Decreased Tissue Damage Research Articles

Amorphous silicon carbide ultramicroelectrode arrays for neural stimulation and recording

Objective. Foreign body response to indwelling cortical microelectrodes limits the reliability of neural stimulation and recording, particularly for extended chronic applications in behaving animals. The extent to which this response compromises the chronic stability of neural devices depends on many factors including the materials used in the electrode construction, the size, and geometry of the indwelling structure. Here, we report on the development of microelectrode arrays (MEAs) based on amorphous silicon carbide (a-SiC). Approach. This technology utilizes a-SiC for its chronic stability and employs semiconductor manufacturing processes to create MEAs with small shank dimensions. The a-SiC films were deposited by plasma enhanced chemical vapor deposition and patterned by thin-film photolithographic techniques. To improve stimulation and recording capabilities with small contact areas, we investigated low impedance coatings on the electrode sites. The assembled devices were characterized in phosphate buffered saline for their electrochemical properties. Main results. MEAs utilizing a-SiC as both the primary structural element and encapsulation were fabricated successfully. These a-SiC MEAs had 16 penetrating shanks. Each shank has a cross-sectional area less than 60 µm2 and electrode sites with a geometric surface area varying from 20 to 200 µm2. Electrode coatings of TiN and SIROF reduced 1 kHz electrode impedance to less than 100 kΩ from ~2.8 MΩ for 100 µm2 Au electrode sites and increased the charge injection capacities to values greater than 3 mC cm−2. Finally, we demonstrated functionality by recording neural activity from basal ganglia nucleus of Zebra Finches and motor cortex of rat. Significance. The a-SiC MEAs provide a significant advancement in the development of microelectrodes that over the years has relied on silicon platforms for device manufacture. These flexible a-SiC MEAs have the potential for decreased tissue damage and reduced foreign body response. The technique is promising and has potential for clinical translation and large scale manufacturing.

Abstract 21206: Manf, a Structurally Unique Redox-Sensitive Chaperone, Restores ER-Protein Folding in the Ischemic Heart

Rationale: In cardiomyocytes, most secreted and membrane proteins are synthesized and folded in the sarcoplasmic/endoplasmic reticulum (SR/ER). We previously showed that during myocardial ischemia, decreased oxygen creates a reducing environment in the SR/ER, preventing protein disulfide isomerases (PDIs) from forming disulfide bonds in nascent proteins, causing ER stress, i.e. the toxic accumulation of unfolded proteins which contributes to cardiomyocyte death. In response to ER stress, the transcription factor, ATF6 induces chaperones that restore SR/ER protein folding. We found that ATF6 also induces mesencephalic astrocyte-derived neurotrophic factor (MANF), a recently identified protein of unknown function. MANF is structurally unique, so its function cannot be inferred from other proteins. Since MANF is induced by ATF6, is ER-localized, and possesses a conserved pattern of cysteines found in all known species of MANF, we hypothesized that MANF is a redox-regulated chaperone that optimizes cardiomyocyte viability during ischemia. Methods: The ability of MANF to bind misfolded proteins during reductive ER stress or ischemia were assessed in neonatal rat ventricular myocytes (NRVM). The ability of recombinant MANF (rMANF) to suppress aggregation of misfolded proteins was examined in an in vitro chaperone assay. Finally, the effects of MANF loss-of-function in the ischemic heart, in vivo , were determined by generating a transgenic mouse model that expresses a cardiomyocyte-specific MANF-targeted microRNA. Results: In NRVM subjected to reductive ER stress or simulated ischemia, MANF formed disulfide-linked complexes with misfolded proteins. Under reducing conditions, rMANF suppressed aggregation of model misfolded proteins in vitro , and mutant rMANF in which the cysteine residues were mutated to alanine did not suppress misfolded protein aggregation. MANF knockdown in the heart, in vivo , increased damage from myocardial infarction, and an AAV9-based gene therapy approach rescued the effects of MANF deficiency, in vivo . Conclusions: MANF is a redox-sensitive SR/ER-resident chaperone that is a critical contributor to SR/ER protein folding during the adaptive ER stress response and decreases tissue damage in the ischemic heart.

MANF, a structurally unique redox-sensitive chaperone, restores ER-protein folding in the ischemic heart

Rationale: In cardiomyocytes, most secreted and membrane proteins are synthesized and folded in the sarcoplasmic/endoplasmic reticulum (SR/ER). We previously showed that during ischemia, decreased oxygen creates a reducing environment in the SR/ER, preventing protein disulfide isomerases (PDIs) from forming disulfide bonds in nascent proteins, causing ER stress– the toxic accumulation of unfolded proteins which contributes to cardiomyocyte death. Upon ER stress, the transcription factor, ATF6 induces chaperones that restore SR/ER protein folding. We found that ATF6 induces mesencephalic astrocyte-derived neurotrophic factor (MANF), a recently identified protein of unknown function. MANF is structurally unique, so its function cannot be inferred from other proteins. Since MANF ATF6-inducible, ER-localized, and contains a conserved redox-sensitive motif found in PDIs, we hypothesized that MANF is a redox-sensitive chaperone that optimizes cardiomyocyte viability during ischemia. Methods: The redox status of MANF during reductive ER stress, and ability to bind misfolded proteins during ischemia were assessed in neonatal rat ventricular myocytes (NRVM). The ability of recombinant MANF to suppress misfolded protein aggregation was examined in an in vitro chaperone assay. Finally, the effects of MANF loss-of-function in the ischemic heart, in vivo, were determined by generating a transgenic mouse that expresses a cardiomyocyte-specific MANF-targeted microRNA. Results: In NRVM subjected to ER stress MANF was as sensitive to changes in ER redox status as the sentinel PDI, PDIA1 and formed disulfide-linked complexes with misfolded proteins during ischemia-mediated ER stress. Under reducing conditions, recombinant MANF suppressed aggregation of model misfolded proteins, in vitro. MANF knockdown in the heart, in vivo, increased damage from myocardial infarction, and an AAV9-based gene therapy approach rescued the effects of MANF deficiency, in vivo. Conclusions: MANF is a redox-sensitive SR/ER-resident chaperone that is a critical contributor to SR/ER protein folding during the adaptive ER stress response and decreases tissue damage in the ischemic heart.

Abstract 2: MANF, A Structurally Unique Redox-Sensitive Chaperone, Restores ER Protein Folding in the Ischemic Heart.

Rationale: In cardiomyocytes, most secreted and membrane proteins are synthesized and folded in the sarcoplasmic/endoplasmic reticulum (SR/ER). We previously showed that during myocardial ischemia, decreased oxygen creates a reducing environment in the SR/ER, preventing protein disulfide isomerases (PDIs) from forming disulfide bonds in nascent proteins, causing ER stress, i.e. the toxic accumulation of unfolded proteins which contributes to cardiomyocyte death. In response to ER stress, the transcription factor, ATF6 induces chaperones that restore SR/ER protein folding. We found that ATF6 also induces mesencephalic astrocyte-derived neurotrophic factor (MANF), a recently identified protein of unknown function. MANF is structurally unique, so its function cannot be inferred from other proteins. Since MANF is induced by ATF6, is ER-localized, and contains a conserved redox-sensitive motif found in PDIs, we hypothesized that MANF is a redox-sensitive chaperone that optimizes cardiomyocyte viability during ischemia. Methods: The redox status of MANF during reductive ER stress and the ability of MANF to bind misfolded proteins during ischemia were assessed in neonatal rat ventricular myocytes (NRVM). The ability of recombinant MANF to suppress aggregation of misfolded proteins was examined in an in vitro chaperone assay. Finally, the effects of MANF loss-of-function in the ischemic heart, in vivo , were determined by generating a transgenic mouse model that expresses a cardiomyocyte-specific MANF-targeted microRNA. Results: In NRVM subjected to ER stress MANF was as sensitive to changes in ER redox status as the sentinel PDI, PDIA1. Moreover, MANF formed disulfide-linked complexes with misfolded proteins during ischemia-mediated ER stress. Under reducing conditions, recombinant MANF suppressed aggregation of model misfolded proteins, in vitro . MANF knockdown in the heart, in vivo , increased damage from myocardial infarction, and an AAV9-based gene therapy approach rescued the effects of MANF deficiency, in vivo. Conclusions: MANF is a redox-sensitive SR/ER-resident chaperone that is a critical contributor to SR/ER protein folding during the adaptive ER stress response and decreases tissue damage in the ischemic heart.

Mycobacterium smegmatis proteoliposome induce protection in a murine progressive pulmonary tuberculosis model

Tuberculosis (TB) remains an important cause of mortality and morbidity. The TB vaccine, BCG, is not fully protective against the adult form of the disease and is unable to prevent its transmission although it is still useful against severe childhood TB. Hence, the search for new vaccines is of great interest. In a previous study, we have shown that proteoliposomes obtained from Mycobacterium smegmatis (PLMs) induced cross reactive humoral and cellular response against Mycobacterium tuberculosis (Mtb) antigens. With the objective to evaluate the protective capability of PLMs, a murine model of progressive pulmonary TB was used. Animals immunized with PLMs with and without alum (PLMs/PLMsAL respectively) showed protection compared to non-immunized animals. Mice immunized with PLMsAL induced similar protection as that of BCG. Animals immunized with BCG, PLMs and PLMsAL showed a significant decrease in tissue damage (percentage of pneumonic area/lung) compared to non-immunized animals, with a more prominent effect in BCG vaccinated mice. The protective effect of the administration of PLMs in mice supports its future evaluation as experimental vaccine candidate against Mtb.

Abstract 291: MANF, a Structurally Unique ER Stress-inducible Protein, Restores ER-protein Folding in ER Stressed Cardiac Myocytes and in the Ischemic Heart

The sarco/endoplasmic reticulum (SR/ER) of cardiomyocytes is a critical site of protein synthesis and folding, as most secreted and membrane proteins including receptors, growth factors, ion channels, and calcium-handling proteins are made at this location. Myocardial ischemia induces ER stress, during which toxic, misfolded proteins accumulate in the SR/ER and contribute to cardiomyocyte death. The branch of the ER stress response mediated by the transcription factor, ATF6, induces ER chaperones that restore SR/ER protein folding. We found that ATF6 also induces mesencephalic astrocyte-derived neurotrophic factor (MANF), a novel, ubiquitously expressed, ER-luminal protein of unknown function. MANF is structurally unique, thus its function cannot be inferred by structural analogy to known proteins. Since it is ATF6-inducible, and resides in the ER lumen, we hypothesized that MANF is an ER chaperone required for optimal viability of cardiac myocytes during ER stresses, including ischemia. The characteristics of MANF gene induction by ER stress, and the effects of MANF knockdown on the ER stress response and cell viability were determined in cultured neonatal rat ventricular myocytes (NRVM). The ability of recombinant MANF to restore structure and function to model misfolded proteins was also examined. Finally, the effects of MANF loss-of-function in the ischemic heart, in vivo , were determined by generating a transgenic mouse model that expresses a cardiomyocyte-specific MANF-targeted microRNA. MANF induction and functional characteristics phenocopied those of a well-studied ER chaperone, glucose-regulated protein 78 (Grp78). Like Grp78, MANF was induced by ER stress in an ATF6-dependent manner. Like knockdown of Grp78, knockdown of MANF in NRVM increased myocyte death in response to ER stress. Like recombinant Grp78, recombinant MANF exhibited a robust ability to restore structure and function to model misfolded proteins, in vitro. Finally, MANF knockdown in the heart, in vivo , increased damage in a mouse model of myocardial infarction. These results suggest that MANF is an SR/ER-resident chaperone required for restoration of SR/ER protein folding during the adaptive ER stress response, and decreasing tissue damage in the ischemic heart.

Reduced Leukocyte Infiltration in Absence of Eosinophils Correlates with Decreased Tissue Damage and Disease Susceptibility in ΔdblGATA Mice during Murine Neurocysticercosis.

Neurocysticercosis (NCC) is one of the most common helminth parasitic diseases of the central nervous system (CNS) and the leading cause of acquired epilepsy worldwide. NCC is caused by the presence of the metacestode larvae of the tapeworm Taenia solium within brain tissues. NCC patients exhibit a long asymptomatic phase followed by a phase of symptoms including increased intra-cranial pressure and seizures. While the asymptomatic phase is attributed to the immunosuppressive capabilities of viable T. solium parasites, release of antigens by dying organisms induce strong immune responses and associated symptoms. Previous studies in T. solium-infected pigs have shown that the inflammatory response consists of various leukocyte populations including eosinophils, macrophages, and T cells among others. Because the role of eosinophils within the brain has not been investigated during NCC, we examined parasite burden, disease susceptibility and the composition of the inflammatory reaction in the brains of infected wild type (WT) and eosinophil-deficient mice (ΔdblGATA) using a murine model of NCC in which mice were infected intracranially with Mesocestoides corti, a cestode parasite related to T. solium. In WT mice, we observed a time-dependent induction of eosinophil recruitment in infected mice, contrasting with an overall reduced leukocyte infiltration in ΔdblGATA brains. Although, ΔdblGATA mice exhibited an increased parasite burden, reduced tissue damage and less disease susceptibility was observed when compared to infected WT mice. Cellular infiltrates in infected ΔdblGATA mice were comprised of more mast cells, and αβ T cells, which correlated with an abundant CD8+ T cell response and reduced CD4+ Th1 and Th2 responses. Thus, our data suggest that enhanced inflammatory response in WT mice appears detrimental and associates with increased disease susceptibility, despite the reduced parasite burden in the CNS. Overall reduced leukocyte infiltration due to absence of eosinophils correlates with attenuated tissue damage and longer survival of ΔdblGATA mice. Therefore, our study suggests that approaches to clear NCC will require strategies to tightly control the host immune response while eradicating the parasite with minimal damage to brain tissue.

Enhancing Fluorogold-based neural tract tracing

BackgroundFluorogold (FG) is used by many groups to retrogradely trace nervous system pathways. Fluorogold, while a robust tracer, also is neurotoxic and causes tissue damage at the injection site and leads to motor deficits. New methodIn the current study, we describe a method for enhancing FG-uptake using Triton™ and an overall procedure for reducing FG-related tissue damage while still allowing effective quantification. ResultsTriton™ decreases the amount of FG, as well as the time required for long-distance transport from the thoracic spinal cord to the motor cortex by >4 fold when this distance is >10in. Although small FG concentrations and injection volumes are ideal for minimizing associated tissue damage and motor deficits, they result in difficult-to-detect fluorescence. This can be solved using FG antiserum paired with an ABC chromogen reaction. This ABC chromogen reaction product can remain stable for at least 9 years. Comparison with existing method(s)This study is the first to collectively address FG-induced tissue damage and describe methods for minimizing this damage. ConclusionsTriton™ enhances the uptake of FG in the nervous system, reduces the FG required, and allows for a substantial decrease in tracing time that limits FG-induced motor deficits. Small FG concentration and volume decreases tissue damage but also decreases FG fluorescent detection. Detection challenges are resolved using FG anti-serum and chromogen reactions.

CD47 regulates renal tubular epithelial cell self-renewal and proliferation following renal ischemia reperfusion

Defects in renal tubular epithelial cell repair contribute to renal ischemia reperfusion injury, cause acute kidney damage, and promote chronic renal disease. The matricellular protein thrombospondin-1 and its receptor CD47 are involved in experimental renal ischemia reperfusion injury, although the role of this interaction in renal recovery is unknown. We found upregulation of self-renewal genes (transcription factors Oct4, Sox2, Klf4 and cMyc) in the kidney of CD47(-/-) mice after ischemia reperfusion injury. Wild-type animals had minimal self-renewal gene expression, both before and after injury. Suggestive of cell autonomy, CD47(-/-) renal tubular epithelial cells were found to increase expression of the self-renewal genes. This correlated with enhanced proliferative capacity compared with cells from wild-type mice. Exogenous thrombospondin-1 inhibited self-renewal gene expression in renal tubular epithelial cells from wild-type but not CD47(-/-) mice, and this was associated with decreased proliferation. Treatment of renal tubular epithelial cells with a CD47 blocking antibody or CD47-targeting small interfering RNA increased expression of some self-renewal transcription factors and promoted cell proliferation. In a syngeneic kidney transplant model, treatment with a CD47 blocking antibody increased self-renewal transcription factor expression, decreased tissue damage, and improved renal function compared with that in control mice. Thus, thrombospondin-1 via CD47 inhibits renal tubular epithelial cell recovery after ischemia reperfusion injury through inhibition of proliferation/self-renewal.

Delayed injection of polypyrrole doped with iodine particle suspension after spinal cord injury in rats improves functional recovery and decreased tissue damage evaluated by 3.0 Tesla in vivo magnetic resonance imaging

Background ContextTraumatic spinal cord injury (SCI) causes irreversible damage with loss of motor, sensory, and autonomic functions. Currently, there is not an effective treatment to restore the lost neurologic functions. PurposeInjection of polypyrrole-iodine(PPy-I) particle suspension is proposed as a therapeutic strategy. Study DesignThis is an in vivo animal study. MethodsThis study evaluates the use of such particles in rats after SCI by examining spared nervous tissue and the Basso, Beattie, and Bresnahan (BBB) scale to evaluate the functional outcome. Diffusive magnetic resonance imaging (MRI) was employed to measure the apparent diffusion coefficient (ADC) and fractional anisotropy (FA) as non-invasive biomarkers of damage after SCI. ResultsFractional anisotropy decreased, whereas ADC increased in all groups after the lesion. There were significant differences in FA when compared with the SCI-PPy-I group versus the SCI group (p<.05). Significant positive correlations between BBB and FA (r2=0.449, p<.05) and between FA and preserved tissue (r2=0.395, p<.05) were observed, whereas significant negative associations between BBB and ADC (r2=0.367, p<.05) and between ADC and preserved tissue (r2=0.421, p<.05) were observed. ConclusionsThe results suggested that PPy-I is neuroprotective as it decreased the amount of damaged tissue while improving the motor function. Non-invasive MRI proved to be useful in the characterization of SCI and recovery.

Cornel Iridoid Glycoside Improves Locomotor Impairment and Decreases Spinal Cord Damage in Rats.

Purpose. This study was to investigate the effects of cornel iridoid glycoside (CIG) on spinal cord injury (SCI) in rats. Methods. The thoracic cord (at T9) of rats was injured by clip compression for 30 sec. Locomotor function was assessed using the Basso, Beattie, and Bresnahan (BBB) rating scale. Neuroanatomic stereological parameters as well as Nogo-A, p75 neurotrophin receptor (p75NTR), and ROCKII expression were measured by histological processing, immunohistochemistry, and stereological analyses. The axons passing through the lesion site were detected by BDA tracing. Results. Intragastric administration of CIG (60 and 180 mg/kg) improved the locomotor impairment at 10, 17, 24, and 31 days post-injury (dpi) compared with untreated SCI model rats. CIG treatment decreased the volume of the lesion epicenter (LEp) and increased the volume of spared tissue and the number of surviving neurons in the injured spinal cord at 31 dpi. CIG promoted the growth of BDA-positive axons and their passage through the lesion site and decreased the expression of Nogo-A, p75NTR, and ROCKII both in and around the LEp. Conclusion. CIG improved the locomotor impairment, decreased tissue damage, and downregulated the myelin-associated inhibition signaling pathway in SCI rats. The results suggest that CIG may be beneficial for SCI therapy.

Comparative study of clonidine versus esmolol on hemodynamic responses during laparoscopic cholecystectomy

BackgroundLaparoscopic surgical procedures have various benefits to the patient in terms of decreased tissue damage, early ambulation, decreased hospital stay and reduced analgesic needs. Pneumoperitoneum with carbon dioxide (CO2) leads to stimulation of the sympathetic nervous system which can be a risk factor in patients with cardiovascular diseases. Moreover, reverse Trendelenburg position affects homeostasis in laparoscopic surgeries. In this study, we compared the efficacy of clonidine (which is α2 adrenergic agonist) versus esmolol (which is ultra short acting cardio-selective β1-receptor antagonist) on the hemodynamic response during laparoscopic cholecystectomy.Materials and methodsA total of 60 patients scheduled to undergo laparoscopic cholecystectomy were randomly assigned into two groups: Group C: received 2 μg/kg of clonidine diluted in 20 ml normal saline, given with slow intravenous infusion over 10 min just before induction of GA and Group E: received 1.5 mg/kg of esmolol as a loading dose over a period of 5 min just before induction of GA followed by 10 μg/kg/min as a maintenance dose throughout the procedure. No hypnotic medication was given on the evening before surgery. Systolic, diastolic, mean arterial blood pressures and heart rate were recorded at (1) baseline, (2) three minutes after endotracheal intubation, (3) before pneumoperitoneum, (4) fifteen minutes after pneumoperitoneum, (5) thirty minutes after pneumoperitoneum, (6) five minutes after release of CO2 and (7) five minutes after extubation. Degree of sedation according to Ramsay sedation score was assessed 15 min after reaching PACU.ResultsBoth groups were similar with respect to demographic data. Clonidine group showed more stability in hemodynamic responses than esmolol group in all hemodynamic variables but with more postoperative sedation.ConclusionThis study concluded that clonidine and esmolol provide hemodynamic stability in laparoscopic cholecystectomy but clonidine provides more stability with postoperative sedation.

Tristetraprolin Mediates Anti-Inflammatory Effects of Carbon Monoxide on Lipopolysaccharide-Induced Acute Lung Injury

Low-dose inhaled carbon monoxide is reported to suppress inflammatory responses and exhibit a therapeutic effect in models of lipopolysaccharide (LPS)-induced acute lung injury (ALI). However, the precise mechanism by which carbon monoxide confers protection against ALI is not clear. Tristetraprolin (TTP; official name ZFP36) exerts anti-inflammatory effects by enhancing decay of proinflammatory cytokine mRNAs. With the use of TTP knockout mice, we demonstrate here that the protection by carbon monoxide against LPS-induced ALI is mediated by TTP. Inhalation of carbon monoxide substantially increased the pulmonary expression of TTP. carbon monoxide markedly enhanced the decay of mRNA-encoding inflammatory cytokines, blocked the expression of inflammatory cytokines, and decreased tissue damage in LPS-treated lung tissue. Moreover, knockout of TTP abrogated the anti-inflammatory and tissue-protective effects of carbon monoxide in LPS-induced ALI. These results suggest that carbon monoxide-induced TTP mediates the protective effect of carbon monoxide against LPS-induced ALI by enhancing the decay of mRNA encoding proinflammatory cytokines.

Protective effect of neutralizing anti-IL-18α monoclonal antibody on a mouse model of acute graft-versus-host disease.

Graft-versus-host disease (GVHD) is a devastating complication of hematopoietic stem cell transplantation (HSCT), and is characterized by systemic inflammation and tissue damage in multiple organs, such as the liver and small intestine. Interleukin-18 (IL-18), an important pro-inflammatory cytokine, is elevated during the course of acute GVHD (aGVHD), and is associated with the severe clinical manifestations of the disease. The biological activity of IL-18 is based on its interaction with the IL-18 receptor (IL-18R) expressed in a variety of cells. The aim of this study was to assess whether blocking the interaction of IL-18 with IL-18R by the anti-IL‑18Rα antibody could attenuate the severity of aGVHD. We used a well-established mouse bone marrow transplantation (BMT) model (B6→BALB/c) to block the IL-18/IL-18R interaction by a neutralizing monoclonal antibody (mAb) against murine IL-18Rα. Administration of anti-IL-18Rα mAb had a significant protective effect on the clinical and pathologic manifestations of aGVHD, resulting in a markedly improved survival rate, modified inflammatory response and decreased tissue damage. Interfering with IL-18/IL-18R interaction affected levels of Th1, Th2 and Th17 subsets in the peripheral blood of the aGVHD animals. Additionally, it led to decreased tissue expression of IL-18 and apoptosis-associated molecules (Fas and FasL), and lower phosphorylation levels of p38MAPK in the liver and small intestine. These changes coincided with the decrease in cell apoptosis in aGVHD target organs. Thus, anti‑IL-18Rα therapy may, therefore, represent a new therapeutic interference approach for treating aGVHD.

WNT Agonist Decreases Tissue Damage and Improves Renal Function After Ischemia-Reperfusion.

Renal ischemia-reperfusion (IR) injury (IRI) after shock states or transplantation causes tissue damage and delayed graft function, respectively. The Wnt/β-catenin signaling pathway plays a critical role in nephrogenesis. We therefore hypothesized that pharmacological activation of the Wnt/β-catenin signaling by the Wnt agonist, a synthetic pyrimidine, could protect kidneys from IRI. Adult male rats were subjected to bilateral clamping of the renal pedicles with microvascular clips for 60 min, followed by reperfusion. The Wnt agonist (5 mg/kg body weight) or vehicle (20% dimethyl sulfoxide in saline) was administered intravenously 1 h before ischemia. Blood and renal tissues were collected 24 h after IR for evaluation. Renal IR caused a significant reduction of β-catenin and its downstream target gene cyclin D1 by 65% and 39%, respectively, compared with the sham, whereas the Wnt agonist restored them to sham levels. The number and intensity of cells staining with the proliferation marker Ki67 in ischematized kidneys were enhanced by the Wnt agonist. The integrity of the renal histological architecture in the Wnt agonist group was better preserved than the vehicle group. The Wnt agonist significantly lowered serum levels of creatinine, aspartate aminotransferase, and lactate dehydrogenase and inhibited the production of interleukin 6 and interleukin 1β and myeloperoxidase activities. Lastly, the Wnt agonist reduced inducible nitric oxide synthase, nitrotyrosine proteins, and 4-hydroxynonenal in the kidneys by 60%, 47%, and 21%, respectively, compared with the vehicle. These results indicate that the Wnt agonist improves renal regeneration and function while attenuating inflammation and oxidative stress in the kidneys after IR. Thus, pharmacologic stimulation of the Wnt/β-catenin signaling provides a beneficial effect on the prevention of renal IRI.

Acetylcholine Inhibits LPS-Induced MMP-9 Production and Cell Migration via the a7 nAChR-JAK2/STAT3 Pathway in RAW264.7 Cells

Background: Excessive activation of matrix metalloproteinase 9 (MMP-9) has been found in several inflammatory diseases. Previous studies have shown that acetylcholine (ACh) reduced the levels of pro-inflammatory cytokines and decreased tissue damage. Therefore, this study was designed to explore the potential effects and mechanisms of ACh on MMP-9 production and cell migration in response to lipopolysaccharide (LPS) stimulation in RAW264.7 cells. Methods: MMP-9 expression and activity were induced by LPS in RAW264.7 cells, and examined by real-time PCR, western blotting and gelatin zymography, respectively. ELISA was used to determine the changes in MMP-9 secretion among the groups. Macrophage migration was evaluated using transwell migration assay. Knockdown of a7 nicotinic acetylcholine receptor (a7 nAChR) expression was performed using siRNA transfection. Results: Pre-treatment with ACh inhibited LPS-induced MMP-9 production and macrophage migration in RAW264.7 cells. These effects were abolished by the a7 nAChR antagonist methyllycaconitine (MLA) and a7 nAChR siRNA. The a7 nAChR agonist PNU282987 was found to have an effect similar to that of ACh. Moreover, ACh enhanced the expression of JAK2 and STAT3, and the JAK2 inhibitor AG490 and the STAT3 inhibitor static restored the effect of ACh. Meanwhile, ACh decreased the phosphorylation and nuclear translocation of NF-κB, and this effect was abrogated in the presence of MLA. In addition, the JAK2 and STAT3 inhibitor abolished the inhibitory effects of ACh on phosphorylation of NF-κB. Conclusions: Activation of a7 nAChR by ACh inhibited LPS-induced MMP-9 production and macrophage migration through the JAK2/STAT3 signaling pathway. These results provide novel insights into the anti-inflammatory effects and mechanisms of ACh.

Surgical management of Giant Lumbar Extradural Schwannoma: Report of 3 cases.

Standard surgical approach for extradural paraspinal tumours has been through a unilateral facetectomy, inter-transverse or retroperitoneal approach. Some of these approaches destabilise the spine and consequently require a fusion procedure. Access to these tumours through a minimal access route can decrease tissue damage, fasten post-operative recovery and obviate the need for a concomitant fusion procedure. However, proper case selection and adequate pre-operative planning are important in choosing cases for a minimally invasive approach. We discuss three cases of giant extradural, paraspinal schwannomas. One case that was associated with listhesis along with the tumour situated more anteriorly and embedded in the psoas muscle was managed by lateral retroperitoneal route with simultaneous interbody graft followed by posterior interspinous stabilising device. The other two cases were managed by minimal access route using a 22mm fixed tubular retractor, one by a paramedian approach and one by far lateral approach. The technique and merits of the procedure are discussed.

Interaction of complement-coagulation factors in ischemia/reperfusion-induced injury in autoimmune prone (B6.MRL/lpr) mice (BA3P.205)

Abstract Studies in the B6.MRL/lpr strain using the ischemia/reperfusion tissue injury (IR) model have demonstrated that complement activation is a key modulator of inflammatory tissue damage. The role of various components of the complement pathway (activation vs. deficiency) and the use of complement inhibition as therapeutic agents are areas of active research for systemic lupus erythematous (SLE). C1q inhibition is acutely effective in attenuating tissue injury in B6.MRL/lpr strain. Studies in our laboratory identified a C3-independent mechanism of complement C5a generation, terminal tissue injury, and systemic inflammation. The studies suggest a pathway which bypasses C3 by interacting with the coagulation cascade. Using the mesenteric IR model, we compared the extent of intestinal injury after IR when C1q inhibitors or coagulation inhibitors were administered. In our model C1q inhibition resulted in decreased tissue damage, thrombin generation, and C5a generation. Tissue Factor pathway inhibitor (TFPI) and anti-thrombin III attenuated tissue injury, thrombin generation, and C5a production but not C3b production. Interaction between thrombin and the complement pathway in the autoimmune mouse has significant clinical implications - especially in autoimmune diseases in which inflammation and hypercoagulability are known manifestations.

Parasitological, histopathological, and immunohistochemical assessment of nitric oxide synthase inhibitor: aminoguanidine versus albendazole in the treatment of experimental murine toxocariasis

Objectives The study was intended to evaluate the effect of aminoguanidine compared with that of albendazole on mice infected with eggs of Toxocara canis by parasitological, histopathological, and immunohistochemical studies. Background T. canis is a widely distributed parasite. The nematode can cause toxocariasis in man. The infection takes place after swallowing fully embryonated eggs with larvae inside. Materials and methods The study was conducted on 117 albino mice classified into four groups: GI (the control group with subgroups Ia, Ib, and Ic); GII (infected with T. canis eggs); GIII (infected with T. canis eggs and treated with albendazole); and GIV (infected with T. canis eggs and treated with aminoguanidine). Sera from different groups of mice were collected for nitrite assay. The lung and brain tissues were taken for larval recovery and histopathological and immunohistochemical [inducible nitric oxide synthase (iNOS)] studies. Results The mean larval count decreased significantly from 7th to 45th day post infection (d.p.i), with albendazole and aminoguanidine, which was more effective in decreasing larval count. Aminoguanidine treatment caused early improvement in histopathological lesions initiating from the second d.p.i.; however, albendazole improvement was observed on the seventh d.p.i. The grade of iNOs expression in GII and GIII was high as compared with that in GIV. There was significantly positive correlation between serum nitrite and the grade of iNOS expression, especially on early d.p.i only in groups II and III. Conclusion Both albendazole and aminoguanidine treatment for T. canis-infected mice caused decrease in larval count and improvement in histopathological lesions. Aminoguanidine was more effective with early response and decreased tissue damage.

Experts convene to improve cardiovascular disease outcomes

Experts convene to improve cardiovascular disease outcomes