Feng Ling

Upper limb ischemic preconditioning prevents recurrent stroke in intracranial arterial stenosis

Objective: This study aims to evaluate protective effects of brief repetitive bilateral arm ischemic preconditioning (BAIPC) on stroke recurrence in patients with symptomatic atherosclerotic intracranial arterial stenosis (IAS). Methods: A total of 68 consecutive cases with symptomatic IAS, diagnosed by imaging, were enrolled in this prospective and randomized study. All patients received standard medical management. Patients in the BAIPC group (n = 38) underwent 5 brief cycles consisting of bilateral upper limb ischemia followed by reperfusion. The BAIPC procedure was performed twice daily over 300 consecutive days. Incidence of recurrent stroke and cerebral perfusion status in BAIPC-treated patients were compared with the untreated control group (n = 30). Results: In the control group, incidence of recurrent stroke at 90 and 300 days were 23.3% and 26.7%, respectively. In the BAIPC group, incidence of recurrent stroke was reduced to 5% and 7.9% at 90 and 300 days (p < 0.01), respectively. The average time to recovery (modified Rankin Scale score 0–1) was also shortened by BAIPC. Cerebral perfusion status, measured by SPECT and transcranial Doppler sonography, improved remarkably in BAIPC-treated brain than in control (p < 0.01). Conclusion: This study provides a proof-of-concept that BAIPC may be an effective way to improve cerebral perfusion and reduce recurrent strokes in patients with IAS. Further investigation of this therapeutic approach is warranted as some patients were excluded after randomization.

β-Catenin overexpression in malignant glioma and its role in proliferation and apoptosis in glioblastma cells

Abstractβ-Catenin, a core component of Wnt/β-catenin signaling, has been shown to be a crucial factor in a broad range of tumors, while its role in glioma is not well understood. In this study, the expression of β-catenin in astrocytic glioma tissues with different grade and human normal cerebral tissues was examined using reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. We found a higher expression level of β-catenin in astrocytic glioma patients with high grade in comparison with the normal controls. Additionally, siRNA was transfected into human U251 glioblastoma cells by liposome after the design of siRNA was confirmed to effectively inhibit the expression of β-catenin by RT-PCR. Compared to the control siRNA group, siRNA-mediated knockdown of β-catenin in human U251 cells inhibited cell proliferation, resulted in cell apoptosis, and arrested cell cycle in G0/G1. Additionally, downregulation of β-catenin decreased the expression level of cyclin D1, c-Myc and c-jun. Taken together, these results indicate that overexpression of β-catenin may be an important contributing factor to glioma progression.

Correlation Between Carotid Intraplaque Hemorrhage and Clinical Symptoms Systematic Review of Observational Studies

Background and Purpose— We sought to investigate the association between carotid intraplaque hemorrhage (IPH) and ipsilateral symptoms of cerebral ischemia. Methods— A search was performed for clinical observational studies comparing the incidence of IPH between symptomatic and asymptomatic patients. Odds ratios (ORs) for IPH as a factor in the pathogenesis of neurologic events were calculated and combined by a meta-analysis. Interstudy heterogeneity, estimated effects, and methodologic quality of the studies were assessed. Results— Thirty-one studies were included for analysis. The reported ORs varied widely. Overall, the incidence of IPH in the symptomatic groups was significantly higher than in the asymptomatic group. However, there was an apparent trend for heterogeneity (P<0.00001) between studies. The random-effects summary estimator of ORs was 2.25 (95% CI, 1.57 to 3.22; P<0.00001). To identify potential sources of heterogeneity, subgroup analyses were performed. The pooled ORs varied greatly by stratification. Major heterogeneity was found among studies with low quality, microscopic methods of examination, significant effects, small sizes, early publication, and unequal severity of carotid stenosis in both groups. Large, recent, macroscopic, or high-quality studies, as well as studies with equal degrees of stenosis, tended to yield insignificant associations. The methods in defining and evaluating hemorrhage were very heterogeneous. Characterizations of the age, size, number, and location of hemorrhages were poorly reported and highly variable. In addition, a lack of control of confounders and selection bias were frequently identified among studies. Conclusions— Statistical inferences have suggested a plausible role in the production of cerebral ischemia; however, reliable interpretation was strongly undermined by poor methodologic quality, substantial heterogeneity, and suspicious publication bias. To preciously estimate the underlying correlation, a well-designed study with uniformity in definition and evaluation for IPH might be warranted.

Inhibition of autophagy induced by proteasome inhibition increases cell death in human SHG-44 glioma cells

AbstractAim:The ubiquitin-proteasome system (UPS) and lysosome-dependent macroautophagy (autophagy) are two major intracellular pathways for protein degradation. Recent studies suggest that proteasome inhibitors may reduce tumor growth and activate autophagy. Due to the dual roles of autophagy in tumor cell survival and death, the effect of autophagy on the destiny of glioma cells remains unclear. In this study, we sought to investigate whether inhibition of the proteasome can induce autophagy and the effects of autophagy on the fate of human SHG-44 glioma cells.Methods:The proteasome inhibitor MG-132 was used to induce autophagy in SHG-44 glioma cells, and the effect of autophagy on the survival of SHG-44 glioma cells was investigated using an autophagy inhibitor 3-MA. Cell viability was measured by MTT assay. Apoptosis and cell cycle were detected by flow cytometry. The expression of autophagy related proteins was determined by Western blot.Results:MG-132 inhibited cell proliferation, induced cell death and cell cycle arrest at G2/M phase, and activated autophagy in SHG-44 glioma cells. The expression of autophagy-related Beclin-1 and LC3-I was significantly up-regulated and part of LC3-I was converted into LC3-II. However, when SHG-44 glioma cells were co-treated with MG-132 and 3-MA, the cells became less viable, but cell death and cell numbers at G2/M phase increased. Moreover, the accumulation of acidic vesicular organelles was decreased, the expression of Beclin-1 and LC3 was significantly down-regulated and the conversion of LC3-II from LC3-I was also inhibited.Conclusion:Inhibition of the proteasome can induce autophagy in human SHG-44 glioma cells, and inhibition of autophagy increases cell death. This discovery may shed new light on the effect of autophagy on modulating the fate of SHG-44 glioma cells.

Celastrol causes apoptosis and cell cycle arrest in rat glioma cells.

Abstract Glioma still remains a major health problem in the world. Celastrol has been proved to be an effective natural proteasome inhibitor and was used for treatment of autoimmune disease, chronic inflammation and neurodegenerative disease. However, its effect on glioma is unclear. In this study, we investigated the therapeutic effects of celastrol on C6 glioma cells. The results demonstrated that celastrol inhibited cell proliferation in a time- and dose-dependent manner, suppressed proteasome chymotrypsin-like activity and induced apoptosis and cell cycle arrest at G2/M phase in C6 cells. Proapoptosis proteins bax and caspase-3 were up-regulated, as well as cell cycle G2/M-related proteins cyclin B1, p21 and p27. Conversely, anti-apoptosis proteins bcl-2 and XIAP and cell cycle regulator cyclin-dependent kinase 2 were down-regulated. Taken together, our data suggest that celastrol can suppress proteasome activity and induce apoptosis and cell cycle arrest in C6 glioma cells, which make it be a potential drug for glioma.

Color Doppler imaging evaluation of proximal vertebral artery stenosis.

OBJECTIVEnThe sonographic diagnostic criteria for vertebral artery stenosis have not been fully investigated. The purpose of this study was to assess hemodynamic parameters at color Doppler imaging and to determine, with digital subtraction angiography as the reference standard, the optimal thresholds for evaluation of proximal vertebral artery stenosis.nnnMATERIALS AND METHODSnAmong 653 patients with symptoms of ischemia of the posterior circulation, 247 subjects with normal arteries or stenosis of the proximal vertebral artery confirmed with digital subtraction angiography were included in the study. Peak systolic velocity at the origin of the vertebral artery (PSV(origin)) and in intervertebral segments of the vertebral artery (PSV(IV)), end-diastolic velocity at the origin and in the intervertebral segments of the vertebral artery, and the diameter of the vascular lumen were measured. The cutoff values for the diagnosis of < 50%, 50-69%, and 70-99% stenosis were determined with receiver operating characteristics analysis.nnnRESULTSnThe optimal cutoff values of hemodynamic parameters in evaluation of stenosis of the proximal vertebral artery for < 50% stenosis were PSV(origin) >or= 85 cm/s, PSV(origin) / PSV(IV) >or= 1.3, and end-diastolic velocity at the origin >or= 27 cm/s; for 50-69% stenosis were PSV(origin) >or= 140 cm/s, PSV(origin) / PSV(IV) >or/= 2.1, and end-diastolic velocity at the origin >or= 35 cm/s; and for 70-99% stenosis were PSV(origin) >or= 210 cm/s, PSV(origin) / PSV(IV) >or= 4.0, and end-diastolic velocity at the origin >or= 50 cm/s. PSV(origin) was the most useful hemodynamic parameter, having accuracy of 94.5%, 96.2%, and 88.7% for the diagnosis of < 50%, 50-69%, and 70-99% stenosis.nnnCONCLUSIONnColor Doppler imaging is a reliable method for evaluation of vertebral artery stenosis. The results derived from this study can be used as a reference for establishing sonographic criteria for proximal vertebral artery stenosis.

Effect of remote ischemic postconditioning on an intracerebral hemorrhage stroke model in rats

Abstract Background and purpose: While recent studies suggest that remote ischemic postconditioning (RIP) therapy may be of benefit to patients with acute ischemic stroke, RIP’s effects on intracerebral hemorrhage (ICH) still remains unclear. In the present study, the use of RIP in a rat model ICH was investigated to elucidate any potential beneficial or detrimental effects as determined by motor testing, blood brain barrier integrity, and brain water content, as well as aquaporin-4 (AQP-4) and matrix metalloproteinase-9 (MMP-9) expression. Methods: ICH was induced in Sprague–Dawley rats and they were randomized into either a control (nu200a=u200a24) or RIP treatment (nu200a=u200a24) group. RIP was performed by repetitive, brief occlusion and release of the bilateral femoral arteries. Functional outcome in each group was assessed by neurologic deficits on vibrissae-elicited forelimb placing test and a 12-point outcome scale. At 72 hours, brain blood volume, water content, blood–brain barrier (BBB) permeability, and protein expression of AQP-4 and MMP-9 were determined. Results: This collagenase model yielded well-defined striatal hematomas. Vibrissae-elicited forelimb placement was significantly (P<0·01) affected by ICH. However, there was no significant difference between the RIP and control groups at either 24 or 72 hours. A 12-point neurological deficit score also failed to differentiate between the RIP and control. There were no significant differences between the two groups in cerebral blood volumes, brain water content, Evans blue extravasations, and expressions of AQP-4 and MMP-9. Conclusions: Although RIP did not show a beneficial effect in our ICH model, treatment with RIP did not exacerbate ICH.

Reduced apoptosis by combining normobaric oxygenation with ethanol in transient ischemic stroke.

BACKGROUND AND PURPOSEnThe effect of normobaric oxygen (NBO) on apoptosis remains controversial. The present study evaluated the effect of NBO on ischemia-induced apoptosis and assessed the potential for improved outcomes by combining NBO administration with another neuroprotective agent, ethanol, in a rat stroke model.nnnMETHODSnRats were subjected to right middle cerebral artery occlusion (MCAO) for 2h. At the onset of reperfusion, ischemic animals received either NBO (2h duration), an intraperitoneal injection of ethanol (1.0g/kg), or both NBO and ethanol. Extent of brain injury was determined by infarct volume, neurological deficit, and apoptotic cell death. Expression of pro- and anti-apoptotic proteins was evaluated through Western immunoblotting.nnnRESULTSnGiven alone, NBO and ethanol each slightly (p<0.05) reduced infarct volume to 38% and 37%, respectively, as compared to the impressive reduction of 51% (p<0.01) seen with combined NBO-ethanol administration. Neurologic deficits were also significantly reduced by 48% with combined NBO-ethanol therapy, as compared to lesser reductions of 24% and 23% with NBO or ethanol, respectively. Combined NBO-ethanol therapy decreased apoptotic cell death by 49%, as compared to 31% with NBO and 30% with ethanol. Similarly, combination therapy significantly increased expression of anti-apoptotic factors (Bcl-2 and Bcl-xL) and significantly reduced expression of pro-apoptotic proteins (BAX, Caspase-3, and AIF), as compared to the minimal or nil protein expression changes elicited by NBO or ethanol alone.nnnCONCLUSIONSnIn rats subjected to ischemic stroke, NBO administration salvages ischemic brain tissue through evidenced decrease in apoptotic cell death. Combined NBO therapy with ethanol administration greatly improves both degree of neuroprotection and associated apoptosis.

Synergetic Neuroprotection of Normobaric Oxygenation and Ethanol in Ischemic Stroke Through Improved Oxidative Mechanism

Background and Purpose— Normobaric oxygenation (NBO) and ethanol both provide neuroprotection in stroke. We evaluated the enhanced neuroprotective effect of combining these 2 treatments in a rat stroke model. Methods— Sprague-Dawley rats were subjected to middle cerebral artery occlusion for 2 hours. Reperfusion was then established and followed by treatment with either (1) an intraperitoneal injection of ethanol (1.0 g/kg), (2) NBO treatment (2-hour duration), or (3) NBO plus ethanol. The extent of brain injury was determined by infarct volume and motor performance. Oxidative metabolism was determined by ADP/ATP ratios, reactive oxygen species levels, nicotinamide adenine dinucleotide phosphate oxidase activity, and pyruvate dehydrogenase activity. Protein expression of major nicotinamide adenine dinucleotide phosphate oxidase subunits (p47phox, gp91phox, and p67phox) and the enzyme pyruvate dehydrogenase was evaluated through Western immunoblotting. Results— NBO and ethanol monotherapies each demonstrated reductions as compared to stroke without treatment in infarct volume (36.7% and 37.9% vs 48.4%) and neurological deficits (score of 6.4 and 6.5 vs 8.4); however, the greatest neuroprotection (18.8% of infarct volume and 4.4 neurological deficit) was found in animals treated with combination therapy. This neuroprotection was associated with the largest reductions in ADP/ATP ratios, reactive oxygen species levels, and nicotinamide adenine dinucleotide phosphate oxidase activity, and the largest increase in pyruvate dehydrogenase activity. Conclusions— Combination therapy with NBO and ethanol enhances the neuroprotective effect produced by each therapy alone. The mechanism behind this synergistic action is related to changes in cellular metabolism after ischemia reperfusion. NBO plus ethanol is attractive for clinical study because of its ease of use, tolerability, and tremendous neuroprotective potential in stroke.

Local mild hypothermia induced by intra-arterial cold saline infusion prolongs the time window of onset of reperfusion injury after transient focal ischemia in rats

Abstract Objectives: The aims of this study were to determine the effects of intra-arterial local hypothermia on infarct volume in rats with different durations of ischemia and to determine whether hypothermia can prolong the therapeutic time window compared with reperfusion without hypothermia. Methods: Adult male Sprague-Dawley rats weighing 260–300 g were divided into control group (permanent MCA occlusion), normothermia groups (NT groups) and hypothermia groups (HT groups). NT groups included rats induced with blood reperfusion for 1.5, 2, 2.5 or 3 hour ischemia. In the HT groups with ischemia of 1.5, 2, 2.5 or 3 hours, 6 ml 20°C normal saline solution was flushed at a speed of 0.6 ml/min, beginning 10 minutes before blood reperfusion. The infarct volumes of brains stained by TTC were observed 48 hours later. Brain temperature, blood flow and neurological scores were also recorded during this procedure. Results: In the 1.5, 2, 2.5 and 3 hour ischemic groups, cold saline (20°C infusion via the MCA) rapidly reduced the temperature of the MCA-supplied ischemic territory in the cortex from 37.0–37.1 to 32.8–33.2°C and in the striatum from 37.3–37.5 to 33.2–33.3°C. In NT groups, the average total infarct volumes of 1.5 and 2 hour ischemia (29.80 ± 2.20 and 34.29 ± 2.14%, respectively) were significantly less than that of the control group (48.41 ± 5.82%), but the average total infarct volumes of the 2.5 and 3 hour ischemia groups (47.31 ± 4.72 and 50.17 ± 8.08%, respectively) did not change. Compared with the ischemia groups without local saline infusion, the average total infarct volumes of 1.5, 2 and 2.5 hours with local saline infusion to the ischemic territory (16.79 ± 2.51, 23.09 ± 4.63% and 25.19 ± 7.82%, respectively) decreased significantly, but the average total infarct volume of 3 hour ischemia with local saline infusion (43.30 ± 2.62%) was not different. Conclusion: Local cold saline infusion to the ischemic territory before reperfusion can lead to mild hypothermia of the ischemic territory and can prolong the therapeutic time window of reperfusion from 2 to 2.5 hours. Refinements of the cooling process, optimal target temperature, duration of the therapy and most importantly, clinical efficacy, require further study.