The expression of the protooncogenes, c-fos, jun B, c-jun, and jun D was investigated in a rat focal cerebral ischemia model by Northern analysis and in situ hybridization. Severe ischemia (reduction of regional blood flow by 88-92%) in this model is confined to cerebral cortex irrigated by the right middle cerebral artery. Ischemia for 30 minutes, which caused only slight cortical damage (infarct size, < 10 mm3), induced both jun B and c-fos mRNAs exclusively in the right cerebral cortex. Ischemia for 90 minutes, which led to large cortical infarction (infarct size, > 140 mm3), also induced the expression of these two genes in the right cerebral cortex as well as the ipsilateral hippocampus. The latter sustained very mild ischemia (reduction of regional blood flow by 10-20%). The coinduction of jun B and c-fos expression occurred immediately after reperfusion and peaked at 60 minutes after reperfusion. The expression of c-jun was enhanced in a similar pattern, but at a much lower magnitude. In contrast, no change in jun D expression was observed. Nuclear run-on assays indicated that the increase in c-fos, jun B, and c-jun mRNA levels was due to the increase of transcription rate in these genes. Mobility shift assays showed a basal DNA binding activity of transcription factor AP-1 in the right cerebral cortex. Ischemia for 30 or 90 minutes followed by reperfusion for 4 hours resulted in a four- to sixfold increase of AP-1 binding activity. The enhanced DNA binding activity persisted for as long as 24 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Background and Purpose-Angiogenesis occurs after cerebral ischemia, but the relationship between angiogenesis and cerebral hemodynamic change is unknown. The aim of the present study was to investigate the relationship between ischemia-induced angiogenesis and hemodynamics in a well-defined 3-vessel occlusion model of the rat by using diffusion-(DWI), perfusion-, and T2-weighted MRI (T2WI). Methods-Rats were subjected to 60 minutes of transient middle cerebral artery occlusion or sham operation. DWI and T2WI were used to characterize the extent of the ischemic lesion from 4.5 hours to 14 days after reperfusion. A flow-sensitive alternating inversion recovery method and dynamic susceptibility contrast MRI were used to evaluate the temporal changes in relative cerebral blood flow (CBF) and cerebral blood volume (CBV), respectively. Rats were randomly selected and killed at each time point for investigation of vascular density and for hematoxylin-eosin staining. Results-Ischemic lesions developed in the ipsilateral cortex, as demonstrated by DWI and T2WI. CBF was significantly increased in the ipsilateral cortex, especially in the cortical outer layer from day 1 to day 14, and peaked on day 7 (PϽ0.05), while CBV was significantly increased on day 7 (PϽ0.01). The vascular density on the ipsilateral brain surface was gradually increased from day 1 to day 5, peaked on day 7, and then decreased on day 14. Histology study showed pannecrosis in the cortex from day 1 to day 5 and partial liquefaction of the necrotic tissues on days 7 and 14. Conclusions-A delayed increase in both CBF and CBV is documented in the ipsilateral cortex after transient focal brain ischemia, and such an increase may be associated with angiogenesis.

To determine the involvement of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) in cytoprotection, we subjected N2-A cells to oxygen-glucose deprivation followed by reoxygenation (H-R). Following H-R insults, H(2)O(2) production was increased while cell viability declined, which was accompanied by loss of mitochondrial membrane potential (MMP), cytochrome c release, caspases 9 and 3 activation, poly(ADP-ribose)polymerase (PARP) cleavage and apoptosis. Rosiglitazone up to 5 microM protected cell viability, normalized MMP, and prevented apoptotic signals. The protective effect of rosiglitazone was abrogated by GW9662, a PPAR-gamma antagonist, or a specific PPAR-gamma small interference RNA (siRNA) but not a control scRNA. PPAR-gamma overexpression alone was effective in maintaining MMP and preventing apoptosis and its protective effect was also abrogated by PPAR-gamma siRNA or GW9662. To elucidate the mechanism by which PPAR-gamma protects MMP and prevents apoptosis, we analyzed Bcl-2, Bcl-xl, and phosphorylated Bad (p-Bad). H-R suppressed them. Rosiglitazone or PPAR-gamma overexpression restored them via PPAR-gamma. Rosiglitazone or PPAR-gamma overexpression preserved phosphorylated Akt and 3-phosphoinositide-dependent kinase-1 (PDK-1) in a PPAR-gamma dependent manner. These results indicate that ligand-activated PPAR-gamma protects N2-A cells against H-R damage by enhancing Bcl-2/Bcl-xl and maintaining p-Bad via preservation of p-Akt.

Stroke is a leading cause of adult disability and mortality. Diabetes is a major risk factor for stroke. Patients with diabetes have a higher incidence of stroke and a poorer prognosis after stroke. Peroxisome proliferator-activated receptor gamma (PPAR-gamma) is a ligand-modulated transcriptional factor and a therapeutic target for treating type II diabetes. It is well-documented that activation of PPAR-gamma can also attenuate postischemic inflammation and damage. In this review, we focus on the newly revealed anti-apoptotic actions of PPAR-gamma against cerebral ischemia. PPAR-gamma, by increasing superoxide dismutase/catalase and decreasing nicotinamide adenine dinucleotide phosphate oxidase levels, attenuated ischemia-induced reactive oxygen species and subsequently alleviated the postischemic degradation of Bcl-2, Bcl-xl, and Akt. The preserved Akt phosphorylated Bad. Meanwhile, PPAR-gamma also promotes the transcription of 14-3-3epsilon. Elevated 14-3-3epsilon binds and sequesters p-Bad and prevents Bad translocation to neutralize the anti-apoptotic function of Bcl-2. This review further supports the notion that PPAR-gamma may serve as a potential therapeutic target for treating ischemic stroke.

Galectins, β-galactoside-binding lectins, play multiple roles in the regulation of immune and inflammatory responses. The major galectins expressed in the CNS are galectins 1, 3, 4, 8, and 9. Under normal physiological conditions, galectins maintain CNS homeostasis by participating in neuronal myelination, neuronal stem cell proliferation, and apical vesicle transport in neuronal cells. In neuronal diseases and different experimental neuroinflammatory disease models, galectins may serve as extracellular mediators or intracellular regulators in controlling the inflammatory response or conferring the remodeling capacity in damaged CNS tissues. In general, galectins 1 and 9 attenuate experimental autoimmune encephalomyelitis (a model of multiple sclerosis), while galectin-3 promotes inflammation in this model. In brain ischemic lesions, both galectins 1 and 3 are induced to help neuronal regeneration. The expression of galectin-1 is required for astrocyte-derived neurotrophic factor secretion, and recombinant galectin-1 promotes neuronal regeneration. Galectin-3 promotes microglial cell proliferation and attenuates ischemic damage and neuronal apoptosis after cerebral ischemia. In amyotrophic lateral sclerosis models, galectin-3 is deleterious to neuroregeneration, while intramuscular administration of oxidized galectin-1 can improve neuromuscular disorders. In axotomy and Wallerian degeneration, galectin-3 helps phagocytosis of macrophages to clear degenerate myelin in the injured PNS or CNS. Thus, galectins are important modulators participating in homeostasis of the CNS and neuroinflammation. Continued investigations of the roles of galectins in neuroinflammation promise to provide a better understanding of the mechanism of this process and lead to new therapeutic approaches.

Hypoxia-inducible factor-1 (HIF-1) takes part in the transcriptional activation of hypoxia-responsive genes. HIF-1␣, a subunit of HIF-1, is rapidly degraded under normoxic conditions by the ubiquitin-proteosome system. Hypoxia up-regulates HIF-1␣ by inhibiting its degradation, thereby allowing it to accumulate to high levels with 3-6 h of hypoxia treatment and decreasing thereafter. In vascular tissues, prostacyclin (prostaglandin I 2 (PGI 2 )) is a potent vasodilator and inhibitor of platelet aggregation and is known as a vasoprotective molecule. However, the role of PGI 2 in HIF-1 activation has not been studied. In the present study, we investigated the effect of PGI 2 on HIF-1 regulation in human umbilical vein endothelial cells under prolonged hypoxia (12 h). Augmentation of PGI 2 via adenovirus-mediated gene transfer of both cyclooxygenase-1 and PGI 2 synthase activated HIF-1 by stabilizing HIF-1␣ in cells under prolonged hypoxia or the hypoxia-normoxia transition but not under normoxia. Exogenous H 2 O 2 abolished PGI 2 -and catalase-induced HIF-1␣ up-regulation, which suggests that degradation of HIF-1␣ under prolonged hypoxia is through a reactive oxygen species-dependent pathway. Moreover, PGI 2 attenuated NADPH oxidase activity by suppressing Rac1 and p47 phox expression under hypoxia. These data demonstrate a novel function of PGI 2 in down-regulating reactive oxygen species production by attenuating NADPH oxidase activity, which stabilizes HIF-1␣ in human umbilical vein endothelial cells exposed to prolonged hypoxia.Hypoxia induces a number of cellular responses, such as angiogenesis, erythropoiesis, and glycolysis, through both gene regulation and post-translational modification of proteins. Hypoxia-inducible factor-1 (HIF-1) 2 takes part in the transcriptional activation of hypoxia-responsive genes through binding to the hypoxia-responsive element (HRE) in the promoter or enhancer regions and activating a number of genes (1-4). HIF-1 is a heterodimer composed of HIF-1␣ and HIF-1␤. HIF-1␣ is rapidly degraded under normoxic conditions by the ubiquitin-proteosome system, whereas HIF-1␤ is constitutively expressed (5). Under hypoxia, HIF-1␣ has been shown to be up-regulated to high levels at 3-6 h and decrease thereafter (6, 7). A number of studies have focused on the mechanism of HIF-1␣ stabilization under hypoxia for 4 -6 h. Recently, a natural antisense HIF-1␣ was suggested to down-regulate HIF-1␣ in A549 cells under prolonged hypoxia (6). However, regulation of HIF-1␣ under prolonged hypoxia (12 h) in endothelial cells remains largely unknown.Under normoxic conditions, HIF-1␣ is regulated through hydroxylation of proline residues by prolyl hydroxylase enzymes (8, 9). The von Hippel-Lindau tumor suppressor protein (pVHL) associates with hydroxylated HIF-1␣ and targets it for ubiquitination and rapid degradation (10). Under hypoxia, prolyl hydroxylase is inactivated through oxygen-sensing mechanisms, and the unmodified HIF-1␣ accumulates, permitting dimerization with HIF-1␤ (3, 11). Various redox-de...

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