DCX表达影响脑胶质瘤生长、转移及放射敏感性的作用机制研究
|
摘要
第一部分构建稳定表达的U87-DCX、U87-NG细胞株
目的：构建针对DCX的质粒载体，转染U87-MG细胞株，通过筛选获得DCX表达的稳定转染的细胞模型。
方法：采用UbC-GFP-L.V.慢病毒感染U87-MG细胞株，选择感染效率高、MOI值高、对细胞毒性低为最佳感染条件，获得U87-DCX和U87-NG细胞株，用PCR和Western Blot方法检测。
结果：U87-DCX细胞为DCX过表达的稳定转染细胞，目的基因DCX融合GFP和FLAG共同表达；Western Blot检测到72-95KD处有阳性条带；其大小和DCX-GFP-FLAG融合蛋白（48KDa+28KDa+2KDa＝78KDa）相吻合。表达克隆中插入目的基因片段大小为1326bp。
结论：U87-DCX、U87-NG细胞株构建成功。
第二部分基因芯片筛选与DCX和放射相关的差异基因
目的：基因芯片筛选转染DCX的U87-MG细胞在照射前后的差异基因，探讨与DCX和电离辐射均具有相关性的下游基因在胶质瘤放射治疗中的作用。
方法：将U87-MG细胞、U87-DCX细胞经60Coγ射线（10Gy，剂量率1.0Gy/min）照射，用基因芯片技术筛选差异基因，均值两两比较，取P <0.05的差异基因分析。
结果：放射和DCX转染均有相关性差异基因有：EIF5A、LYN、SPN；与放射相关的差异基因有：IMAA、LDHA、WIBG、UBC；与DCX相关的差异基因有：PLS3、IL11、NOV。U87-MG细胞、U87-DCX细胞分别经60Coγ射线（10Gy，剂量率1.0Gy/min）照射，选取SPN与DCX免疫共沉淀实验，结果表明，U87-MG细胞照射组与对照组均无DCX和SPN蛋白表达，而U87-DCX细胞照射组与对照组DCX和SPN蛋白均表达，并且DCX和SPN的蛋白表达水平在10Gy照射组明显高于对照组。
结论：表达谱基因芯片技术能快速、灵敏地筛选出U87-DCX细胞在放疗前后的差异基因，通过上述实验证实，在放射条件下DCX蛋白表达水平增高，且与SPN相互作用。
第三部分体外试验研究DCX的放射作用机制
目的：研究DCX基因高表达联合γ-射线照射对胶质瘤细胞生长、浸润转移、DNA损伤修复及凋亡的影响。
方法：1.U87-MG、U87-DCX、U87-NG三种细胞经60Coγ射线照射，剂量分别为0，2，4，6，8，10Gy。收集细胞提取蛋白，用Western Blot方法测定不同放射剂量的DCX蛋白量，并绘制剂量效应曲线。2.测定DCX下游基因在不同放射剂量下的蛋白量，并绘制剂量效应曲线。3.激光共聚焦显微镜观察U87-MG、U87-DCX、U87-NG三种细胞的DCX及SPN蛋白表达水平，并进行数据分析。4.彗星试验研究照射后U87-MG、U87-DCX、U87-NG三种细胞的双链DNA损伤，并进行数据分析。5.流式细胞仪测定细胞凋亡。6.细胞计数方法测定不同细胞的生存曲线以及划痕试验检测细胞的浸润情况。
结果：1.2，4，6，8，10Gy照射条件下DCX、SPN蛋白显示表达水平较对照组高；2.DCX、SPN蛋白在胞浆内共定位表达；3.彗星试验表明U87-DCX细胞的尾长在0、5、10Gy剂量组均较U87-MG、U87-NG细胞长（P<0.05）；4.在辐射作用下，U87-DCX和U87-NG细胞的凋亡率均较低,两者凋亡率比较无统计学意义，U87-DCX细胞的凋亡率照射组与对照组比较无统计学意义（P>0.05）；6.U87-DCX细胞生长、浸润能力较未转染组低。
结论：电离辐射诱导下DCX蛋白表达水平增高，随着照射剂量的增加蛋白表达水平呈上升趋势，并且与下游基因SPN相互作用，DCX在射线所致的细胞损伤过程中发挥作用；DCX能够通过辐射诱导DNA损伤，证实DCX基因转染能够增强U87胶质瘤细胞的放射敏感性；细胞的放射损伤与凋亡之间无明显相关性；转染DCX的U87细胞生长缓慢，缺乏侵袭能力。
第四部分体内试验究DCX对胶质瘤放射敏感性的影响
目的：构建U87-DCX转染细胞的荷瘤鼠模型，研究DCX转染的细胞在体内生长、转移的生物学特性，探讨其放射增敏机制。
方法：构建U87-DCX和U87-NG裸鼠脑胶质瘤原位种植模型；成瘤后以60Coγ射线照射，正常组织以铅板屏蔽，暴露头部照射野，剂量率0.5Gy/min，单次剂量200cGy，每日一次，共五天，总剂量为10Gy。小动物MRI评估小鼠在放射治疗前后的肿瘤大小；Micro PET测量裸鼠接种部位的肿瘤大小及微血管密度SUVmax值；免疫组化观察放疗前后的脑组织内DCX和SPN表达水平。
结果：小动物MRI测量结果表明U87-DCX实验组放疗后肿瘤体积比U87-NG实验组肿瘤体积缩小明显。Micro PET显示经过放射治疗的裸小鼠SUVmax值较放疗前明显下降，其中DCX转染组的裸小鼠下降更为明显，均为60%以上；阴性对照组小鼠在放射治疗后SUVmax值亦略有下降；免疫组化显示两组细胞放射治疗后较对照组微血管密度（MVD）值均有下降，但相同剂量的两组细胞之间无明显差异（P>0.05）。说明放疗可以降低SUVmax。
结论：放射治疗能够有效抑制肿瘤增殖、MVD降低。DCX转染组对放射治疗更为敏感，即肿瘤体积缩小较对照组下降明显；Micro PET可测量到放射治疗后早期SUVmax变化，DCX转染组的SUVmax较对照组明显降低。
Abstract Part One
Objectives：Establishment of doublecortin (DCX) stably expressed U87cells.
Methods: U87cells were infected with Ubc-GFP-Lentivirus. The multiplicity ofnfection (MOI) is10.
Results: PSB276(pLV-UbC-DCX) clone highly express fusion gene DCX in U87ells. Enzyme digestion analysis and sequencing analysis showed that target gene hadeen cloned into recombinant vector and molecular size is1326bp. Then theecombinant was transfected into U87cell by techniques of gene transfection andetected expression by fluoroscopy, DCX-GFP-FLAG was identified by Westernlot(48KDa+28KDa+2KDa＝78KDa).
Conclusion: U87-DCX、U87-NG were established to fulfill.
Abstract Part Two
Objectives: Radiation therapy plays important roles in the treatment ofneurogliocytoma. It is also known that DCX transfected U87-MG cells can inhibittumor cell growth. To determine if there is a relationship between these cells andradiation treatment, microarray analysis was performed to screen for differentiallyexpressed genes in DCX-transfected U87cells before and after radiation in order todiscover DCX-related genes and to reveal the functions of DCX and downstream genesin radiation therapy of neurogliocytoma.
Methods: Stably transfected U87cells were constructed (U87-DCX) and thedifferentially expressed genes were screened via microarray analysis to compare U87cells to U87-DCX cells in both non-irradiated and irradiated conditions. Cells wereirradiated using60Coγ-ray at a dose rate of1.0Gy/min. Mean values were subject topaired comparison analysis, and genes with a p-value less than0.05were analyzed.
Results: Differentially expressed genes that correlated with radiation sensitivityand DCX transfection included eukaryotic initiation factor5A (EIF5A), LYN (v-yes-1Yamaguchi sarcoma viral related oncogene homolog), and neurabin II （SPN）.Differentially expressed genes that only correlated with radiation sensitivity includedIMAA, LDHA, WIBG, and UBC, while differentially expressed genes that were onlyrelated to DCX included PLS3, IL11, and NOV. SPN and DCX were selected forimmunoprecipitation to compare binding activity in both U87-DCX cells and U87cellsin the0and10Gy groups. Results showed that DCX and SPN proteins were notdetected in the two dosage groups in the U87cells, while DCX and SPN proteins weredetected in U87-DCX cells in both two dosage groups, with expression levels of DCXand SPN in the10Gy group higher than those of the0Gy group.
Conclusion: The differential gene expression in U87-DCX cells before and afterradiation provides a rationale for further investigations of the mechanism for radiationtherapy in neurogliocytoma cells. This method can also provide a potential solution forradiation resistance in neurogliocytoma, using gene expression panels as a predictor ofradiation sensitivity. Expression profile microarray technology can rapidly andsensitively predict the radiation sensitivity of neurogliocytoma and screen for relatedradiation sensitive genes.
Abstract Part Three
Objectives：Relationship of expression of DCX overexpressed with60Co γradiation to cell growth, DNA damage and repair, invasion and metastasis，cell cycle distribution and apoptosis.
Methods: U87-DCX, U87-MG and U87-NG cells were treated with radiated by γray of different doses. Immunocoprecipitation and Western Blotting were performed toanalyze the interactive action between DCX and SPN protein in U87-DCX cells after60Coγ radiation. The curve of dosage effect was obtained. The localizations andexpressions of DCX, SPN proteins were detected in U87-MG, U87-DCX and U87-NGcells by immunofluorescence staining and laser scanning confocal microscopy after0and10Gy60Coγ radiation. Quantitative analysis was also processed. Alkaline cometassay also named alkaline single gel electrophoresis assay were applied to determinecomet rate and comet tail length. Apoptosis ratio was calculated by flow cytometry. Wedrew the survival curves of the different cells and invasion.
Results: DCX and SPN protein were expressed in U87-DCX cells, not U87-MGand U87-NG cells. The expressions of DCX,SPN proteins in10Gy dose is higher than0Gy（P<0.05）. Comet tail length significantly increased with the doses of radiationincreasing in three kinds of cells (P<0.05) and with the different cells ranking asU87-DCX cells, U87-MG and U87-NG cells (P<0.05). Two different cells apoptosisratio is low by flow cytometry. U87-DCX cells survival and invasion is lower than othertwo cells.
Conclusion: Three kinds of cell damage enhanced as the doses of radiation wereincreased up to10Gy. DCX is followed by interactive action with SPN. DCX react withsignaling pathway of DNA injury repair after induction by radiation, DCX gene whichexists in U87-DCX cells could activate its downstream genes SPN, and could promoteor increase the expressions of and interactions between these two proteins to increaseradiation injury. The radiation injury of U87-DCX cell is higher than other cells.U87-DCX cells survival and invasion is lower than other two cells.
Abstract Part Four
Objectives：With the in vivo test, analysis of DCX function of U87-DCX cells intumor-bearing mouse model by measuring tumor growth, metastasis andradiosensitivity.
Methods: Construction tumor-bearing mouse model with U87-DCX cells andU87-NG cells. Nude brain were irradiated with60Coγ radioactive rays in total10Gydoses.(dose rate:0.5Gy/min, single dose:200cGy, once a day,5days).Using the microanimals Magnetic Resonance Imaging scanner, the volume size of the tumor in nude’sbrain was analysed. Using the micro animals PET scanner, SUV of the tumor in nude’sbrain was measeured. The expressing of DCX and SPN was evaluated onimmunohistochemical slide image in radiation and post-radiation.
Results: After radiation treatment, the SUVmax values of two groups decreasedsignificantly. The SUVmax values of U87-DCX group (decreased60%) weresignificantly lower than the control seed group while there was no significant differencebetween the same cells tumor-bearing mouse. The value of microvessel density (MVD)decreased in two groups after radiation treatment, but not related to DCX gene. It isindicated that MVD is related of radiation.
Conclusion: Radiation can decrease MVD, and DCX gene increased theradiosensitivity. But MVD is not related to DCX gene.
目的：构建针对DCX的质粒载体，转染U87-MG细胞株，通过筛选获得DCX表达的稳定转染的细胞模型。
方法：采用UbC-GFP-L.V.慢病毒感染U87-MG细胞株，选择感染效率高、MOI值高、对细胞毒性低为最佳感染条件，获得U87-DCX和U87-NG细胞株，用PCR和Western Blot方法检测。
结果：U87-DCX细胞为DCX过表达的稳定转染细胞，目的基因DCX融合GFP和FLAG共同表达；Western Blot检测到72-95KD处有阳性条带；其大小和DCX-GFP-FLAG融合蛋白（48KDa+28KDa+2KDa＝78KDa）相吻合。表达克隆中插入目的基因片段大小为1326bp。
结论：U87-DCX、U87-NG细胞株构建成功。
第二部分基因芯片筛选与DCX和放射相关的差异基因
目的：基因芯片筛选转染DCX的U87-MG细胞在照射前后的差异基因，探讨与DCX和电离辐射均具有相关性的下游基因在胶质瘤放射治疗中的作用。
方法：将U87-MG细胞、U87-DCX细胞经60Coγ射线（10Gy，剂量率1.0Gy/min）照射，用基因芯片技术筛选差异基因，均值两两比较，取P <0.05的差异基因分析。
结果：放射和DCX转染均有相关性差异基因有：EIF5A、LYN、SPN；与放射相关的差异基因有：IMAA、LDHA、WIBG、UBC；与DCX相关的差异基因有：PLS3、IL11、NOV。U87-MG细胞、U87-DCX细胞分别经60Coγ射线（10Gy，剂量率1.0Gy/min）照射，选取SPN与DCX免疫共沉淀实验，结果表明，U87-MG细胞照射组与对照组均无DCX和SPN蛋白表达，而U87-DCX细胞照射组与对照组DCX和SPN蛋白均表达，并且DCX和SPN的蛋白表达水平在10Gy照射组明显高于对照组。
结论：表达谱基因芯片技术能快速、灵敏地筛选出U87-DCX细胞在放疗前后的差异基因，通过上述实验证实，在放射条件下DCX蛋白表达水平增高，且与SPN相互作用。
第三部分体外试验研究DCX的放射作用机制
目的：研究DCX基因高表达联合γ-射线照射对胶质瘤细胞生长、浸润转移、DNA损伤修复及凋亡的影响。
方法：1.U87-MG、U87-DCX、U87-NG三种细胞经60Coγ射线照射，剂量分别为0，2，4，6，8，10Gy。收集细胞提取蛋白，用Western Blot方法测定不同放射剂量的DCX蛋白量，并绘制剂量效应曲线。2.测定DCX下游基因在不同放射剂量下的蛋白量，并绘制剂量效应曲线。3.激光共聚焦显微镜观察U87-MG、U87-DCX、U87-NG三种细胞的DCX及SPN蛋白表达水平，并进行数据分析。4.彗星试验研究照射后U87-MG、U87-DCX、U87-NG三种细胞的双链DNA损伤，并进行数据分析。5.流式细胞仪测定细胞凋亡。6.细胞计数方法测定不同细胞的生存曲线以及划痕试验检测细胞的浸润情况。
结果：1.2，4，6，8，10Gy照射条件下DCX、SPN蛋白显示表达水平较对照组高；2.DCX、SPN蛋白在胞浆内共定位表达；3.彗星试验表明U87-DCX细胞的尾长在0、5、10Gy剂量组均较U87-MG、U87-NG细胞长（P<0.05）；4.在辐射作用下，U87-DCX和U87-NG细胞的凋亡率均较低,两者凋亡率比较无统计学意义，U87-DCX细胞的凋亡率照射组与对照组比较无统计学意义（P>0.05）；6.U87-DCX细胞生长、浸润能力较未转染组低。
结论：电离辐射诱导下DCX蛋白表达水平增高，随着照射剂量的增加蛋白表达水平呈上升趋势，并且与下游基因SPN相互作用，DCX在射线所致的细胞损伤过程中发挥作用；DCX能够通过辐射诱导DNA损伤，证实DCX基因转染能够增强U87胶质瘤细胞的放射敏感性；细胞的放射损伤与凋亡之间无明显相关性；转染DCX的U87细胞生长缓慢，缺乏侵袭能力。
第四部分体内试验究DCX对胶质瘤放射敏感性的影响
目的：构建U87-DCX转染细胞的荷瘤鼠模型，研究DCX转染的细胞在体内生长、转移的生物学特性，探讨其放射增敏机制。
方法：构建U87-DCX和U87-NG裸鼠脑胶质瘤原位种植模型；成瘤后以60Coγ射线照射，正常组织以铅板屏蔽，暴露头部照射野，剂量率0.5Gy/min，单次剂量200cGy，每日一次，共五天，总剂量为10Gy。小动物MRI评估小鼠在放射治疗前后的肿瘤大小；Micro PET测量裸鼠接种部位的肿瘤大小及微血管密度SUVmax值；免疫组化观察放疗前后的脑组织内DCX和SPN表达水平。
结果：小动物MRI测量结果表明U87-DCX实验组放疗后肿瘤体积比U87-NG实验组肿瘤体积缩小明显。Micro PET显示经过放射治疗的裸小鼠SUVmax值较放疗前明显下降，其中DCX转染组的裸小鼠下降更为明显，均为60%以上；阴性对照组小鼠在放射治疗后SUVmax值亦略有下降；免疫组化显示两组细胞放射治疗后较对照组微血管密度（MVD）值均有下降，但相同剂量的两组细胞之间无明显差异（P>0.05）。说明放疗可以降低SUVmax。
结论：放射治疗能够有效抑制肿瘤增殖、MVD降低。DCX转染组对放射治疗更为敏感，即肿瘤体积缩小较对照组下降明显；Micro PET可测量到放射治疗后早期SUVmax变化，DCX转染组的SUVmax较对照组明显降低。
Abstract Part One
Objectives：Establishment of doublecortin (DCX) stably expressed U87cells.
Methods: U87cells were infected with Ubc-GFP-Lentivirus. The multiplicity ofnfection (MOI) is10.
Results: PSB276(pLV-UbC-DCX) clone highly express fusion gene DCX in U87ells. Enzyme digestion analysis and sequencing analysis showed that target gene hadeen cloned into recombinant vector and molecular size is1326bp. Then theecombinant was transfected into U87cell by techniques of gene transfection andetected expression by fluoroscopy, DCX-GFP-FLAG was identified by Westernlot(48KDa+28KDa+2KDa＝78KDa).
Conclusion: U87-DCX、U87-NG were established to fulfill.
Abstract Part Two
Objectives: Radiation therapy plays important roles in the treatment ofneurogliocytoma. It is also known that DCX transfected U87-MG cells can inhibittumor cell growth. To determine if there is a relationship between these cells andradiation treatment, microarray analysis was performed to screen for differentiallyexpressed genes in DCX-transfected U87cells before and after radiation in order todiscover DCX-related genes and to reveal the functions of DCX and downstream genesin radiation therapy of neurogliocytoma.
Methods: Stably transfected U87cells were constructed (U87-DCX) and thedifferentially expressed genes were screened via microarray analysis to compare U87cells to U87-DCX cells in both non-irradiated and irradiated conditions. Cells wereirradiated using60Coγ-ray at a dose rate of1.0Gy/min. Mean values were subject topaired comparison analysis, and genes with a p-value less than0.05were analyzed.
Results: Differentially expressed genes that correlated with radiation sensitivityand DCX transfection included eukaryotic initiation factor5A (EIF5A), LYN (v-yes-1Yamaguchi sarcoma viral related oncogene homolog), and neurabin II （SPN）.Differentially expressed genes that only correlated with radiation sensitivity includedIMAA, LDHA, WIBG, and UBC, while differentially expressed genes that were onlyrelated to DCX included PLS3, IL11, and NOV. SPN and DCX were selected forimmunoprecipitation to compare binding activity in both U87-DCX cells and U87cellsin the0and10Gy groups. Results showed that DCX and SPN proteins were notdetected in the two dosage groups in the U87cells, while DCX and SPN proteins weredetected in U87-DCX cells in both two dosage groups, with expression levels of DCXand SPN in the10Gy group higher than those of the0Gy group.
Conclusion: The differential gene expression in U87-DCX cells before and afterradiation provides a rationale for further investigations of the mechanism for radiationtherapy in neurogliocytoma cells. This method can also provide a potential solution forradiation resistance in neurogliocytoma, using gene expression panels as a predictor ofradiation sensitivity. Expression profile microarray technology can rapidly andsensitively predict the radiation sensitivity of neurogliocytoma and screen for relatedradiation sensitive genes.
Abstract Part Three
Objectives：Relationship of expression of DCX overexpressed with60Co γradiation to cell growth, DNA damage and repair, invasion and metastasis，cell cycle distribution and apoptosis.
Methods: U87-DCX, U87-MG and U87-NG cells were treated with radiated by γray of different doses. Immunocoprecipitation and Western Blotting were performed toanalyze the interactive action between DCX and SPN protein in U87-DCX cells after60Coγ radiation. The curve of dosage effect was obtained. The localizations andexpressions of DCX, SPN proteins were detected in U87-MG, U87-DCX and U87-NGcells by immunofluorescence staining and laser scanning confocal microscopy after0and10Gy60Coγ radiation. Quantitative analysis was also processed. Alkaline cometassay also named alkaline single gel electrophoresis assay were applied to determinecomet rate and comet tail length. Apoptosis ratio was calculated by flow cytometry. Wedrew the survival curves of the different cells and invasion.
Results: DCX and SPN protein were expressed in U87-DCX cells, not U87-MGand U87-NG cells. The expressions of DCX,SPN proteins in10Gy dose is higher than0Gy（P<0.05）. Comet tail length significantly increased with the doses of radiationincreasing in three kinds of cells (P<0.05) and with the different cells ranking asU87-DCX cells, U87-MG and U87-NG cells (P<0.05). Two different cells apoptosisratio is low by flow cytometry. U87-DCX cells survival and invasion is lower than othertwo cells.
Conclusion: Three kinds of cell damage enhanced as the doses of radiation wereincreased up to10Gy. DCX is followed by interactive action with SPN. DCX react withsignaling pathway of DNA injury repair after induction by radiation, DCX gene whichexists in U87-DCX cells could activate its downstream genes SPN, and could promoteor increase the expressions of and interactions between these two proteins to increaseradiation injury. The radiation injury of U87-DCX cell is higher than other cells.U87-DCX cells survival and invasion is lower than other two cells.
Abstract Part Four
Objectives：With the in vivo test, analysis of DCX function of U87-DCX cells intumor-bearing mouse model by measuring tumor growth, metastasis andradiosensitivity.
Methods: Construction tumor-bearing mouse model with U87-DCX cells andU87-NG cells. Nude brain were irradiated with60Coγ radioactive rays in total10Gydoses.(dose rate:0.5Gy/min, single dose:200cGy, once a day,5days).Using the microanimals Magnetic Resonance Imaging scanner, the volume size of the tumor in nude’sbrain was analysed. Using the micro animals PET scanner, SUV of the tumor in nude’sbrain was measeured. The expressing of DCX and SPN was evaluated onimmunohistochemical slide image in radiation and post-radiation.
Results: After radiation treatment, the SUVmax values of two groups decreasedsignificantly. The SUVmax values of U87-DCX group (decreased60%) weresignificantly lower than the control seed group while there was no significant differencebetween the same cells tumor-bearing mouse. The value of microvessel density (MVD)decreased in two groups after radiation treatment, but not related to DCX gene. It isindicated that MVD is related of radiation.
Conclusion: Radiation can decrease MVD, and DCX gene increased theradiosensitivity. But MVD is not related to DCX gene.
引文
1. Jeremy N. Rich，Christopher Hans，Beatrix Jones et al.Gene Expression Profilingand Genetic Markers in Glioblastoma Survival. Cancer Res;2005;65:4051－4058.
2. Santra M, Santra S, Buller B, et al. Effect of doublecortin on self-renewal anddifferentiation in brain tumor stem cells.Cancer Sci.2011Jul;102(7):1350-7.
3. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
4. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009Jan;108(1):231-45.
5. Santra M, Zhang X, Santra S, et al. Ectopic Doublecortin Gene ExpressionSuppresses the Malignant Phenotype in Glioblastoma Cells. Cancer Res.2006Dec15;66(24):11726-35.
6. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation. Neuroscience.2006Oct27;142(3):739-52.
7. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51-61.
8. Tiziano Pramparo, Yong Ha Youn, Jessica Yingling，et al. Novel embryonicneuronal migration and proliferation defects in Dcx mutant mice are exacerbated byLis1reduction. J Neurosci.2010February24;30(8):3002–3012.
9. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a putativesignaling protein. Cell.1998;92:63-72.
10. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron.1999;23:257–71.
11. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet,1999;8:1599–610.
12. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247–56.
13. Nutt CL, Mani DR, Betensky RA, et al. Gene expression-based classification ofmalignant gliomas correlates better with survival than histological classification.Cancer Res.2003,63(7):1602-1607.
14. Wall M, Rechtsteiner A, Rocha LM. Singular value decomposition and principalcomponent analysis.. Springer US.2003,91-109.
15. Draminski M, Rada Iglesias A, Enroth S, et al. Monte Carlo feature selection forsupervised classification.bioinformatics.2008,24(1):110-117.
16.来海锋,韩斌,厉力华等，基于集成类随机森林方法的神经胶质瘤特征基因选择的研究.生物物理学报2010：Vol.26，No.9：833-845.
1. Burmester T, Weich B,Reinhardt S,et al.A vertebrate globin ex-pressed in thebrain[J].Nature.2000,407:520-52.
2. Li RC, Guo SZ, Lee SK, et al.Neuroglobin protects neurons against ox-idative stressin global ischemia.J Cereb Blood Flow Metab.2010,30:1874-1882.
3. GreenbergDA,Jin K,Khan AA.Neuroglobin:an endogenous neuro-protectant[J].CurrOpin Pharmacol.2008,8:20-24.
4. Escors D, Breckpot K.Lentiviral vectors in gene therapy: their currentstatus andfuture potential[J].Arch Immunol Ther Exp(Warsz).2010,58:107-119.
5. Nanou A, Azzouz M.Gene therapy for neurodegenerative diseasesbased on lentiviralvectors[J].Prog Brain Res.2009,175:187-200.
6. Das S, Srikanth M, Kessler JA. Cancer stem cells and glioma. Nat ClinPract.Neurol.2008;4:427–35.
1. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51–61.
2. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a putativesignaling protein. Cell.1998;92:63–72.
3. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron.1999;23:257–71.
4. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet.1999(8):1599–610.
5. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247–56.
6. Manoranjan Santra, Xuepeng Zhang, Sutapa Santra, et al. Ectopic DoublecortinGene Expression Suppresses the Malignant Phenotype in Glioblastoma Cells.Cancer Res.2006;66:11726-11735.
7. Jeremy N. Rich，Christopher Hans，Beatrix Jones et al.Gene Expression Profilingand Genetic Markers in Glioblastoma Survival. Cancer Res.2005;65:4051－4058
8. Nutt CL, Mani DR, Betensky RA,el. Gene expression-based classification ofmalignant gliomas correlates better with survival than histological classification.Cancer Res.2003,63(7):1602-1607.
9. Wall M, Rechtsteiner A, Rocha LM. Singular value decomposition and principalcomponent analysis Springer US.2003,91-109.
10. Draminski M, Rada-Iglesias A, Enroth S,el. Monte Carlo feature selection forsupervised classification.bioinformatics.2008,24(1):110-117.
11.来海锋,韩斌,厉力华等，基于集成类随机森林方法的神经胶质瘤特征基因选择的研究.生物物理学报.2010：Vol.26，No.9：833-845.
12.杨吉成，宋礼华，周建华等.医用细胞工程,上海交通大学出版社，2000:76.
13. Illumina, Normalization and differential Analysis. Bead Studio Gene ExpressionModule User Guide.2008,92-100.
14. Park, M. H., Lee, Y. B., and Joe, Y. A. Hypusine Formation on Eukaryotic InitiationFactor5A Biol. Signals.1997(6),115-123.
15.李爱玲.eIF-5A调控p53和p53依赖的细胞凋亡.中国人民解放军军事医学科学院,2004,(11)
16. Ai-Ling Li el，A Novel eIF5A Complex Functions As a Regulator of p53andp53-dependent Apoptosis.THE JOURNAL OF BIOLOGICAL CHEMISTRY.2004.279,49251-49258.
17. M Murphy et al. Comparison of the expression of p53, p21, Bax and the inductionof apoptosis between patients with basal cell carcinoma and normal controls inresponse to ultraviolet irradiation. J Clin Pathol.2002;55:829-833.
1. Nanou A,Azzouz M.Gene therapy for neurodegenerative diseasesbased on lentiviralvectors[J].Prog Brain Res.2009,175:187-200.
2. Chunsheng Liu, Jann N. Sarkaria, CoryA. Petell， et al. Combination ofMeaslesVirusVirotherapy and Radiation Therapy Has Synergistic Activity intheTreatmentof Glioblastoma Multiforme. Clin Cancer Res.2007;715513(23).
3. Jeremy N. Rich，Christopher Hans，Beatrix Jones et al.Gene Expression Profilingand Genetic Markers in Glioblastoma Survival. Cancer Res.2005;65:4051－4058.
4. Santra M, Santra S, Buller B, et al. Effect of doublecortin on self-renewal anddifferentiation in brain tumor stem cells.Cancer Sci.2011Jul;102(7):1350-7.
5. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
6. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009Jan;108(1):231-45.
7. Santra M, Zhang X, Santra S, et al. Ectopic Doublecortin Gene ExpressionSuppresses the Malignant Phenotype in Glioblastoma Cells. Cancer Res.2006Dec15;66(24):11726-35.
8. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation. Neuroscience.2006Oct27;142(3):739-52.
9. Campo L, Turley H, Han C, et al. Angiogenin is up-regulated in the nucleus andcytoplasm in human primary breast carcinoma and is associated with markers ofhypoxia but not survival. J Pathol.2005;205:585-91.
10. Hartmann A, Kunz M, Kostlin S, et al. Hypoxiainduced up-regulation of angiogeninin human malignant melanoma. Cancer Res.1999;59:1578-83.
11. Nakamura M,Yamabe H, Osawa H, et al. Hypoxic conditions stimulate theproduction of angiogenin and vascular endothelial growth factor by human renalproximal tubular epithelial cells in culture. Nephrol Dial Transplant2006;21:1489-95.
12. Pilch H, Schlenger K, Steiner E, et al. Hypoxiastimulated expression of angiogenicgrowth factors in cervical cancer cells and cervical cancer-derived fibroblasts.Int JGynecol Cancer.2001;11:137-42.
13. Agda K. Lucio-Eterovic,Yuji Piao, et al. Mediators of Glioblastoma Resistance andInvasion during Antivascular Endothelial Growth FactorTherapy. Clin CancerRes.2009;15(14)：4588-4599.
14. Singh N P, McCoy M T, Tice R R et al. A simple tech2nique for quantitation oflow levels of DNA damage in individual cells. Expti Cell Res.1988,175(1):184～191
15.朱志良,庄志雄,黄钰,籍涛.单细胞凝胶电泳图象分析系统的研制与应用.中华劳动卫生职业病杂志.2001;19(4):298-300.
16. Fairbairn DW, Olive DL,O’Neill Kim L. The comet assay: a comprehensive review.Mutat Res.1995;339:37-59
17. McKelvey， Martin VJ, Green MHL, Schmezer P, et al. The single gelelectrophoresis assay (comet assay): a European review. Mutat Res.1993;288:47-63
18. Gedik CM, Ewen Wb, et al. Collins AR. Single cell gel electrophoresis applied tothe analysis of UV2C damage and its repair in human Cells. Int J Radiat Biol.1992;62:313-320
19. Singh NP, Banner DB, Tice RR, et al. DNA damage and repair with age inindividual human lymphocytes.Mutat Res.1990;237:123-130.
20. Su JH, Anderson AJ, Cribbs DH, et al. Fas and Fas ligand are associated withneuritic degeneration in the AD brain and participate in beta-amyloid-inducedneuronal death [J]. Neurobiol Dis.2003,12(3):182-193.
1. Pantelouris EM.Absence of thymus in a mouse mutant. Nature，1968，271：370-371.
2. LaＭont JT，O’Gorman TA.Experimental colon cancer.Gastroenterology，1978，75：1157-1169.
3. Sharkey FE, Fogh J. Considerations in the use of nude mice for cancer research.Cancer and Metastasis Reviews.1984,3:341-360.
4.王舒楠,王斌,张伟国,等.1.5T磁共振成像在检测裸小鼠胶质瘤原位成瘤模型中的应用[J].第三军医大学学报.2010,32(02):131-134.
5.李如军、黄强、沈钧康等，Balb/c小鼠颅内植入物MR显像的初步研究立体定向和功能性神经外科杂志.Chin J Stereot act Funct Neurosurg.Jun.2005,Vol.18,No.3.163-166.
6.王建交等.Wistar大鼠C6胶质瘤模型中的肿瘤消退现象研究.现代生物医学进展Progress in Modern Biomedicine.2009年Vol.9No.11.2064-66.
7.孙建军等.应用磁共振成像技术实时监测小鼠瘤体的初步实验研究.医学研究杂志JM ed Res, Jun2011,Vo.l40.No.6
8. Jan Thies Deichen, Olaf Prante, Michaela Gack, et al.Uptake of[18F]fluorodeoxyglucose in humanmonocyte-macrophages in vitro. EuropeanJournal of Nuclear Medicine and Molecular Imaging Vol.30, No.2, February2003
9. Xiujuan Zhenga, Chin-Lung Yub, Wei Shab, et al. Study of an image-derivedSUVmaxand a modified SUVmaxusing mouse FDG-PET. Nuclear Medicine andBiology.38(2011):353–362.
10. Dong-Pyo Jang, So-Hee Lee, Chan-Woong Park, et al. Effects of fluoxetine on therat brain in the forced swimming test:A [F-18]FDG micro-PET imagingstudyNeuroscience Letters.451(2009):60–64
11. Kang-Keyng Sung, Dong-Pyo Jang, Sangkwan Lee, et al.Neural responses in ratbrain during acute immobilization stress: A [F-18]FDG micro PET imagingstudy.NeuroImage.44(2009):1074–1080
12. Stephen P Povoski1, Nathan C Hall, et al. Douglas A Murrey Jr.Multimodalimaging and detection approach to18F-FDG-directed surgery for patients withknown or suspected malignancies: a comprehensive description of the specificmethodology utilized in a single-institution cumulative retrospective experience.
13. Heike Endepols, Susanne Sommer, Heiko Backes, et al.Effort-Based DecisionMaking in the rat: An [18F]Fluorodeoxyglucose Micro Positron EmissionTomography Study. The Journal of Neuroscience.July21,2010,30(29):9708-9714.
14. Beer AJ, Lorenzen S, Metz S, et al.Comparison of integrin alphaVbeta3expressionand glucose metabolism in primary and metastatic lesions in cancer patients: a PETstudy using18F-galacto-RGD and18F-FDG.J Nucl Med.2008Jan;49(1):22-9.
15. Gaertner FC, Kessler H, Wester HJ, et al. Radiolabelled RGD peptides for imagingand therapy.Eur J Nucl Med Mol Imaging.2012Feb;39Suppl1:126-38.
16. Xiaoyuan Chena, Ryan Parka, Anthony H.18F-labeled RGD peptide: initialevaluation for imaging brain tumor Angiogenesis. Nuclear Medicine andBiology:31(2004):179-189.
17. Matthew S. Morrison, Sally-Ann Ricketts, Jon Barnett, et al. Use of a NovelArg-Gly-Asp Radioligand,18F-AH111585, to Determine Changes in TumorVascularity After Antitumor Therapy. J Nucl Med.2009;50:116–122.
18. Cappelli A, Matarrese M, Moresco RM, et al.Synthesis, labeling, and biologicalevaluation of halogenated2-quinolinecarboxamides as potential radioligands for thevisualization of peripheral benzodiazepine receptors. Bioorg Med Chem.2006,Jun15;14(12):4055-66.
19.杨明等，地龙蛋白组分Ⅲ对鼻咽癌裸鼠移植瘤微血管密度的影响湖南中医药大学学报.Journal of TCM Univ. of Hunan.2008年8月第28卷第4期.
20. Borgstrom P,Hillan KJ,Sriramarao P,et al.Complete inhibition of angiogenesis andgrowth of microtumors by anti VEGF neutralizing antibody:novel concepts ofangiostatic therapy from intranital videomicroscopy[J].Cancer Res.1996,56(17):4032-4039
21. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51-61.
22. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a
23. putative signaling protein.Cell.1998;92:63-72.
24. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron1999;23:257–71.
25. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet1999;8:1599–610.
26. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247-56.
27. Manoranjan Santra, Xuepeng Zhang, Sutapa Santra, et al. Ectopic DoublecortinGene Expression Suppresses the Malignant Phenotype in Glioblastoma Cells.Cancer Res.2006;66:11726-11735.
28. Manoranjan Santra, et al. Effect of doublecortin on self-renewal and differentiationin brain tumor stem cells. Cancer science, Accepted manuscript online.April7,2011
29. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
30. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009,Jan;108(1):231-45.
31. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation.Neuroscience.2006Oct27;142(3):739-52.
1.Manoranjan Santra1, Sutapa Santra1, Cindi Roberts1，el.Doublecortin inducesmitotic microtubule catastrophe and inhibits glioma cell invasion.J Neurochem.2009January;108(1):231–245.）
2. Legler JM, Gloeckler Ries LA, Smith MA, et al. Brain and other central nervoussystem cancers: recent trends in incidence and mortality. J Natl Cancer Inst1999;91:1382–90.
3. Jemal A, Murray T, Ward E, et al. Cancer statistics,2005. CA Cancer J Clin2005;55:10–30.
4. Rich JN, Hans C, Jones B, et al. Gene expression profiling and genetic markersin glioblastoma survival.Cancer Res2005;6510:4051–8.
5. Francis F, Koulakoff A, Boucher D, Chafey P, Schaar B, Vinet MC, Friocourt G,McDonnell N, Reiner O, Kahn A, et al.: Doublecortin is a developmentally regulated,microtubule-associated protein expressed in migrating and differentiating neurons.Neuron1999,23(2):247-256.
6. des Portes V, Pinard JM, Billuart P, Vinet MC, Koulakoff A, Carrie A, GelotA,Dupuis E, Motte J, Berwald-Netter Y, et al.: A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar heterotopia and lissencephalysyndrome. Cell1998,92(1):51-61.
7. Walker TL, Yasuda T, Adams DJ, Bartlett PF: The doublecortin-expressingpopulation in the developing and adult brain contains multipotential precursors inaddition to neuronal-lineage cells. Journal of Neuroscience2007,27(14):3734-3742.
8. Friocourt G, Koulakoff A, Chafey P, Boucher D, Fauchereau F, Chelly J,FrancisF: Doublecortin functions at the extremities of growing neuronal processes. CerebCortex2003,13(6):620-626.
9. Brown JP, Couillard-Despres S, Cooper-Kuhn CM, Winkler J, Aigner L, KuhnHG: Transient expression of doublecortin during adult neurogenesis. JComp Neurol2003,467(1):1-10.
10. Mattar P, Britz O, Johannes C, Nieto M, Ma L, Rebeyka A, Klenin N, PolleuxF, Guillemot F, Schuurmans C: A screen for downstream effectors of Neurogenin2inthe embryonic neocortex. Developmental Biology2004,273(2):373-389.
11. Couillard-Despres S, Winner B, Schaubeck S, Aigner R, Vroemen M,WeidnerN, Bogdahn U, Winkler J, Kuhn H, Aigner L: Doublecortin expression levels in adultbrain reflect neurogenesis. Eur J Neurosci2005,21(1):1-14.
12. Gleeson JG, Allen KM, Fox JW, Lamperti ED, Berkovic S, Scheffer I, CooperEC, Dobyns WB, Minnerath SR, Ross ME, et al.: Doublecortin, a brainspecific genemutated in human X-linked lissencephaly and double cortex syndrome, encodes aputative signaling protein. Cell1998,92(1):63-72.
13. Schaar, B. T., Kinoshita, K., and McConnell, S. K.(2004) Neuron41,203–2131
14. Tanaka, T., Serneo, F. F., Tseng, H-C., Kulkarni, A. B., Tsai, L-H., andGleeson, J. G.(2004) Neuron41,215–227
15.RAes esahrcho arrttic leupstream promoter region mediates transcriptionalregulation of the mouse doublecortin gene in differentiating neuronsPiens et al. BMCNeuroscience2010,11:64）
16.Doublecortin Association with Actin Filaments Is Regulated by Neurabin II*THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol.280, No.12, Issue of March25,pp.11361–11368,2005）
17. Zhou Q, Anderson DJ: The bHLH transcription factors OLIG2and OLIG1couple neuronal and glial subtype specification. Cell2002,109(1):61-73
18. Ross ME, Allen KM, Srivastava AK, et al. Linkage and physical mapping ofX-linked lissencephaly/SBH (XLIS):a gene causing neuronal migration defects inhuman brain. Hum Mol Genet1997;64:555–62.
19. Santra M, Lui XS, Zhang J, Zhang RL, Zhang ZG,Chopp M. Ectopicexpression of doublecortin protects adult mouse progenitor cells and human gliomacells from severe oxygen and glucose deprivation. Neuroscience2006;92:7016–20.
20. Tsuiki H, Tnani M, Okamoto I, et al. Constitutively active forms of c-JunNH2-terminal kinase are expressed in primary glial tumors. Cancer Res2003;631:250–5.
21.Phosphoproteomic Analysis of the Developing Mouse Brain*Molecular&Cellular Proteomics3:1093–1101,2004.）
22. Tiziano Pramparo1, Yong Ha Youn1, Jessica Yingling2el. Novel embryonicneuronal migration and proliferation defects in Dcx mutant mice are exacerbated byLis1reduction.J Neurosci.2010February24;30(8):3002–3012.
23. Cohen P. The origins of protein phosphorylation. Nat Cell Biol2002;45:E127–30.
24. Inui S, Sanjo H, Maeda K, Yamamoto H, Miyamoto E,Sakaguchi N. Igreceptor binding protein1(a4) is associated with a rapamycin-sensitive signaltransduction in lymphocytes through direct binding to the catalytic subunit of proteinphosphatase2A. Blood1998;922:539–46.
25. MacMillan LB, Bass MA, Cheng N, et al. Brain actinassociated proteinphosphatase1holoenzymes containing spinophilin, neurabin, and selected catalyticsubunit isoforms. J Biol Chem1999;27450:35845–54.31.
26. Shmueli A, Gdalyahu A, Sapoznik S, Sapir T, Tsukada M, Reiner O.Site-specific dephosphorylation of doublecortin (DCX) by protein phosphatase1(PP1).Mol Cell Neurosci2006. Epub ahead of print.
27. Manoranjan Santra1, Sutapa Santra1, Cindi Roberts1，el.Doublecortin inducesmitotic microtubule catastrophe and inhibits glioma cell invasion.J Neurochem.2009January;108(1):231–245.
2. Santra M, Santra S, Buller B, et al. Effect of doublecortin on self-renewal anddifferentiation in brain tumor stem cells.Cancer Sci.2011Jul;102(7):1350-7.
3. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
4. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009Jan;108(1):231-45.
5. Santra M, Zhang X, Santra S, et al. Ectopic Doublecortin Gene ExpressionSuppresses the Malignant Phenotype in Glioblastoma Cells. Cancer Res.2006Dec15;66(24):11726-35.
6. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation. Neuroscience.2006Oct27;142(3):739-52.
7. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51-61.
8. Tiziano Pramparo, Yong Ha Youn, Jessica Yingling，et al. Novel embryonicneuronal migration and proliferation defects in Dcx mutant mice are exacerbated byLis1reduction. J Neurosci.2010February24;30(8):3002–3012.
9. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a putativesignaling protein. Cell.1998;92:63-72.
10. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron.1999;23:257–71.
11. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet,1999;8:1599–610.
12. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247–56.
13. Nutt CL, Mani DR, Betensky RA, et al. Gene expression-based classification ofmalignant gliomas correlates better with survival than histological classification.Cancer Res.2003,63(7):1602-1607.
14. Wall M, Rechtsteiner A, Rocha LM. Singular value decomposition and principalcomponent analysis.. Springer US.2003,91-109.
15. Draminski M, Rada Iglesias A, Enroth S, et al. Monte Carlo feature selection forsupervised classification.bioinformatics.2008,24(1):110-117.
16.来海锋,韩斌,厉力华等，基于集成类随机森林方法的神经胶质瘤特征基因选择的研究.生物物理学报2010：Vol.26，No.9：833-845.
1. Burmester T, Weich B,Reinhardt S,et al.A vertebrate globin ex-pressed in thebrain[J].Nature.2000,407:520-52.
2. Li RC, Guo SZ, Lee SK, et al.Neuroglobin protects neurons against ox-idative stressin global ischemia.J Cereb Blood Flow Metab.2010,30:1874-1882.
3. GreenbergDA,Jin K,Khan AA.Neuroglobin:an endogenous neuro-protectant[J].CurrOpin Pharmacol.2008,8:20-24.
4. Escors D, Breckpot K.Lentiviral vectors in gene therapy: their currentstatus andfuture potential[J].Arch Immunol Ther Exp(Warsz).2010,58:107-119.
5. Nanou A, Azzouz M.Gene therapy for neurodegenerative diseasesbased on lentiviralvectors[J].Prog Brain Res.2009,175:187-200.
6. Das S, Srikanth M, Kessler JA. Cancer stem cells and glioma. Nat ClinPract.Neurol.2008;4:427–35.
1. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51–61.
2. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a putativesignaling protein. Cell.1998;92:63–72.
3. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron.1999;23:257–71.
4. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet.1999(8):1599–610.
5. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247–56.
6. Manoranjan Santra, Xuepeng Zhang, Sutapa Santra, et al. Ectopic DoublecortinGene Expression Suppresses the Malignant Phenotype in Glioblastoma Cells.Cancer Res.2006;66:11726-11735.
7. Jeremy N. Rich，Christopher Hans，Beatrix Jones et al.Gene Expression Profilingand Genetic Markers in Glioblastoma Survival. Cancer Res.2005;65:4051－4058
8. Nutt CL, Mani DR, Betensky RA,el. Gene expression-based classification ofmalignant gliomas correlates better with survival than histological classification.Cancer Res.2003,63(7):1602-1607.
9. Wall M, Rechtsteiner A, Rocha LM. Singular value decomposition and principalcomponent analysis Springer US.2003,91-109.
10. Draminski M, Rada-Iglesias A, Enroth S,el. Monte Carlo feature selection forsupervised classification.bioinformatics.2008,24(1):110-117.
11.来海锋,韩斌,厉力华等，基于集成类随机森林方法的神经胶质瘤特征基因选择的研究.生物物理学报.2010：Vol.26，No.9：833-845.
12.杨吉成，宋礼华，周建华等.医用细胞工程,上海交通大学出版社，2000:76.
13. Illumina, Normalization and differential Analysis. Bead Studio Gene ExpressionModule User Guide.2008,92-100.
14. Park, M. H., Lee, Y. B., and Joe, Y. A. Hypusine Formation on Eukaryotic InitiationFactor5A Biol. Signals.1997(6),115-123.
15.李爱玲.eIF-5A调控p53和p53依赖的细胞凋亡.中国人民解放军军事医学科学院,2004,(11)
16. Ai-Ling Li el，A Novel eIF5A Complex Functions As a Regulator of p53andp53-dependent Apoptosis.THE JOURNAL OF BIOLOGICAL CHEMISTRY.2004.279,49251-49258.
17. M Murphy et al. Comparison of the expression of p53, p21, Bax and the inductionof apoptosis between patients with basal cell carcinoma and normal controls inresponse to ultraviolet irradiation. J Clin Pathol.2002;55:829-833.
1. Nanou A,Azzouz M.Gene therapy for neurodegenerative diseasesbased on lentiviralvectors[J].Prog Brain Res.2009,175:187-200.
2. Chunsheng Liu, Jann N. Sarkaria, CoryA. Petell， et al. Combination ofMeaslesVirusVirotherapy and Radiation Therapy Has Synergistic Activity intheTreatmentof Glioblastoma Multiforme. Clin Cancer Res.2007;715513(23).
3. Jeremy N. Rich，Christopher Hans，Beatrix Jones et al.Gene Expression Profilingand Genetic Markers in Glioblastoma Survival. Cancer Res.2005;65:4051－4058.
4. Santra M, Santra S, Buller B, et al. Effect of doublecortin on self-renewal anddifferentiation in brain tumor stem cells.Cancer Sci.2011Jul;102(7):1350-7.
5. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
6. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009Jan;108(1):231-45.
7. Santra M, Zhang X, Santra S, et al. Ectopic Doublecortin Gene ExpressionSuppresses the Malignant Phenotype in Glioblastoma Cells. Cancer Res.2006Dec15;66(24):11726-35.
8. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation. Neuroscience.2006Oct27;142(3):739-52.
9. Campo L, Turley H, Han C, et al. Angiogenin is up-regulated in the nucleus andcytoplasm in human primary breast carcinoma and is associated with markers ofhypoxia but not survival. J Pathol.2005;205:585-91.
10. Hartmann A, Kunz M, Kostlin S, et al. Hypoxiainduced up-regulation of angiogeninin human malignant melanoma. Cancer Res.1999;59:1578-83.
11. Nakamura M,Yamabe H, Osawa H, et al. Hypoxic conditions stimulate theproduction of angiogenin and vascular endothelial growth factor by human renalproximal tubular epithelial cells in culture. Nephrol Dial Transplant2006;21:1489-95.
12. Pilch H, Schlenger K, Steiner E, et al. Hypoxiastimulated expression of angiogenicgrowth factors in cervical cancer cells and cervical cancer-derived fibroblasts.Int JGynecol Cancer.2001;11:137-42.
13. Agda K. Lucio-Eterovic,Yuji Piao, et al. Mediators of Glioblastoma Resistance andInvasion during Antivascular Endothelial Growth FactorTherapy. Clin CancerRes.2009;15(14)：4588-4599.
14. Singh N P, McCoy M T, Tice R R et al. A simple tech2nique for quantitation oflow levels of DNA damage in individual cells. Expti Cell Res.1988,175(1):184～191
15.朱志良,庄志雄,黄钰,籍涛.单细胞凝胶电泳图象分析系统的研制与应用.中华劳动卫生职业病杂志.2001;19(4):298-300.
16. Fairbairn DW, Olive DL,O’Neill Kim L. The comet assay: a comprehensive review.Mutat Res.1995;339:37-59
17. McKelvey， Martin VJ, Green MHL, Schmezer P, et al. The single gelelectrophoresis assay (comet assay): a European review. Mutat Res.1993;288:47-63
18. Gedik CM, Ewen Wb, et al. Collins AR. Single cell gel electrophoresis applied tothe analysis of UV2C damage and its repair in human Cells. Int J Radiat Biol.1992;62:313-320
19. Singh NP, Banner DB, Tice RR, et al. DNA damage and repair with age inindividual human lymphocytes.Mutat Res.1990;237:123-130.
20. Su JH, Anderson AJ, Cribbs DH, et al. Fas and Fas ligand are associated withneuritic degeneration in the AD brain and participate in beta-amyloid-inducedneuronal death [J]. Neurobiol Dis.2003,12(3):182-193.
1. Pantelouris EM.Absence of thymus in a mouse mutant. Nature，1968，271：370-371.
2. LaＭont JT，O’Gorman TA.Experimental colon cancer.Gastroenterology，1978，75：1157-1169.
3. Sharkey FE, Fogh J. Considerations in the use of nude mice for cancer research.Cancer and Metastasis Reviews.1984,3:341-360.
4.王舒楠,王斌,张伟国,等.1.5T磁共振成像在检测裸小鼠胶质瘤原位成瘤模型中的应用[J].第三军医大学学报.2010,32(02):131-134.
5.李如军、黄强、沈钧康等，Balb/c小鼠颅内植入物MR显像的初步研究立体定向和功能性神经外科杂志.Chin J Stereot act Funct Neurosurg.Jun.2005,Vol.18,No.3.163-166.
6.王建交等.Wistar大鼠C6胶质瘤模型中的肿瘤消退现象研究.现代生物医学进展Progress in Modern Biomedicine.2009年Vol.9No.11.2064-66.
7.孙建军等.应用磁共振成像技术实时监测小鼠瘤体的初步实验研究.医学研究杂志JM ed Res, Jun2011,Vo.l40.No.6
8. Jan Thies Deichen, Olaf Prante, Michaela Gack, et al.Uptake of[18F]fluorodeoxyglucose in humanmonocyte-macrophages in vitro. EuropeanJournal of Nuclear Medicine and Molecular Imaging Vol.30, No.2, February2003
9. Xiujuan Zhenga, Chin-Lung Yub, Wei Shab, et al. Study of an image-derivedSUVmaxand a modified SUVmaxusing mouse FDG-PET. Nuclear Medicine andBiology.38(2011):353–362.
10. Dong-Pyo Jang, So-Hee Lee, Chan-Woong Park, et al. Effects of fluoxetine on therat brain in the forced swimming test:A [F-18]FDG micro-PET imagingstudyNeuroscience Letters.451(2009):60–64
11. Kang-Keyng Sung, Dong-Pyo Jang, Sangkwan Lee, et al.Neural responses in ratbrain during acute immobilization stress: A [F-18]FDG micro PET imagingstudy.NeuroImage.44(2009):1074–1080
12. Stephen P Povoski1, Nathan C Hall, et al. Douglas A Murrey Jr.Multimodalimaging and detection approach to18F-FDG-directed surgery for patients withknown or suspected malignancies: a comprehensive description of the specificmethodology utilized in a single-institution cumulative retrospective experience.
13. Heike Endepols, Susanne Sommer, Heiko Backes, et al.Effort-Based DecisionMaking in the rat: An [18F]Fluorodeoxyglucose Micro Positron EmissionTomography Study. The Journal of Neuroscience.July21,2010,30(29):9708-9714.
14. Beer AJ, Lorenzen S, Metz S, et al.Comparison of integrin alphaVbeta3expressionand glucose metabolism in primary and metastatic lesions in cancer patients: a PETstudy using18F-galacto-RGD and18F-FDG.J Nucl Med.2008Jan;49(1):22-9.
15. Gaertner FC, Kessler H, Wester HJ, et al. Radiolabelled RGD peptides for imagingand therapy.Eur J Nucl Med Mol Imaging.2012Feb;39Suppl1:126-38.
16. Xiaoyuan Chena, Ryan Parka, Anthony H.18F-labeled RGD peptide: initialevaluation for imaging brain tumor Angiogenesis. Nuclear Medicine andBiology:31(2004):179-189.
17. Matthew S. Morrison, Sally-Ann Ricketts, Jon Barnett, et al. Use of a NovelArg-Gly-Asp Radioligand,18F-AH111585, to Determine Changes in TumorVascularity After Antitumor Therapy. J Nucl Med.2009;50:116–122.
18. Cappelli A, Matarrese M, Moresco RM, et al.Synthesis, labeling, and biologicalevaluation of halogenated2-quinolinecarboxamides as potential radioligands for thevisualization of peripheral benzodiazepine receptors. Bioorg Med Chem.2006,Jun15;14(12):4055-66.
19.杨明等，地龙蛋白组分Ⅲ对鼻咽癌裸鼠移植瘤微血管密度的影响湖南中医药大学学报.Journal of TCM Univ. of Hunan.2008年8月第28卷第4期.
20. Borgstrom P,Hillan KJ,Sriramarao P,et al.Complete inhibition of angiogenesis andgrowth of microtumors by anti VEGF neutralizing antibody:novel concepts ofangiostatic therapy from intranital videomicroscopy[J].Cancer Res.1996,56(17):4032-4039
21. Des Portes V, Pinard JM, Billuart P, et al. A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar hetrotropia andlissencephaly syndrome. Cell.1998;92:51-61.
22. Gleeson JG, Allen KM, Fox JW, et al. Doublecortin, a brain-specific gene mutatedin human X-linked lissencephaly and double cortex syndrome, encodes a
23. putative signaling protein.Cell.1998;92:63-72.
24. Gleeson JG, Lin PT, Flanagan LA, Walsh CA. Doublecortin is amicrotubule-associated protein and is expressed widely by migrating neurons.Neuron1999;23:257–71.
25. Horesh D, Sapir T, Francis F, et al. Doublecortin, a stabilizer of microtubules. HumMol Genet1999;8:1599–610.
26. Francis F, Koulakoff A, Boucher D, et al. Doublecortin is a developmentallyregulated, microtubule-associated protein expressed in migrating and differentiatingneurons. Neuron.1999;23:247-56.
27. Manoranjan Santra, Xuepeng Zhang, Sutapa Santra, et al. Ectopic DoublecortinGene Expression Suppresses the Malignant Phenotype in Glioblastoma Cells.Cancer Res.2006;66:11726-11735.
28. Manoranjan Santra, et al. Effect of doublecortin on self-renewal and differentiationin brain tumor stem cells. Cancer science, Accepted manuscript online.April7,2011
29. Santra M, Zheng X, Roberts C, et al. Single doublecortin gene therapy significantlyreduces glioma tumor volume.J Neurosci Res.2010Feb1;88(2):304-14.
30. Santra M, Santra S, Roberts C, et al. Doublecortin induces mitotic microtubulecatastrophe and inhibits glioma cell invasion. J Neurochem.2009,Jan;108(1):231-45.
31. Santra M, Liu XS, Santra S, et al. Ectopic expression of doublecortin protects adultrat progenitor cells and human glioma cells from severe oxygen and glucosedeprivation.Neuroscience.2006Oct27;142(3):739-52.
1.Manoranjan Santra1, Sutapa Santra1, Cindi Roberts1，el.Doublecortin inducesmitotic microtubule catastrophe and inhibits glioma cell invasion.J Neurochem.2009January;108(1):231–245.）
2. Legler JM, Gloeckler Ries LA, Smith MA, et al. Brain and other central nervoussystem cancers: recent trends in incidence and mortality. J Natl Cancer Inst1999;91:1382–90.
3. Jemal A, Murray T, Ward E, et al. Cancer statistics,2005. CA Cancer J Clin2005;55:10–30.
4. Rich JN, Hans C, Jones B, et al. Gene expression profiling and genetic markersin glioblastoma survival.Cancer Res2005;6510:4051–8.
5. Francis F, Koulakoff A, Boucher D, Chafey P, Schaar B, Vinet MC, Friocourt G,McDonnell N, Reiner O, Kahn A, et al.: Doublecortin is a developmentally regulated,microtubule-associated protein expressed in migrating and differentiating neurons.Neuron1999,23(2):247-256.
6. des Portes V, Pinard JM, Billuart P, Vinet MC, Koulakoff A, Carrie A, GelotA,Dupuis E, Motte J, Berwald-Netter Y, et al.: A novel CNS gene required for neuronalmigration and involved in X-linked subcortical laminar heterotopia and lissencephalysyndrome. Cell1998,92(1):51-61.
7. Walker TL, Yasuda T, Adams DJ, Bartlett PF: The doublecortin-expressingpopulation in the developing and adult brain contains multipotential precursors inaddition to neuronal-lineage cells. Journal of Neuroscience2007,27(14):3734-3742.
8. Friocourt G, Koulakoff A, Chafey P, Boucher D, Fauchereau F, Chelly J,FrancisF: Doublecortin functions at the extremities of growing neuronal processes. CerebCortex2003,13(6):620-626.
9. Brown JP, Couillard-Despres S, Cooper-Kuhn CM, Winkler J, Aigner L, KuhnHG: Transient expression of doublecortin during adult neurogenesis. JComp Neurol2003,467(1):1-10.
10. Mattar P, Britz O, Johannes C, Nieto M, Ma L, Rebeyka A, Klenin N, PolleuxF, Guillemot F, Schuurmans C: A screen for downstream effectors of Neurogenin2inthe embryonic neocortex. Developmental Biology2004,273(2):373-389.
11. Couillard-Despres S, Winner B, Schaubeck S, Aigner R, Vroemen M,WeidnerN, Bogdahn U, Winkler J, Kuhn H, Aigner L: Doublecortin expression levels in adultbrain reflect neurogenesis. Eur J Neurosci2005,21(1):1-14.
12. Gleeson JG, Allen KM, Fox JW, Lamperti ED, Berkovic S, Scheffer I, CooperEC, Dobyns WB, Minnerath SR, Ross ME, et al.: Doublecortin, a brainspecific genemutated in human X-linked lissencephaly and double cortex syndrome, encodes aputative signaling protein. Cell1998,92(1):63-72.
13. Schaar, B. T., Kinoshita, K., and McConnell, S. K.(2004) Neuron41,203–2131
14. Tanaka, T., Serneo, F. F., Tseng, H-C., Kulkarni, A. B., Tsai, L-H., andGleeson, J. G.(2004) Neuron41,215–227
15.RAes esahrcho arrttic leupstream promoter region mediates transcriptionalregulation of the mouse doublecortin gene in differentiating neuronsPiens et al. BMCNeuroscience2010,11:64）
16.Doublecortin Association with Actin Filaments Is Regulated by Neurabin II*THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol.280, No.12, Issue of March25,pp.11361–11368,2005）
17. Zhou Q, Anderson DJ: The bHLH transcription factors OLIG2and OLIG1couple neuronal and glial subtype specification. Cell2002,109(1):61-73
18. Ross ME, Allen KM, Srivastava AK, et al. Linkage and physical mapping ofX-linked lissencephaly/SBH (XLIS):a gene causing neuronal migration defects inhuman brain. Hum Mol Genet1997;64:555–62.
19. Santra M, Lui XS, Zhang J, Zhang RL, Zhang ZG,Chopp M. Ectopicexpression of doublecortin protects adult mouse progenitor cells and human gliomacells from severe oxygen and glucose deprivation. Neuroscience2006;92:7016–20.
20. Tsuiki H, Tnani M, Okamoto I, et al. Constitutively active forms of c-JunNH2-terminal kinase are expressed in primary glial tumors. Cancer Res2003;631:250–5.
21.Phosphoproteomic Analysis of the Developing Mouse Brain*Molecular&Cellular Proteomics3:1093–1101,2004.）
22. Tiziano Pramparo1, Yong Ha Youn1, Jessica Yingling2el. Novel embryonicneuronal migration and proliferation defects in Dcx mutant mice are exacerbated byLis1reduction.J Neurosci.2010February24;30(8):3002–3012.
23. Cohen P. The origins of protein phosphorylation. Nat Cell Biol2002;45:E127–30.
24. Inui S, Sanjo H, Maeda K, Yamamoto H, Miyamoto E,Sakaguchi N. Igreceptor binding protein1(a4) is associated with a rapamycin-sensitive signaltransduction in lymphocytes through direct binding to the catalytic subunit of proteinphosphatase2A. Blood1998;922:539–46.
25. MacMillan LB, Bass MA, Cheng N, et al. Brain actinassociated proteinphosphatase1holoenzymes containing spinophilin, neurabin, and selected catalyticsubunit isoforms. J Biol Chem1999;27450:35845–54.31.
26. Shmueli A, Gdalyahu A, Sapoznik S, Sapir T, Tsukada M, Reiner O.Site-specific dephosphorylation of doublecortin (DCX) by protein phosphatase1(PP1).Mol Cell Neurosci2006. Epub ahead of print.
27. Manoranjan Santra1, Sutapa Santra1, Cindi Roberts1，el.Doublecortin inducesmitotic microtubule catastrophe and inhibits glioma cell invasion.J Neurochem.2009January;108(1):231–245.