脑卒中后抑郁大鼠海马原位增殖的新生细胞存活和分化状态

目的 观察脑卒中后抑郁(post-stroke depression,PSD)大鼠海马原位增殖新生细胞的存活和分化.方法 采用左侧大脑中动脉阻塞(MCAO)联合慢性不可预见温和应激刺激(chronicmild stress,CMS)及孤养法建立PSD模型,将雄性SD大鼠分为假手术、脑卒中、CMS和PSD组.每组均为18只.采取免疫组织化学、荧光双标染色及共聚焦成像动态检测,比较各研究组大鼠左侧海马齿状回溴脱氧尿苷嘧啶(BrdU)及其与神经元特异性核蛋白(NeuN)或胶质纤维酸性蛋白(GFAP)共表达.结果 与脑卒中组(232.2±8.6、123.7±2.6、136.2±2.6)相比,PSD组大鼠左侧(伤侧)海马齿状回BrdU+细胞数在脑梗死后第21(156.2±2.5)、30(70.2±2.0)和45天(81.2±1.1)均明显减少(t=28.83、52.2、62.08,均P<0.01),但仍高于假手术组.与脑卒中组(79.3%±2.8%、87.7%±4.6%)相比,PSD组大鼠左侧(伤侧)海马齿状回BrdU+/NeuN+细胞比例在脑梗死后第30(69.0%±3.4%)和45天(78.3%±2.4%)均明显减少(t=5.871、4.403,均P<0.01).BrdU+/GFAP+细胞比例在脑梗死后30和45 d均明显增加(t=4.226、8.945,P<0.01).结论 PSD大鼠脑卒中后海马齿状回原位增殖的新生细胞存活降低,分化为神经元的比例下降,胶质细胞比例增加.     

Temporal changes of neurogenesis in the mouse hippocampus after experimental subarachnoid hemorrhage

Recent studies indicate the existence of progenitor cells and their potential for neurogenesis in the subventricular zone (SVZ) and the hippocampus dentate gyrus (DG) of normal adult mammalian brain. Increased neurogenesis has been shown following cerebral ischemia and traumatic brain injury; however, the involvement of neurogenesis in subarachnoid hemorrhage (SAH) has not been examined. Adult male CD-1 mice were subjected to SAH by endovascular perforation of the left anterior cerebral artery. Mice received intraperitoneal injections of the cell proliferation-specific marker 5'-bromodeoxyuridine (BrdU) after SAH induction. BrdU incorporation was examined from 1 to 30 days after SAH by immunohistochemistry. The BrdU-positive cells were detected in SVZ and DG of normal control brain, and were significantly decreased in both areas three days after SAH. The number of these cells had recovered to its control level seven days after SAH. Double staining with BrdU and NeuN indicated that the majority of the BrdU-positive cells migrating into the granular cell layer of the DG became NeuN-positive 30 days after SAH. In conclusion, temporal changes of the neurogenesis as shown in the present study suggest that neurogenesis in the hippocampus may affect functional outcome after SAH. The induction of the neurogenesis can provide therapeutic value against SAH.     

Temporal changes of neurogenesis in the mouse hippocampus after experimental subarachnoid hemorrhage

Abstract

Recent studies indicate the existence of progenitor cells and their potential for neurogenesis in the subventricular zone (SVZ) and the hippocampus dentate gyrus (DG) of normal adult mammalian brain. Increased neurogenesis has been shown following cerebral ischemia and traumatic brain injury; however, the involvement of neurogenesis in subarachnoid hemorrhage (SAH) has not been examined. Adult male CD-1 mice were subjected to SAH by endovascular perforation of the left anterior cerebral artery. Mice received intraperitoneal injections of the cell proliferation-specific marker 5 ′ -bromodeoxyuridine (BrdU) after SAH induction. BrdU incorporation was examined from 1 to 30 days after SAH by immunohistochemistry. The BrdU-positive cells were detected in SVZ and DG of normal control brain, and were significantly decreased in both areas three days after SAH. The number of these cells had recovered to its control level seven days after SAH. Double staining with BrdU and NeuN indicated that the majority of the BrdU-positive cells migrating into the granular cell layer of the DG became NeuN-positive 30 days after SAH. In conclusion, temporal changes of the neurogenesis as shown in the present study suggest that neurogenesis in the hippocampus may affect functional outcome after SAH. The induction of the neurogenesis can provide therapeutic value against SAH.     

Ischemia-stimulated neurogenesis is regulated by proliferation, migration, differentiation and caspase activation of hippocampal precursor cells

A brief ischemic injury to the gerbil forebrain that caused selective damage in the CA1 region of the hippocampus also enhanced the production of new cells in the hippocampal neurogenic area. When evaluated 1 week after bromodeoxyuridine (BrdU) injection, approximately ten times more labeled cells were detected in the hippocampal dentate gyrus in ischemic animals than controls, indicating a stimulation of mitotic activity. To assess the temporal course of the survival and fate of these newborn cells, we monitored BrdU labeling and cell marker expression up to 60 days after ischemia (DAI). Loss of BrdU-positive cells was observed from both control and ischemic animals, but at 30 DAI and afterward, the ischemic group maintained more than 3 times as many BrdU-positive cells as the control group. In addition, ischemic injury also fostered the neuronal differentiation of these cells beyond the capacity observed in control animals and facilitated the migration of developing neurons to a neuronal cellular layer. The establishment of a temporal correlation between differentiation and migration provides evidence of the functional maturation of these cells. Surprisingly, we found that ischemic injury induced activation of caspase-3, not only in the CA1 region as expected, but also in the dentate subgranular zone (SGZ). Active caspase-3 immunoreactivity in the subgranular layer was co-localized with an early neuronal marker, suggesting that caspase-mediated apoptosis could mediate the loss of neurogenic cells in the SGZ. Inhibiting caspase-3 in the context of ischemia-induced neurogenesis might provide an opportunity for functional repair and a therapeutic outcome in the wake of ischemic injury.     

Cortical neurogenesis in adult rats after reversible photothrombotic stroke.

Neurogenesis occurs throughout life in the dentate gyrus of hippocampus and subventricular zone, but this phenomenon has rarely been observed in other brain regions of adult mammals. The aim of the current study was to investigate the cell proliferation process in the ischemically challenged region-at-risk after focal cerebral ischemia in the adult rat brain. A reversible photothrombotic ring stroke model was used, which features sustained hypoperfusion followed by late spontaneous reperfusion and a remarkable morphologic tissue recovery in the anatomically well defined somatosensory cortical region-at-risk. Twelve-week-old male Wistar rats received repeated intraperitoneal injections of the cell proliferation specific marker 5-bromodeoxyuridine (BrdU) after stroke induction. Immunocytochemistry of coronal brain sections revealed that the majority of BrdU-positive cells were of glial, macrophage, and endothelial origin, whereas 3% to 6% of the BrdU-positive cells were double-labeled by BrdU and the neuronspecific marker Map-2 at 7 and 100 days after stroke onset in the region-at-risk. They were distributed randomly in cortical layers II-VI. Three-dimensional confocal analyses of BrdU and the neuronal-specific marker Neu N by double immunofluorescence confirmed their colocalization within the same cells at 72 hours and 30 days after stroke induction. This study suggests that, as a potential pathway for brain repair, new neurons can be generated in the cerebral cortex of adult rats after sublethal focal cerebral ischemia.     

Temporal profile of neurogenesis in the subventricular zone,dentate gyrus and cerebral cortex following transient focal cerebral ischemia

Abstract

Background: In the adult mammalian brain, it is considered that neurogenesis persists in limited regions such as the hippocampal dentate gyrus (DG) and the subventricular zone (SVZ) of the lateral ventricle. On the other hand, neurogenesis in the cortex after cerebral ischemia and its role in post-stroke recovery have not been clarified yet. In this study, we investigated neurogenesis in the cortex and the spatiotemporal profile of neural progenitors in SVZ and DG of rats subjected to transient focal cerebral ischemia.

Materials and methods: Male Sprague–Dawley rats (270–300 g) were subjected to 60 minute middle cerebral artery occlusion. Proliferating cells were labeled by the cumulative administration of BrdU 1, 2, 3, 4, 6 and 8 weeks after ischemia induction (at weeks 1–4, 6 and 8). Double labeling was also performed with antibodies against BrdU and NeuN.

Results: BrdU-positive cells proliferated in DG and SVZ of the bilateral hemispheres, and their proliferation peaked at week 3 in SVZ and at week 4 in DG. In the peri-infarct zone of cerebral cortex, BrdU-positive cells co-expressed NeuN from weeks 3 to 8.

Conclusion: Neurogenesis was observed in the cerebral cortex and proliferation of neural progenitors occurred in SVZ and DG of rats subjected to transient focal cerebral ischemia. Our data might indicate that endogenous dormant neural stem cells residing in the cortex were activated by ischemic insult to induce the proliferation of neural progenitors and differentiation into mature neurons.     

Enforced physical training promotes neurogenesis in the subgranular zone after focal cerebral ischemia

BACKGROUND: Cerebral ischemia increases neurogenesis in the subventricular zone (SVZ) and in the subgranular zone (SGZ) of the dentate gyrus, and this might be modulated by an enriched environment including voluntary physical activity. We examined whether enforced physical training (EPT) influences neurogenesis in the SVZ and SGZ after cerebral ischemia. METHODS: Adult male Sprague-Dawley rats were subjected to focal cerebral ischemia for 2 h, and divided into an EPT and a non-EPT group. All rats in the EPT group were trained using a rota-rod for 14 days. 5-bromo-2'-deoxyuridine (BrdU) was injected to determine levels of cell proliferation. Functional recovery was assessed using a set of behavioral test batteries. Extents of endogenous neurogenesis in the SVZ and SGZ were quantified by immunofluorescence staining. Although final infarction volumes were not significantly different in the groups, functional recovery was better in the EPT group at 10 and 17 days after ischemia. In the SVZ, BrdU labeling and double labeling of BrdU/Dcx and of BrdU/NeuN were not significantly different in the two groups. However, in the SGZ, EPT significantly increased the number of BrdU-positive cell numbers (EPT vs. non-EPT: 159.1+/-19.9 vs. 101.8+/-7.8, p=0.04), and the number of BrdU/Dcx double-labeled cells (130.6+/-16.9 vs. 73.6+/-7.2, p=0.01). CONCLUSIONS: The results obtained indicate that EPT promotes neurogenesis in the SGZ of the dentate gyrus after ischemia, but not in the SVZ. The biochemical mechanism that determines the differential effects of EPT remains to be clarified.     

Enhanced neurogenesis during trimethyltin-induced neurodegeneration in the hippocampus of the adult rat

The occurrence of neurogenesis in the hippocampus of the adult rat during trimethyltin (TMT)-induced neurodegeneration was investigated using bromodeoxyuridine (BrdU). Fifteen days after TMT intoxication, BrdU-labeled cells were significantly more numerous in the hippocampus of treated animals, gradually decreasing towards the control value 21 days after intoxication in the dentate gyrus (DG), while in the CA3/hilus region BrdU-labeled cells were still more numerous in TMT-treated rats. In order to investigate the fate of newly-generated cells double labeling experiments using neuronal or glial markers were performed. Colocalization of the neuronal marker NeuN was detected in many BrdU-positive cells in the DG, while in the CA3/hilus region no colocalization of NeuN and BrdU could be observed. No colocalization of BrdU and the astroglial marker GFAP or the microglial marker OX-42 was detected either in the DG and or in the CA3/hilus region. The results indicate an enhancement of endogenous neurogenesis in the hippocampus during TMT-induced neurodegeneration, with the development of a subpopulation of regenerated cells into neurons in the DG, while in the CA3/hilus region the population of newly-generated cells should be regarded as undifferentiated.     

Functional and morphological effects of NG2 proteoglycan deletion on hippocampal neurogenesis

NG2-expressing cells are the largest proliferating cell population in the adult central nervous system. The function of NG2 proteoglycan or NG2-expressing cells in the adult brain, however, is unknown. So far, NG2-positive cells are thought to be mainly oligodendrocyte precursor cells. This view was recently challenged when NG2+/CNP-EGFP-positive cells were identified as multipotent progenitor cells in the postnatal and adult CNS (e.g., [Belachew, S., Chittajallu, R., Aguirre, A.A., Yuan, X., Kirby, M., Anderson, S., Gallo, V., 2003. Postnatal NG2 proteoglycan-expressing progenitor cells are intrinsically multipotent and generate functional neurons. J. Cell Biol. 161, 169-186]). In addition, purified NG2-expressing progenitor cells, were shown to differentiate into neurons and astrocytes in vitro [Sellers, D.L., Horner, P.J., 2005. Instructive niches: environmental instructions that confound NG2 proteoglycan expression and the fate-restriction of CNS progenitors J. Anat. 207, 727-734]. In this study, we focus on the influence of NG2 ablation on neurogenesis in the hippocampus, where putative multipotent NG2-positive cells reside, and on hippocampus-dependent behavior using NG2 knockout mice. Using the thymidine analogue bromodeoxyuridine (BrdU) to label dividing cells in vivo we show that the number of BrdU-positive cells was unchanged in the hippocampus of NG2 knockout mice 1 day after a series of BrdU injections. This finding suggests that the proliferation rate of hippocampal progenitor cells is not influenced by NG2. A few BrdU-positive cells were found in deeper layers of the granule zone 1 day after a series of BrdU injections, which is different from the wild type. The presence and the phenotype of newborn hippocampal cells were studied 4 weeks after a series of BrdU injections. The survival and differentiation of BrdU-positive cells in NG2 knockout hippocampus did not significantly differ from wild-type mice. Concurrently, the water maze task did not reveal obvious differences compared to wild-type animals. These results suggest that the null mutation for NG2 does not influence adult hippocampal neurogenesis or hippocampal-dependent behavioral tasks.     

Human neural stem cells promote proliferation of endogenous neural stem cells and enhance angiogenesis in ischemic rat brain

Transplantation of human neural stem cells into the dentate gyrus or ventricle of rodents has been reportedly to enhance neurogenesis. In this study, we examined endogenous stem cell proliferation and angiogenesis in the ischemic rat brain after the transplantation of human neural stem cells. Focal cerebral ischemia in the rat brain was induced by middle cerebral artery occlusion. Human neural stem cells were transplanted into the subventricular zone. The behavioral performance of human neural stem cells-treated ischemic rats was significantly improved and cerebral infarct volumes were reduced compared to those in untreated animals. Numerous transplanted human neural stem cells were alive and preferentially localized to the ipsilateral ischemic hemisphere. Furthermore, 5-bromo-2′-deoxyuridine-labeled endogenous neural stem cells were observed in the subventricular zone and hippocampus, where they differentiated into cells immunoreactive for the neural markers doublecortin, neuronal nuclear antigen Neu N, and astrocyte marker glial fibrillary acidic protein in human neural stem cells-treated rats, but not in the untreated ischemic animals. The number of 5-bromo-2′-deoxyuridine-positive ? anti-von Willebrand factor-positive proliferating endothelial cells was higher in the ischemic boundary zone of human neural stem cells-treated rats than in controls. Finally, transplantation of human neural stem cells in the brains of rats with focal cerebral ischemia promoted the proliferation of endogenous neural stem cells and their differentiation into mature neural-like cells, and enhanced angiogenesis. This study provides valuable insights into the effect of human neural stem cell transplantation on focal cerebral ischemia, which can be applied to the development of an effective therapy for stroke.     

Chronic fluoxetine treatment improves ischemia-induced spatial cognitive deficits through increasing hippocampal neurogenesis after stroke

Cognitive deficits, including spatial memory impairment, are very common after ischemic stroke. Neurogenesis in the dentate gyrus (DG) contributes to forming spatial memory in the ischemic brain. Fluoxetine, a selective serotonin reuptake inhibitor, can enhance neurogenesis in the hippocampus in physiological situations and some neurological diseases. However, whether it has effects on ischemia-induced spatial cognitive impairment and hippocampal neurogenesis has not been determined. Here we report that fluoxetine treatment (10 mg kg(-1), i.p.) for 4 weeks promoted the survival of newborn cells in the ischemic hippocampus and, consequently, attenuated spatial memory impairment of mice after focal cerebral ischemia. Disrupting hippocampal neurogenesis blocked the beneficial effect of fluoxetine on ischemia-induced spatial cognitive impairment. These results suggest that chronic fluoxetine treatment benefits spatial cognitive function recovery following ischemic insult, and the improved cognitive function is associated with enhanced newborn cell survival in the hippocampus. Our results raise the possibility that fluoxetine can be used as a drug to treat poststroke spatial cognitive deficits.     

慢性应激抑制卒中后大鼠海马内源性神经干细胞增殖和分化研究

目的 观察慢性应激对卒中后海马内源性神经干细胞增殖和分化的影响。方法 将雄性SD大鼠分为正常、卒中、慢性应激组。建立左侧大脑中动脉闭塞卒中动物模型并予以慢性不可预见温和应激刺激结合孤养。经溴脱氧尿苷嘧啶（BrdU）标记，免疫组化、荧光双标染色及共聚焦成像动态检测并比较各研究组大鼠左侧海马齿状回BrdU及其与神经元核性蛋白（NeuN）共表达。结果 与卒中组相比，慢性应激组大鼠病灶侧海马齿状回BrdU+细胞数在脑梗死后第7天未见减少，第21天明显减少（28.5±1.9 vs 72.2±1.4），差异有统计学意义（P<0.001）。与卒中组相比，慢性应激组大鼠病灶侧海马齿状回BrdU+/NeuN+细胞比例在脑梗死后30 d[（69.0±3.4）%）]和45 d[（78.3±2.4）%]均明显减少，差异有统计学意义（P<0.001； P<0.01）。结论 慢性应激能明显抑制卒中后海马内源性神经干细胞的增殖和分化，可能是卒中后抑郁发病的因素之一。     

Bcl2 enhances survival of newborn neurons in the normal and ischemic hippocampus

Neuronal progenitors in the adult hippocampus continually proliferate and differentiate to the neuronal lineage, and ischemic insult promotes hippocampal neurogenesis. However, newborn neurons show a progressive reduction in numbers during the initial few weeks, therefore, enhanced survival of newborn neurons seems to be essential for therapeutic strategy. Bcl-2 is a crucial regulator of programmed cell death in CNS development and in apoptotic and necrotic cell death. Therefore, we tested whether Bcl-2 overexpression enhances survival of newborn neurons in the adult mouse hippocampus under normal and ischemic conditions. Many newborn neurons in the hippocampal dentate gyrus undergo apoptosis. Human Bcl-2 expression in NSE-bcl-2 transgenic mice began at the immature neuronal stage and remained constant in surviving mature neurons. Bcl-2 significantly increased survival of newborn neurons under both conditions, but particularly after ischemia, with decreased cell death of newborn neurons in NSE-bcl-2 transgenic mice. We also clarified the effect by Bcl-2 overexpression of enhanced survival of newborn neurons in primary hippocampal cultures with BrdU labeling. These findings suggest that Bcl-2 plays a crucial role in adult hippocampal neurogenesis under normal and ischemic conditions.     

Proliferation of neural and neuronal progenitors after global brain ischemia in young adult macaque monkeys

To investigate the effect of global cerebral ischemia on brain cell proliferation in young adult macaques, we infused 5-bromo-2'-deoxyuridine (BrdU), a DNA replication indicator, into monkeys subjected to ischemia or sham-operated. Subsequent quantification by BrdU immunohistochemistry revealed a significant postischemic increase in the number of BrdU-labeled cells in the hippocampal dentate gyrus, subventricular zone of the temporal horn of the lateral ventricle, and temporal neocortex. In all animals, 20-40% of the newly generated cells in the dentate gyrus and subventricular zone expressed the neural progenitor cell markers Musashi1 or Nestin. A few BrdU-positive cells in postischemic monkeys were double-stained for markers of neuronal progenitors (class III beta-tubulin, TUC4, doublecortin, or Hu), neurons (NeuN), or glia (S100beta or GFAP). Our results suggest that ischemia activates endogenous neuronal and glial precursors residing in diverse locations of the adult primate central nervous system.     

脑梗死大鼠神经前体细胞增殖水平的研究

目的研究脑梗死病灶周围及海马处神经前体细胞增殖水平的动态变化。方法采用易卒中型肾性高血压大鼠(RHRSP)，电凝大脑中动脉(MCA)主干制成脑梗死(MCAO)模型。行大鼠神经功能评定，免疫组化观察并计数梗死灶边缘、对侧镜区及双侧海马5-溴脱氧尿核苷(Bromodeoxyuridine，BrdU)标记的细胞。结果MCAO后大鼠神经功能评分减低，5d时恢复正常。MCAO后梗死灶边缘、对侧镜区及双侧海马均有BrdU阳性细胞分布，且病灶侧多于病灶对侧，集中分布于病灶周围。结论脑缺血可诱导神经前体细胞增殖并移向病灶，可能成为脑梗死恢复的重要物质基础。     

Vascular adventitia generates neuronal progenitors in the monkey hippocampus after ischemia

In the adult hippocampus, neurogenesis proceeds in the subgranular zone (SGZ) of the dentate gyrus (DG), but not in the cornu Ammonis (CA). Recently, we demonstrated in monkeys that transient brain ischemia induces an increase of the neuronal progenitor cells in the SGZ, but not in CA1, in the second week after the insult. To identify the origin of primary neuronal progenitors in vivo, we compared the postischemic monkey DG and CA1, using light and electron microscopy, focusing on specific phenotype markers, as well as the expression of neurotrophic factors. Laser confocal microscopy showed that 1-3% of 5-bromo-2'-deoxyuridine (BrdU)-positive cells in the SGZ after 2-96 h labeling were also positive for neuronal markers such as TUC4, betaIII tubulin, and NeuN on days 9 and 15. In contrast, despite the presence of numerous BrdU-positive cells, CA1 showed no neurogenesis at any time points, and all the progenitors were positive for glial markers: Iba1 or S-100beta on days 4, 9, and 15. Highly polysialylated neural cell adhesion molecule (PSA-NCAM)-positive cells were abundant in the SGZ, but were absent in CA1. On day 9, most of the immature neurons positive for betaIII-tubulin in SGZ showed an increase in PSA-NCAM immunoreactivity. The immunoreactivity of brain-derived neurotrophic factor (BDNF) was abundant at the vascular adventitia of the SGZ, but was absent at the adventitia of CA1. BrdU-positive progenitor cells were frequently seen in the vicinity of proliferating blood vessels. Ultrastructural analysis indicated that most of the neuronal progenitor cells and microglia originated from the pericytes of capillaries and/or adventitial cells of arterioles (called vascular adventitia). The detaching adventitial cells showed mitotic figures in the perivascular space, and the resultant neuronal progenitor cells made contact with dendritic spines associated with synaptic vesicles or boutons. These data implicate the vascular adventitia as a novel potential source of neuronal progenitor cells in the postischemic primate SGZ.     

Focal transient ischemia increases APP-BP1 expression in neural progenitor cells

Amyloid precursor protein binding protein-1 (APP-BP1) binds to the carboxyl terminus of APP. In this study, we explored whether APP-BP1 expression is affected by focal transient cerebral ischemia induced by middle cerebral artery occlusion in Wistar rats. APP-BP1 expression was increased in the dentate gyrus of the hippocampus and in the subventricular zone of rats exposed to focal transient cerebral ischemia. In addition, APP-BP1 immunoreactivity overlapped with antidoublecortin and anti-5-bromo-2-deoxyuridine labeling. Focal transient cerebral ischemia has been reported earlier to induce neurogenesis in adult brains. The upregulation of APP-BP1 expression in neural progenitor cells after focal transient ischemia suggests that this protein contributes to the neurogenesis induced by transient ischemia and reperfusion.     

Sema4C Expression in Neural Stem/Progenitor Cells and in Adult Neurogenesis Induced by Cerebral Ischemia

Sema4C is a transmembrane protein that belongs to axon guidance molecules of semaphorin family. Previous reports have shown that Sema4C could interact with postsynaptic protein PSD95, etc, but the expression and the role of Sema4C in neurogenesis remains unknown. In this study, whole-mount in situ hybridization result showed that Sema4C was expressed abundantly in the areas of lateral ventricle, the striatum, the wall of midbrain, and the pons/midbrain junction of E11.5 embryos brain. Neural stem/progenitor cells (NSPs) obtained from E13.5 embryonic rat midbrain are also positive for Sema4C immunoreactivity. Sema4C expression was dramatically downregulated during induction of NSP differentiation. In order to confirm the involvement of Sema4C in neurogenesis, we used the rat global cerebral ischemia model to make adult neurogenesis in vivo. The robust proliferative NSPs were monitored by labeling with bromodeoxyuridine (BrdU) within the subventricular zone and dentate gyrus that continues for at least 2 weeks. Immunohistochemistry and Western blot analysis showed that Sema4C expression was dramatically upregulated during neurogenesis after cerebral ischemia–perfusion injury. Double immunostaining and stereologic counting analysis indicated that a high proportion of BrdU-positive proliferative cells were Nestin-positive NSPs, and also, Sema4C was highly expressed in these proliferative populations at specific stages after ischemic injury. These observations provide the evidence to support a putative role of Sema4C during neurogenesis both in vivo and in vitro.     

Differential proliferative response in the postischemic hippocampus,temporal cortex,and olfactory bulb of young adult macaque monkeys

We investigated the fate of proliferating cells in the adult monkey brain after global ischemia. We used the thymidine analogue bromodeoxyuridine (BrdU) to label S-phase cells and their progeny in Japanese macaques subjected to global cerebral ischemia for 20 min or to a sham operation. Subsequently, newly generated cells were identified by BrdU immunohistochemistry, and their immunophenotype was determined quantitatively, using specific markers. The ischemic insult significantly increased the number of proliferating cells in the hippocampus and temporal neocortex, where the majority BrdU-labeled cells expressed markers for microglia (Iba1, CD68, and Ham56) or astrocytes (S-100beta and glial fibrillary acidic protein [GFAP]). In contrast, the proliferation level in the parahippocampal region remained unchanged. This discrepancy prompted us to investigate the postischemic response in the olfactory bulb, a well-known site of adult cell generation that is anatomically distant from the above-mentioned regions but that is also subjected to the global ischemic insult. The olfactory bulb contained clusters of proliferating cells expressing markers for neural (Musashi1 and Nestin) and/or neuronal (class III beta-tubulin) progenitors; these were immunophenotypically distinct from other cell types. Their number and distribution were unaltered by ischemia. Our results demonstrate that cell proliferation and differentiation in the adult macaque brain and olfactory bulb are differentially affected by a common insult.