Application of a Novel Anti-Adhesive Membrane, E8002, in a Rat Laminectomy Model.

Neuropathic pain after spinal surgery, so-called failed back surgery syndrome, is a frequently observed common complication. One cause of the pain is scar tissue formation, observed as post-surgical epidural adhesions. These adhesions may compress surrounding spinal nerves, resulting in pain, even after successful spinal surgery. E8002 is an anti-adhesive membrane. In Japan, a clinical trial of E8002 is currently ongoing in patients undergoing abdominal surgery. However, animal experiments have not been performed for E8002 in spinal surgery. We assessed the anti-adhesive effect of E8002 in a rat laminectomy model. The dura matter was covered with an E8002 membrane or left uncovered as a control. Neurological evaluations and histopathological findings were compared at six weeks postoperatively. Histopathological analyses were performed by hematoxylin⁻eosin and aldehyde fuchsin-Masson Goldner staining. Three assessment areas were selected at the middle and margins of the laminectomy sites, and the numbers of fibroblasts and inflammatory cells were counted. Blinded histopathological evaluation revealed that adhesions and scar formation were reduced in the E8002 group compared with the control group. The E8002 group had significantly lower numbers of fibroblasts and inflammatory cells than the control group. The present results indicate that E8002 can prevent epidural scar adhesions after laminectomy.

Preconditioning Exercise in Rats Attenuates Early Brain Injury Resulting from Subarachnoid Hemorrhage by Reducing Oxidative Stress, Inflammation, and Neuronal Apoptosis.

Subarachnoid hemorrhage (SAH) is a catastrophic form of stroke responsible for significant morbidity and mortality. Oxidative stress, inflammation, and neuronal apoptosis are important in the pathogenesis of early brain injury (EBI) following SAH. Preconditioning exercise confers neuroprotective effects, mitigating EBI; however, the basis for such protection is unknown. We investigated the effects of preconditioning exercise on brain damage and sensorimotor function after SAH. Male rats were assigned to either a sham-operated (Sham) group, exercise (Ex) group, or no-exercise (No-Ex) group. After a 3-week exercise program, they underwent SAH by endovascular perforation. Consciousness level, neurological score, and sensorimotor function were studied. The expression of nuclear factor erythroid 2 p45-related factor 2 (Nrf2), heme oxygenase 1 (HO-1), 4-hydroxynonenal (4HNE), nitrotyrosine (NT), ionized calcium-binding adaptor molecule 1 (Iba1), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), interleukin 1ß (IL-1ß), 14-3-3γ, p-ß-catenin Ser37, Bax, and caspase-3 were evaluated by immunohistochemistry or western blotting. The terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick end labeling (TUNEL) assay was also performed. After SAH, the Ex group had significantly reduced neurological deficits, sensorimotor dysfunction, and consciousness disorder compared with the No-Ex group. Nrf2, HO-1, and 14-3-3γ were significantly higher in the Ex group, while 4HNE, NT, Iba1, TNF-α, IL-6, IL-1ß, Bax, caspase-3, and TUNEL-positive cells were significantly lower. Our findings suggest that preconditioning exercise ameliorates EBI after SAH. The expression of 4HNE and NT was reduced by Nrf2/HO-1 pathway activation; additionally, both oxidative stress and inflammation were reduced. Furthermore, preconditioning exercise reduced apoptosis, likely via the 14-3-3γ/p-ß-catenin Ser37/Bax/caspase-3 pathway.

Preconditioning exercise reduces brain damage and neuronal apoptosis through enhanced endogenous 14-3-3γ after focal brain ischemia in rats.

14-3-3γ is an important early ischemia-inducible protective factor against ischemic cell death in cerebral cortical neurons. We investigated the anti-apoptosis mechanism of enhanced 14-3-3γ mediated by preconditioning exercise-induced brain ischemic tolerance after stroke. Rats were assigned to four groups: exercise and ischemia (Ex group), ischemia and no exercise (No-Ex group), exercise and no ischemia (Ex-only group), and no exercise and ischemia (control group). Rats were trained on a treadmill for 5 days a week for 3 weeks (running speed, 25 m/min; running duration, 30 min/day). After the exercise program, stroke was induced by left middle cerebral artery occlusion. The infarct volume, neurological deficits, and motor function, as well as expression levels of hypoxia-induced factor-1α (HIF-1α), 14-3-3γ, P2X7 receptors, p-ß-catenin Ser37, Bax, and caspase 3 were evaluated by immunohistochemistry and western blotting. The expression of HIF-1α and 14-3-3γ significantly increased in neurons and astrocytes in the Ex-only group. HIF-1α was co-expressed with P2X7 receptor- and GFAP-positive astrocytes. After stroke, the Ex group had significantly reduced brain infarction. HIF-1α and 14-3-3γ significantly increased in the Ex group compared to the No-Ex group. In addition, p-ß-catenin Ser37 significantly increased following elevated 14-3-3γ; in contrast, Bax and caspase 3 were significantly reduced in the Ex group. Our findings suggest that preconditioning exercise prior to ischemia induces neuron- and astrocyte-mediated brain ischemic tolerance through increased expression of HIF-1α and 14-3-3γ, which are intrinsic protective factors; the upregulated 14-3-3γ induced by preconditioning exercise reduces ischemic neuronal cell death through the 14-3-3γ/p-ß-catenin Ser37/Bax/caspase 3 anti-apoptotic pathway.

Correction to: Preconditioning exercise reduces brain damage and neuronal apoptosis through enhanced endogenous 14-3-3γ after focal brain ischemia in rats.

In the original publication of the article, both the Fig. 2e and Fig. 7e have been published incorrectly and the correct figures are given below.

Neuroprotective effects of different frequency preconditioning exercise on neuronal apoptosis after focal brain ischemia in rats.

OBJECTIVE: Preconditioning exercise can exert neuroprotective effects after stroke; however, the effects of exercise intensity, frequency, duration are unknown. We investigated the neuroprotective effect of different frequency preconditioning exercise on neuronal apoptosis after cerebral ischemia in rats. METHODS: Rats were divided into the following five groups: 5 times a week of exercise (5/w-Ex) group, 3 times a week of exercise (3/w-Ex) group, once a week of exercise (1/w-Ex) group, no exercise (No-Ex) group, and intact control (control) group. Rats were made to run on a treadmill for 30 min per day at a speed of 25 m/min for 3 weeks. After the running program, the rats were subjected to 60-min left middle cerebral artery occlusion. Two days after ischemia, the cerebral infarct volume, neurological and motor function, Bcl-2-associated X protein (Bax)/B-cell lymphoma 2 (Bcl-2) ratio, expression of caspase-3, and TUNEL positive cells were examined in the cerebral cortex surrounding the ischemic zone. RESULTS: The 3/w-Ex and 5/w-Ex groups showed significantly reduced infarct volumes compared with the No-Ex group, but the 1/w-Ex group did not. In addition, the 3/w-Ex and 5/w-Ex groups had improved neurological scores and sensorimotor function compared with the No-Ex group. The Bax/Bcl-2 ratio, expression of caspase-3, and TUNEL-positive cells significantly decreased in the penumbra area in the 3/w-Ex or 5/w-Ex groups compared with the No-Ex group. DISCUSSION: Our findings suggested that three times or more per week of high-intensity preconditioning exercise exert neuroprotective effects through the downregulation of the Bax/Bcl-2 ratio and caspase-3 activation after stroke. ABBREVIATIONS: TUNEL: terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick and labeling; MCAO:middle cerebral artery occlusion; BAX:Bcl-2-associated X protein; Bcl-2: B-cell lymphoma 2; TTC: 2,3,5-triphenyltetrazorlium chloride.

The effect of exercise frequency on neuropathic pain and pain-related cellular reactions in the spinal cord and midbrain in a rat sciatic nerve injury model.

BACKGROUND: Exercise regimens are established methods that can relieve neuropathic pain. However, the relationship between frequency and intensity of exercise and multiple cellular responses of exercise-induced alleviation of neuropathic pain is still unclear. We examined the influence of exercise frequency on neuropathic pain and the intracellular responses in a sciatic nerve chronic constriction injury (CCI) model. MATERIALS AND METHODS: Rats were assigned to four groups as follows: CCI and high-frequency exercise (HFE group), CCI and low-frequency exercise (LFE group), CCI and no exercise (No-Ex group), and naive animals (control group). Rats ran on a treadmill, at a speed of 20 m/min, for 30 min, for 5 (HFE) or 3 (LFE) days a week, for a total of 5 weeks. The 50% withdrawal threshold was evaluated for mechanical sensitivity. The activation of glial cells (microglia and astrocytes), expression of brain-derived neurotrophic factor (BDNF) and µ-opioid receptor in the spinal dorsal horn and endogenous opioid in the midbrain were examined using immunohistochemistry. Opioid receptor antagonists (naloxone) were administered using intraperitoneal injection. RESULTS: The development of neuropathic pain was related to the activation of glial cells, increased BDNF expression, and downregulation of the µ-opioid receptor in the ipsilateral spinal dorsal horn. In the No-Ex group, neuropathic pain showed the highest level of mechanical hypersensitivity at 2 weeks, which improved slightly until 5 weeks after CCI. In both exercise groups, the alleviation of neuropathic pain was accelerated through the regulation of glial activation, BDNF expression, and the endogenous opioid system. The expression of BDNF and endogenous opioid in relation to exercise-induced alleviation of neuropathic pain differed in the HFE and LFE groups. The effects of exercise-induced alleviation of mechanical hypersensitivity were reversed by the administration of naloxone. CONCLUSION: The LFE and HFE program reduced neuropathic pain. Our findings indicated that aerobic exercise-induced alleviated neuropathic pain through the regulation of glial cell activation, expression of BDNF in the ipsilateral spinal dorsal horn, and the endogenous opioid system.

Uric acid enhances alteplase-mediated thrombolysis as an antioxidant.

Uric acid (UA) therapy may prevent early ischemic worsening after acute stroke in thrombolysis patients. The aim of this study was to examine the influence of UA on the thrombolytic efficacy of alteplase in human blood samples by measuring thrombolysis under flow conditions using a newly developed microchip-based flow-chamber assay. Human blood samples from healthy volunteers were exposed to UA, alteplase, or a combination of UA and alteplase. Whole blood and platelet-rich plasma were perfused over a collagen- and thromboplastin-coated microchip, and capillary occlusion was monitored with a video microscope and flow-pressure sensor. The area under the curve (extent of thrombogenesis or thrombolysis) at 30 minutes was 92% lower in the UA-alteplase-treated group compared with the alteplase-treated group. D-dimers were measured to evaluate these effects in human platelet-poor plasma samples. Although hydrogen peroxide significantly decreased the elevation of D-dimers by alteplase, UA significantly inhibited the effect of hydrogen peroxide. Meanwhile, rat models of thromboembolic cerebral ischemia were treated with either alteplase or UA-alteplase combination therapy. Compared with alteplase alone, the combination therapy reduced the infarct volume and inhibited haemorrhagic transformation. UA enhances alteplase-mediated thrombolysis, potentially by preventing oxidative stress, which inhibits fibrinolysis by alteplase in thrombi.

Edaravone, a Synthetic Free Radical Scavenger, Enhances Alteplase-Mediated Thrombolysis.

The combination of alteplase, a recombinant tissue plasminogen activator, and edaravone, an antioxidant, reportedly enhances recanalization after acute ischemic stroke. We examined the influence of edaravone on the thrombolytic efficacy of alteplase by measuring thrombolysis using a newly developed microchip-based flow-chamber assay. Rat models of embolic cerebral ischemia were treated with either alteplase or alteplase-edaravone combination therapy. The combination therapy significantly reduced the infarct volume and improved neurological deficits. Human blood samples from healthy volunteers were exposed to edaravone, alteplase, or a combination of alteplase and edaravone or hydrogen peroxide. Whole blood was perfused over a collagen- and thromboplastin-coated microchip; capillary occlusion was monitored with a video microscope and flow-pressure sensor. The area under the curve (extent of thrombogenesis or thrombolysis) at 30 minutes was 69.9% lower in the edaravone-alteplase- than alteplase-treated group. The thrombolytic effect of alteplase was significantly attenuated in the presence of hydrogen peroxide, suggesting that oxidative stress might hinder thrombolysis. D-dimers were measured to evaluate these effects in human platelet-poor plasma samples. Although hydrogen peroxide significantly decreased the elevation of D-dimers by alteplase, edaravone significantly inhibited the decrease. Edaravone enhances alteplase-mediated thrombolysis, likely by preventing oxidative stress, which inhibits fibrinolysis by alteplase in thrombi.

The neuroprotective effects of preconditioning exercise on brain damage and neurotrophic factors after focal brain ischemia in rats.

Preconditioning exercise can exert neuroprotective effects after stroke. However, the mechanism underlying these neuroprotective effects by preconditioning exercise remains unclear. We investigated the neuroprotective effects of preconditioning exercise on brain damage and the expression levels of the midkine (MK) and brain-derived neurotrophic factor (BDNF) after brain ischemia. Animals were assigned to one of 4 groups: exercise and ischemia (Ex), no exercise and ischemia (No-Ex), exercise and no ischemia (Ex-only), and no exercise and intact (Control). Rats ran on a treadmill for 30 min once a day at a speed of 25 m/min for 5 days a week for 3 weeks. After the exercise program, stroke was induced by a 60 min left middle cerebral artery occlusion using an intraluminal filament. The infarct volume, motor function, neurological deficits, and the cellular expressions levels of MK, BDNF, GFAP, PECAM-1, caspase 3, and nitrotyrosine (NT) were evaluated 48 h after the induction of ischemia. The infarct volume, neurological deficits and motor function in the Ex group were significantly improved compared to that of the No-Ex group. The expression levels of MK, BDNF, GFAP, and PECAM-1 were enhanced in the Ex group compared to the expression levels in the No-Ex group after brain ischemia, while the expression levels of activated caspase 3 and NT were reduced in the area surrounding the necrotic lesion. Our findings suggest that preconditioning exercise reduced the infract volume and ameliorated motor function, enhanced expression levels of MK and BDNF, increased astrocyte proliferation, increased angiogenesis, and reduced neuronal apoptosis and oxidative stress.