Ischemic penumbra in retina endures： vascular neuropathology is reconciled

A physiological state of＂normoxia＂obtains when tissue oxygen tension（pO_2）is sufficient to drive mitochondrial respiration throughout a volume of cells.Between 30-40%of the available oxygen is normally extracted from hemoglobin as it passes through neural tissue.

Involvement of Apoptosis in 3-nitropropionic Acid-induced Ischemic Tolerance to Transient Focal Cerebral Ischemia in Rats

The involvement of apoptosis in mitochondrial toxin 3 nitropropionic acid (3 NPA) induced ischemic tolerance to transient focal cerebral ischemia in rats and the mechanism was investigated. 3 NPA at a dose of 20 mg/kg or vehicle control was intraperitoneally into the rats. Three days later, rats were exposed to 2 h of middle cerebral artery occlusion followed by 24 h of reperfusion. Infarct volumes were assessed by 2,3,5 triphenyltetrazolinm chloride (TTC) staining 24 h after reperfusion. Neural cell apoptosis in cerebral ischemic penumbra was detected by terminal deoxynucleotidyl transferase mediated dUTP biotin in situ nick end labeling (TUNEL) and flow cytometry methods (FCM). The results showed that as compared to the vehicle treated group, pretreatment with 3 NPA could reduce the infarct volume by 23.3 % and decrease the number of TUNEL positive neural cells and apoptotic percentage by 47 % ( P< 0.05) and 44.9 % ( P< 0 01), respectively. It was concluded that the development of 3 NPA induced ischemic tolerance in brain might be related to the decreases in neural cell apoptosis.

URB597 exerts neuroprotective effect against transient cerebral ischemic injury via restor⁃ing autophagy flux and inhibiting neuronal necroptosis in mice

OBJECTIVE URB597(KDS-4103)is a potent and selective inhibitor of the enzyme fatty acid amide hydrolase(FAAH)and can ele⁃vate the level of oleoylethanolamide(OEA),a naturally occurring endocannabinoid in the brain.However,the effect of URB597 on cerebral isch⁃emic injury in mice remains unclear.METHODS Focal cerebral ischemia was induced by middle cerebral artery occlusion for 30 min followed by reperfusion for 24 h in mice.To observe the dose-dependent effect,URB597(0.04-5.00 mg·kg-1,ip)was administered at the same time of reperfu⁃sion.To determine the time-dependent effect,URB597(1.00 mg·kg-1,ip)was administered as a single dose at 0,1,3 or 5 h after reperfusion.Twenty-four hours after brain ischemia,Beder⁃son scoring test and grip strength test were used to evaluate the neurological function;brain in⁃farct volume was assayed by 2,3,5-triphenyltetra⁃zolium chloride(TTC)staining or diffusion-weighted magnetic resonance imaging(MRI).Laser speckle imaging(LSI)technique was used to assay the regional cerebral blood flow(rCBF);NeuN immunofluorescence staining was used to observe the neuron survival in the penumbra.To further investigate the underlying mechanism,au⁃tophagy flux related proteins(LC3-Ⅱ,P62 and LAMP2)and necroptosis related proteins(pRIPK3 and pMLKL)were detected by Western blotting and immunofluorescence staining.RESULTS Twenty-four hours after brain ischemia,URB597 dose-dependently improved neurological func⁃tion and reduced brain infarct volume.The most effective dose was 1.00 mg·kg-1;the therapeutic time window was within 1 h after ischemic stroke.The protective effect is further confirmed by the results that post-ischemic treatment with URB597(1.00 mg·kg-1)significantly increased neurons survival,promoted autophagy flux and reduced cell necroptosis in cortical penumbra after cerebral I/R.CONCLUSION URB597 dose-and time-dependently exerts a neuroprotective effect against acute cerebral I/R injury.This neu⁃roprotective effect of URB597 may be associated with its restoration of autophagy flux and inhibi⁃tion of neuronal necroptosis in the cortical penumbra.

Computed tomography perfusion and computed tomography angiography for prediction of clinical outcomes in ischemic stroke patients after thrombolysis

Cerebral blood perfusion and cerebrovascular lesions are important factors that can affect the therapeutic efficacy of thrombolysis.At present,the majority of studies focus on assessing the accuracy of lesion location using imaging methods before treatment,with less attention to predictions of outcomes after thrombolysis.Thus,in the present study,we assessed the efficacy of combined computed tomography（CT） perfusion and CT angiography in predicting clinical outcomes after thrombolysis in ischemic stroke patients.The study included 52 patients who received both CT perfusion and CT angiography.Patients were grouped based on the following criteria to compare clinical outcomes：（1） thrombolytic and non-thrombolytic patients,（2） thrombolytic patients with CT angiography showing the presence or absence of a vascular stenosis,（3） thrombolytic patients with CT perfusion showing the presence or absence of hemodynamic mismatch,and（4） different CT angiography and CT perfusion results.Short-term outcome was assessed by the 24-hour National Institution of Health Stroke Scale score change.Long-term outcome was assessed by the 3-month modified Rankin Scale score.Of 52 ischemic stroke patients,29 were treated with thrombolysis and exhibited improved short-term outcomes compared with those without thrombolysis treatment（23 patients）.Patients with both vascular stenosis and blood flow mismatch（13 patients） exhibited the best short-term outcome,while there was no correlation of long-term outcome with CT angiography or CT perfusion findings.These data suggest that combined CT perfusion and CT angiography are useful for predicting short-term outcome,but not long-term outcome,after thrombolysis.

Cerebroprotection with recombinant neuroglobin plasmid in a rat model of focal cerebral ischemia

BACKGROUND： Adenovirus has been used to develop neuroglobin （Ngb） vectors. Although transfection efficiency is high, induced gene mutation, cytotoxicity, inflammation, and low exogenous gene content have limited its application. OBJECTIVE： To observe the effects of recombinant Ngb plasmid in a rat model of focal cerebral ischemia. DESIGN, TIME AND SETTING： Genetically engineered, randomized, controlled, animal experiment was performed at the Laboratory of Chongqing Medical University from May 2006 and January 2007. MATERIALS： 2, 3, 5-triphenyltetrazolium chloride was purchased from Shanghai Sangon Biological Engineering Technology and Services. Rabbit anti-rat Bcl-2 polyclonal antibody, rabbit anti-rat β-actin monoclonal antibody, and FITC-labeled goat anti-rabbit IgG were purchased from Sigma, USA. TUNEL apoptosis kit was purchased from Roche, Germany. METHODS： A total of 54 male, adult, Wistar rats were randomly assigned to 3 groups （n=18）： normal saline, plasmid control, and recombinant Ngb （pCDNA3.1 （＋）/Ngb）. Normal saline, plasmid pCDNA3.1 （＋）, and recombinant plasmid pCDNA3.1 （＋）/Ngb were separately injected into two sites in the rat cerebral cortex, and models of focal ischemia were established by occlusion of the right middle cerebral artery after 24 hours. MAIN OUTCOME MEASURES： Local ischemic damage was detected by 2, 3, 5- triphenyltetrazolium chloride staining, apoptosis in the penumbra was confirmed using the TUNEL method, and Bcl-2 protein expression in the penumbra was determined by indirect immunofluorescent staining and Western blot analysis. RESULTS： Compared with the normal saline and plasmid control groups, cerebral infarction size and the number of apoptotic cells in the pCDNA3.1 （＋）/Ngb group were significantly reduced （P 〈 0.01）. The percentage of Bcl-2-positive cells in the penumbra of the pCDNA3.1 （＋）/Ngb group was significantly increased （P 〈 0.01）. The relative expression level of Bcl-2 protein was increased by 40%-50%. CONCLUSION： Recombinant plasmid pCDNA3.1/Ngb provides neuroprotection by upregulating Bcl-2 expression and inhibiting cell apoptosis in the penumbra.

Rethinking the necessity of low glucose intervention for cerebral ischemia/reperfusion injury

Glucose is the essential and almost exclusive metabolic fuel for the brain.Ischemic stroke caused by a blockage in one or more cerebral arteries quickly leads to a lack of regional cerebral blood supply resulting in severe glucose deprivation with subsequent induction of cellular homeostasis disturbance and eventual neuronal death.To make up ischemiamediated adenosine 5′-triphosphate depletion,glucose in the ischemic penumbra area rapidly enters anaerobic metabolism to produce glycolytic adenosine 5′-triphosphate for cell survival.It appears that an increase in glucose in the ischemic brain would exert favorable effects.This notion is supported by in vitro studies,but generally denied by most in vivo studies.Clinical studies to manage increased blood glucose levels after stroke also failed to show any benefits or even brought out harmful effects while elevated admission blood glucose concentrations frequently correlated with poor outcomes.Surprisingly,strict glycaemic control in clinical practice also failed to yield any beneficial outcome.These controversial results from glucose management studies during the past three decades remain a challenging question of whether glucose intervention is needed for ischemic stroke care.This review provides a brief overview of the roles of cerebral glucose under normal and ischemic conditions and the results of managing glucose levels in non-diabetic patients.Moreover,the relationship between blood glucose and cerebral glucose during the ischemia/reperfusion processes and the potential benefits of low glucose supplements for non-diabetic patients are discussed.

Therapeutic imaging window of cerebral infarction revealed by multisequence magnetic resonance imaging An animal and clinical study

In this study, we established a Wistar rat model of right middle cerebral artery occlusion and observed pathological imaging changes （T2-weighted imaging [T2WI], T2FLAIR, and diffusion-weighted imaging [DWI]） following cerebral infarction. The pathological changes were divided into three phases： early cerebral infarction, middle cerebral infarction, and late cerebral infarction. In the early cerebral infarction phase （less than 2 hours post-infarction）, there was evidence of intracellular edema, which improved after reperfusion. This improvement was defined as the ischemic penumbra. In this phase, a high DWI signal and a low apparent diffusion coefficient were observed in the right basal ganglia region. By contrast, there were no abnormal T2WI and T2FLAIR signals. For the middle cerebral infarction phase （2-4 hours post-infarction）, a mixed edema was observed. After reperfusion, there was a mild improvement in cell edema, while the angioedema became more serious. A high DWI signal and a low apparent diffusion coefficient signal were observed, and some rats showed high T2WI and T2FLAIR signals. For the late cerebral infarction phase （4-6 hours post-infarction）, significant angioedema was visible in the infarction site. After reperfusion, there was a significant increase in angioedema, while there was evidence of hemorrhage and necrosis. A mixed signal was observed on DWI, while a high apparent diffusion coefficient signal, a high T2WI signal, and a high T2FLAIR signal were also observed. All 86 cerebral infarction patients were subjected to T2WI, T2FLAIR, and DWI. MRI results of clinic data similar to the early infarction phase of animal experiments were found in 51 patients, for which 10 patients （10/51） had an onset time greater than 6 hours. A total of 35 patients had MRI results similar to the middle and late infarction phase of animal experiments, of which eight patients （8/35） had an onset time less than 6 hours. These data suggest that defining the ＂therapeutic time window＂ as the time 6 hours after infarction may not be suitable for all patients. Integrated application of MRI sequences including T2WI, T2FLAIR, DW-MRI, and apparent diffusion coefficient mapping should be used to examine the ischemic penumbra, which may provide valuable information for identifying the ＂therapeutic time window＂.

Progesterone is neuroprotective by inhibiting cerebral edema after ischemia

Ischemic edema can alter the structure and permeability of the blood-brain barrier. Recent studies have reported that progesterone reduces cerebral edema after cerebral ischemia. However, the underlying mechanism of this effect has not yet been elucidated. In the present study, progesterone effectively reduced Evans blue extravasation in the ischemic penumbra, but not in the ischemic core, 48 hours after cerebral ischemia in rats. Progesterone also inhibited the down-regulation of gene and protein levels of occludin and zonula occludens-1 in the penumbra. These results indicate that progesterone may effectively inhibit the down-regulation of tight junctions, thereby maintaining the integrity of the blood-brain barrier and reducing cerebral edema.

Resveratrol provides neuroprotection by regulating the JAK2/STAT3/PI3K/AKT/mTOR pathway after stroke in rats

Ischemic stroke is a common disease with high mortality and morbidity worldwide.One of the important pathophysiological effects of ischemic stroke is apoptosis.A neuroprotective effect is defined as the inhibition of neuronal apoptosis to rescue or delay the infarction in the surviving ischemic penumbra.Resveratrol is a natural polyphenol that reportedly prevents cerebral ischemia injury by regulating the expression of PI3K/AKT/mTOR.Therefore,this study aimed to elucidate the neuroprotective effect of resveratrol on cerebral ischemia/reperfusion injury and to investigate the signaling pathways and mechanisms through which resveratrol regulates apoptosis in the ischemic penumbra.Rats were subjected to middle cerebral artery occlusion for 2 h followed by 24 h reperfusion.Cerebral infarct volume was measured using 2%TTC staining.TUNEL staining was conducted to evaluate neuronal apoptosis.Western blotting and immunohistochemistry were used to detect the proteins involved in the JAK2/STAT3/PI3K/AKT/mTOR pathway.The results suggested that resveratrol significantly improved neurological function,reduced cerebral infarct volume,decreased neuronal damage,and markedly attenuated neuronal apoptosis;these effects were attenuated by the inhibition of PI3K/AKT with LY294002 and JAK2/STAT3 with AG490.We also found that resveratrol significantly upregulated the expression of p-JAK2,p-STAT3,p-AKT,p-mTOR,and BCL-2 and downregulated expression of cleaved caspase-3 and BAX,which was partially reversed by LY294002 and AG490.These results suggested that resveratrol provides a neuroprotective effect against cerebral ischemia/reperfusion injury,which is partially mediated by the activation of JAK2/STAT3 and PI3K/AKT/mTOR.Resveratrol may indirectly upregulate the PI3K/AKT/mTOR pathway by activating JAK2/STAT3.

Quantitative Measurement of Cerebral Perfusion with Intravoxel Incoherent Motion in Acute Ischemia Stroke： Initial Clinical Experience

Background：Intravoxel incoherent motion （IVIM） has the potential to provide both diffusion and perfusion information without an exogenous contrast agent,its application for the brain is promising,however,feasibility studies on this are relatively scarce.The aim of this study is to assess the feasibility of IVIM perfusion in patients with acute ischemic stroke （AIS）.Methods：Patients with suspected AIS were examined by magnetic resonance imaging within 24 h of symptom onset.Fifteen patients （mean age was 68.7 ± 8.0 years） who underwent arterial spin labeling （ASL） and diffusion-weighted imaging （DWI） were identified as having AIS with ischemic penumbra were enrolled,where ischemic penumbra referred to the mismatch areas of ASL and DWI.Eleven different b-values were applied in the biexponential model.Regions of interest were selected in ischemic penumbras and contralateral normal brain regions.Fast apparent diffusion coefficients （ADCs） and ASL cerebral blood flow （CBF） were measured.The paired t-test was applied to compare ASL CBF,fast ADC,and slow ADC measurements between ischemic penumbras and contralateral normal brain regions.Linear regression and Pearson＇s correlation were used to evaluate the correlations among quantitative results.Results：The fast ADCs and ASL CBFs of ischemic penumbras were significantly lower than those of the contralateral normal brain regions （1.93 ± 0.78 μm2/ms vs.3.97 ± 2.49 μm2/ms,P =0.007;13.5 ± 4.5 ml· 100 g-1 ·min-1 vs.29.1 ± 12.7 ml·100 g-1 ·min-1,P ＜ 0.001,respectively）.No significant difference was observed in slow ADCs between ischemic penumbras and contralateral normal brain regions （0.203 ± 0.090 μm2/ms vs.0.198 ± 0.100 μm2/ms,P =0.451）.Compared with contralateral normal brain regions,both CBFs and fast ADCs decreased in ischemic penumbras while slow ADCs remained the same.A significant correlation was detected between fast ADCs and ASL CBFs （r =0.416,P ＜ 0.05）.No statistically significant correlation was observed between ASL CBFs and slow ADCs,or between fast ADCs and slow ADCs （r =0.111,P =0.558;r =0.200,P =0.289,respectively）.Conclusions：The decrease in cerebral blood perfusion primarily results in the decrease in fast ADC in ischemic penumbras;therefore,fast ADC can reflect the perfusion situation in cerebral tissues.