缺血神经元细胞周期进程与凋亡机制研究
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摘要
第一部分糖氧剥夺诱导培养的皮层神经元细胞周期再进入及凋亡机制的研究
目的:观察糖氧剥夺处理后培养的皮层神经元细胞周期再进入和神经元凋亡情况,探讨糖氧剥夺诱导神经元凋亡的可能机制。
方法:建立体外培养的皮层神经元糖氧剥夺(Oxygen-glucose deprivation, OGD)损伤实验,分为对照组、OGD1h后恢复正常培养6h、12h、24h组、roscovitine干预组。应用流式细胞仪检测神经元凋亡,免疫荧光细胞化学染色观察神经元BrdU摄取、磷酸化Rb蛋白(p-Rb,ser 795)表达及其与TUNEL染色的关系;利用western blot检测神经元p-Rb和E2F1的表达情况。
结果:与对照组相比,OGD1h恢复正常培养6h、12h、24h组神经元BrdU摄取的细胞比例、TUNEL染色的细胞比例和两者共定位比例依次增高;p-Rb的表达在OGD后各组均明显增高,与BrdU摄取相平行;p-Rb与TUNEL共定位表达在OGD后6h和12h明显,24h共定位细胞比例显著下降;给予roscovitine干预后,OGD处理的神经元p-Rb和E2F1的表达减少,凋亡神经元比例下降。
结论:糖氧剥夺后神经元尝试通过Rb磷酸化再进入细胞周期,并诱导神经元凋亡,抑制Rb磷酸化减少OGD处理后神经元的凋亡;磷酸化Rb介导的凋亡途径可能是OGD诱导的神经元凋亡机制之一。
第二部分磷酸化Rb蛋白与大鼠局灶性脑缺血后神经元凋亡关系的研究
目的:观察大鼠局灶性脑缺血后磷酸化Rb蛋白(p-Rb,ser 795)的表达、定位及其与神经元凋亡的时空关系。
方法:制备大鼠大脑中动脉梗塞(MCAO)模型,分为假手术对照组、缺血1小时再灌注12h, 1d, 3d, 7d组。利用TUNEL法检测缺血周边区神经元凋亡情况;免疫荧光组织化学染色方法观察MCAO后各时间点p-Rb的表达与定位;TUNEL与p-Rb荧光双标观察神经元凋亡与p-Rb表达、定位的关系。
结果:与对照组相比,大鼠MCAO再灌注12h缺血灶周边开始检测到凋亡细胞,神经元内p-Rb表达增加,部分细胞胞核胞浆均有p-Rb表达;再灌注1d大部分神经元p-Rb从胞核转移到胞浆,TUNEL阳性细胞数达到高峰;再灌注12h和1d, TUNEL与p-Rb分别以重叠和镶嵌的方式共定位;再灌注3d, 7d发生p-Rb核浆转移的神经元与TUNEL染色细胞仍然分别维持在高水平,但是两者却没有明显的共定位关系。
结论:p-Rb可能参与短暂局灶脑缺血后神经元早期凋亡过程,间接或者不参与神经元晚期凋亡过程。
第三部分流式细胞仪分选thy1-GFP-J转基因小鼠皮层及海马神经元
目的:建立一种快速高效获得活体哺乳动物脑内单一纯化神经元群的方法。
方法:用RT-PCR和激光共聚焦显微镜扫描脑片鉴定thy1 -GFP-J转基因小鼠(神经元内转入绿色荧光蛋白),取2月龄鉴定阳性小鼠皮层和海马组织,应用番木瓜蛋白酶分离系统制备脑组织单细胞悬液,PI标记死亡细胞,流式细胞仪荧光激活细胞分选技术(FACS)分选收集GFP~+和PI~-的存活神经元,并抽提获得细胞的RNA和蛋白。
结果:阳性转基因小鼠DNA的PCR扩增鉴定显示为内参(324bp)和所转基因(173bp)两条带,脑片的激光共聚焦扫描结果显示海马和皮层均可见大小较均一的圆点状绿色荧光。经FACS分选后的细胞几乎全都发出绿色荧光,每次产量约为0.5-1×10~6个存活神经元。获得的纯化神经元中抽提的RNA和蛋白可用于RT-PCR和western blot检测。
结论:利用FACS可以从thy1 -GFP-J转基因小鼠脑内获得单一纯化的存活神经元群。该技术的应用将为活体神经元细胞特异性的基因表达和蛋白定量分析研究提供一个可行方案。
Part One Cortical neurons re-enter cell cycle and undergo apoptosis Induced by oxygen-glucose deprivation in vitro
Objective: To definite the relationship between cell cyler re-entery and neuronal apoptosis in cultured cortical neurons following oxygen-glucose deprivation (OGD) and to explore the potential mechanism of apoptosis in post-mitotic neurons induced by OGD.
Methods: The model of OGD injury on cultured cortical neurons was established. The cultured neurons were randomly divided into 5 groups: control group, the group of 6h, 12h, 24h reoxygenation after 1h OGD and roscovitine treated group. Flow cytometry was used to detect the percent of neuronal apoptosis after 1h OGD. Immunofluorescence staining was used to detect uptake of BrdU and expression of phosphorylated Rb protein (p-Rb, ser 795) in neurons, and their co-localizated relationship with the TUNEL staining. Western Bolt was used to detect the expression of p-Rb and E2F1.
Results: Compared with the control group, the percentage of neurons with BrdU uptake ,TUNEL staining and co-localization of both were increased in the 6h, 12h, 24h reoxygenation after 1h OGD. The expression of p-Rb in each group after OGD were significantly increased, in parallel with BrdU uptake. The co-localization of p-Rb and TUNEL in 6h and 12h after OGD was apparent, but the percent of co-localization between p-Rb and TUNEL was strikingly decreased in 24h after OGD. The expression of p-Rb and E2F1 and the percent of neuronal apoptosis were decreased in roscovitine treated group.
Conclusion: Attempted cell cycle re-entry with phosphorylation of Rb in neurons induces neuronal apoptosis after OGD treatment. Inhibition of phosphorylation of Rb following OGD reduces neuronal apoptosis, phosphorylated Rb pathway is one of the mechanisms of neuronal apoptosis induced by OGD.
Part Two The involvement of upregulation and translocation of phospho-Rb in early neuronal apoptosis following focal cerebral ischemia in rats
Objective: To investigate the expression and subcellular localization of phospho-Rb (ser 795), and temporal and spatial relationship between it and neuronal apoptotic death in rats subjected to transient focal cerebral ischemia.
Methods: The model of middle cerebral artery occlusion (MCAO) was carried out. All animals were divided into five groups: sham-operated controls, 12 h, 1 d, 3 d, and 7 d after MCAO/reperfusion. By analyzing DNA fragmentation with a TUNEL assay, apoptotic cells were examined in the peri-infarct area of the ischemic cerebral cortex. Immunofluorescence staining was used to detect the expression and subcellular localization of phospho-Rb (ser 795) in the each time points after MCAO/reperfusion. Double-labeling analysis of TUNEL and phospho-Rb staining was used to detect the temporal and spatial relationship between phospho-Rb and neuronal apoptotic death.
Results: Compared with sham-operated controls, at 12 h after MCAO/ reperfusion TUNEL-positive cells began to be detected, and the immunoreactivity of phospho-Rb in neurons was increased. One day after the transient ischemic injury, it was showed that subcellular localization of phospho-Rb translocated from the nucleus to the cytoplasm in many neurons in the peri-infarct area and the number of TUNEL-positive cells peaked at this time point. Most phospho-Rb appeared to be localized within TUNEL staining cells at 12 h and 1 d after MCAO/reperfusion in an overlapping and mosaic pattern. However, there was no apparent colocalization of phospho-Rb and TUNEL staining at 3 d and 7 d, although the number of phospho-Rb translocated neurons and TUNEL stained cells were still at high levels at these time points.
Conclusion: phospho-Rb may be involved in the early stages of neuronal apoptosis after transient cerebral ischemia, and it may not be involved or in an indirect way in the late stages of neuronal apoptosis.
Part Three Sorting of neurons in cortex and hippocampus from thy1-GFP-J transgenic mice by flow cytometry
Objective: To establish an effective method fast getting purified neuronal populations from mammalian brain in vivo.
Methods: Thy1-GFP-J transgenic mice, which expressed green fluorescent protein (GFP) in neurons, were confirmed by RT-PCR and confocal laser scan microscopy. The cortex and hippocampus of 2-months-old transgenic mice were dissected and then dissociated using the papin dissociation system. Single cells of brain tissue were treated with PI to label dead cells and sorted on a FACS-Aria cell sorter. Cells with high GFP signals and negative PI signals were selected, then RNA and protein were extracted from the sorted cells.
Results: PCR amplification of DNA from tail of transgenic mice indicated that it appears two bands inculding transgene band (173bp) and internal control band (324bp), and the pictures of brain tissue captured by confocal microscopy showed that green fluorescent spots with similar round patterns can be seen in the cortex and hippocampus of these mice. After sorting, almost all cells seemed brightly fluorescent in their soma, and about 0.5-1×10~6 lived neurons were obtained in each sort. RNA and protein extracted from purified neurons were detected by RT-PCR and western blot.
Conclusion: It is a reliable and practical method for study of neuronal cell type-specific gene expression profiling and quantitative analysis of proteins extracted from purified neuronal populations, which obtained from mammalian brain of adult thy1-GFP-J transgenic mice by FACS in vivo.
目的:观察糖氧剥夺处理后培养的皮层神经元细胞周期再进入和神经元凋亡情况,探讨糖氧剥夺诱导神经元凋亡的可能机制。
方法:建立体外培养的皮层神经元糖氧剥夺(Oxygen-glucose deprivation, OGD)损伤实验,分为对照组、OGD1h后恢复正常培养6h、12h、24h组、roscovitine干预组。应用流式细胞仪检测神经元凋亡,免疫荧光细胞化学染色观察神经元BrdU摄取、磷酸化Rb蛋白(p-Rb,ser 795)表达及其与TUNEL染色的关系;利用western blot检测神经元p-Rb和E2F1的表达情况。
结果:与对照组相比,OGD1h恢复正常培养6h、12h、24h组神经元BrdU摄取的细胞比例、TUNEL染色的细胞比例和两者共定位比例依次增高;p-Rb的表达在OGD后各组均明显增高,与BrdU摄取相平行;p-Rb与TUNEL共定位表达在OGD后6h和12h明显,24h共定位细胞比例显著下降;给予roscovitine干预后,OGD处理的神经元p-Rb和E2F1的表达减少,凋亡神经元比例下降。
结论:糖氧剥夺后神经元尝试通过Rb磷酸化再进入细胞周期,并诱导神经元凋亡,抑制Rb磷酸化减少OGD处理后神经元的凋亡;磷酸化Rb介导的凋亡途径可能是OGD诱导的神经元凋亡机制之一。
第二部分磷酸化Rb蛋白与大鼠局灶性脑缺血后神经元凋亡关系的研究
目的:观察大鼠局灶性脑缺血后磷酸化Rb蛋白(p-Rb,ser 795)的表达、定位及其与神经元凋亡的时空关系。
方法:制备大鼠大脑中动脉梗塞(MCAO)模型,分为假手术对照组、缺血1小时再灌注12h, 1d, 3d, 7d组。利用TUNEL法检测缺血周边区神经元凋亡情况;免疫荧光组织化学染色方法观察MCAO后各时间点p-Rb的表达与定位;TUNEL与p-Rb荧光双标观察神经元凋亡与p-Rb表达、定位的关系。
结果:与对照组相比,大鼠MCAO再灌注12h缺血灶周边开始检测到凋亡细胞,神经元内p-Rb表达增加,部分细胞胞核胞浆均有p-Rb表达;再灌注1d大部分神经元p-Rb从胞核转移到胞浆,TUNEL阳性细胞数达到高峰;再灌注12h和1d, TUNEL与p-Rb分别以重叠和镶嵌的方式共定位;再灌注3d, 7d发生p-Rb核浆转移的神经元与TUNEL染色细胞仍然分别维持在高水平,但是两者却没有明显的共定位关系。
结论:p-Rb可能参与短暂局灶脑缺血后神经元早期凋亡过程,间接或者不参与神经元晚期凋亡过程。
第三部分流式细胞仪分选thy1-GFP-J转基因小鼠皮层及海马神经元
目的:建立一种快速高效获得活体哺乳动物脑内单一纯化神经元群的方法。
方法:用RT-PCR和激光共聚焦显微镜扫描脑片鉴定thy1 -GFP-J转基因小鼠(神经元内转入绿色荧光蛋白),取2月龄鉴定阳性小鼠皮层和海马组织,应用番木瓜蛋白酶分离系统制备脑组织单细胞悬液,PI标记死亡细胞,流式细胞仪荧光激活细胞分选技术(FACS)分选收集GFP~+和PI~-的存活神经元,并抽提获得细胞的RNA和蛋白。
结果:阳性转基因小鼠DNA的PCR扩增鉴定显示为内参(324bp)和所转基因(173bp)两条带,脑片的激光共聚焦扫描结果显示海马和皮层均可见大小较均一的圆点状绿色荧光。经FACS分选后的细胞几乎全都发出绿色荧光,每次产量约为0.5-1×10~6个存活神经元。获得的纯化神经元中抽提的RNA和蛋白可用于RT-PCR和western blot检测。
结论:利用FACS可以从thy1 -GFP-J转基因小鼠脑内获得单一纯化的存活神经元群。该技术的应用将为活体神经元细胞特异性的基因表达和蛋白定量分析研究提供一个可行方案。
Part One Cortical neurons re-enter cell cycle and undergo apoptosis Induced by oxygen-glucose deprivation in vitro
Objective: To definite the relationship between cell cyler re-entery and neuronal apoptosis in cultured cortical neurons following oxygen-glucose deprivation (OGD) and to explore the potential mechanism of apoptosis in post-mitotic neurons induced by OGD.
Methods: The model of OGD injury on cultured cortical neurons was established. The cultured neurons were randomly divided into 5 groups: control group, the group of 6h, 12h, 24h reoxygenation after 1h OGD and roscovitine treated group. Flow cytometry was used to detect the percent of neuronal apoptosis after 1h OGD. Immunofluorescence staining was used to detect uptake of BrdU and expression of phosphorylated Rb protein (p-Rb, ser 795) in neurons, and their co-localizated relationship with the TUNEL staining. Western Bolt was used to detect the expression of p-Rb and E2F1.
Results: Compared with the control group, the percentage of neurons with BrdU uptake ,TUNEL staining and co-localization of both were increased in the 6h, 12h, 24h reoxygenation after 1h OGD. The expression of p-Rb in each group after OGD were significantly increased, in parallel with BrdU uptake. The co-localization of p-Rb and TUNEL in 6h and 12h after OGD was apparent, but the percent of co-localization between p-Rb and TUNEL was strikingly decreased in 24h after OGD. The expression of p-Rb and E2F1 and the percent of neuronal apoptosis were decreased in roscovitine treated group.
Conclusion: Attempted cell cycle re-entry with phosphorylation of Rb in neurons induces neuronal apoptosis after OGD treatment. Inhibition of phosphorylation of Rb following OGD reduces neuronal apoptosis, phosphorylated Rb pathway is one of the mechanisms of neuronal apoptosis induced by OGD.
Part Two The involvement of upregulation and translocation of phospho-Rb in early neuronal apoptosis following focal cerebral ischemia in rats
Objective: To investigate the expression and subcellular localization of phospho-Rb (ser 795), and temporal and spatial relationship between it and neuronal apoptotic death in rats subjected to transient focal cerebral ischemia.
Methods: The model of middle cerebral artery occlusion (MCAO) was carried out. All animals were divided into five groups: sham-operated controls, 12 h, 1 d, 3 d, and 7 d after MCAO/reperfusion. By analyzing DNA fragmentation with a TUNEL assay, apoptotic cells were examined in the peri-infarct area of the ischemic cerebral cortex. Immunofluorescence staining was used to detect the expression and subcellular localization of phospho-Rb (ser 795) in the each time points after MCAO/reperfusion. Double-labeling analysis of TUNEL and phospho-Rb staining was used to detect the temporal and spatial relationship between phospho-Rb and neuronal apoptotic death.
Results: Compared with sham-operated controls, at 12 h after MCAO/ reperfusion TUNEL-positive cells began to be detected, and the immunoreactivity of phospho-Rb in neurons was increased. One day after the transient ischemic injury, it was showed that subcellular localization of phospho-Rb translocated from the nucleus to the cytoplasm in many neurons in the peri-infarct area and the number of TUNEL-positive cells peaked at this time point. Most phospho-Rb appeared to be localized within TUNEL staining cells at 12 h and 1 d after MCAO/reperfusion in an overlapping and mosaic pattern. However, there was no apparent colocalization of phospho-Rb and TUNEL staining at 3 d and 7 d, although the number of phospho-Rb translocated neurons and TUNEL stained cells were still at high levels at these time points.
Conclusion: phospho-Rb may be involved in the early stages of neuronal apoptosis after transient cerebral ischemia, and it may not be involved or in an indirect way in the late stages of neuronal apoptosis.
Part Three Sorting of neurons in cortex and hippocampus from thy1-GFP-J transgenic mice by flow cytometry
Objective: To establish an effective method fast getting purified neuronal populations from mammalian brain in vivo.
Methods: Thy1-GFP-J transgenic mice, which expressed green fluorescent protein (GFP) in neurons, were confirmed by RT-PCR and confocal laser scan microscopy. The cortex and hippocampus of 2-months-old transgenic mice were dissected and then dissociated using the papin dissociation system. Single cells of brain tissue were treated with PI to label dead cells and sorted on a FACS-Aria cell sorter. Cells with high GFP signals and negative PI signals were selected, then RNA and protein were extracted from the sorted cells.
Results: PCR amplification of DNA from tail of transgenic mice indicated that it appears two bands inculding transgene band (173bp) and internal control band (324bp), and the pictures of brain tissue captured by confocal microscopy showed that green fluorescent spots with similar round patterns can be seen in the cortex and hippocampus of these mice. After sorting, almost all cells seemed brightly fluorescent in their soma, and about 0.5-1×10~6 lived neurons were obtained in each sort. RNA and protein extracted from purified neurons were detected by RT-PCR and western blot.
Conclusion: It is a reliable and practical method for study of neuronal cell type-specific gene expression profiling and quantitative analysis of proteins extracted from purified neuronal populations, which obtained from mammalian brain of adult thy1-GFP-J transgenic mice by FACS in vivo.
引文
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7. Dick, FA and Dyson, N. pRB contains an E2F1-specific binding domain that allows E2F1-induced apoptosis to be regulated separately from other E2F activities. Mol Cell. 2003, 12(3): 639-649.
8. Denchi, EL and Helin, K. E2F1 is crucial for E2F-dependent apoptosis. EMBO Rep. 2005, 6(7): 661-668.
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6. Hata R, Mies G, Wiessner C et al. A reproducible model of middle cerebral artery occlusion in mice: hemodynamic, biochemical, and magnetic resonance imaging. J Cereb Blood Flow Metab. 1998,18:367-375
7. Bederson JB, Pitts LH, Tsuji M, et al. Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination. Stroke. 1986, 17(3): 472-476
8. Wen Y, Yang S, Liu R et al. Estrogen attenuates nuclear factor-kappa B activation induced by transient cerebral ischemia. Brain Res. 2004, 1008:147-154
9. Weinberg RA. The retinoblastoma protein and cell cycle control. Cell. 1995, 81:323-330
10. Doonan, F and Cotter, TG. Apoptosis: A potential therapeutic target for retinal degenerations. Curr Neurovasc Res. 2004, 1(1): 41-53
11. Ferretti, P. Neural stem cell plasticity: Recruitment of endogenous populations for regeneration. Curr Neurovasc Res. 2004, 1(3): 215-229
12. Koyama, R and Ikegaya, Y. Mossy fiber sprouting as a potential therapeutic target for epilepsy. Curr Neurovasc Res. 2004, 1(1): 3-10
13. Li, F, Chong, ZZ and Maiese, K. Erythropoietin on a Tightrope: Balancing Neuronal and Vascular Protection between Intrinsic and Extrinsic Pathways. Neurosignals. 2004,13(6): 265-289
14. El-Khodor BF, Oo TF, Kholodilov N, et al. Ectopic expression of cell cycle markers in models of induced programmed cell death in dopamine neurons of the rat substantia nigrapars compacta. Exp Neurol. 2003,179(l):17-27
15. Rideout HJ, Wang Q, Park DS, et al. Cyclin-dependent kinase activity is required for apoptotic death but not inclusion formationin cortical neurons after proteasomal inhibition. J Neurosci. 2003,23(4):1237-1245
16. Ino H, Chiba T. Cyclin-dependent kinase 4 and cyclin D1 are required for excitotoxin-induced neuronal cell death in vivo. J Neurosci. 2001,21(16): 6086-6094
17. Padmanabhan J, Park DS, Greene LA et al. Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. J Neurosci. 1999,19:8747-8756
18. Park DS, Morris EJ, Padmanabhan J et al. Cyclin-dependent kinases participate in death of neurons evoked by DNA-damaging agents. J Cell Biol. 1998,143:457-467
19. Wen Y, Yang S, Liu R et al. Transient Cerebral Ischemia Induces Aberrant Neuronal Cell Cycle Re-entry and Alzheimer's Disease-like Tauopathy in Female Rats. J Biol Chem. 2004, 279:22684-22692
20. Kuan CY, Schloemer AJ, Lu A, et al. Hypoxia-ischemia induces DNA synthesis without cell proliferation in dying neurons in adult rodent brain. J Neurosci.2004,24(47):10763-10772
21. Chen B, Wang W. The expression of Cyclins in neurons of rats after focal cerebral ischemia. J Huazhong Univ Sci Technolog Med Sci. 2008, 28:60-64
22. Wen Y, Yang S, Liu R et al. Cell-cycle regulators are involved in transient cerebral ischemia induced neuronal apoptosis in female rats. FEBS Lett. 2005, 79:4591- 4599
23. Sears RC, Nevins JR. Signaling networks that link cell proliferation and cell fate. J Biol Chem. 2002, 277:11617-11620
24. O'Hare M, Wang F, Park DS. Cyclin-dependent kinases as potential targets to improve stroke outcome. Pharmacol Ther. 2002, 93: 135-143
25. Nguyen MD, Mushynski WE, Julien JP. Cycling at the interface between neurodevelopment and neurodegeneration. Cell Death Differ. 2002, 9:1294-1306
26. Byrnes KR, Faden AI. Role of cell cycle proteins in CNS injury. Neurochem Res. 2007, 32(10):1799-1807
27. Wang W, Redecker C, Bidmon HJ et al. Delayed neuronal death and damage of GDNF family receptors in CA1 following focal cerebral ischemia. Brain Res. 2004, 1023:92-101
28. Cho BB, Toledo-Pereyra LH. Caspase-independent programmed cell death following ischemic stroke. J Invest Surg. 2008,21:141-14
29. Park DS, Morris EJ, Bremner R et al. Involvement of retinoblastoma family members and E2F/DP complexes in the death of neurons evoked by DNA damage. J Neurosci.2000,20:3104-3114
30. Liu DX, Greene LA. Regulation of neuronal survival and death by E2F-dependent gene repression and derepression. Neuron. 2001, 32:425-438
1. Cahoy JD, Emery B, Kaushal A, et al. A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci, 2008, 28(1):264-278
2. Lobo MK, Karsten SL, Gray M, et al. FACS-array profiling of striatal projection neuron subtypes in juvenile and adult mouse brains. Nat Neurosci, 2006, 9(3):443-452
3. Georgious G, Stathopoulos C, Daugherty PS, et al. Display of heterologous proteins on the surface of microorganisms: From the screening of combinatorial libraries to live recombinant vaccines. Nat Biotechnol, 1997,13(1):29-34
4．曹云新，胡金涛，杨安钢．FACS-Aria流式细胞分选调试参数和最佳条件的设定．第四军医大学学报，2007，28(19)：1813-1815
5. Sergent-Tanguy S, Chagneau C, Neveu I, et al. Fluorescent activated cell sorting (FACS): a rapid and reliable method to estimate the number of neurons in a mixed population. J Neurosci Methods, 2003, 129(1):73-79
6. Ford AL, Goodsall AL, Hickey WF, et al. Normal adult ramified microglia separated from other central nervous system macrophages by flow cytometric sorting. Phenotypic differences defined and direct ex vivo antigen presentation to myelin basic protein-reactive CD4+ T cells compared. J Immunol, 1995, 154(9):4309-4321
7. 7 Feng G, Mellor RH, Bernstein M, et al. Imaging neuronal subsets in transgenic mice expressing multiple spectral variants of GFP. Neuron, 2000, 28(1):41-51
8. McLaren FH, Svendsen CN, Van der Meide P, Jolv E. Analysis of neural stem cells by flow cytometry: cellular differentiation modifies patterns of MHC expression. J Neuroimmunol. 2001,112 ( 1-2):35-46
9. Fujikawa T, Hirose T, Fujii H, et al. Purification of adult hepatic progenitor cells using green fluorescent protein (GFP)-transgenic mice and fluorescence- activated cell sorting. J Hepatol. 2003;39(2): 162-70
10. Tomomura M, Rice DS, Morgan JI, et al. Purification of Purkinje cells by fluorescence-activated cell sorting from transgenic mice that express green fluorescent protein. Eur J Neurosci. 2001 Jul;14(1):57-63
11. Malatesta P, Hartfuss E, Gotz M, et al. Isolation of radial glial cells by fluorescent-activated cell sorting reveals a neuronal lineage. Development. 2000 Dec;127(24):5253-63
12. Hedlund E, Pruszak J, Lardaro T, et al. Embryonic stem cell-derived Pitx3-enhanced green fluorescent protein midbrain dopamine neurons survive enrichment by fluorescence-activated cell sorting and function in an animal model of Parkinson's disease. Stem Cells. 2008;26(6):1526-36
1. Yang Y, Varvel NH, Lamb BT, et al. Ectopic cell cycle events link human Alzheimer's disease and amyloid precursor protein transgenic mouse models. J Neurosci. 2006, 26(3):775-784
2. Hoqlinger GU, Breunig JJ, Depboylu C, et al. The pRb/E2F cell-cycle pathway mediates cell death in Parkinson's disease. Proc Natl Acad Sci USA. 2007,104(9):3585-3590
3. Lee SS, Kim YM, Junn E, et al. Cell cycle aberrations by alpha-synuclein over-expression and cyclin B immunoreactivity in Lewy bodies. Neurobiol Aging. 2003,24(5):687-696
4. Lopes JP, Oliveira CR, Aqostinho P. Cdk5 acts as a mediator of neuronal cell cycle re-entry triggered by amyloid-beta and prion peptides. Cell Cycle. 2009, 8(1): 97-104
5. Timsit S, Menn B. Cerebral ischemia, cell cycle elements and cdk5. Biotechnol J. 2007,2(8): 958-966
6. Vermeulen K, Van Bockstaele DR and Berneman ZN. The cell cycle: a review of regulation, deregulation and therapeutic targets in cancer. Cell Prolif. 2003, 36:131-149
7. Hulleman E , Boonsha J. Regulation of Gl phase progressionby growt h factors and the extracellular mat rix . Cell Mol Life Sci. 2001 ,58 (1): 80-93
8. Vermeulen K, Berneman ZN and Van Bockstaele DR. Cell cycle and apoptosis. Cell Prolif. 2003, 36: 165-175
9. Herrup K and Busser JC. The induction of multiple cell cycle events precedes target-related neuronal death. Development. 1995, 121: 2385-2395
10. Schmetsdorf S, Gartner U, Arendt T. Expression of cell cycle-related proteins in developing and adult mouse hippocampus. Int J Dev Neurosci. 2005, 23(1):101-112
11. Legrier ME, Ducray A, Propper A, et al. Region-specific expression of cell cycle inhibitors in the adult brain. Neuroreport. 2001, 12: 3127-3131
12. Akashiba H, Matsuki N, Nishiyama N. p27 small interfering RNA induces cell death through elevating cell cycle activity in cultured cortical neurons: a proof-of-concept study. Cell Mol Life Sci. 2006, 63(19-20):2397-2404
13. Schmetsdorf S, Gartner U, Arendt T. Constitutive Expression of Functionally Active Cyclin-Dependent Kinases and Their Binding Partners Suggests Noncanonical Functions of Cell Cycle Regulators in Differentiated Neurons. Cereb Cortex. 2007,17(8): 1821-1829
14. Yoshikawa K. Cell cycle regulators in neural stem cells and postmitotic neurons. Neurosci Res. 2000, 37(1):1-14
15. Nquyen MD, Mushynski WE, Julien JP. Cycling at the interface between neurodevelopment and neurodegeneration. Cell Death Differ. 2002, 9(12): 1294-1306
16. Panickar KS, Nonner D, White MG, et al. Overexpression of Cdk5 or non-phosphorylatable retinoblastorma protein protects septal neurons from oxygen-glucose deprivation. Neurochem Res. 2008, 33(9): 1852-1858
17. Verdaguer E, Jimenez A, Canudas AM et al. Inhibition of cell cycle pathway by flavopiridol promotes survival of cerebellar granule cells after an excitotoxic treatment. J Pharmacol Exp Ther. 2004, 308:609-616
18. Kranenburg O, van der Eb AJ, Zantema A. Cyclin D1 is an essential mediator of apoptotic neuronal cell death. Embo J. 1996, 15:46-54
19. Padmanabhan J, Park DS, Greene LA et al. Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. J Neurosci. 1999, 19: 8747-8756
20. Rashidian J, Iyirhiaro G, Aleyasin H, et al. Multiple cyclindependent kinases signals are critical mediators of ischemia/hypoxic neuronal death in vitro and in vivo. Proc Natl Acad Sci USA. 2005, 102:14080-14085
21. Park DS, Morris EJ, Bremner R, et al. Involvement of retinoblastoma family members and E2F/DP complexes in the death of neurons evoked by DNA damage. J Neurosci.2000,20:3104-3114
22. Chong ZZ, Li F, Maiese K. Attempted cell cycle induction in post-mitotic neurons occurs in early and late apoptotic programs through Rb, E2F1, and caspase 3. Curr Neurovasc Res. 2006; 3(1):25-39
23. Liu DX, Biswas SC, Greene LA. B-myb and C-myb play required roles in neuronal apoptosis evoked by nerve growth factor deprivation and DNA damage. J Neurosci. 2004, 24:8720-8725
24. Nahle Z, Polakoff J, Davuluri RV, et al. Direct coupling of the cell cycle and cell death machinery by E2F. Nat Cell Biol. 2002,4:859-864
25. Greene LA, Biswas SC, Liu DX. Cell cycle molecules and vertebrate neuron death: E2F at the hub. Cell Death Differ. 2004,11:49-60
26. Honma H, Gross L, Windebank AJ. Hypoxia-induced apoptosis of dorsal root ganglion neurons is associated with DNA damage recognition and cell cycle disruption in rats. Neurosci Lett. 2004, 354(2):95-98
27. Nguyen MD, Boudreau M, Kriz J, et al. Cell cycle regulators in the neuronal death pathway of amyotrophic lateral sclerosis caused by mutant superoxide dismutase 1. J Neurosci. 2003, 23: 2131-2140
28. Konishi Y, Bonni A. The E2F-Cdc2 cell-cycle pathway specifically mediates activity deprivation-induced apoptosis of postmitotic neurons. J Neurosci. 2003,23(5):1649-1658
29. Nakatsuji Y, Miller RH. Density dependent modulation of cell cycle protein expression in astrocytes. J Neurosci Res. 2001, 66(3):487-496
30. Kato H, Takahashi A, Itoyama Y. Cell cycle protein expression in proliferating microglia and astrocytes following transient global cerebral ischemia in the rat. Brain Res Bull. 2003, 60(3): 215-221
31. Luo X, Yu Z, Feng Y, et al. Effects of ischemia and anoxia on cell activation and cell cycle of cultured astrocytes in vitro. J Huazhong Univ Sci Technolog Med Sci. 2006,26(l):21-24
2. Osuga H, Osuga S, Wang F, et al. Cyclin-dependent kinases as a therapeutic target for stroke. Proc Natl Acad Sci USA. 2000, 97: 10254-10259.
3. Wang F, Corbett D, Osuga H, et al. Inhibition of cyclin-dependent kinases improves CA1 neuronal survival and behavioral performance after global ischemia in the rat. J Cereb Blood Flow Metab. 2002,22: 171-182.
4. Rashidian J, Iyirhiaro G, Aleyasin H, et al. Multiple cyclin-dependent kinases signals are critical mediators of ischemia/hypoxic neuronal death in vitro and in vivo. Proc Natl Acad Sci USA. 2005, 102:14080-14085.
5. Yoshikawa K. Cell cycle regulators in neural stem cells and postmitotic neurons. Neurosci Res. 2000, 37(1): 1-14.
6. Liu DX and Greene LA. Neuronal apoptosis at the G1/S cell cycle checkpoint. Cell Tissue Res. 2001, 305: 217-228.
7. Dick, FA and Dyson, N. pRB contains an E2F1-specific binding domain that allows E2F1-induced apoptosis to be regulated separately from other E2F activities. Mol Cell. 2003, 12(3): 639-649.
8. Denchi, EL and Helin, K. E2F1 is crucial for E2F-dependent apoptosis. EMBO Rep. 2005, 6(7): 661-668.
1. Copani A, Uberti D, Sortino MA, et al. Activation of cell-cycle-associated proteins in neuronal death: a mandatory or dispensable path? Trends Neurosci. 2001,24:25-31
2. Rashidian J, Iyirhiaro G, Aleyasin H, et al. Multiple cyclin-dependent kinases signals are critical mediators of ischemia/hypoxic neuronal death in vitro and in vivo. Proc Natl Acad Sci USA. 2005,102:14080-14085
3. Wang F, Corbett D, Osuga H, et al. Inhibition of cyclin-dependent kinases improves CA1 neuronal survival and behavioral performance after global ischemia in the rat. J Cereb Blood Flow Metab. 2002,22:171-182
4. Wen Y, Yang S, Liu R, et al. Cell-cycle regulators are involved in transient cerebral ischemia induced neuronal apoptosis in female rats. FEBS Lett. 2005, 579:4591-4599
5. Harbour JW, Dean DC. The Rb/E2F pathway: expanding roles and emerging paradigms.Genes Dev. 2000,14:2393-2409
6. Liu DX, Greene LA. Regulation of neuronal survival and death by E2F-dependent gene repression and derepression. Neuron. 2001, 32:425-438
7. Liu DX, Greene LA. Neuronal apoptosis at the G1/S cell cycle checkpoint. Cell Tissue Res. 2001, 305:217-228
8. Greene LA, Liu DX, Troy CM et al. Cell cycle molecules define a pathway required for neuron death in development and disease. Biochim Biophys Acta. 2007,1772:392-401
9. Padmanabhan J, Park DS, Greene LA et al. Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. J Neurosci. 1999,19:8747 -8756
10. Park DS, Morris EJ, Padmanabhan J et al. Cyclin-dependent kinases participate in death of neurons evoked by DNA-damaging agents. J Cell Biol. 1998,143:457-467
11. Copani A, Condorelli F, Caruso A et al. Mitotic signaling by beta-amyloid causes neuronal death. FASEB J. 1999, 13:2225-2234
12. Park DS, Obeidat A, Giovanni A et al. Cell cycle regulators in neuronal death evoked by excitotoxic stress: implications for neurodegeneration and its treatment. Neurobiol Aging. 2000,21:771-781
13. Park DS, Morris EJ, Bremner R et al. Involvement of retinoblastoma family members and E2F/DP complexes in the death of neurons evoked by DNA damage. J Neurosci. 2000,20:3104-3114
14. Katchanov J, Harms C, Gertz K, et al. Mild Cerebral Ischemia Induces Loss of Cyclin-Dependent Kinase Inhibitors and Activation of Cell Cycle Machinery before Delayed Neuronal Cell Death. J Neurosci. 2001,21(14): 5045-5053
15. Manthrope M, Adler R, Varon S. Development, reactivity and GFAP immunofluorescence of astroglia-containing monolayer cultures from rat cerebrum. J neurocyto,1979, 8:605
16. Gwag BJ, Lobner D, Koh JY, et al. Blockade of glutamate receptors unmasks neuronal apoptosis after oxygen glucose deprivation in vitro. Neuroscience. 1995,68: 615-619
17. Gong QH, Wang Q, Shi JS, et al. Inhibition of caspases and intracellular free Ca2+concentrations are involved in resveratrol protection against apoptosis in rat primary neuron cultures. Acta Pharmacol Sin. 2007,28(11): 1724-1730
18. Dick, FA and Dyson, N. pRB contains an E2F1-specific binding domain that allows E2F1-induced apoptosis to be regulated separately from other E2F activities. Mol Cell.2003, 12(3): 639-649
19. Denchi, EL and Helin, K. E2F1 is crucial for E2F-dependent apoptosis. EMBO Rep.2005, 6(7): 661-668
20. Osuga H, Osuga S, Wang F, et al. Cyclin-dependent kinases as a therapeutic target for stroke. Proc Natl Acad Sci USA. 2000, 97: 10254-10259
21. Nguyen MD, Boudreau M, Kriz J, et al. Cell cycle regulators in the neuronal death pathway of amyotrophic lateral sclerosis caused by mutant superoxide dismutase 1. J Neurosci. 2003, 23: 2131-2140
22. Jordan-Sciutto KL, Malaiyandi LM and Bowser R. Altered distribution of cell cycle transcriptional regulators during Alzheimer disease. J. Neuropathol. Exp. Neurol. 2002,61:358-367
23. Ranganathan S and Bowser R. Alterations in G(l) to S phase cell-cycle regulators during amyotrophic lateral sclerosis. Am. J. Pathol. 2003, 162: 823-835
1. Dimagl U, Iadecola C, Moskowitz MA. Pathobiology of ischaemic stroke: an integrated view. Trends Neurosci. 1999,22:391-397
2. Hayashi T, Sakai K, Sasaki C et al. Phosphorylation of retinoblastoma protein in rat brain after transient middle cerebral artery occlusion. Neuropathol Appl Neurobiol.2000,26:390-397
3. Katchanov J, Harms C, Gertz K et al. Mild cerebral ischemia induces loss of cyclin-dependent kinase inhibitors and activation of cell cycle machinery before delayed neuronal cell death. J Neurosci. 2001, 21:5045-5053
4. Osuga H, Osuga S, Wang F et al. Cyclin-dependent kinases as a therapeutic target for stroke. Proc Natl Acad Sci USA. 2000, 97:10254-10259
5. Connolly ES Jr, Winfree CJ, Stern DM et al. Procedural and strain-related variables significantly affect outcome in a murine model of focal cerebral ischemia. Neurosurgery.1996,38:523-531
6. Hata R, Mies G, Wiessner C et al. A reproducible model of middle cerebral artery occlusion in mice: hemodynamic, biochemical, and magnetic resonance imaging. J Cereb Blood Flow Metab. 1998,18:367-375
7. Bederson JB, Pitts LH, Tsuji M, et al. Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination. Stroke. 1986, 17(3): 472-476
8. Wen Y, Yang S, Liu R et al. Estrogen attenuates nuclear factor-kappa B activation induced by transient cerebral ischemia. Brain Res. 2004, 1008:147-154
9. Weinberg RA. The retinoblastoma protein and cell cycle control. Cell. 1995, 81:323-330
10. Doonan, F and Cotter, TG. Apoptosis: A potential therapeutic target for retinal degenerations. Curr Neurovasc Res. 2004, 1(1): 41-53
11. Ferretti, P. Neural stem cell plasticity: Recruitment of endogenous populations for regeneration. Curr Neurovasc Res. 2004, 1(3): 215-229
12. Koyama, R and Ikegaya, Y. Mossy fiber sprouting as a potential therapeutic target for epilepsy. Curr Neurovasc Res. 2004, 1(1): 3-10
13. Li, F, Chong, ZZ and Maiese, K. Erythropoietin on a Tightrope: Balancing Neuronal and Vascular Protection between Intrinsic and Extrinsic Pathways. Neurosignals. 2004,13(6): 265-289
14. El-Khodor BF, Oo TF, Kholodilov N, et al. Ectopic expression of cell cycle markers in models of induced programmed cell death in dopamine neurons of the rat substantia nigrapars compacta. Exp Neurol. 2003,179(l):17-27
15. Rideout HJ, Wang Q, Park DS, et al. Cyclin-dependent kinase activity is required for apoptotic death but not inclusion formationin cortical neurons after proteasomal inhibition. J Neurosci. 2003,23(4):1237-1245
16. Ino H, Chiba T. Cyclin-dependent kinase 4 and cyclin D1 are required for excitotoxin-induced neuronal cell death in vivo. J Neurosci. 2001,21(16): 6086-6094
17. Padmanabhan J, Park DS, Greene LA et al. Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. J Neurosci. 1999,19:8747-8756
18. Park DS, Morris EJ, Padmanabhan J et al. Cyclin-dependent kinases participate in death of neurons evoked by DNA-damaging agents. J Cell Biol. 1998,143:457-467
19. Wen Y, Yang S, Liu R et al. Transient Cerebral Ischemia Induces Aberrant Neuronal Cell Cycle Re-entry and Alzheimer's Disease-like Tauopathy in Female Rats. J Biol Chem. 2004, 279:22684-22692
20. Kuan CY, Schloemer AJ, Lu A, et al. Hypoxia-ischemia induces DNA synthesis without cell proliferation in dying neurons in adult rodent brain. J Neurosci.2004,24(47):10763-10772
21. Chen B, Wang W. The expression of Cyclins in neurons of rats after focal cerebral ischemia. J Huazhong Univ Sci Technolog Med Sci. 2008, 28:60-64
22. Wen Y, Yang S, Liu R et al. Cell-cycle regulators are involved in transient cerebral ischemia induced neuronal apoptosis in female rats. FEBS Lett. 2005, 79:4591- 4599
23. Sears RC, Nevins JR. Signaling networks that link cell proliferation and cell fate. J Biol Chem. 2002, 277:11617-11620
24. O'Hare M, Wang F, Park DS. Cyclin-dependent kinases as potential targets to improve stroke outcome. Pharmacol Ther. 2002, 93: 135-143
25. Nguyen MD, Mushynski WE, Julien JP. Cycling at the interface between neurodevelopment and neurodegeneration. Cell Death Differ. 2002, 9:1294-1306
26. Byrnes KR, Faden AI. Role of cell cycle proteins in CNS injury. Neurochem Res. 2007, 32(10):1799-1807
27. Wang W, Redecker C, Bidmon HJ et al. Delayed neuronal death and damage of GDNF family receptors in CA1 following focal cerebral ischemia. Brain Res. 2004, 1023:92-101
28. Cho BB, Toledo-Pereyra LH. Caspase-independent programmed cell death following ischemic stroke. J Invest Surg. 2008,21:141-14
29. Park DS, Morris EJ, Bremner R et al. Involvement of retinoblastoma family members and E2F/DP complexes in the death of neurons evoked by DNA damage. J Neurosci.2000,20:3104-3114
30. Liu DX, Greene LA. Regulation of neuronal survival and death by E2F-dependent gene repression and derepression. Neuron. 2001, 32:425-438
1. Cahoy JD, Emery B, Kaushal A, et al. A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci, 2008, 28(1):264-278
2. Lobo MK, Karsten SL, Gray M, et al. FACS-array profiling of striatal projection neuron subtypes in juvenile and adult mouse brains. Nat Neurosci, 2006, 9(3):443-452
3. Georgious G, Stathopoulos C, Daugherty PS, et al. Display of heterologous proteins on the surface of microorganisms: From the screening of combinatorial libraries to live recombinant vaccines. Nat Biotechnol, 1997,13(1):29-34
4．曹云新，胡金涛，杨安钢．FACS-Aria流式细胞分选调试参数和最佳条件的设定．第四军医大学学报，2007，28(19)：1813-1815
5. Sergent-Tanguy S, Chagneau C, Neveu I, et al. Fluorescent activated cell sorting (FACS): a rapid and reliable method to estimate the number of neurons in a mixed population. J Neurosci Methods, 2003, 129(1):73-79
6. Ford AL, Goodsall AL, Hickey WF, et al. Normal adult ramified microglia separated from other central nervous system macrophages by flow cytometric sorting. Phenotypic differences defined and direct ex vivo antigen presentation to myelin basic protein-reactive CD4+ T cells compared. J Immunol, 1995, 154(9):4309-4321
7. 7 Feng G, Mellor RH, Bernstein M, et al. Imaging neuronal subsets in transgenic mice expressing multiple spectral variants of GFP. Neuron, 2000, 28(1):41-51
8. McLaren FH, Svendsen CN, Van der Meide P, Jolv E. Analysis of neural stem cells by flow cytometry: cellular differentiation modifies patterns of MHC expression. J Neuroimmunol. 2001,112 ( 1-2):35-46
9. Fujikawa T, Hirose T, Fujii H, et al. Purification of adult hepatic progenitor cells using green fluorescent protein (GFP)-transgenic mice and fluorescence- activated cell sorting. J Hepatol. 2003;39(2): 162-70
10. Tomomura M, Rice DS, Morgan JI, et al. Purification of Purkinje cells by fluorescence-activated cell sorting from transgenic mice that express green fluorescent protein. Eur J Neurosci. 2001 Jul;14(1):57-63
11. Malatesta P, Hartfuss E, Gotz M, et al. Isolation of radial glial cells by fluorescent-activated cell sorting reveals a neuronal lineage. Development. 2000 Dec;127(24):5253-63
12. Hedlund E, Pruszak J, Lardaro T, et al. Embryonic stem cell-derived Pitx3-enhanced green fluorescent protein midbrain dopamine neurons survive enrichment by fluorescence-activated cell sorting and function in an animal model of Parkinson's disease. Stem Cells. 2008;26(6):1526-36
1. Yang Y, Varvel NH, Lamb BT, et al. Ectopic cell cycle events link human Alzheimer's disease and amyloid precursor protein transgenic mouse models. J Neurosci. 2006, 26(3):775-784
2. Hoqlinger GU, Breunig JJ, Depboylu C, et al. The pRb/E2F cell-cycle pathway mediates cell death in Parkinson's disease. Proc Natl Acad Sci USA. 2007,104(9):3585-3590
3. Lee SS, Kim YM, Junn E, et al. Cell cycle aberrations by alpha-synuclein over-expression and cyclin B immunoreactivity in Lewy bodies. Neurobiol Aging. 2003,24(5):687-696
4. Lopes JP, Oliveira CR, Aqostinho P. Cdk5 acts as a mediator of neuronal cell cycle re-entry triggered by amyloid-beta and prion peptides. Cell Cycle. 2009, 8(1): 97-104
5. Timsit S, Menn B. Cerebral ischemia, cell cycle elements and cdk5. Biotechnol J. 2007,2(8): 958-966
6. Vermeulen K, Van Bockstaele DR and Berneman ZN. The cell cycle: a review of regulation, deregulation and therapeutic targets in cancer. Cell Prolif. 2003, 36:131-149
7. Hulleman E , Boonsha J. Regulation of Gl phase progressionby growt h factors and the extracellular mat rix . Cell Mol Life Sci. 2001 ,58 (1): 80-93
8. Vermeulen K, Berneman ZN and Van Bockstaele DR. Cell cycle and apoptosis. Cell Prolif. 2003, 36: 165-175
9. Herrup K and Busser JC. The induction of multiple cell cycle events precedes target-related neuronal death. Development. 1995, 121: 2385-2395
10. Schmetsdorf S, Gartner U, Arendt T. Expression of cell cycle-related proteins in developing and adult mouse hippocampus. Int J Dev Neurosci. 2005, 23(1):101-112
11. Legrier ME, Ducray A, Propper A, et al. Region-specific expression of cell cycle inhibitors in the adult brain. Neuroreport. 2001, 12: 3127-3131
12. Akashiba H, Matsuki N, Nishiyama N. p27 small interfering RNA induces cell death through elevating cell cycle activity in cultured cortical neurons: a proof-of-concept study. Cell Mol Life Sci. 2006, 63(19-20):2397-2404
13. Schmetsdorf S, Gartner U, Arendt T. Constitutive Expression of Functionally Active Cyclin-Dependent Kinases and Their Binding Partners Suggests Noncanonical Functions of Cell Cycle Regulators in Differentiated Neurons. Cereb Cortex. 2007,17(8): 1821-1829
14. Yoshikawa K. Cell cycle regulators in neural stem cells and postmitotic neurons. Neurosci Res. 2000, 37(1):1-14
15. Nquyen MD, Mushynski WE, Julien JP. Cycling at the interface between neurodevelopment and neurodegeneration. Cell Death Differ. 2002, 9(12): 1294-1306
16. Panickar KS, Nonner D, White MG, et al. Overexpression of Cdk5 or non-phosphorylatable retinoblastorma protein protects septal neurons from oxygen-glucose deprivation. Neurochem Res. 2008, 33(9): 1852-1858
17. Verdaguer E, Jimenez A, Canudas AM et al. Inhibition of cell cycle pathway by flavopiridol promotes survival of cerebellar granule cells after an excitotoxic treatment. J Pharmacol Exp Ther. 2004, 308:609-616
18. Kranenburg O, van der Eb AJ, Zantema A. Cyclin D1 is an essential mediator of apoptotic neuronal cell death. Embo J. 1996, 15:46-54
19. Padmanabhan J, Park DS, Greene LA et al. Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. J Neurosci. 1999, 19: 8747-8756
20. Rashidian J, Iyirhiaro G, Aleyasin H, et al. Multiple cyclindependent kinases signals are critical mediators of ischemia/hypoxic neuronal death in vitro and in vivo. Proc Natl Acad Sci USA. 2005, 102:14080-14085
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