of ischemia. To address this, here we propose an unsupervised ischemia detection (UID) method which relies on the inherent spatio-temporal correlation between oxygenation and wall motion to formalize a joint learning and detection problem based on dictionary decomposition. Considering input data of a single subject, it treats ischemia as an anomaly and iteratively learns dictionaries to represent only normal observations (corresponding to myocardial territories remote to ischemia). Anomaly detection is based on a modified version of One-class Support Vector Machines (OCSVM) to regulate directly the margins by incorporating the dictionary-based representation errors. A measure of ischemic extent (IE) is estimated, reflecting the relative portion of the myocardium affected by ischemia. For visualization purposes an ischemia likelihood map is created by estimating posterior probabilities from the OCSVM outputs, thus obtaining how likely the classification is correct. UID is evaluated on synthetic data and in a 2D CP-BOLD data set from a canine experimental model emulating acute coronary syndromes. Comparing early ischemic territories identified with UID against infarct territories (after several hours of ischemia), we find that IE, as measured by UID, is highly correlated (Pearson's r=0.84) with respect to infarct size. When advances in automated registration and segmentation of CP-BOLD images and full coverage 3D acquisitions become available, we hope that this method can enable pixel-level assessment of ischemia with this truly non-invasive imaging technique.

Metabolism-Guided Bowel Resection: Potential Role and Accuracy of Instant Capillary Lactates to Identify the Optimal Resection Site.

Strip-based handheld devices can measure lactatemia on capillary blood obtained by needle puncturing. We aimed to assess the kinetic of bowel capillary lactates, metabolomics profiling, and mitochondria respiratory rate in a prolonged model of bowel hypoperfusion. In 6 pigs, a 3- to 4-cm ischemic segment was created in 6 small bowel loops (total = 36 loops) by clamping the vascular supply, for a duration of 1 to 6 hours. Hourly, 5 blood samples were obtained by puncturing the serosa, and lactates were measured using a handheld analyzer. Samples were made at the following regions of interest (ROIs): center of the ischemic area (1), proximal and distal clinical margins of resection (2a-2b), and vascularized zones (3a-3b). Every hour, surgical biopsies of ROIs were sampled. Activity of bowel mitochondria complexes was measured after 1, 3, and 5 hours of ischemia. Quantification of metabolites was performed on all samples (total N = 180). Capillary lactates were significantly higher at ROI 1 versus ROI 3ab at all time points. After 1 hour lactates at the margins were significantly higher than those at vascularized areas (P = .0095), showing a mismatch between visual assessment and actual perfusion status. From 2 to 6 hours, there was no difference in lactates between ROIs 2a-2b and 3a-3b. Maximal tissue respiration decreased significantly after 1 hour (ROI 1 vs ROI 3ab). Seven metabolites (lactate, glucose, aspartate, choline, creatine, taurine, and tyrosine) expressed significantly different evolutions between ROIs. Capillary lactates could help precisely estimate local bowel perfusion status.

Machine perfusion enhances hepatocyte isolation yields from ischemic livers

BackgroundHigh-quality human hepatocytes form the basis of drug safety and efficacy tests, cell-based therapies, and bridge-to-transplantation devices. Presently the only supply of cells derives from an inadequate pool of suboptimal disqualified donor livers. Here we evaluated whether machine perfusion could ameliorate ischemic injury that many of these livers experience prior to hepatocyte isolation. MethodsNon-heparinized female Lewis rat livers were exposed to an hour of warm ischemia (34°C) and then perfused for 3h. Five different perfusion conditions that utilized the cell isolation apparatus were investigated, namely: (1) modified Williams Medium E and (2) Lifor, both with active oxygenation (95%O2/5%CO2), as well as (3) Lifor passively oxygenated with ambient air (21%O2/0.04%CO2), all at ambient temperatures (20±2°C). At hypothermic temperatures (5±1°C) and under passive oxygenation were (4) University of Wisconsin solution (UW) and (5) Vasosol. Negative and positive control groups comprised livers that had ischemia (WI) and livers that did not (Fresh) prior to cell isolation, respectively. ResultsFresh livers yielded 32±9million cells/g liver while an hour of ischemia reduced the cell yield to 1.6±0.6million cells/g liver. Oxygenated Williams Medium E and Lifor recovered yields of 39±11 and 31±2.3million cells/g liver, respectively. The passively oxygenated groups produced 15±7 (Lifor), 13±7 (Vasosol), and 10±6 (UW)million cells/g liver. Oxygenated Williams Medium E was most effective at sustaining pH values, avoiding the accumulation of lactate, minimizing edematous weight gain and producing bile during perfusion. ConclusionsMachine perfusion results in a dramatic increase in cell yields from livers that have had up to an hour of warm ischemia, but perfusate choice significantly impacts the extent of recovery. Oxygenated Williams Medium E at room temperature is superior to Lifor, UW and Vasosol, largely facilitated by its high oxygen content and low viscosity.