A polymorphism in the promoter region of the survivin gene is related to hemorrhagic transformation in patients with acute ischemic stroke.

Hemorrhagic transformation (HT) of cerebral infarction is a common and serious occurrence following acute ischemic stroke. The expression of survivin, a member of the inhibitor of apoptosis protein family, has been shown to increase after cerebral ischemia. This protein has been mainly located at the microvasculature within the infarcted and peri-infarcted area, so we aimed to investigate whether survivin gene polymorphisms, also known as BIRC5 gene, were associated with HT of cerebral infarction. Polymorphism screening of the BIRC5 gene was performed in 107 patients with a hemispheric ischemic stroke and 93 controls by polymerase chain reaction, single-strand conformation polymorphism and sequencing analysis. Genotype-phenotype correlation was performed in patients. MRI was carried out within 12 h of symptoms onset and at 72 ± 12 h. The presence of HT was determined on the second DWI sequence and classified according to ECASS II criteria. MMP-9 levels were analyzed at admission. Forty-nine patients (45.8%) had HT. The -241 C/T (rs17878467) polymorphism was identified in the promoter region of the survivin gene. The prevalence of the mutant allele (T) was similar in patients and controls (14 vs. 16%, respectively; P = 0.37). However, 9 (29%) patients with allele T had HT compared to 40 (52.6%) of wild-type (P = 0.021). Logistic regression analysis showed that the polymorphism was associated with a lower risk of HT (OR 0.16; 95% CI 0.04-0.65; P = 0.01). The -241 C/T polymorphism in the promoter region of the survivin gene is associated with a lower risk of HT in patients with ischemic stroke. It has recently been reported that the -241 C/T polymorphism increases survivin promoter activity, reinforcing the hypothesis that patients with the mutant allele may have increased survivin expression in the brain. Different mechanisms, including BBB protection by the inhibition or activation of different angiogenic growth factors and the inhibition of apoptosis during angiogenesis, may explain the protective effect of this polymorphism on HT development in ischemic stroke. Further studies are needed to confirm our results and elucidate the mechanisms explaining this effect.

Oct-2 transcription factor binding activity and expression up-regulation in rat cerebral ischaemia is associated with a diminution of neuronal damage in vitro.

Brain plasticity provides a mechanism to compensate for lesions produced as a result of stroke. The present study aims to identify new transcription factors (TFs) following focal cerebral ischaemia in rat as potential therapeutic targets. A transient focal cerebral ischaemia model was used for TF-binding activity and TF-TF interaction profile analysis. A permanent focal cerebral ischaemia model was used for the transcript gene analysis and for the protein study. The identification of TF variants, mRNA analysis, and protein study was performed using conventional polymerase chain reaction (PCR), qPCR, and Western blot and immunofluorescence, respectively. Rat cortical neurons were transfected with small interfering RNA against the TF in order to study its role. The TF-binding analysis revealed a differential binding activity of the octamer family in ischaemic brain in comparison with the control brain samples both in acute and late phases. In this study, we focused on Oct-2 TF. Five of the six putative Oct-2 transcript variants are expressed in both control and ischaemic rat brain, showing a significant increase in the late phase of ischaemia. Oct-2 protein showed neuronal localisation both in control and ischaemic rat brain cortical slices. Functional studies revealed that Oct-2 interacts with TFs involved in important brain processes (neuronal and vascular development) and basic cellular functions and that Oct-2 knockdown promotes neuronal injury. The present study shows that Oct-2 expression and binding activity increase in the late phase of cerebral ischaemia and finds Oct-2 to be involved in reducing ischaemic-mediated neuronal injury.

miRNA expression is modulated over time after focal ischaemia: up-regulation of miR-347 promotes neuronal apoptosis.

Despite the large number of molecules reported as being over-expressed after ischaemia, little is known regarding their regulation. miRNAs are potent post-transcriptional regulators of gene expression, and reports have shown differentially miRNA expression in response to focal cerebral ischaemia. The present study analysed miRNA expression from acute to late phases of ischaemia to identify specific ischaemia-related miRNAs, elucidate their role, and identify potential targets involved in stroke pathophysiology. Of 112 miRNAs, 32 showed significant changes and different expression profiles. In addition to the previously reported differentially expressed miRNAs, new ischaemia-regulated miRNAs have been found, including miR-347. Forty-seven genes involved in brain functions or related to ischaemia are predicted to be potential targets of the differentially expressed miRNAs after middle cerebral artery occlusion. Analysis of four of these targets (Acsl4, Arf3, Btg2 and Dpysl5) showed them to be differentially regulated by ischaemia at the transcriptional or post-transcriptional level. Acsl4, Bnip3l and Phyhip, potential targets of miR-347, were up-regulated after miR-347 over-expression, inducing neuronal apoptotic death. Our findings suggest that miR-347 plays an important role in regulating neuronal cell death, identify Acsl4 as a new protein requiring study in ischaemia, and provide an important resource for future functional studies of miRNAs after ischaemia.

Experimental models for assaying microvascular endothelial cell pathophysiology in stroke.

It is important to understand the molecular mechanisms underlying neuron death following stroke in order to develop effective neuroprotective strategies. Since studies on human stroke are extremely limited due to the difficulty in collecting post-mortem tissue at different time points after the onset of stroke, brain ischaemia research focuses on information derived from in-vitro models of neuronal death through ischaemic injury [1]. This review aims to provide an update on the different in-vitro stroke models with brain microvascular endothelial cells that are currently being used. These models provide a physiologically relevant tool to screen potential neuroprotective drugs in stroke and to study the molecular mechanisms involved in brain ischaemia.