Galectin-1 regulates neurogenesis in the subventricular zone and promotes functional recovery after stroke

Galectin-1 (Gal-1) has recently been identified as a key molecule that plays important roles in the regulation of neural progenitor cell proliferation in two neurogenic regions: the subventricular zone (SVZ) of the lateral ventricle and the subgranular zone of the hippocampal dentate gyrus. To test the hypothesis that Gal-1 contributes to adult neurogenesis after focal ischemia, we studied the temporal profile of endogenous Gal-1 expression and the effects of human recombinant Gal-1 on neurogenesis and neurological functions in an experimental focal ischemic model. In the normal brain, Gal-1 expression was observed only in the SVZ. In the ischemic brain, Gal-1 expression was markedly upregulated in the SVZ and the area of selective neuronal death around the infarct in the striatum. The temporal profile of Gal-1 expression was correlated with that of neural progenitor cell proliferation in the SVZ of the ischemic hemisphere. Double-labeling studies revealed that Gal-1 was localized predominantly in both reactive astrocytes and SVZ astrocytes. Administration of Gal-1, which is known to have carbohydrate-binding ability, into the lateral ventricle increased neurogenesis in the ipsilateral SVZ and improved sensorimotor dysfunction after focal ischemia. By contrast, blockade of Gal-1 in the SVZ by the administration of anti-Gal-1 neutralizing antibody strongly inhibited neurogenesis and diminished neurological function. These results suggest that Gal-1 is one of the principal regulators of adult SVZ neurogenesis through its carbohydrate-binding ability and provide evidence that Gal-1 protein has a role in the improvement of sensorimotor function after stroke.     

Effects of CXCR7-neutralizing antibody on neurogenesis in the hippocampal dentate gyrus and cognitive function in the chronic phase of cerebral ischemia

Stromal cell-derived factor-1 and its receptor CXCR4 are essential regulators of the neurogenesis that occurs in the adult hippocampal dentate gyrus.However,the effects of CXCR7,a new atypical receptor of stromal cell-derived factor-1,on hippocampal neurogenesis after a stroke remain largely unknown.Our study is the first to investigate the effect of a CXCR7-neutralizing antibody on neurogenesis in the dentate gyrus and the associated recovery of cognitive function of rats in the chronic stage of cerebral ischemia.The rats were randomly divided into sham,sham+anti-CXCR7,ischemia and ischemia+anti-CXCR7 groups.Endothelin-1 was injected in the ipsilateral motor cortex and striatum to induce focal cerebral ischemia.Sham group rats were injected with saline instead of endothelin-1 via intracranial injection.Both sham and ischemic rats were treated with intraventricular infusions of CXCR7-neutralizing antibodies for 6 days 1 week after surgery.Immunofluorescence staining with doublecortin,a marker for neuronal precursors,was performed to assess the neurogenesis in the dentate gyrus.We found that anti-CXCR7 antibody infusion enhanced the proliferation and dendritic development of doublecortin-labeled cells in the dentate gyrus in both ischemic and sham-operated rats.Spatial learning and memory functions were assessed by Morris water maze tests 30-32 days after ischemia.CXCR7-neutralizing antibody treatment significantly reduced the escape latency of the spatial navigation trial and increased the time spent in the target quadrant of spatial probe trial in animals that received ischemic insult,but not in sham operated rats.These results suggest that CXCR7-neutralizing antibody enhances the neurogenesis in the dentate gyrus and improves the cognitive function after cerebral ischemia in rats.All animal experimental protocols and procedures were approved by the Institutional Animal Care and Use Committee of China Medical University(CMU16089 R)on December 8,2016.     

Forebrain neurogenesis after focal Ischemic and traumatic brain injury

Neural stem cells persist in the adult mammalian forebrain and are a potential source of neurons for repair after brain injury. The two main areas of persistent neurogenesis, the subventricular zone (SVZ)-olfactory bulb pathway and hippocampal dentate gyrus, are stimulated by brain insults such as stroke or trauma. Here we focus on the effects of focal cerebral ischemia on SVZ neural progenitor cells in experimental stroke, and the influence of mechanical injury on adult hippocampal neurogenesis in models of traumatic brain injury (TBI). Stroke potently stimulates forebrain SVZ cell proliferation and neurogenesis. SVZ neuroblasts are induced to migrate to the injured striatum, and to a lesser extent to the peri-infarct cortex. Controversy exists as to the types of neurons that are generated in the injured striatum, and whether adult-born neurons contribute to functional restoration remains uncertain. Advances in understanding the regulation of SVZ neurogenesis in general, and stroke-induced neurogenesis in particular, may lead to improved integration and survival of adult-born neurons at sites of injury. Dentate gyrus cell proliferation and neurogenesis similarly increase after experimental TBI. However, pre-existing neuroblasts in the dentate gyrus are vulnerable to traumatic insults, which appear to stimulate neural stem cells in the SGZ to proliferate and replace them, leading to increased numbers of new granule cells. Interventions that stimulate hippocampal neurogenesis appear to improve cognitive recovery after experimental TBI. Transgenic methods to conditionally label or ablate neural stem cells are beginning to further address critical questions regarding underlying mechanisms and functional significance of neurogenesis after stroke or TBI. Future therapies should be aimed at directing appropriate neuronal replacement after ischemic or traumatic injury while suppressing aberrant integration that may contribute to co-morbidities such as epilepsy or cognitive impairment.     

Directed migration of neuronal precursors into the ischemic cerebral cortex and striatum

Pathological processes, including cerebral ischemia, can enhance neurogenesis in the adult brain, but the fate of the newborn neurons that are produced and their role in brain repair are obscure. To determine if ischemia-induced neuronal proliferation is associated with migration of nascent neurons toward ischemic lesions, we mapped the migration of cells labeled by cell proliferation markers and antibodies against neuronal marker proteins, for up to 2 weeks after a 90-min episode of focal cerebral ischemia caused by occlusion of the middle cerebral artery. Doublecortin-immunoreactive cells in the rostral subventricular zone, but not the dentate gyrus, migrated into the ischemic penumbra of the adjacent striatum and, via the rostral migratory stream and lateral cortical stream, into the penumbra of ischemic cortex. These results indicate that after cerebral ischemia, new neurons are directed toward sites of brain injury, where they might be in a position to participate in brain repair and functional recovery.     

Increased proliferation of neural progenitor cells but reduced survival of newborn cells in the contralateral hippocampus after focal cerebral ischemia in rats.

Recent studies demonstrated that neurogenesis in the adult hippocampus increased after transient global ischemia; however, the molecular mechanism underlying increased neurogenesis after ischemia remains unclear. The finding that proliferation of progenitor cells occurred at least a week after ischemic insult suggests that the stimulus was not an ischemic insult to progenitor cells. To clarify whether focal ischemia increases the rate of neurogenesis in the remote area, the authors examined the contralateral hemisphere in rats subjected to permanent occlusion of the middle cerebral artery. In the subgranular zone of the hippocampal dentate gyrus, the numbers of bromodeoxyuridine (BrdU)-positive cells increased approximately sixfold 7 days after ischemia. In double immunofluorescence staining, more than 80% of newborn cells expressed Musashi1, a marker of neural stem/progenitor cells, but only approximately 10% of BrdU-positive cells expressed glial fibrillary acidic protein (GFAP), a marker of astrocytes. The number of BrdU-positive cells markedly decreased 28 days after BrdU administration after ischemia, but it was still elevated compared with that of sham-operated rats. In double immunofluorescence staining, 80% of newborn cells expressed NeuN, a marker of differentiated neurons, and 10% of BrdU-positive cells expressed GFAP. However, in the other areas of the contralateral hemisphere including the rostral subventricular zone, the number of BrdU-positive cells remained unchanged. These results showed that focal ischemia stimulated the proliferation of neuronal progenitor cells, but did not support survival of newborn cells in the contralateral hippocampus.     

Post-ischemic administration of heparin-binding epidermal growth factor-like growth factor (HB-EGF) reduces infarct size and modifies neurogenesis after focal cerebral ischemia in the rat.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a hypoxia-inducible, neuroprotective protein that also stimulates proliferation of neuronal precursor cells. Accordingly, HB-EGF may contribute to recovery from cerebral injury through direct neuroprotective effects, by enhancing neurogenesis, or both. When administered by the intracerebroventricular route 1-3 days after focal cerebral ischemia in adult rats, HB-EGF decreased the volume of the resulting infarcts and reduced post-ischemic neurological deficits. HB-EGF also increased the incorporation of bromodeoxyuridine into cells expressing the immature neuronal marker protein TUC-4 in the dentate subgranular and rostral subventricular zones, consistent with increased proliferation of neuronal precursors. However, HB-EGF decreased the number of newborn neurons that migrated into the ischemic striatum, perhaps partly because reduction of infarct size by HB-EGF also reduced the stimulus to migration. To determine if HB-EGF might also directly inhibit migration of neuronal precursors, we co-cultured subventricular zone (SVZ) explants treated with HB-EGF or vehicle together with hypoxic cerebral cortical explants, and measured cell migration from the former toward the latter. HB-EGF reduced directed migration of SVZ cells toward the cortical explants, possibly due to a local chemoattractant effect on neuronal precursor cells, which may be mediated through the HB-EGF-specific receptor, N-arginine dibasic convertase. The delayed neuroprotective effect of HB-EGF may have implications for efforts to prolong the therapeutic window for intervention in stroke.     

VEGF-overexpressing transgenic mice show enhanced post-ischemic neurogenesis and neuromigration

New neurons are generated continuously in the subventricular zone and dentate gyrus of the adult brain. Neuropathologic processes, including cerebral ischemia, can enhance neurogenesis, as can growth factors and other physiologic stimuli. Vascular endothelial growth factor (VEGF) is an angiogenic and neuroprotective growth factor that can promote neurogenesis, but it is unknown whether VEGF can enhance migration of newborn neurons toward sites of ischemic injury, where they might be able to replace neurons that undergo ischemic death. In the present study we produced permanent focal cerebral ischemia in transgenic (Tg) mice that overexpress VEGF. Cell proliferation and neurogenesis were assessed with bromodeoxyuridine (Brdu) labeling and immunostaining for cell type-specific markers. In VEGF-Tg mice, brains examined 7-28 days after cerebral ischemia showed markedly increased subventricular zone (SVZ) neurogenesis, chains of neuroblasts extending from the SVZ to the peri-infarct cortex, and an increase in the number of newly generated cortical neurons at 14-28 days after ischemia. In concert with these effects, VEGF overexpression reduced infarct volume and improved postischemic motor function. These findings provide evidence that VEGF increases SVZ neurogenesis and neuromigration, consistent with a possible role in repair. Our data suggest that in addition to its neuroprotective effects, which are associated with improved outcome in the acute phase after cerebral ischemia, VEGF enhances postischemic neurogenesis, which could provide a therapeutic target for more chronic brain repair.     

Implication of cyclooxygenase-2 on enhanced proliferation of neural progenitor cells in the adult mouse hippocampus after ischemia

Global ischemia promotes neurogenesis in the dentate gyrus of the adult mouse hippocampus. Cyclooxygenase (COX)-2, the principal isoenzyme in the brain, modulates inflammation, glutamate-mediated cytotoxicity, and synaptic plasticity. We demonstrated that delayed treatment with different classes of COX inhibitor significantly blunted enhancement of dentate gyrus proliferation of neural progenitor cells after ischemia. COX-2 immunoreactivity was observed in both neurons and astrocytes in the dentate gyrus, but not in neural progenitor cells in the subgranular zone. Moreover, in the postischemic dentate gyrus of heterozygous and homozygous COX-2 knockout mice, proliferating bromodeoxyuridine-positive cells were significantly fewer than in wild-type littermates. These results demonstrate that COX-2 is an important modulator in enhancement of proliferation of neural progenitor cells after ischemia.     

A nitric oxide donor induces neurogenesis and reduces functional deficits after stroke in rats.

The adult rodent brain is capable of generating neuronal progenitor cells in the subventricular zone, and in the dentate gyrus of the hippocampus, throughout the life of the animal. Signals that regulate progenitor cell proliferation, differentiation, and migration are not well known. We report that administration of a nitric oxide donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) aminio]diazen-1-ium-1,2-diolate (DETA/NONOate), to young adult rats significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus. Treatment with DETA/ NONOate also increases neurogenesis in the dentate gyrus. Furthermore, administration of DETA/NONOate to rats subjected to embolic middle cerebral artery occlusion significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus, and these rats exhibit significant improvements of neurological outcome during recovery from ischemic stroke. Administration of DETA/NONOate significantly increases cortical levels of guanosine monophosphate both in ischemic and nonischemic rats, supporting the role of nitric oxide in promoting cell proliferation and neurogenesis. Thus, our data indicate that nitric oxide is involved in the regulation of progenitor cells and neurogenesis in the adult brain. This suggests that nitric oxide delivered to the brain well after stroke may have therapeutic benefits.