9-Cis-retinoic acid reduces ischemic brain injury in rodents via bone morphogenetic protein.

Retinoic acid (RA), a biologically active derivative of vitamin A, has protective effects against damage caused by H(2)O(2) or oxygen-glucose deprivation in mesangial and PC12 cells. In cultured human osteosarcoma cells, RA enhances the expression of bone morphogenetic protein-7 (BMP7), a trophic factor that reduces ischemia- or neurotoxin-mediated neurodegeneration in vivo. The purpose of this study is to examine whether RA reduces ischemic brain injury through a BMP7 mechanism. We found that intracerebroventricular administration of 9-cis-retinoic acid (9cRA) enhanced BMP7 mRNA expression, detected by RT-PCR, in rat cerebral cortex at 24 hr after injection. Rats were also subjected to transient focal ischemia induced by ligation of the middle cerebral artery (MCA) at 1 day after 9cRA injection. Pretreatment with 9cRA increased locomotor activity and attenuated neurological deficits 2 days after MCA ligation. 9cRA also reduced cerebral infarction and TUNEL labeling. These protective responses were antagonized by the BMP antagonist noggin given 1 day after 9cRA injection. Taken together, our data suggest that 9cRA has protective effects against ischemia-induced injury, and these effects involve BMPs.

Neuroregenerative effects of BMP7 after stroke in rats.

Previous reports have indicated that the expression of bone morphogenetic protein-7 (BMP7) is enhanced after ischemic injury in brain. This upregulation may induce endogenous neurorepair in the ischemic brain. The purpose of this study was to examine neuroregenerative effects of BMP7 after ischemia-reperfusion injury. Adult Sprague-Dawley rats were anesthetized with chloral hydrate. Right middle cerebral artery (MCA) was transiently ligated with 10-O suture for 1 h. One day after MCA occlusion, vehicle or BMP7 was infused to the contralateral cerebral ventricle. To identify possible neurogenesis, bromodeoxyurindine (BrdU) was systemically injected on the fourth and fifth days after MCA occlusion. Animals treated with BMP7 showed a rapid correction of body asymmetry and neurological deficits, suggesting BMP7 facilitates recovery after stroke. Animals were sacrificed at 1 month after stroke and brains were analyzed using immunohistological techniques. BMP7 treatment enhanced immunoreactivity of BrdU in the subventricular zone, lesioned cortex, and corpus callosum. These BrdU-positive cells co-labeled with nestin and NeuN. Our behavioral and anatomical data suggest that BMP7 promotes neuroregeneration in stroke animals, possibly through the proliferation of new neuronal precursors after ischemia.

Dietary supplementation with blueberries, spinach, or spirulina reduces ischemic brain damage.

Free radicals are involved in neurodegenerative disorders, such as ischemia and aging. We have previously demonstrated that treatment with diets enriched with blueberry, spinach, or spirulina have been shown to reduce neurodegenerative changes in aged animals. The purpose of this study was to determine if these diets have neuroprotective effects in focal ischemic brain. Adult male Sprague-Dawley rats were fed with equal amounts of diets (blueberry, spinach, and spirulina) or with control diet. After 4 weeks of feeding, all animals were anesthetized with chloral hydrate. The right middle cerebral artery was ligated with a 10-O suture for 60 min. The ligature was later removed to allow reperfusional injury. Animals were sacrificed and brains were removed for caspase-3 enzymatic assays and triphenyltetrazolium chloride staining at 8 and 48 h after the onset of reperfusion. A subgroup of animals was used for locomotor behavior and biochemical assays. We found that animals which received blueberry, spinach, or spirulina enriched diets had a significant reduction in the volume of infarction in the cerebral cortex and an increase in post-stroke locomotor activity. There was no difference in blood biochemistry, blood CO2, and electrolyte levels among all groups, suggesting that the protection was not indirectly mediated through the changes in physiological functions. Animals treated with blueberry, spinach, or spirulina had significantly lower caspase-3 activity in the ischemic hemisphere. In conclusion, our data suggest that chronic treatment with blueberry, spinach, or spirulina reduces ischemia/reperfusion-induced apoptosis and cerebral infarction.

Diadenosine tetraphosphate protects against injuries induced by ischemia and 6-hydroxydopamine in rat brain.

Diadenosine tetraphosphate (AP4A), an endogenous diadenosine polyphosphate, reduces ischemic injury in the heart. In this study, we report the potent and protective effects of AP4A in rodent models of stroke and Parkinson's disease. AP4A, given intracerebroventricularly before middle cerebral artery (MCA) ligation, reduced cerebral infarction size and enhanced locomotor activity in adult rats. The intravenous administration of AP4A also induced protection when given early after MCA ligation. AP4A suppressed terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling (TUNEL) induced by hypoxia/reperfusion in primary cortical cultures, and reduced both ischemia-induced translocation of mitochondrial cytochrome c and the increase in cytoplasmic caspase-3 activity in vivo. The purinergic P2/P4 antagonist di-inosine pentaphosphate or P1-receptor antagonist sulfonylphenyl theophylline, but not the P2-receptor antagonist suramin, antagonized the effect of AP4A, suggesting that the observed protection is mediated through an anti-apoptotic mechanism and the activation of P1- and P4-purinergic receptors. AP4A also afforded protection from toxicity induced by unilateral medial forebrain bundle injection of 6-hydroxydopamine (6-OHDA). One month after lesioning, vehicle-treated rats exhibited amphetamine-induced rotation. Minimal tyrosine hydroxylase immunoreactivity was detected in the lesioned nigra or striatum. No KCl-induced dopamine release was found in the lesioned striatum. All of these indices of dopaminergic degeneration were attenuated by pretreatment with AP4A. In addition, AP4A reduced TUNEL in the lesioned nigra 2 d after 6-OHDA administration. Collectively, our data suggest that AP4A is protective against neuronal injuries induced by ischemia or 6-OHDA through the inhibition of apoptosis. We propose that AP4A may be a potentially useful target molecule in the therapy of stroke and Parkinson's disease.

Intravenous administration of bone morphogenetic protein-7 after ischemia improves motor function in stroke rats.

BACKGROUND AND PURPOSE: We and others have previously reported that bone morphogenetic protein-7 (BMP-7), given before middle cerebral artery occlusion (MCAO), reduces ischemic injury in brain. Recent studies have indicated that receptors for BMP are upregulated after brain ischemia. It is possible that this upregulation may facilitate endogenous neurorepair in the ischemic brain. The purpose of this study was to determine the neuroregenerative effects of BMP-7 given parenterally after ischemia/reperfusion injury. METHODS: Adult Sprague-Dawley rats were anesthetized with chloral hydrate. The middle cerebral artery was transiently occluded by a filament inserted through the right internal carotid artery. The filament was removed after 60-minute ischemia to allow reperfusion. Some animals were killed 24 hours after MCAO to examine BMP-7 mRNA expression. Other animals received a single dose of intravenous BMP-7 or vehicle at 24 hours after MCAO and were used for subsequent behavioral studies and BMP-7 immunostaining. RESULTS: BMP-7 mRNA was upregulated 24 hours after MCAO in untreated animals. BMP-7 immunoreactivity was dose-dependently increased on the ischemic side of the hippocampus/dentate on day 6 after MCAO in animals receiving intravenous injection of BMP-7. Animals receiving BMP-7 also showed a decrease in body asymmetry from day 7 to day 14 and an increase in locomotor activity on day 14 after MCAO. CONCLUSIONS: Our data indicate that BMP-7, given parenterally after stroke, can pass through the blood-brain barrier on the ischemic side and induce behavioral recovery in stroke animals at longer testing times.

Effects of cerebral ischemia in mice deficient in Persephin.

Persephin (Pspn), a recently cloned member of the transforming growth factor-beta superfamily (TGF-beta) and glial cell line-derived neurotrophic factor (GDNF) subfamily, is distributed throughout the nervous system at extremely low levels and is thought to function as a survival factor for midbrain dopaminergic and spinal motor neurons in vivo. Here, we report that mice lacking Pspn by homologous recombination show normal development and behavior, but are hypersensitive to cerebral ischemia. A 300% increase in infarction volume was observed after middle cerebral artery occlusion. We find that glutamate-induced Ca(2+) influx, thought to be a major component of ischemic neuronal cell death, can be regulated directly by the Persephin protein (PSP) and that PSP can reduce hypoxia/reperfusion cell death in vitro. Neuronal cell death can be prevented or markedly attenuated by administration of recombinant human PSP in vivo before ischemia in both mouse and rat models. Taken together, these data indicate that PSP is a potent modulator of excitotoxicity in the central nervous system with pronounced neuroprotective activity. Our findings support the view that PSP signaling can exert an important control function in the context of stroke and glutamate-mediated neurotoxicity, and also suggest that future therapeutic approaches may involve this novel trophic protein.