Quercetin postconditioning attenuates myocardial ischemia/reperfusion injury in rats through the PI3K/Akt pathway

Quercetin (Que), a plant-derived flavonoid, has multiple benefical actions on the cardiovascular system. The current study investigated whether Que postconditioning has any protective effects on myocardial ischemia/reperfusion (I/R) injury in vivo and its potential cardioprotective mechanisms. Male Sprague-Dawley rats were randomly allocated to 5 groups (20 animals/group): sham, I/R, Que postconditioning, Que+LY294002 [a phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway inhibitor], and LY294002+I/R. I/R was produced by 30-min coronary occlusion followed by 2-h reperfusion. At the end of reperfusion, myocardial infarct size and biochemical changes were compared. Apoptosis was evaluated by both TUNEL staining and measurement of activated caspase-3 immunoreactivity. The phosphorylation of Akt and protein expression of Bcl-2 and Bax were determined by Western blotting. Que postconditioning significantly reduced infarct size and serum levels of creatine kinase and lactate dehydrogenase compared with the I/R group (all P<0.05). Apoptotic cardiomyocytes and caspase-3 immunoreactivity were also suppressed in the Que postconditioning group compared with the I/R group (both P<0.05). Akt phosphorylation and Bcl-2 expression increased after Que postconditioning, but Bax expression decreased. These effects were inhibited by LY294002. The data indicate that Que postconditioning can induce cardioprotection by activating the PI3K/Akt signaling pathway and modulating the expression of Bcl-2 and Bax proteins.

Diacylglycerol-induced protection against injury during ischemia and reperfusion can be achieved without activation of protein kinase C in the rat heart: Comparative studies with ischemic preconditioning

The role of protein kinase C (PKC) activation in ischemic preconditioning remains controversial. Since diacylglycerol is the endogenous activator of PKC and as such might be expected cardioprotective, we have investigated whether: (i) the diacylglycerol analog 1,2-dioctanoyl-sn-glycerol (DOG) can protect against injury during ischemia and reperfusion; (ii) any effect is mediated via PKC activation; and (iii) the outcome is influenced by the time of administration. Isolated rat hearts were perfused with buffer at 37°C and paced at 400 bpm. In Study 1, hearts (n=6/group) were subjected to one of the following: (1) 36 min aerobic perfusion (controls); (2) 20 min aerobic perfusion plus ischemic preconditioning (3 min ischemia/3 min reperfusion+5 min ischemia/5 min reperfusion); (3) aerobic perfusion with buffer containing DOG (10 μM) given as a substitute for ischemic preconditioning; (4) aerobic perfusion with DOG (10 μM) during the last 2 min of aerobic perfusion. All hearts then were subjected to 35 min of global ischemia and 40 min reperfusion. A further group (5) were perfused with DOG (10 μM) for the first 2 min of reperfusion. Ischemic preconditioning improved postischemic recovery of LVDP from 24±3% in controls to 71±2% (P<0.05). Recovery of LVDP also was enhanced by DOG when given just before ischemia (54±4%), however, DOG had no effect on the recovery of LVDP when used as a substitute for ischemic preconditioning (22±5%) or when given during reperfusion (29±6%). In Study 2, the first four groups of study were repeated (n=4–5/group) without imposing the periods of ischemia and reperfusion, instead hearts were taken for the measurement of PKC activity (pmol/min/mg protein±SEM). PKC activity after 36 min in groups (1), (2), (3) and (4) was: 332±102, 299±63, 521±144, and 340±113 and the membrane:cytosolic PKC activity ratio was: 5.6±1.5, 5.3±1.8, 6.6±2.7, and 3.9±2.1 (P=NS in each instance). In conclusion, DOG is cardioprotective but under the conditions of the present study is less cardioprotective than ischemic preconditioning, furthermore the protection does not appear to necessitate PKC activation prior to ischemia.

Effects of contrast material on computed tomographic measurements of lung volumes in patients with acute lung injury

Background. Intravenous injection of contrast material is routinely performed in order to differentiate nonaerated lung parenchyma from pleural effusion in critically ill patients undergoing thoracic computed tomography (CT). The aim of the present study was to evaluate the effects of contrast material on CT measurement of lung volumes in 14 patients with acute lung injury. Method. A spiral thoracic CT scan, consisting of contiguous axial sections of 10 mm thickness, was performed from the apex to the diaphragm at end-expiration both before and 30 s (group 1; n=7) or 15 min (group 2; n=7) after injection of 80 ml contrast material. Volumes of gas and tissue, and volumic distribution of CT attenuations were measured before and after injection using specially designed software (Lungview®; Institut National des Télécommunications, Evry, France). The maximal artifactual increase in lung tissue resulting from a hypothetical leakage within the lung of the 80 ml contrast material was calculated. Results. Injection of contrast material significantly increased the apparent volume of lung tissue by 83 ± 57 ml in group 1 and 102 ± 80 ml in group 2, whereas the corresponding maximal artifactual increases in lung tissue were 42 ± 52 ml and 31 ± 18 ml. Conclusion. Because systematic injection of contrast material increases the amount of extravascular lung water in patients with acute lung injury, it seems prudent to avoid this procedure in critically ill patients undergoing a thoracic CT scan and to reserve its use for specific indications.

False positivity of gamma-glutamyl transpeptidase measurement in urine

Although enzymuria tends to be associated to renal injury, there are no studies that have evaluated the presence of the enzyme gamma-glutamyl transpeptidase (GGT) spectrophotometry in the urine using a non-nephrotoxic agent (Nerium oleander) in order to evaluate the possibility of false positive results. The urinary GGT/urinary creatinine concentration ratio (uGGT/uCr) of 10 healthy dogs was calculated and posteriorly confronted with data from clinical evaluation, hematological and serum biochemical profiles, creatinine clearance (CrC), urinalysis, urine protein/creatinine ratio (UPC), electrocardiogram, systemic blood pressure (SBP) and light and electron microscopy. The results for kidney histology, SBP, UPC and CrC were not significantly different in any of the time-points analyzed. However, uGGT/uCr was significantly higher when measured 4 hours and 24 hours after administration of N. oleander. The measurement of the urinary GGT enzyme, as performed in many studies, yielded false positive results in dogs poisoned by a non-nephrotoxic agent.

Nitric oxide is responsible for oxidative skin injury and modulation of cell proliferation after 24 hours of UVB exposures

Nitric oxide (NO) is produced by various mammalian cells and plays a variety of regulatory roles in normal physiology and in pathological processes. This article provides evidence regarding the participation of NO in UVB-induced skin lesions and in the modulation of skin cell proliferation following UVB skin irradiation. Hairless mice were subjected to UVB irradiation for 3 hours and the skin evaluated immediately, 6 and 24 hours postirradiation. The skin lipid peroxidation, and NO levels evaluated by chemiluminescence and inducible nitric oxide synthase (iNOS) and nitrotyrosine immunolabelling increased significantly 24 hours after irradiation and decreased under the treatment with aminoguanidine (AG). On the other hand, cell proliferation markers, PCNA and VEGF showed a strong labelling index when AG was used. The data indicate that NO mediates, at least in part, the lipid peroxidation and protein nitration and also promotes the down regulation of factors involved in cell proliferation. This work shows that the NO plays an important role in the oxidative stress damage and on modulation of cell proliferation pathways in UVB irradiated skin.

Oxidative Stress and Modification of Renal Vascular Permeability Are Associated with Acute Kidney Injury during P. berghei ANKA Infection

Malaria associated-acute kidney injury (AKI) is associated with 45% of mortality in adult patients hospitalized with severe form of the disease. However, the causes that lead to a framework of malaria-associated AKI are still poorly characterized. Some clinical studies speculate that oxidative stress products, a characteristic of Plasmodium infection, as well as proinflammatory response induced by the parasite are involved in its pathophysiology. Therefore, we aimed to investigate the development of malaria-associated AKI during infection by P. berghei ANKA, with special attention to the role played by the inflammatory response and the involvement of oxidative stress. For that, we took advantage of an experimental model of severe malaria that showed significant changes in the renal pathophysiology to investigate the role of malaria infection in the renal microvascular permeability and tissue injury. Therefore, BALB/c mice were infected with P. berghei ANKA. To assess renal function, creatinine, blood urea nitrogen, and ratio of proteinuria and creatininuria were evaluated. The products of oxidative stress, as well as cytokine profile were quantified in plasma and renal tissue. The change of renal microvascular permeability, tissue hypoxia and cellular apoptosis were also evaluated. Parasite infection resulted in renal dysfunction. Furthermore, we observed increased expression of adhesion molecule, proinflammatory cytokines and products of oxidative stress, associated with a decrease mRNA expression of HO-1 in kidney tissue of infected mice. The measurement of lipoprotein oxidizability also showed a significant increase in plasma of infected animals. Together, our findings support the idea that products of oxidative stress, as well as the immune response against the parasite are crucial to changes in kidney architecture and microvascular endothelial permeability of BALB/c mice infected with P. berghei ANKA.

Near-Term Fetal Hypoxia-Ischemia in Rabbits MRI Can Predict Muscle Tone Abnormalities and Deep Brain Injury

Background and Purpose-The pattern of antenatal brain injury varies with gestational age at the time of insult. Deep brain nuclei are often injured at older gestational ages. Having previously shown postnatal hypertonia after preterm fetal rabbit hypoxia-ischemia, the objective of this study was to investigate the causal relationship between the dynamic regional pattern of brain injury on MRI and the evolution of muscle tone in the near-term rabbit fetus. Methods-Serial MRI was performed on New Zealand white rabbit fetuses to determine equipotency of fetal hypoxia-ischemia during uterine ischemia comparing 29 days gestation (E29, 92% gestation) with E22 and E25. E29 postnatal kits at 4, 24, and 72 hours after hypoxia-ischemia underwent T2- and diffusion-weighted imaging. Quantitative assessments of tone were made serially using a torque apparatus in addition to clinical assessments. Results-Based on the brain apparent diffusion coefficient, 32 minutes of uterine ischemia was selected for E29 fetuses. At E30, 58% of the survivors manifested hind limb hypotonia. By E32, 71% of the hypotonic kits developed dystonic hypertonia. Marked and persistent apparent diffusion coefficient reduction in the basal ganglia, thalamus, and brain stem was predictive of these motor deficits. Conclusions-MRI observation of deep brain injury 6 to 24 hours after near-term hypoxia-ischemia predicts dystonic hypertonia postnatally. Torque-displacement measurements indicate that motor deficits in rabbits progressed from initial hypotonia to hypertonia, similar to human cerebral palsy, but in a compressed timeframe. The presence of deep brain injury and quantitative shift from hypo-to hypertonia may identify patients at risk for developing cerebral palsy. (Stroke. 2012;43:2757-2763.)

Effects of cerebral perfusion pressure and increased fraction of inspired oxygen on brain tissue oxygen, lactate and glucose in patients with severe head injury

OBJECTIVE: The purpose of the study was to measure the effects of increased inspired oxygen on patients suffering severe head injury and consequent influences on the correlations between CPP and brain tissue oxygen (PtiO2) and the effects on brain microdialysate glucose and lactate. METHODS: In a prospective, observational study 20 patients suffering severe head injury (GCS< or =8) were studied between January 2000 and December 2001. Each patient received an intraparenchymal ICP device and an oxygen sensor and, in 17 patients brain microdialysis was performed at the cortical-subcortical junction. A 6 h 100% oxygen challenge (F IO2 1.0) ( Period A) was performed as early as possible in the first 24 hours after injury and compared with a similar 6 hour period following the challenge ( Period B). Statistics were performed using the linear correlation analysis, one sample t-test, as well as the Lorentzian peak correlation analysis. RESULTS: F IO2 was positively correlated with PtiO2 (p < 0.0001) over the whole study period. PtiO2 was significantly higher (p < 0.001) during Period A compared to Period B. CPP was positively correlated with PtiO2 (p < 0.001) during the whole study. PtiO2 peaked at a CPP value of 78 mmHg performing a Lorentzian peak correlation analysis of all patients over the whole study. During Period A the brain microdialysate lactate was significantly lower (p = 0.015) compared with Period B. However the brain microdialysate glucose remained unchanged. CONCLUSION: PtiO2 is significantly positively correlated with F IO2, meaning that PtiO2 can be improved by the simple manipulation of increasing F IO2 and ABGAO2. PtiO2 is positively correlated with CPP, peaking at a CPP value of 78 mmHg. Brain microdialysate lactate can be lowered by increasing PtiO2 values, as observed during the oxygen challenge, whereas microdialysate glucose is unchanged during this procedure. Extension of the oxygen challenge time and measurement of the intermediate energy metabolite pyruvate may clarify the metabolic effects of the intervention. Prospective comparative studies, including analysis of outcome on a larger multicenter basis, are necessary to assess the long term clinical benefits of this procedure.

rCBF in hemorrhagic, non-hemorrhagic and mixed contusions after severe head injury and its effect on perilesional cerebral blood flow

Intracerebral contusions can lead to regional ischemia caused by extensive release of excitotoxic aminoacids leading to increased cytotoxic brain edema and raised intracranial pressure. rCBF measurements might provide further information about the risk of ischemia within and around contusions. Therefore, the aim of the presented study was to compare the intra- and perilesional rCBF of hemorrhagic, non-hemorrhagic and mixed intracerebral contusions. In 44 patients, 60 stable Xenon-enhanced CT CBF-studies were performed (EtCO2 30 +/- 4 mmHg SD), initially 29 hours (39 studies) and subsequent 95 hours after injury (21 studies). All lesions were classified according to localization and lesion type using CT/MRI scans. The rCBF was calculated within and 1-cm adjacent to each lesion in CT-isodens brain. The rCBF within all contusions (n = 100) of 29 +/- 11 ml/100 g/min was significantly lower (p < 0.0001, Mann-Whitney U) compared to perilesional rCBF of 44 +/- 12 ml/100 g/min and intra/perilesional correlation was 0.4 (p < 0.0005). Hemorrhagic contusions showed an intra/perilesional rCBF of 31 +/- 11/44 +/- 13 ml/100 g/min (p < 0.005), non-hemorrhagic contusions 35 +/- 13/46 +/- 10 ml/100 g/min (p < 0.01). rCBF in mixed contusions (25 +/- 9/44 +/- 12 ml/100 g/min, p < 0.0001) was significantly lower compared to hemorrhagic and non-hemorrhagic contusions (p < 0.02). Intracontusional rCBF is significantly reduced to 29 +/- 11 ml/100 g/min but reduced below ischemic levels of 18 ml/100 g/min in only 16% of all contusions. Perilesional CBF in CT normal appearing brain closed to contusions is not critically reduced. Further differentiation of contusions demonstrates significantly lower rCBF in mixed contusions (defined by both hyper- and hypodense areas in the CT-scan) compared to hemorrhagic and non-hemorrhagic contusions. Mixed contusions may evolve from hemorrhagic contusions with secondary increased perilesional cytotoxic brain edema leading to reduced cerebral blood flow and altered brain metabolism. Therefore, the treatment of ICP might be individually modified by the measurement of intra- and pericontusional cerebral blood.

Intravenous mesenchymal stem cell therapy for traumatic brain injury.

OBJECT: Cell therapy has shown preclinical promise in the treatment of many diseases, and its application is being translated to the clinical arena. Intravenous mesenchymal stem cell (MSC) therapy has been shown to improve functional recovery after traumatic brain injury (TBI). Herein, the authors report on their attempts to reproduce such observations, including detailed characterizations of the MSC population, non-bromodeoxyuridine-based cell labeling, macroscopic and microscopic cell tracking, quantification of cells traversing the pulmonary microvasculature, and well-validated measurement of motor and cognitive function recovery. METHODS: Rat MSCs were isolated, expanded in vitro, immunophenotyped, and labeled. Four million MSCs were intravenously infused into Sprague-Dawley rats 24 hours after receiving a moderate, unilateral controlled cortical impact TBI. Infrared macroscopic cell tracking was used to identify cell distribution. Immunohistochemical analysis of brain and lung tissues 48 hours and 2 weeks postinfusion revealed transplanted cells in these locations, and these cells were quantified. Intraarterial blood sampling and flow cytometry were used to quantify the number of transplanted cells reaching the arterial circulation. Motor and cognitive behavioral testing was performed to evaluate functional recovery. RESULTS: At 48 hours post-MSC infusion, the majority of cells were localized to the lungs. Between 1.5 and 3.7% of the infused cells were estimated to traverse the lungs and reach the arterial circulation, 0.295% reached the carotid artery, and a very small percentage reached the cerebral parenchyma (0.0005%) and remained there. Almost no cells were identified in the brain tissue at 2 weeks postinfusion. No motor or cognitive functional improvements in recovery were identified. CONCLUSIONS: The intravenous infusion of MSCs appeared neither to result in significant acute or prolonged cerebral engraftment of cells nor to modify the recovery of motor or cognitive function. Less than 4% of the infused cells were likely to traverse the pulmonary microvasculature and reach the arterial circulation, a phenomenon termed the "pulmonary first-pass effect," which may limit the efficacy of this therapeutic approach. The data in this study contradict the findings of previous reports and highlight the potential shortcomings of acute, single-dose, intravenous MSC therapy for TBI.

Chronic spontaneous activity generated in the somata of primary nociceptors is associated with pain-related behavior after spinal cord injury.

Mechanisms underlying chronic pain that develops after spinal cord injury (SCI) are incompletely understood. Most research on SCI pain mechanisms has focused on neuronal alterations within pain pathways at spinal and supraspinal levels associated with inflammation and glial activation. These events might also impact central processes of primary sensory neurons, triggering in nociceptors a hyperexcitable state and spontaneous activity (SA) that drive behavioral hypersensitivity and pain. SCI can sensitize peripheral fibers of nociceptors and promote peripheral SA, but whether these effects are driven by extrinsic alterations in surrounding tissue or are intrinsic to the nociceptor, and whether similar SA occurs in nociceptors in vivo are unknown. We show that small DRG neurons from rats (Rattus norvegicus) receiving thoracic spinal injury 3 d to 8 months earlier and recorded 1 d after dissociation exhibit an elevated incidence of SA coupled with soma hyperexcitability compared with untreated and sham-treated groups. SA incidence was greatest in lumbar DRG neurons (57%) and least in cervical neurons (28%), and failed to decline over 8 months. Many sampled SA neurons were capsaicin sensitive and/or bound the nociceptive marker, isolectin B4. This intrinsic SA state was correlated with increased behavioral responsiveness to mechanical and thermal stimulation of sites below and above the injury level. Recordings from C- and Aδ-fibers revealed SCI-induced SA generated in or near the somata of the neurons in vivo. SCI promotes the entry of primary nociceptors into a chronic hyperexcitable-SA state that may provide a useful therapeutic target in some forms of persistent pain.

Reactive oxygen intermediates contribute to necrotic and apoptotic neuronal injury in an infant rat model of bacterial meningitis due to group B streptococci.

Reactive oxygen intermediates (ROI) contribute to neuronal injury in cerebral ischemia and trauma. In this study we explored the role of ROI in bacterial meningitis. Meningitis caused by group B streptococci in infant rats led to two distinct forms of neuronal injury, areas of necrosis in the cortex and neuronal loss in the dentate gyrus of the hippocampus, the latter showing evidence for apoptosis. Staining of brain sections with diaminobenzidine after perfusion with manganese buffer and measurement of lipid peroxidation products in brain homogenates both provided evidence that meningitis led to the generation of ROI. Treatment with the radical scavenger alpha-phenyl-tert-butyl nitrone (PBN) (100 mg/kg q8h i.p.) beginning at the time of infection completely abolished ROI detection and the increase in lipidperoxidation. Cerebral cortical perfusion was reduced in animals with meningitis to 37.5+/-21.0% of uninfected controls (P < 0.05), and PBN restored cortical perfusion to 72.0+/-8.1% of controls (P < 0.05 vs meningitis). PBN also completely prevented neuronal injury in the cortex and hippocampus, when started at the time of infection (P < 0.02), and significantly reduced both forms of injury, when started 18 h after infection together with antibiotics (P < 0.004 for cortex and P < 0.001 for hippocampus). These data indicate that the generation of ROI is a major contributor to cerebral ischemia and necrotic and apoptotic neuronal injury in this model of neonatal meningitis.

Modeling and Simulation of Upper Brachial Plexus Injury

This article presents the first musculoskeletal model and simulation of upper plexus brachial injury. From this model is possible to analyse forces and movement ranges in order to develop a robotic exoskeleton to improve rehabilitation. The software that currently exists for musculoskeletal modeling is varied and most have advanced features for proper analysis and study of motion simulations. Whilst more powerful computer packages are usually expensive, there are other free and open source packages available which offer different tools to perform animations and simulations and which obtain forces and moments of inertia. Among them, Musculoskeletal Modeling Software was selected to construct a model of the upper limb, which has 7 degrees of freedom and 10 muscles. These muscles are important for two of the movements simulated in this article that are part of the post-surgery rehabilitation protocol. We performed different movement animations which are made using the inertial measurement unit to capture real data from movements made by a human being. We also performed the simulation of forces produced in elbow flexion-extension and arm abduction-adduction of a healthy subject and one with upper brachial plexus injury in a postoperative state to compare the force that is capable of being produced in both cases.

Overexpression of human copper,zinc-superoxide dismutase (SOD1) prevents postischemic injury

Superoxide and superoxide-derived oxidants have been hypothesized to be important mediators of postischemic injury. Whereas copper,zinc-superoxide dismutase, SOD1, efficiently dismutates superoxide, there has been controversy regarding whether increasing intracellular SOD1 expression would protect against or potentiate cellular injury. To determine whether increased SOD1 protects the heart from ischemia and reperfusion, studies were performed in a newly developed transgenic mouse model in which direct measurement of superoxide, contractile function, bioenergetics, and cell death could be performed. Transgenic mice with overexpression of human SOD1 were studied along with matched nontransgenic controls. Immunoblotting and immunohistology demonstrated that total SOD1 expression was increased 10-fold in hearts from transgenic mice compared with nontransgenic controls, with increased expression in both myocytes and endothelial cells. In nontransgenic hearts following 30 min of global ischemia a reperfusion-associated burst of superoxide generation was demonstrated by electron paramagnetic resonance spin trapping. However, in the transgenic hearts with overexpression of SOD1 the burst of superoxide generation was almost totally quenched, and this was accompanied by a 2-fold increase in the recovery of contractile function, a 2.2-fold decrease in infarct size, and a greatly improved recovery of high energy phosphates compared with that in nontransgenic controls. These results demonstrate that superoxide is an important mediator of postischemic injury and that increasing intracellular SOD1 dramatically protects the heart from this injury. Thus, increasing intracellular SOD1 expression may be a highly effective approach to decrease the cellular injury that occurs following reperfusion of ischemic tissues.

The Utility of the CVLT-C as an Embedded Validity Indicator After Pediatric Mild Traumatic Brain Injury.

Results of neuropsychological examinations depend on valid data. Whereas clinicians previously believed that clinical skill was sufficient to identify non-credible performance by examinees on standard tests, research demonstrates otherwise. Consequently, studies on measures to detect suspect effort in adults have received tremendous attention in the previous twenty years, and incorporation of validity indicators into neuropsychological examinations is now seen as integral. Few studies exist that validate methods appropriate for the measurement of effort in pediatric populations. Of extant studies, most evaluate standalone measures originally developed for use with adults. The present study examined the utility of indices from the California Verbal Learning Test – Children's Version (CVLT-C) as embedded validity indicators in a pediatric sample. Participants were 225 outpatients aged 8 to 16 years old referred for clinical assessment after mild traumatic brain injury (mTBI). Non-credible performance (n = 39) was defined as failure of the Medical Symptom Validity Test (MSVT). Logistic regression demonstrated that only the Recognition Discriminability index was predictive of MSVT failure (OR = 2.88, p < .001). A cutoff of z ≤ -1.0 was associated with sensitivity of 51% and specificity of 91%. In the current study, CVLT-C Recognition Discriminability was useful in the identification of non-credible performance in a sample of relatively high-functioning pediatric outpatients with mTBI. Thus, this index can be added to the short list of embedded validity indicators appropriate for pediatric neuropsychological assessment.