Interest in histone deacetylase (HDAC)-based therapeutics as a potential treatment for stroke has grown dramatically. The neuroprotection of HDAC inhibition may involve multiple mechanisms, including modulation of transcription factor acetylation independent of histones. The transcription factor Nrf2 has been shown to be protective in stroke as a key regulator of antioxidant-responsive genes. Here, we hypothesized that HDAC inhibition might provide neuroprotection against mouse cerebral ischemia by activating the Nrf2 pathway. We determined that the classic HDAC inhibitor trichostatin A increased neuronal cell viability after oxygen-glucose deprivation (from an OD value of 0.10±0.01 to 0.25±0.08) and reduced infarct volume in wild-type mice with stroke (from 49.1±3.8 to 21.3±4.6%). In vitro studies showed that HDAC inhibition reduced Nrf2 suppressor Keap1 expression, induced Keap1/Nrf2 dissociation, Nrf2 nuclear translocation, and Nrf2 binding to antioxidant response elements in heme oxygenase 1 (HO1), and caused HO1 transcription. Furthermore, we demonstrated that HDAC inhibition upregulated proteins downstream of Nrf2, including HO1, NAD(P)H:quinone oxidoreductase 1, and glutamate-cysteine ligase catalytic subunit in neuron cultures and brain tissue. Finally, unlike wild-type mice, Nrf2-deficient mice were not protected by pharmacologic inhibition of HDAC after cerebral ischemia. Our studies suggest that activation of Nrf2 might be an important mechanism by which HDAC inhibition provides neuroprotection.

The clinical side effects associated with the inhibition of cyclooxygenase enzymes under pathologic conditions have recently raised concerns. A better understanding of neuroinflammatory mechanisms and neuronal survival requires knowledge of cyclooxygenase downstream pathways, especially PGE2 and its G-protein-coupled receptors. In this study, we postulate that EP1 receptor is one of the mechanisms that propagate neurotoxicity and could be a therapeutic target in brain injury. This hypothesis was tested by pretreating C57BL/6 wildtype mice with the EP1 receptor selective agonist ONO-DI-004 and the selective antagonist ONO-8713, followed by striatal unilateral NMDA injection. Results revealed that ONO-DI-004 increased NMDA-induced lesion volume up to 128.7 +/- 12.0%, while ONO-8713 significantly decreased lesion volume to 71.3 +/- 10.9%, as compared to the NMDA-control group. Neurotoxic EP1 receptor properties were also studied using C57BL/6 EP1 receptor knockout (EP1-/-) mice, which revealed a significant decrease to 74.5 +/- 8.2%, as compared to wildtype controls. The protective effect of the antagonist ONO-8713 was also tested in the EP1-/- mice, revealing no additional protection in these mice. Together, these results support the selectivity of ONO-8713 toward EP1 receptor and suggest the neurotoxic role of EP1 receptor. Furthermore, the EP1 receptor role in ischemic brain damage was investigated using a model of middle cerebral artery occlusion (MCAO) and reperfusion. The infarct volume was significantly reduced to 56.9 +/- 11.5% in EP1-/- mice, as compared to wildtype controls. This is the first study that demonstrates that EP1 receptor aggravates neurotoxicity and that modulation of this receptor can determine the outcomes in both excitotoxic and focal ischemic neuronal damage.

No abstract

No abstract

Hemoproteins undergo degradation during hypoxic/ischemic conditions, but the pro-oxidant free heme that is released cannot be recycled and must be degraded. The extracellular heme associates with its high-affinity binding protein, hemopexin (HPX). Hemopexin is shown here to be expressed by cortical neurons and it is present in mouse cerebellum, cortex, hippocampus, and striatum. Using the transient ischemia model (90-min middle cerebral artery occlusion followed by 96-h survival), we provide evidence that HPX is protective in the brain, as neurologic deficits and infarct volumes were significantly greater in HPX−/− than in wild-type mice. Addressing the potential protective HPX cellular pathway, we observed that exogenous free heme decreased cell survival in primary mouse cortical neuron cultures, whereas the heme bound to HPX was not toxic. Heme–HPX complexes induce HO1 and, consequently, protect primary neurons against the toxicity of both heme and prooxidant tert-butyl hydroperoxide; such protection was decreased in HO1−/− neuronal cultures. Taken together, these data show that HPX protects against heme-induced toxicity and oxidative stress and that HO1 is required. We propose that the heme–HPX system protects against stroke-related damage by maintaining a tight balance between free and bound heme. Thus, regulating extracellular free heme levels, such as with HPX, could be neuroprotective.

6-Hydroxydopamine (6-OHDA) is one of the most widely used rat models for Parkinson's disease. There is ample evidence in the literature that 6-OHDA elicits its toxic manifestations through oxidant stress. In the present study, we evaluated the anti-parkinsonian effects of Withania somnifera extract, which has been reported to have potent anti-oxidant, anti-peroxidative and free radical quenching properties in various diseased conditions. Rats were pretreated with 100, 200 and 300 mg/kg b.w. of the W. somnifera extract orally for 3 weeks. On day 21, 2 mL of 6-OHDA (10 mg in 0.1% in ascorbic acidsaline) was infused into the right striatum while sham operated group received 2 mL of the vehicle. Three weeks after 6-OHDA injections, rats were tested for neurobehavioral activity and were killed 5 weeks after lesioning for the estimation of lipidperoxidation, reduced glutathione content, activities of glutathione-S-transferase, glutathione reductase, glutathione peroxidase, superoxide dismutase and catalase, catecholamine content, dopaminergic D2 receptor binding and tyrosine hydroxylase expression. W. somnifera extract was found to reverse all the parameters significantly in a dose-dependent manner. Thus, the study demonstrates that the extract of W. somnifera may be helpful in protecting the neuronal injury in Parkinson's disease.

Background and Purpose Ginkgo biloba extracts are now prescribed in several countries for their reported health benefits, particularly for medicinal properties in the brain. The standardized Ginkgo extract, EGb761, has been reported to protect neurons against oxidative stress, but the underlying mechanisms are not fully understood. Methods To characterize the oral consumption of EGb761 in transient ischemia, we performed the middle cerebral artery occlusion (MCAO) filament model in wild-type and heme oxygenase 1 (HO-1) knockouts. Mice were pretreated for 7 days before the transient occlusion or posttreated acutely during reperfusion; then neurobehavioral scores and infarct volumes were assessed. Furthermore, primary cortical neuronal cultures were used to investigate the contribution of the antioxidant enzyme HO-1 in the EGb761-associated cytoprotection. Results Mice that were pretreated with EGb761 had 50.9±5.6% less neurological dysfunction and 48.2±5.3% smaller infarct volumes than vehicle-treated mice; this effect was abolished in HO-1 knockouts. In addition to the prophylactic properties of EGb761, acute posttreatment 5 minutes and 4.5 hours after reperfusion also led to significant reduction in infarct size (P<0.01). After our previous demonstration that EGb761 significantly induced HO-1 levels in a dose- and time-dependent manner in neuronal cultures, here we revealed that this de novo HO-1 induction was required for neuroprotection against free radical damage and excitotoxicity as it was significantly attenuated by the enzyme inhibitor. Conclusion These results demonstrate that EGb761 could be used as a preventive or therapeutic agent in cerebral ischemia and suggest that HO-1 contributes, at least in part, to EGb761 neuroprotection.

SUMMARY Redox-mediated posttranslational modifications represent a molecular switch that controls major mechanisms of cell function. Nitric oxide (NO) can mediate redox reactions via S-nitrosylation, representing transfer of an NO group to a critical protein thiol. NO is known to modulate neurogenesis and neuronal survival in various brain regions in disparate neurodegenerative conditions. However, a unifying molecular mechanism linking these phenomena remains unknown. Here we report that S-nitrosylation of myocyte enhancer factor 2 (MEF2) transcription factors acts as a redox switch to inhibit both neurogenesis and neuronal survival. Structure-based analysis reveals that MEF2 dimerization creates a pocket, facilitating S-nitrosylation at an evolutionally conserved cysteine residue in the DNA binding domain. S-Nitrosylation disrupts MEF2-DNA binding and transcriptional activity, leading to impaired neurogenesis and survival in vitro and in vivo. Our data define a novel molecular switch whereby redox-mediated posttranslational modification controls both neurogenesis and neurodegeneration via a single transcriptional signaling cascade.