转染hVEGF165-GFP基因的内皮祖细胞对兔MODS模型微血栓形成影响的实验性研究
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摘要
内皮祖细胞(endothelial progenitor cell, EPC)是存在于外周血、骨髓及组织中,具备定向分化为成熟血管内皮潜能的前体细胞,是一类干细胞群体的总称。在创伤、炎症及失血所导致的组织损伤过程中,骨髓中的EPC可被大量动员,释放入外周血,在VEGF、SDF-1等趋化因子的作用下富集于损伤部位,修复血管内皮,改善局部血供,对组织的修复有着重要的意义。近年来,体外培养EPC用于治疗缺血性疾病及炎症损伤已得到广泛的研究。
多器官功能障碍综合征(MODS)是指机体遭受严重创伤、休克或感染24h后同时或序贯出现2个或2个以上的系统或器官的功能障碍,是多种疾病最严重的并发症和最终结局。创伤及感染是发生MODS的两个重要诱因,两者均可激活炎症细胞及因子,使机体产生过度的应激及炎症反应,内皮细胞是炎症介质首先打击的靶细胞,其损伤和功能紊乱,导致炎症反应的放大,内环境无法保持稳态,导致MODS的发生。
本实验通过以慢病毒载体介导hVEGF165-GFP基因转染EPC,并观察其对双相迟发MODS动物模型脏器功能、微循环改变的影响,研究异体移植EPC对MODS的治疗作用及相关机制,并探寻其更优方案。
本实验共分为三个部分：一、体外分离、培养及鉴定兔骨髓来源的EPC,检测其增殖、分化及成血管能力,为异体移植提供可靠的技术平台和细胞来源：二、构建慢病毒载体,将VEGF-GFP目的基因转染入EPC,监测细胞增殖功能变化；三、异体移植EPC用于治疗MODS动物,观察移植后重要脏器功能改变、微循环中血栓形成情况以及内皮祖细胞相关细胞因子(MCP-1,ANG-1)的表达情况,评价转染VEGF-GFP双表达基因的EPC对MODS的治疗效果。
第一部分家兔骨髓来源内皮祖细胞的培养及鉴定
本部分旨在建立起家兔骨髓来源的EPC的标准化分离、培养、扩增及鉴定的方法,为下一步病毒转染及EPC移植治疗MODS的研究提供技术平台及细胞来源。
密度梯度离心法分离家兔骨髓,提取、纯化单个核细胞,按照1×105/cm2的密度接种于培养皿内,以含有多种促生长因子(VEGF、bFGF)及胎牛血清的内皮祖细胞专用培养液培养及诱导分化,观察细胞形态、数量变化,并进行传代扩增。对培养的P3代EPC进行形态学特征、超微结构、表面抗原(CD133、CD31、KDR)及吞噬实验(FITC-UEA-1、Dil-ac-LDL)、体外血管生成功能等检测及鉴定,以确认培养的细胞即是EPC,为后文的移植治疗做好准备工作。
结果显示：培养24-48小时后即出现梭形贴壁细胞,培养6-8天为细胞增殖高峰,可观察到集落形成。消化传代后,检测P3代细胞,电镜下可见内皮细胞特异性标志—Weibel-Palade小体；细胞表面抗原检测表明CD133阳性率为4%,,CD31及KDR阳性率超过90%；贴壁细胞可特异性摄取了Dil-ac-LDL和FITC-UEA-1,表现出内皮细胞的特征。P3代细胞体外生成血管功能检测提示细胞间可相互连接,生成毛细血管网状结构。本部分实验从兔骨髓中分离出单个核细胞,并成功诱导分化出带有干细胞表型及血管内皮细胞特征,能有效形成毛细血管结构的EPC。
第二部分慢病毒载体的构建和内皮祖细胞的转染
本实验部分构建hVEGF/GFP双表达基因慢病毒载体,并转染体外培养的内皮祖细胞,为下一步的细胞移植治疗MODS提供相关的细胞来源。
将预先构建的hVEGF165-GFP双表达盒连接至穿梭质粒pDC315的多克隆位点上。得到重组穿梭质粒pDC315-hVEGF165-GFP。利用酶切技术,将hVEGF165-GFP目的片段连接于慢病毒载体的转移质粒上,构建FUGW/hVEGF165-GFP转移质粒,将慢病毒转移质粒FUGW/hVEGF165-GFP,包装质粒pCMVR-delta8.9及包膜质粒VSVG通过磷酸钙法三质粒瞬时共转染293 FT细胞,得到LV/hVEGF165-GFP,将LV/hVEGF165-GFP与EPC共培养以感染EPC。检测转染后细胞增殖能力。结果表明,目的基因插入慢病毒过表达载体pWPXL-MOD后PCR扩增载体片段与目的片段均可表达；EPC经转染hVEGF-GFP双表达基因后均能稳定表达VEGF及GFP,且细胞增殖能力较未转染前显著提高。
第三部分转染hVEGF165-GFP基因内皮祖细胞对兔MODS模型微血栓形成的影响
本部分实验在构建MODS动物模型的基础上,予异体移植体外培养的EPC及转染hVEGF-GFP双表达基因的EPC,观察重要脏器的功能变化及微循环中血栓的形成情况,并检测EPC旁分泌功能相关细胞因子,以了解EPC移植对MODS的治疗作用及相关机制,并探索其更优方案。
健康家兔36只,随机分为三组,予失血性休克及内毒素打击,构建MODS模型：未治疗动物12只作为对照组(M组)；12只动物行单纯EPC移植(ET组)；12只动物予转染hVEGF-GFP双表达基因内皮祖细胞。移植时间点为内毒素打击后1小时,移植数量为1×107/公斤体重,监测脏器功能变化,动物死亡或观察96h后处死,取脏器行病理学检查,Masson及HE染色后对组织中微血栓进行计数,并采用ELISA法检测组织中MCP-1及ANG-1水平差异。
实验结果提示：移植EPC均能有效改善动物各重要脏器功能,减少微循环中血栓形成,并通过旁分泌机制,促进组织修复,降低MODS发生率及动物死亡率,对MODS有积极的治疗效果。
EPC(endothelial progenitor cell) is a kind of cells which resides in the peripheral blood, bone marrow and organs and could differentiate into mature endothelial cells, working as stem cells. When the tissue damages are caused by trauma, inflammation or hemorrhagic shock, EPC could be released from bone marrow and released into the peripheral blood. Induced by chemokines such as VEGF, SDF-1, EPC assemble in the injury area and repair vascular endothelial which could improve the local blood supply and make a repairment of organizations. In recent years, studies on cultivation and applications of EPC to treat ischemic diseases have become a focus in medical researches.
When body suffers serious trauma, shock or infection, at least two system or organ dysfunctions could be induced which is described as multiple organ dysfunction syndromes (MODS). MODS is the most serious complications of many diseases and the final outcome. Trauma and infection are two important causes of MODS, both of which could activate inflammatory cells, inducing excessive inflammatory response. Endothelial cell is the first target of inflammatory mediators. The injury and dysfunction of endothelial cells could lead to amplification of inflammatory response and ruin steady-state of internal environment, resulting in the occurrence of MODS.
In this study, EPCs were cultured in vitro and transfecetd with VEGF165-GFP gene by lentiviral vector. After that, the cells were transplanted into rabbits which suffered MODS induced by hemorrhagic shock and LPS. The changes of microcirculation were observed and cytokines including ANG-1 and MCP-1 were measured which may reveal the therapeutic effect and related mechanism of EPC in the treatment of MODS
The experiment included four parts:First, marrow-derived mononuclear cells were cultured in vitro; Then, lentiviral vectors were constructed and the VEGF-GFP gene was transfected into EPC. The functions of cell proliferation were observed; in the end, EPCs were allotransplanted into the rabbits suffering MODS which were induced by hemorrhagic shock and LPS. The changes in post-transplant function of organs were observed and micro thrombi were counted as well as the level f EPC related cytokines (MCP-1, ANG-1).
PartⅠCulture and Identification of Marrow-derived Endothelial Progenitor Cells
In this part, Marrow-derived mononuclear cells were cultured and amplified in vitro, so were the identification of EPC functions.
Mononuclear cells were got and purified by density gradient centrifugations, inoculated in petri dishes in density of 1×105/cm2 using a culture media containing a variety of growth-promoting factors. Generations, morphological features, ultra structure, surface antigens (CD133, CD31, and KDR) and phagocytic experiments (FITC-UEA-1, Dil-ac-LDL), angiogenic functions in vitro were detected and identified in the tert cell.
The results showed that:fusiform adherent cells emergenced in 24-48 hours after being cultured. The generation reached peak level at 6-10 days and the colony formation could be observed. The Weibel-Palade body was shown by electron microscope; positive rate of CD133-rate was 4%, the positive rate of CD34, and KDR was above 90%; adherent cells could specially uptake Dil-ac-LDL and FITC-UEA-1, showing the characteristics of endothelial cells. Tert cells could establish capillary network structures in vitro.
PartⅡConstruction of a lent viral vector and Transfection of VEGF-GFP Gene into EPC in vitro
In this part, hVEGF/GFP fusion gene lent viral vectors were made and transfected into endothelial progenitor cells in vitro. The experiments provided a cell source for the next step of study.
The VEGF PCR primers were designed, synthesized and amplified. The combination and transformation were made after these PCR products were cut, then inserted into the lent viral vector--pWPXL-MOD, which contains GFP gene. The positive clones formation were picked and cut by the restriction enzymes of BamH I、Mlu I, and electrophoresis checking were done to confirm that the sequence was correct.
The expression plasmids--pWPXL-MOD were used in the experiment, which contained VEGF-GFP gene as well as the package plasmids--pRsv-REV, pMDlg-pRRE, the envelope plasmids-pMD2G. Then a four-plasmids system of lent viral vector were constructed. The LV-VEGF-GFP could be got in a high concentration when the vectors were concentred. Lent virus and EPC were co-cultured in the MOI for 50 and transfect ion could be accomplished very well. Then the functions of proliferation were identified. The results showed that when the objective gene of VEGF-GEP was inserted into the over-expression carrier of pWPXL-MOD, both the carrier fragment and objective fragment could express. Furthermore the report of gene sequencing showed that the sequence of insertion element in the recombinant plasmids was accordant to the sequence of objective gene of VEGF. In a word, we constructed and packaged the lent viral vectors successfully and proved that the ability of proliferation of post-transfected EPC is significantly enhanced contrast to the normal.
PartⅢThe treatment of MODS by allotransplanting Endothelial Progenitors Cells transfected with Gene of ThVEGF165-GFP
In the parts, the MODS model was prepared and the EPCs were allogeneic transplanted as a treatment. Organ functions and micro-thrombosis in tissues were observed, as well as detection of EPC related cytokine (MCP-1, ANG-1).
36 rabbits were randomly assigned into three groups and MODS model was constructed. The group without treatment was used as the control (M); group treated with MODS and transplantation of EPCs was the EPC treatment group (ET); the other 12 animals treated with MODS and transplatation of EPC transfected with hVEGF-GFP fusion gene was assigned as VT group. The point of transplantation was 1 hour after infusion of LPS and the dosage of cells was 1 x 107/kg weight. Organ functions were monitored. Animals were executed after 96h. Pathological examination and micro thrombus count were made. The ELISA method was used to detect levels of MCP-1 and ANG-1 in tissues.
These results suggested that EPC transplantation effectively improved the functions of important organs, inhibited the micro thrombosis in circulation and reduce the mortality of MODS.
多器官功能障碍综合征(MODS)是指机体遭受严重创伤、休克或感染24h后同时或序贯出现2个或2个以上的系统或器官的功能障碍,是多种疾病最严重的并发症和最终结局。创伤及感染是发生MODS的两个重要诱因,两者均可激活炎症细胞及因子,使机体产生过度的应激及炎症反应,内皮细胞是炎症介质首先打击的靶细胞,其损伤和功能紊乱,导致炎症反应的放大,内环境无法保持稳态,导致MODS的发生。
本实验通过以慢病毒载体介导hVEGF165-GFP基因转染EPC,并观察其对双相迟发MODS动物模型脏器功能、微循环改变的影响,研究异体移植EPC对MODS的治疗作用及相关机制,并探寻其更优方案。
本实验共分为三个部分：一、体外分离、培养及鉴定兔骨髓来源的EPC,检测其增殖、分化及成血管能力,为异体移植提供可靠的技术平台和细胞来源：二、构建慢病毒载体,将VEGF-GFP目的基因转染入EPC,监测细胞增殖功能变化；三、异体移植EPC用于治疗MODS动物,观察移植后重要脏器功能改变、微循环中血栓形成情况以及内皮祖细胞相关细胞因子(MCP-1,ANG-1)的表达情况,评价转染VEGF-GFP双表达基因的EPC对MODS的治疗效果。
第一部分家兔骨髓来源内皮祖细胞的培养及鉴定
本部分旨在建立起家兔骨髓来源的EPC的标准化分离、培养、扩增及鉴定的方法,为下一步病毒转染及EPC移植治疗MODS的研究提供技术平台及细胞来源。
密度梯度离心法分离家兔骨髓,提取、纯化单个核细胞,按照1×105/cm2的密度接种于培养皿内,以含有多种促生长因子(VEGF、bFGF)及胎牛血清的内皮祖细胞专用培养液培养及诱导分化,观察细胞形态、数量变化,并进行传代扩增。对培养的P3代EPC进行形态学特征、超微结构、表面抗原(CD133、CD31、KDR)及吞噬实验(FITC-UEA-1、Dil-ac-LDL)、体外血管生成功能等检测及鉴定,以确认培养的细胞即是EPC,为后文的移植治疗做好准备工作。
结果显示：培养24-48小时后即出现梭形贴壁细胞,培养6-8天为细胞增殖高峰,可观察到集落形成。消化传代后,检测P3代细胞,电镜下可见内皮细胞特异性标志—Weibel-Palade小体；细胞表面抗原检测表明CD133阳性率为4%,,CD31及KDR阳性率超过90%；贴壁细胞可特异性摄取了Dil-ac-LDL和FITC-UEA-1,表现出内皮细胞的特征。P3代细胞体外生成血管功能检测提示细胞间可相互连接,生成毛细血管网状结构。本部分实验从兔骨髓中分离出单个核细胞,并成功诱导分化出带有干细胞表型及血管内皮细胞特征,能有效形成毛细血管结构的EPC。
第二部分慢病毒载体的构建和内皮祖细胞的转染
本实验部分构建hVEGF/GFP双表达基因慢病毒载体,并转染体外培养的内皮祖细胞,为下一步的细胞移植治疗MODS提供相关的细胞来源。
将预先构建的hVEGF165-GFP双表达盒连接至穿梭质粒pDC315的多克隆位点上。得到重组穿梭质粒pDC315-hVEGF165-GFP。利用酶切技术,将hVEGF165-GFP目的片段连接于慢病毒载体的转移质粒上,构建FUGW/hVEGF165-GFP转移质粒,将慢病毒转移质粒FUGW/hVEGF165-GFP,包装质粒pCMVR-delta8.9及包膜质粒VSVG通过磷酸钙法三质粒瞬时共转染293 FT细胞,得到LV/hVEGF165-GFP,将LV/hVEGF165-GFP与EPC共培养以感染EPC。检测转染后细胞增殖能力。结果表明,目的基因插入慢病毒过表达载体pWPXL-MOD后PCR扩增载体片段与目的片段均可表达；EPC经转染hVEGF-GFP双表达基因后均能稳定表达VEGF及GFP,且细胞增殖能力较未转染前显著提高。
第三部分转染hVEGF165-GFP基因内皮祖细胞对兔MODS模型微血栓形成的影响
本部分实验在构建MODS动物模型的基础上,予异体移植体外培养的EPC及转染hVEGF-GFP双表达基因的EPC,观察重要脏器的功能变化及微循环中血栓的形成情况,并检测EPC旁分泌功能相关细胞因子,以了解EPC移植对MODS的治疗作用及相关机制,并探索其更优方案。
健康家兔36只,随机分为三组,予失血性休克及内毒素打击,构建MODS模型：未治疗动物12只作为对照组(M组)；12只动物行单纯EPC移植(ET组)；12只动物予转染hVEGF-GFP双表达基因内皮祖细胞。移植时间点为内毒素打击后1小时,移植数量为1×107/公斤体重,监测脏器功能变化,动物死亡或观察96h后处死,取脏器行病理学检查,Masson及HE染色后对组织中微血栓进行计数,并采用ELISA法检测组织中MCP-1及ANG-1水平差异。
实验结果提示：移植EPC均能有效改善动物各重要脏器功能,减少微循环中血栓形成,并通过旁分泌机制,促进组织修复,降低MODS发生率及动物死亡率,对MODS有积极的治疗效果。
EPC(endothelial progenitor cell) is a kind of cells which resides in the peripheral blood, bone marrow and organs and could differentiate into mature endothelial cells, working as stem cells. When the tissue damages are caused by trauma, inflammation or hemorrhagic shock, EPC could be released from bone marrow and released into the peripheral blood. Induced by chemokines such as VEGF, SDF-1, EPC assemble in the injury area and repair vascular endothelial which could improve the local blood supply and make a repairment of organizations. In recent years, studies on cultivation and applications of EPC to treat ischemic diseases have become a focus in medical researches.
When body suffers serious trauma, shock or infection, at least two system or organ dysfunctions could be induced which is described as multiple organ dysfunction syndromes (MODS). MODS is the most serious complications of many diseases and the final outcome. Trauma and infection are two important causes of MODS, both of which could activate inflammatory cells, inducing excessive inflammatory response. Endothelial cell is the first target of inflammatory mediators. The injury and dysfunction of endothelial cells could lead to amplification of inflammatory response and ruin steady-state of internal environment, resulting in the occurrence of MODS.
In this study, EPCs were cultured in vitro and transfecetd with VEGF165-GFP gene by lentiviral vector. After that, the cells were transplanted into rabbits which suffered MODS induced by hemorrhagic shock and LPS. The changes of microcirculation were observed and cytokines including ANG-1 and MCP-1 were measured which may reveal the therapeutic effect and related mechanism of EPC in the treatment of MODS
The experiment included four parts:First, marrow-derived mononuclear cells were cultured in vitro; Then, lentiviral vectors were constructed and the VEGF-GFP gene was transfected into EPC. The functions of cell proliferation were observed; in the end, EPCs were allotransplanted into the rabbits suffering MODS which were induced by hemorrhagic shock and LPS. The changes in post-transplant function of organs were observed and micro thrombi were counted as well as the level f EPC related cytokines (MCP-1, ANG-1).
PartⅠCulture and Identification of Marrow-derived Endothelial Progenitor Cells
In this part, Marrow-derived mononuclear cells were cultured and amplified in vitro, so were the identification of EPC functions.
Mononuclear cells were got and purified by density gradient centrifugations, inoculated in petri dishes in density of 1×105/cm2 using a culture media containing a variety of growth-promoting factors. Generations, morphological features, ultra structure, surface antigens (CD133, CD31, and KDR) and phagocytic experiments (FITC-UEA-1, Dil-ac-LDL), angiogenic functions in vitro were detected and identified in the tert cell.
The results showed that:fusiform adherent cells emergenced in 24-48 hours after being cultured. The generation reached peak level at 6-10 days and the colony formation could be observed. The Weibel-Palade body was shown by electron microscope; positive rate of CD133-rate was 4%, the positive rate of CD34, and KDR was above 90%; adherent cells could specially uptake Dil-ac-LDL and FITC-UEA-1, showing the characteristics of endothelial cells. Tert cells could establish capillary network structures in vitro.
PartⅡConstruction of a lent viral vector and Transfection of VEGF-GFP Gene into EPC in vitro
In this part, hVEGF/GFP fusion gene lent viral vectors were made and transfected into endothelial progenitor cells in vitro. The experiments provided a cell source for the next step of study.
The VEGF PCR primers were designed, synthesized and amplified. The combination and transformation were made after these PCR products were cut, then inserted into the lent viral vector--pWPXL-MOD, which contains GFP gene. The positive clones formation were picked and cut by the restriction enzymes of BamH I、Mlu I, and electrophoresis checking were done to confirm that the sequence was correct.
The expression plasmids--pWPXL-MOD were used in the experiment, which contained VEGF-GFP gene as well as the package plasmids--pRsv-REV, pMDlg-pRRE, the envelope plasmids-pMD2G. Then a four-plasmids system of lent viral vector were constructed. The LV-VEGF-GFP could be got in a high concentration when the vectors were concentred. Lent virus and EPC were co-cultured in the MOI for 50 and transfect ion could be accomplished very well. Then the functions of proliferation were identified. The results showed that when the objective gene of VEGF-GEP was inserted into the over-expression carrier of pWPXL-MOD, both the carrier fragment and objective fragment could express. Furthermore the report of gene sequencing showed that the sequence of insertion element in the recombinant plasmids was accordant to the sequence of objective gene of VEGF. In a word, we constructed and packaged the lent viral vectors successfully and proved that the ability of proliferation of post-transfected EPC is significantly enhanced contrast to the normal.
PartⅢThe treatment of MODS by allotransplanting Endothelial Progenitors Cells transfected with Gene of ThVEGF165-GFP
In the parts, the MODS model was prepared and the EPCs were allogeneic transplanted as a treatment. Organ functions and micro-thrombosis in tissues were observed, as well as detection of EPC related cytokine (MCP-1, ANG-1).
36 rabbits were randomly assigned into three groups and MODS model was constructed. The group without treatment was used as the control (M); group treated with MODS and transplantation of EPCs was the EPC treatment group (ET); the other 12 animals treated with MODS and transplatation of EPC transfected with hVEGF-GFP fusion gene was assigned as VT group. The point of transplantation was 1 hour after infusion of LPS and the dosage of cells was 1 x 107/kg weight. Organ functions were monitored. Animals were executed after 96h. Pathological examination and micro thrombus count were made. The ELISA method was used to detect levels of MCP-1 and ANG-1 in tissues.
These results suggested that EPC transplantation effectively improved the functions of important organs, inhibited the micro thrombosis in circulation and reduce the mortality of MODS.
引文
[1]Suda T, Takakura. Role of hematopoietic stem cells in angiogenesis. Int J Hematol.2001 Oct;74(3):266-71.
[2]Hristov M, Erl W, Weber PC. Endothelial progenitor cells:mobilization, differentiation, and homing. Arterioscler Thromb Vasc Biol.2003 Jul 1;23(7):1185-9. Epub 2003 Apr 24.
[3]Rabelink TJ, de Boer HC, de Koning EJ, et al. Endothelial progenitor cells:more than an inflammatory response? Arterioscler Thromb Vasc Biol.2004 May;24(5):834-8.
[4]Heissig B, Hattori K, Dias S, Friedrich M, et al. Recruitment of stem and progenitor cells from the bone marrow niche requires MMP-9 mediated release of kit-ligand. Cell.2002 May 31;109(5):625-37.
[5]Asahara T, Murohara T, Sullivan A,et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science.1997 Feb 14;275(5302):964-7.
[6]Tateishi Yuyama E, Matsubara H, Murohara T, et al.Therapeutic angio genesis for patients with limb ischaemia by autologous transplantation of bone marrow cells:a pilot study and a randomized controled trial. Lancet,2002,360:427-35.
[7]Yamada M, Kubo H, Ishizawa K, et al. Increased circulating endothelial progenitor cells in patients with bacterial pneumonia:evidence that bone marrow derived cells contribute to lung repair. Thorax.2005 May;60(5):410-3.
[8]盛志勇,胡森.多器官功能障碍综合征[M].北京：科学出版社,2000：186-187.
[9]Tiney NL, et al. Sequential system failure after rupture of abdominal aortic aneumysms. Ann Surg,1973; 178(2):117-122.
[10]WELCH CE. The treatment of combined intestinal obstruction and peritonitis by refunctionalization of the intestine. Ann Surg.1955 Oct; 142(4):739-49; discussion,749-51.
[11]Weber E, Rossi A, Solito R, et al. Focal adhesion molecules expression and fibrillin deposition by lymphatic and blood vessel endothelial cells in culture. Microvasc Res.2002 Jul;64(1):47-55.
[1]Li B, Sharpe EE, Maupin AB, et al. VEGF and PlGF promote adult vasculogenesis by enhancing EPC recruitment and vessel formation at the site of tumor neovascularization. FASEB J.2006 Jul; 20(9):1495-7.
[2]Zhao Y, Glesne D, Huberman E. A human peripheral blood monocytederived subset acts as pluripotent stem cells. Proc Natl Acad Sci 2003;100:2426-31.
[3]Chapel A, Bertho JM, Hartley RS, et al. Mesenchymal stem cells home to injured tissues when co-infused with hematopoietic cells to treat a radiation-induced multi-organ failure syndrome. J Gene Med 2003;5(12):1028-38
[4]李崇剑,高润霖,杨跃进等.猪心肌缺血再灌注损伤区自体骨髓单个核细胞和外周血内皮祖细胞移植后细胞分化探讨.中华心血管杂志.2007；35(4)：350-4.
[5]孙龙,胡波,朱洪生.自体血管内皮祖细胞植入与心肌血管新生.中华心血管外科杂志.2003；19(5)：315-6.
[6]Asahara T, Murohara T, Sullivan A, et al. Isolation of putative endothelial progenitorcells for angiogenesis. Science 1997;275:964-7.
[7]Hill JM, Gloria Zalos, Guleserian KJ et al. Circulating Endothelial Progenitor Cells,Vascular Function, and Cardiovascular Risk. N. Engl. J. Med.2003; 348:593-600.
[8]Vale PR, Isner JM, Rosenfield K. Therapeutic angiogenesis in critical limb and myocardial ischemia. J Interv Cardiol.2001 Oct;14(5):511-28.
[9]Li XQ, Meng QY, Wu HR. Effects of bone marrow-derived endothelial progenitor cell transplantation on vein microenvironment in a rat model of chronic thrombosis. Chin Med J (Engl).2007 Dec 20;120(24):2245-9.
[10]Walter DH, Rittig K, Bahlmann FH, et al. Statin therapy accelerates reendothelialization:a novel effect involving mobilization and incorporation of bone marrow-derived endothelial progenitor cells. Circulation.2002 Jun 25;105(25):3017-24.
[11]Ott I, Keller U, Knoedler M, et al. Endothelial-like cells expanded from CD34+blood cells improve left ventricular function after experimental myocardial infarction. FASEB J 2005, 19:992-4.
[12]Akashi M, Hisashi I, et al. Platelet-derived growth factor-BB(PDGF-BB) induces differentiation of bone marrow endothelial progenitor cell-derived cell line TR BM E2 into mural cells, and changes the phenotype. Celil Physiol 2005,204:948-955.
[13]Friedrich EB, Walenta K, Scharlau J.et al. CD34/CD133+/VEGFR-2+endothelial progenitor cell subpopulation with potent Vasoregenerative capacities. Circ Res 2006,98:20-25.
[1]Sinn PL, Sauter SL, Mccrayjrpb. Gene therapy progress and prospects:development and improvement of lentiviral and retroviral vectors-design, biosafetyandproduction. Gene Therapy 2005,12:1089-1098.
[2]Luttun A, Carmeliet G, Carmeliet P. Vascular progenitors:from biology to treatment. Trends Cardiovasc Med 2002,12:88-96.
[3]孙小杰,胡何节.血管内皮祖细胞与调控性细胞因子.国际外科学杂志.2007；34(12)：831-3.
[4]Mancuso P, Peccatori F, Rocca A. Circulating endothelial cell number and viability are reduced by exposure to high altitude. Endothelium 2008,15:53-58.
[5]You D, Cochain C, Loinard C. Hypertension impairs postnatal vasculogenesis:role of antihypertensive agents. Hypertension 2008,51:1537-44.
[6]Chang El, Loh SA, Ceradini DJ. Age decreases endothelial progenitor cell recruitment through decreases in hypoxia-inducible factor 1alpha stabilization during ischemia Circulation 2007, 116:2818-29.
[7]Martin K, Stanchina M, Kouttab N. Circulating endothelial cells and endothelial progenitor cells in obstructive sleep apnea. Lung 2008,186:145-50.
[8]罗天航,方国恩.自体移植内皮祖细胞防治多器官功能障碍的研究.优秀硕博论文库.2007,87-8.
[9]Shintani S. Murohara T, Ikeda H. et al. Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation.2001.103(23):2776-9.
[10]Gill M, Dias S, Hattori K, et al. Vascular trauma induces rapid but transient mobilization of VEGFR2(+)AC133(+)endothelial precursor cells. Circ Res.2001.88(2):167—174.
[11]Ackah E, Yu J, Zoellner S, et al. Akt1/protein kinase Balpha is critical for ischemic and VEGF-mediated angiogenesis. J Clin Invest.2005 Aug; 115(8):2119-27.
[1]del Pozo M A, Sanchez-Mateos P, Sanchez-Madrid F. Cellularpolarization induced by chemokines:a mechanism for leukocyte recruitment? Immunol Today,1996,17(3):127-31.
[2]Mann K G, van't Veer C, Cawthern K, et al. The role of the tissue factor pathway in initiation of coagulation. Blood Coagul Fibrinolysis,1998,9 Suppl 1:3～7.
[3]Anrather D, Millan M T, Palmetshofer A, et al. Thrombin activates nuclear factor-κB and potentiates endothelial cellactivation by TNF. J Immunol,1997,159(11):5620～8.
[4]Laszik Z, Mitro A, Taylor F B Jr, et al. Human protein Creceptor is present primarily on endothelium of large blood vessels:implications for the control of the protein C pathway. Circulation,1997,96(10):3633～40.
[5]Sandset P M, Warn-Cramer B J, Rao L V, et al. Depletion of extrinsic pathway inhibitor (EPI) sensitizes rabbits to disseminated intravascular coagulation induced with tissue factor: evidence supporting a physiologic role for EPI as a natural anticoagulant. Proc Natl Acad Sci USA,1991,88(3):708～12.
[6]Oeth P A, Parry G C, Kunsch C, et al. Lipopolysaccharide induction of tissue factor gene expression in monocytic cells is mediated by binding of c-Rel/p65 heterodimers to a κB-like site. Mol Cell Biol,1994,14(6):3772～81.
[7]Ohsawa M, Koyama T, Nara N, et al. Induction of tissue factor expression in human monocytic cells by protease inhibitors through activating activator protein-1 (AP-1) with phosphorylation of Jun-N-terminal kinase and p38. Thromb Res,2003,112(5-6):313～20.
[8]Marshall J C. Inflammation, coagulopathy, and the pathogenesis of multiple organ dysfunction syndrome. Crit Care Med,2001,29(7 Suppl):99～106.
[9]Asahara T, Murohara T, Sullivan A, et al. Isolation of putative endothelial progenitorcells for angiogenesis. Science 1997;275:964-7.
[10]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002;360(9331):427-35
[11]Yamada M, Kubo H, Ishizawa K, et al. Increased circulating endothelial progenitor cells in patients with bacterial pneumonia:evidence that bone marrow derived cells contribute to lung repair. Thorax.2005 May;60(5):410-3.
[12]Ishizawa K, Kubo H, Yamada M, Kobayashi S, et al. Bone marrow-derived cells contribute to lung regeneration after elastase-induced pulmonary emphysema. FEBS Lett.2004 Jan 2;556(1-3):249-52.
[13]Suratt BT, Cool CD, Serls AE, et al. Hunman pulmonary chimerism after hematopoietic stem cell transplantation. Am J Res p ir Crit CareMed,2003,168:318-2.
[14]罗天航,方国恩.自体移植内皮祖细胞防治多器官功能障碍的研究.优秀硕博论文库.2007,87-8.
[15]Asahara T, Takahashi T, Masuda H, Iwaguro H, et al. VEGF contributes to postnatal neovascularization by mobilizing bone marrow-derived endothelial progenitor cells.EMBO J. 1999 Jul 15;18(14):3964-72.
[16]Murayama T, Tepper OM, Silver M, et al. Determination of bone marrow-derived endothelial progenitor cell significance in angiogenic growth factor-induced neovascularization in vivo. Exp Hematol,2002,30 (8):967-72.
[17]SuriC. Requisite role of Angio Poietin-1, a ligand for the TIE-2 receptor, during embryonic angiogenesis. Cell,1996,87:1171-1180.
[18]Gamble, JR. Angiopoietin-1 is a antipermeability and anti-in-flammatory agent in vitro and targets cell junctions. Cir. Res,2000,87:603-607.
[19]Kim I, MoonSO, ParkSK et al. AngioPoietin-1 reduces VEGF stimulated leukocyte adhesion to endothelial cells by reducing ICAM-1, VCAM-1, and E selectin expression. Circ Res.2001, 89:477-479.
[20]WitzenbichlerB, WestermannD, KnueppelS, et al.Protective role of angiopoietin-1 in endotoxic shock. Circulation,2005,111:97-105.
[1]Rafii S, Oz MC, Seldomridge JA, Ferris B, et al. Characterization of hematopoietic cells arising on the textured surface of left ventricular assist devices. Ann Thorac Surg.1995 Dec;60(6):1627-32.
[2]Asahara T,Mumhara T,Suilivan A,et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science.199714;275(5302):964-967
[3]Akashi M, Hisashi I, et al. Platelet-derived growth factor-BB(PDGF-BB) induces differentiation of bone marrow endothelial progenitor cell-derived cell line TR BM E2 into mural cells, and changes the phenotype. Celil Physiol 2005,204:948-955.
[4]Friedrich EB, Walenta K, Scharlau J.et al. CD34/CD133+/VEGFR-2+endothelial progenitor cell subpopulation with potent Vasoregenerative capacities. Circ Res 2006,98:20-25.
[5]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002 Aug 10;360(9331):427-435
[6]He T, Peterson TE, Holmuhamedov EL, et al. Human endothelial progenitor cells tolerate oxidative stress due to intrinsically high expression of manganese superoxide dismutase. Arterioscler Thromb Vasc Biol.2004 Nov;24(11):2021-2027
[7]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002 Aug 10;360(9331):427-435
[8]Hofer T, Desbaillets I, Hopfl G, et al. Dissecting hypoxiadependent and hypoxia-independent steps in the HIF-1 a activation cascade:implications for HIF-1 a gene therapy. FASEB J, 2001,15(14):2715-7.
[9]Heissig B, Hattori K, Dias S, et al. Recruitment of stem and progenitor cells from the bone marrow niche requires MMP-9 mediated release of kit-ligand. Cell,2002,109 (5):625-37.
[10]Zheng JH, Pyatt DW, Gross SA, et al. Hydroquinone modulates the GM-CSF signaling pathway inTF-lcells. L eukemia.2004; 18:1296-304.
[11]Levesque JP, Takamatsu Y, Nilsson SK, et al. Vascular cell adhesion molecule-1 (CD106) is cleaved by neutrophil proteases in the bone marrow following hematopoietic progenitor cell mobilization by granulocyte colony-stimulating factor. Blood,2001,98(5):1289-97
[12]Takahashi T, Kalka C, Masuda H, et al. Ischemia-and cytokine-induced mobilization of bone marrow-derived endothelial progenitor cells for neovascularization. Nat Med.1999 Apr;5(4):434-8.
[13]Tian SS, Tapley P, Sincich C, et al. Multiple Signaling Pathways Induced by Granulocyte Colony-Stimulating Factor Involving Activation of JAKs, STATS, and/or STAT3 Are Required for Regulation of Three Distinct Classes of Immediate Early Genes.Blood,1996; 88:4435-44.
[14]Fuste B, Mazzara R, Escolar G, et al.Granulocyte colony-stimulating factor increases expression of adhesion receptors on cells through activation of p38 MAPK. Haematologica. 2004;89:578-85.
[15]Askari AT, Unzek S, Popovic ZB, et al. Effect of stromal cell-derived factor-1 on stem cell homing and tissue r generation in ischemic cardiomyopathy. Lancet,2003,362:697-703.
[16]Schober A,Knarren S, Lietz M, et al. Crucial role of stromal cell-derived factor-1 alpha in neointima formation after vascular injury in apolipoporte in E-deficient mice. Circulation, 2003,108(20):2491-7.
[17]Mallat Z, Silvestre JS, Le-Ricousse-RoussanneS, et al. Interleukin-18/interleukin-18 binding protein signaling modulates ischemia induced neovasculairzation in mice hindlimb. Circ Res, 2002; 91:441-9.
[18]Quesenberry PJ, Becker PS. Stem cell homing:Rolling,crawling, and nesting. Proc Natl Acad Sci USA,1998,95(26):15155-7.
[19]Levesque JP, Takamatsu Y, Nilsson SK, et al. Vascular cell adhesion molecule-1 (CD106) is cleaved by neutrophil proteases in the bone marrow following hematopoietic progenitor cell mobilization by granulocyte colony-stimulating factor. Blood,2001,98(5):1289-97.
[20]Butler JM, Guthrie SM, Koc M, et al. SDF-1 is both necessary and sufficient to promote proliferative retinopathy. J Clin Invest,2005,115(1):86-93.
[21]Fujii H, Li SH, Szmitko PE, et al. C-reactive protein alters antioxidant defenses and promotes apoptosis in endothelial progenitor cells. Arterioscler Thromb Vasc Biol,2006,26(11): 2476-82.
[2]Hristov M, Erl W, Weber PC. Endothelial progenitor cells:mobilization, differentiation, and homing. Arterioscler Thromb Vasc Biol.2003 Jul 1;23(7):1185-9. Epub 2003 Apr 24.
[3]Rabelink TJ, de Boer HC, de Koning EJ, et al. Endothelial progenitor cells:more than an inflammatory response? Arterioscler Thromb Vasc Biol.2004 May;24(5):834-8.
[4]Heissig B, Hattori K, Dias S, Friedrich M, et al. Recruitment of stem and progenitor cells from the bone marrow niche requires MMP-9 mediated release of kit-ligand. Cell.2002 May 31;109(5):625-37.
[5]Asahara T, Murohara T, Sullivan A,et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science.1997 Feb 14;275(5302):964-7.
[6]Tateishi Yuyama E, Matsubara H, Murohara T, et al.Therapeutic angio genesis for patients with limb ischaemia by autologous transplantation of bone marrow cells:a pilot study and a randomized controled trial. Lancet,2002,360:427-35.
[7]Yamada M, Kubo H, Ishizawa K, et al. Increased circulating endothelial progenitor cells in patients with bacterial pneumonia:evidence that bone marrow derived cells contribute to lung repair. Thorax.2005 May;60(5):410-3.
[8]盛志勇,胡森.多器官功能障碍综合征[M].北京：科学出版社,2000：186-187.
[9]Tiney NL, et al. Sequential system failure after rupture of abdominal aortic aneumysms. Ann Surg,1973; 178(2):117-122.
[10]WELCH CE. The treatment of combined intestinal obstruction and peritonitis by refunctionalization of the intestine. Ann Surg.1955 Oct; 142(4):739-49; discussion,749-51.
[11]Weber E, Rossi A, Solito R, et al. Focal adhesion molecules expression and fibrillin deposition by lymphatic and blood vessel endothelial cells in culture. Microvasc Res.2002 Jul;64(1):47-55.
[1]Li B, Sharpe EE, Maupin AB, et al. VEGF and PlGF promote adult vasculogenesis by enhancing EPC recruitment and vessel formation at the site of tumor neovascularization. FASEB J.2006 Jul; 20(9):1495-7.
[2]Zhao Y, Glesne D, Huberman E. A human peripheral blood monocytederived subset acts as pluripotent stem cells. Proc Natl Acad Sci 2003;100:2426-31.
[3]Chapel A, Bertho JM, Hartley RS, et al. Mesenchymal stem cells home to injured tissues when co-infused with hematopoietic cells to treat a radiation-induced multi-organ failure syndrome. J Gene Med 2003;5(12):1028-38
[4]李崇剑,高润霖,杨跃进等.猪心肌缺血再灌注损伤区自体骨髓单个核细胞和外周血内皮祖细胞移植后细胞分化探讨.中华心血管杂志.2007；35(4)：350-4.
[5]孙龙,胡波,朱洪生.自体血管内皮祖细胞植入与心肌血管新生.中华心血管外科杂志.2003；19(5)：315-6.
[6]Asahara T, Murohara T, Sullivan A, et al. Isolation of putative endothelial progenitorcells for angiogenesis. Science 1997;275:964-7.
[7]Hill JM, Gloria Zalos, Guleserian KJ et al. Circulating Endothelial Progenitor Cells,Vascular Function, and Cardiovascular Risk. N. Engl. J. Med.2003; 348:593-600.
[8]Vale PR, Isner JM, Rosenfield K. Therapeutic angiogenesis in critical limb and myocardial ischemia. J Interv Cardiol.2001 Oct;14(5):511-28.
[9]Li XQ, Meng QY, Wu HR. Effects of bone marrow-derived endothelial progenitor cell transplantation on vein microenvironment in a rat model of chronic thrombosis. Chin Med J (Engl).2007 Dec 20;120(24):2245-9.
[10]Walter DH, Rittig K, Bahlmann FH, et al. Statin therapy accelerates reendothelialization:a novel effect involving mobilization and incorporation of bone marrow-derived endothelial progenitor cells. Circulation.2002 Jun 25;105(25):3017-24.
[11]Ott I, Keller U, Knoedler M, et al. Endothelial-like cells expanded from CD34+blood cells improve left ventricular function after experimental myocardial infarction. FASEB J 2005, 19:992-4.
[12]Akashi M, Hisashi I, et al. Platelet-derived growth factor-BB(PDGF-BB) induces differentiation of bone marrow endothelial progenitor cell-derived cell line TR BM E2 into mural cells, and changes the phenotype. Celil Physiol 2005,204:948-955.
[13]Friedrich EB, Walenta K, Scharlau J.et al. CD34/CD133+/VEGFR-2+endothelial progenitor cell subpopulation with potent Vasoregenerative capacities. Circ Res 2006,98:20-25.
[1]Sinn PL, Sauter SL, Mccrayjrpb. Gene therapy progress and prospects:development and improvement of lentiviral and retroviral vectors-design, biosafetyandproduction. Gene Therapy 2005,12:1089-1098.
[2]Luttun A, Carmeliet G, Carmeliet P. Vascular progenitors:from biology to treatment. Trends Cardiovasc Med 2002,12:88-96.
[3]孙小杰,胡何节.血管内皮祖细胞与调控性细胞因子.国际外科学杂志.2007；34(12)：831-3.
[4]Mancuso P, Peccatori F, Rocca A. Circulating endothelial cell number and viability are reduced by exposure to high altitude. Endothelium 2008,15:53-58.
[5]You D, Cochain C, Loinard C. Hypertension impairs postnatal vasculogenesis:role of antihypertensive agents. Hypertension 2008,51:1537-44.
[6]Chang El, Loh SA, Ceradini DJ. Age decreases endothelial progenitor cell recruitment through decreases in hypoxia-inducible factor 1alpha stabilization during ischemia Circulation 2007, 116:2818-29.
[7]Martin K, Stanchina M, Kouttab N. Circulating endothelial cells and endothelial progenitor cells in obstructive sleep apnea. Lung 2008,186:145-50.
[8]罗天航,方国恩.自体移植内皮祖细胞防治多器官功能障碍的研究.优秀硕博论文库.2007,87-8.
[9]Shintani S. Murohara T, Ikeda H. et al. Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation.2001.103(23):2776-9.
[10]Gill M, Dias S, Hattori K, et al. Vascular trauma induces rapid but transient mobilization of VEGFR2(+)AC133(+)endothelial precursor cells. Circ Res.2001.88(2):167—174.
[11]Ackah E, Yu J, Zoellner S, et al. Akt1/protein kinase Balpha is critical for ischemic and VEGF-mediated angiogenesis. J Clin Invest.2005 Aug; 115(8):2119-27.
[1]del Pozo M A, Sanchez-Mateos P, Sanchez-Madrid F. Cellularpolarization induced by chemokines:a mechanism for leukocyte recruitment? Immunol Today,1996,17(3):127-31.
[2]Mann K G, van't Veer C, Cawthern K, et al. The role of the tissue factor pathway in initiation of coagulation. Blood Coagul Fibrinolysis,1998,9 Suppl 1:3～7.
[3]Anrather D, Millan M T, Palmetshofer A, et al. Thrombin activates nuclear factor-κB and potentiates endothelial cellactivation by TNF. J Immunol,1997,159(11):5620～8.
[4]Laszik Z, Mitro A, Taylor F B Jr, et al. Human protein Creceptor is present primarily on endothelium of large blood vessels:implications for the control of the protein C pathway. Circulation,1997,96(10):3633～40.
[5]Sandset P M, Warn-Cramer B J, Rao L V, et al. Depletion of extrinsic pathway inhibitor (EPI) sensitizes rabbits to disseminated intravascular coagulation induced with tissue factor: evidence supporting a physiologic role for EPI as a natural anticoagulant. Proc Natl Acad Sci USA,1991,88(3):708～12.
[6]Oeth P A, Parry G C, Kunsch C, et al. Lipopolysaccharide induction of tissue factor gene expression in monocytic cells is mediated by binding of c-Rel/p65 heterodimers to a κB-like site. Mol Cell Biol,1994,14(6):3772～81.
[7]Ohsawa M, Koyama T, Nara N, et al. Induction of tissue factor expression in human monocytic cells by protease inhibitors through activating activator protein-1 (AP-1) with phosphorylation of Jun-N-terminal kinase and p38. Thromb Res,2003,112(5-6):313～20.
[8]Marshall J C. Inflammation, coagulopathy, and the pathogenesis of multiple organ dysfunction syndrome. Crit Care Med,2001,29(7 Suppl):99～106.
[9]Asahara T, Murohara T, Sullivan A, et al. Isolation of putative endothelial progenitorcells for angiogenesis. Science 1997;275:964-7.
[10]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002;360(9331):427-35
[11]Yamada M, Kubo H, Ishizawa K, et al. Increased circulating endothelial progenitor cells in patients with bacterial pneumonia:evidence that bone marrow derived cells contribute to lung repair. Thorax.2005 May;60(5):410-3.
[12]Ishizawa K, Kubo H, Yamada M, Kobayashi S, et al. Bone marrow-derived cells contribute to lung regeneration after elastase-induced pulmonary emphysema. FEBS Lett.2004 Jan 2;556(1-3):249-52.
[13]Suratt BT, Cool CD, Serls AE, et al. Hunman pulmonary chimerism after hematopoietic stem cell transplantation. Am J Res p ir Crit CareMed,2003,168:318-2.
[14]罗天航,方国恩.自体移植内皮祖细胞防治多器官功能障碍的研究.优秀硕博论文库.2007,87-8.
[15]Asahara T, Takahashi T, Masuda H, Iwaguro H, et al. VEGF contributes to postnatal neovascularization by mobilizing bone marrow-derived endothelial progenitor cells.EMBO J. 1999 Jul 15;18(14):3964-72.
[16]Murayama T, Tepper OM, Silver M, et al. Determination of bone marrow-derived endothelial progenitor cell significance in angiogenic growth factor-induced neovascularization in vivo. Exp Hematol,2002,30 (8):967-72.
[17]SuriC. Requisite role of Angio Poietin-1, a ligand for the TIE-2 receptor, during embryonic angiogenesis. Cell,1996,87:1171-1180.
[18]Gamble, JR. Angiopoietin-1 is a antipermeability and anti-in-flammatory agent in vitro and targets cell junctions. Cir. Res,2000,87:603-607.
[19]Kim I, MoonSO, ParkSK et al. AngioPoietin-1 reduces VEGF stimulated leukocyte adhesion to endothelial cells by reducing ICAM-1, VCAM-1, and E selectin expression. Circ Res.2001, 89:477-479.
[20]WitzenbichlerB, WestermannD, KnueppelS, et al.Protective role of angiopoietin-1 in endotoxic shock. Circulation,2005,111:97-105.
[1]Rafii S, Oz MC, Seldomridge JA, Ferris B, et al. Characterization of hematopoietic cells arising on the textured surface of left ventricular assist devices. Ann Thorac Surg.1995 Dec;60(6):1627-32.
[2]Asahara T,Mumhara T,Suilivan A,et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science.199714;275(5302):964-967
[3]Akashi M, Hisashi I, et al. Platelet-derived growth factor-BB(PDGF-BB) induces differentiation of bone marrow endothelial progenitor cell-derived cell line TR BM E2 into mural cells, and changes the phenotype. Celil Physiol 2005,204:948-955.
[4]Friedrich EB, Walenta K, Scharlau J.et al. CD34/CD133+/VEGFR-2+endothelial progenitor cell subpopulation with potent Vasoregenerative capacities. Circ Res 2006,98:20-25.
[5]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002 Aug 10;360(9331):427-435
[6]He T, Peterson TE, Holmuhamedov EL, et al. Human endothelial progenitor cells tolerate oxidative stress due to intrinsically high expression of manganese superoxide dismutase. Arterioscler Thromb Vasc Biol.2004 Nov;24(11):2021-2027
[7]Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells:a pilot study and a randomised controlled trial. Lancet.2002 Aug 10;360(9331):427-435
[8]Hofer T, Desbaillets I, Hopfl G, et al. Dissecting hypoxiadependent and hypoxia-independent steps in the HIF-1 a activation cascade:implications for HIF-1 a gene therapy. FASEB J, 2001,15(14):2715-7.
[9]Heissig B, Hattori K, Dias S, et al. Recruitment of stem and progenitor cells from the bone marrow niche requires MMP-9 mediated release of kit-ligand. Cell,2002,109 (5):625-37.
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