摘要
目的:研究miR-126调控内皮祖细胞及对静脉血栓溶解、机化和再通的影响，为慢性深静脉血栓的治疗提供一种新的治疗思路。
方法：首先分离、培养及鉴定骨髓源性内皮祖细胞，将miR-126模拟物、抑制剂或阴性对照物用电转的方法转染到内皮祖细胞中，采用MTT、流式细胞术检测miR-126对内皮祖细胞增殖及细胞周期的影响；采用划痕实验、穿膜实验检测miR-126对内皮祖细胞运动迁移能力的影响；采用matrigel管腔形成实验检测miR-126对内皮祖细胞成小管能力的影响。进一步预测miR-126靶基因PIK3R2，并构建包含野生型PIK3R23’UTR和突变体PIK3R23’UTR的Luciferase报告基因载体，检测miR-126对PIK3R2的直接调控作用，Western blot分析PIK3R2蛋白以及PI3K/AKT信号通路中PI3K、Akt和p-Akt蛋白的变化。再构建miR-126慢病毒表达载体（pLVX-IRES-ZsGreenl-miR-126），HEK293T细胞包装、重组、扩增慢病毒颗粒，荧光显微镜观察绿色荧光表达， PCR、双酶切、基因测序进行鉴定。pLVX-IRES-ZsGreenl-miR-126体外转染内皮祖细胞，用实时荧光定量PCR(qPCR)方法检测转染后的内皮祖细胞中miR-126的表达。然后结扎大鼠肾下段下腔静脉建立深静脉血栓模型，将慢病毒转染的内皮祖细胞移植到静脉血栓模型中。分为三组：A组(12只)，空白对照组，经尾静脉注射1ml PBS；B组(12只)，EPCs/pLVX-IRES-ZsGreenVector (EPCs/vector)，经尾静脉注入含有1.0×106EPCs/vector的PBS细胞悬液。C组(12只)，EPCs/pLVX-IRES-ZsGreen-miR-126(EPCs/miR-126)，经尾静脉注入含有1.0×106EPCs/miR-126的PBS细胞悬液。移植后7天、14天取出血栓段下腔静脉及血栓，通过称重评价血栓溶解情况，荧光示踪观察内皮祖细胞归巢到静脉血栓情况，HE染色评价静脉血栓机化及CD34免疫组化观察血栓再通变化。采用SPSS15.0软件进行分析，P<0.05为差异，有统计学意义。
结果:成功培养、鉴定了大鼠骨髓源性内皮祖细胞。电转成功后，miR-126对内皮祖细胞早期具有促进增殖及改变细胞周期的作用，后期没有影响。划痕实验及穿膜实验都证实了miR-126模拟物能够显著促进内皮祖细胞的迁移运动能力；miR-126抑制剂则能抑制内皮祖细胞的迁移运动能力。miR-126模拟物可以增强内皮祖细胞的成小管能力，miR-126抑制剂能抑制内皮祖细胞的成小管能力。野生型PIK3R23’UTR和突变体PIK3R23’UTR的Luciferase报告基因载体构建及鉴定成功，共转染miR-126模拟物和pMIR/PIK3R2报告基因载体后，萤光素酶活性明显降低，而共转染miR-126抑制剂和pMIR/PIK3R2报告基因载体后，萤光素酶活性变化不大，共转染miR-126模拟物或抑制剂和pMIR/PIK3R2/mut报告基因载体后则不影响信号强度。上调内皮祖细胞中miR-126表达水平后，其PIK3R2蛋白表达水平明显下降，PI3K和磷酸化的Akt（p-Akt）蛋白表达上调，而下调内皮祖细胞中miR-126表达水平后，其PIK3R2蛋白表达水平明显增加，PI3K和磷酸化的Akt（p-Akt）蛋白表达下调。经酶切及测序结果提示慢病毒表达载体pLVX-IRES-ZsGreenl-miR-126构建正确，能有效转染内皮祖细胞，转染miR-126后在内皮祖细胞的表达较对照组高216倍。内皮祖细胞被移植到静脉血栓后，在7天和14天两个时间段内，EPCs/miR-126组中静脉血栓的重量最低，萤光阳性细胞数目最多，HE染色和CD34免疫组化提示更能促进血栓的机化和再通，均有统计学意义。
结论: miR-126能够促进内皮祖细胞成血管能力，包括增殖、迁移和成小管能力，miR-126能够负性调控其靶基因PIK3R2，激活PI3K/AKT信号通路。上调内皮祖细胞的miR-126表达后能够促进静脉血栓溶解、机化和再通。这可能是内皮祖细胞介导的治疗深静脉血栓的一种新思路。
Objective: This research is to study of miR-126regulating endothelial progenitor cells（EPCs）and promoting venous thrombosis resolution,organization and recanalization.And these findings indicate a potential therapeutic intervention of deep venous thrombosis.
Methods: Firstly, rat bone marrow-derived EPCs were isolated, cultivated andidentificated. Secondly,EPCs were transfected with control oligoes and miR-126mimicsor inhibitor by electroporation. Proliferation capacity and cell cycle were analyzed by MTTand flow cytometry.Cell migration analysis was done by wound healing and transwellassay.And tubulogenic activity test was performed by matrigel tube formation assay.Thirdly,by gain-of-function examination, its putative target genes were searched for usingonline search tool. PIK3R2was selected as the candidate target gene of rno-miR-126inEPCs.And further Luciferase reporters were constructed containing either a wild-typePIK3R23’UTR sequence (pMIR/PIK3R2/wt), or a mutated PIK3R23’UTR(pMIR/PIK3R2/mut). Luciferase activity was assessed by co-transfecting the luciferasereporter vectors with the miR-126mimics, inhibitor or NC. The luciferase activity ofreporter was observed. PIK3R2expression was detected by Western blot assays,and PIK3,Akt and p-Akt proteins in PI3K/AKT signal channel by Western blot assays,too.Fourthly,to stably express miR-126in EPCs, the lentiviral expression vectorpLVX-IRES-ZsGreen-miR-126was constructed.They was then transfected into293Tcells. The supernatant containing the lentivirus was harvested at72h. EPCs were infectedwith1ml lentivirus suspension. Green fluorescence was observed to indicate thetransduction efficiency at48h post transduction. miR-126expression in EPCs was detected by real-time quantitative PCR (qPCR).Forthermore, Experimental rat models ofdeep vein thrombosis were obtained by complete ligation of inferior vena cava below therenal veins. When deep venous thrombosis was formed after IVC ligation, the alive ratswere divided as3groups for cells transplantion via tail intravenous injection: A (n=12),blank control group (blank control), which received1ml PBS; B (n=12),EPCs/pLVX-IRES-ZsGreen Vector group (EPCs/vector), which received1.0×106EPCstransfected with lentivirus particle of pLVX-IRES-ZsGreen vector; C (n=12),EPCs/pLVX-IRES-ZsGreen-miR-126group (EPCs/miR-126), which received1.0×106EPCs transfected with lentivirus particle of pLVX-IRES-ZsGreen-miR-126. On the7th and14th day post operation, the rats were sacrificed and the thrombus segment of inferior venacava and thrombus were acquired,then were weighed. The homing of EPCs was shown byGFP expression and observed using fluorescence microscope. And the samples weretreated by dimethybenzene and embedded in paraffin. In order to observe thrombusorganization and recanalization, hematoxylin and eosin (HE) staining andimmunohistochemical staining for CD34and were performed. All statistical analyses wereperformed using SPSS15.0software. A two-tailed value of P <0.05was consideredstatistically significant.
Results: Rat bone marrow-derived EPCs were isolated, cultivated and identificatedsuccessfully.After EPCs were transfected by electroporation successfully, proliferationcapacity of EPCs was enhanced and cell cycle was changed in early stage when transfectedwith miR-126mimics.The relative wound size of EPCs transfected with NC or miR-126oligonucleotides was analyzed at0,24,48and72h post wounding. Compared with NCmimics, miR-126mimics significantly enhanced the migration of EPCs across the woundspace at24,48and72h. In contrast, miR-126inhibitor significantly suppressed themigration of EPCs at the same time. Consistent with the wound healing assay, migrationchamber assay also showed that miR-126mimics significantly promoted while miR-126inhibitor significantly inhibited EPCs migration into the chamber membrane comparedwith control group. EPCs transfected with miR-126mimics showed a significant improvement of capillary tube formation compared with NC by matrigel tube formationassay. In contrast, miR-126inhibitor-infected cells showed a significant impairment ofcapillary tube formation compared with NC. After an extensive review of onlinemicroRNA database (that is, TargetScan, Microrna.org and miRanda), we selected PIK3R2as the candidate target gene of rno-miR-126in EPCs. Luciferase reporter assays firstperformed to verify a direct interaction between miR-126and the3’UTR of PIK3R2.Luciferase reporters were constructed containing either a wild-type PIK3R23’UTRsequence (pMIR/PIK3R2/wt), or a mutated PIK3R23’UTR (pMIR/PIK3R2/mut).Luciferase activity was assessed by co-transfecting the luciferase reporter vectors with themiR-126mimics, inhibitor or NC. Luciferase activity of pMIR/PIK3R2/wt was markedlydecreased in cells transfected with miR-126mimics, compared to luciferase activity ofpMIR/PIK3R2/mut. Conversely, the luciferase activity of reporter plasmid was notinterfered after transfection with miR-126inhibitor. miR-126mimics decreased thePIK3R2expression compared to NC mimics, while miR-126inhibitor increased PIK3R2protein levels. PI3K and phospho-Akt (p-Akt) levels were significantly enhanced in theEPCs transfected with miR-126mimics compared to negative control mimics. A reverseresult was observed when miR-126was knocked down in EPCs transfected with miR-126inhibitor compared to negative control inhibitor.Restriction enzyme digestion andsequencing results suggest that lentiviral vector pLVX-IRES-ZsGreenl-miR-126wasconstructed correctly.And EPCs were effectively transfected by them.Aftertransfection,the miR-126expression in EPCs was higher216times than the control group.
On the7thand14thday after EPCs transplantation, Compared with control group, theweight of thrombus decreased in EPCs/vector and EPCs/miR-126groups. Furthermore, thethrombus weight of EPCs/miR-126group was even lower than that of EPCs/vectorgroup.EPCs homing were observed in section by using fluorescence microscope. Andtransplanted EPCs were recruited into the thrombus in both EPCs/vector andEPCs/miR-126group. In contrast, No GFP was observed in control group. We can alsofind that the larger number of GFP positive cells was appeared in EPCs/miR-126group compared with EPCs/vector group. HE staining indicated there were more degree oforganization in EPCs/vector and EPCs/miR-126group,even higher in EPCs/miR-126group. Compared with control group, more neovascularition were formed in EPCs/vectorand EPCs/miR-126group,and were maximum in EPCs/miR-126group.All werestatistically significant.
Conclusions: MiR-126enhances the ability of proangiogenic properties of EPCs,including proliferation, migration and tubulogenic activity, by targeting PIK3R2directly.The mechanism of miR-126effecting on EPCs is through PI3K/Akt signal pathway. EPCsvia upregulation of miR-126expression can promote venous thrombosisresolution,organization and recanalization. Based on the results in this research,regulationof miR-126in EPCs may represent a potential therapeutic intervention in EPCs-mediatedtherapy for deep venous thrombosis.
引文
[1] Wakefield TW, Myers DD, Henke PK. Mechanisms of venous thrombosis andresolution. Arteriosclerosis, thrombosis, and vascular biology.2008;28:387-91.
[2] Humphries J, McGuinness CL, Smith A, Waltham M, Poston R, Burnand KG.Monocyte chemotactic protein-1(MCP-1) accelerates the organization and resolutionof venous thrombi. Journal of vascular surgery.1999;30:894-9.
[3] Waltham M, Burnand K, Fenske C, Modarai B, Humphries J, Smith A. Vascularendothelial growth factor naked DNA gene transfer enhances thrombus recanalizationand resolution. Journal of vascular surgery.2005;42:1183-9.
[4] Henke PK, Wakefield TW, Kadell AM, Linn MJ, Varma MR, Sarkar M, et al.Interleukin-8administration enhances venous thrombosis resolution in a rat model.The Journal of surgical research.2001;99:84-91.
[5] Asahara T, Murohara T, Sullivan A, Silver M, van der Zee R, Li T, et al. Isolation ofputative progenitor endothelial cells for angiogenesis. Science.1997;275:964-7.
[6] Fadini GP, Losordo D, Dimmeler S. Critical reevaluation of endothelial progenitor cellphenotypes for therapeutic and diagnostic use. Circulation research.2012;110:624-37.
[7] Hur J, Yoon CH, Kim HS, Choi JH, Kang HJ, Hwang KK, et al. Characterization oftwo types of endothelial progenitor cells and their different contributions toneovasculogenesis. Arteriosclerosis, thrombosis, and vascular biology.2004;24:288-93.
[8] Moldovan NI, Asahara T. Role of blood mononuclear cells in recanalization andvascularization of thrombi: past, present, and future. Trends in cardiovascularmedicine.2003;13:265-9.
[9] Santo SD, Tepper OM, von Ballmoos MW, Diehm N, Volzmann J, Baumgartner I, et al.Cell-based therapy facilitates venous thrombus resolution. Thrombosis andhaemostasis.2009;101:460-4.
[10] Li XQ, Meng QY, Wu HR. Effects of bone marrow-derived endothelial progenitor celltransplantation on vein microenvironment in a rat model of chronic thrombosis.Chinese medical journal.2007;120:2245-9.
[11] Meng Q, Li X, Yu X, Lei F, Jiang K, Li C. Transplantation of ex vivo expanded bonemarrow-derived endothelial progenitor cells enhances chronic venous thrombusresolution and recanalization. Clinical and applied thrombosis/hemostasis: officialjournal of the International Academy of Clinical and Applied Thrombosis/Hemostasis.2011;17:E196-201.
[12] Meng QY, Li XQ, Yu XB, Lei FR, Jiang K, Li CY. Transplantation ofVEGF165-gene-transfected endothelial progenitor cells in the treatment of chronicvenous thrombosis in rats. Chinese medical journal.2010;123:471-7.
[13] Ott I, Keller U, Knoedler M, Gotze KS, Doss K, Fischer P, et al. Endothelial-like cellsexpanded from CD34+blood cells improve left ventricular function afterexperimental myocardial infarction. FASEB journal: official publication of theFederation of American Societies for Experimental Biology.2005;19:992-4.
[14] Iwaguro H, Yamaguchi J, Kalka C, Murasawa S, Masuda H, Hayashi S, et al.Endothelial progenitor cell vascular endothelial growth factor gene transfer forvascular regeneration. Circulation.2002;105:732-8.
[15] Urbich C, Heeschen C, Aicher A, Dernbach E, Zeiher AM, Dimmeler S. Relevance ofmonocytic features for neovascularization capacity of circulating endothelialprogenitor cells. Circulation.2003;108:2511-6.
[16] Latronico MV, Catalucci D, Condorelli G. Emerging role of microRNAs incardiovascular biology. Circulation research.2007;101:1225-36.
[17] Cordes KR, Srivastava D. MicroRNA regulation of cardiovascular development.Circulation research.2009;104:724-32.
[18] Suarez Y, Sessa WC. MicroRNAs as novel regulators of angiogenesis. Circulationresearch.2009;104:442-54.
[19] Bonauer A, Boon RA, Dimmeler S. Vascular microRNAs. Current drug targets.2010;11:943-9.
[20] Nicoli S, Standley C, Walker P, Hurlstone A, Fogarty KE, Lawson ND.MicroRNA-mediated integration of haemodynamics and Vegf signalling duringangiogenesis. Nature.2010;464:1196-200.
[21] van Solingen C, Seghers L, Bijkerk R, Duijs JM, Roeten MK, van Oeveren-RietdijkAM, et al. Antagomir-mediated silencing of endothelial cell specific microRNA-126impairs ischemia-induced angiogenesis. Journal of cellular and molecular medicine.2009;13:1577-85.
[22] Fish JE, Santoro MM, Morton SU, Yu S, Yeh RF, Wythe JD, et al. miR-126regulatesangiogenic signaling and vascular integrity. Developmental cell.2008;15:272-84.
[23] Wang S, Aurora AB, Johnson BA, Qi X, McAnally J, Hill JA, et al. Theendothelial-specific microRNA miR-126governs vascular integrity and angiogenesis.Developmental cell.2008;15:261-71.
[24] Zernecke A, Bidzhekov K, Noels H, Shagdarsuren E, Gan L, Denecke B, et al.Delivery of microRNA-126by apoptotic bodies induces CXCL12-dependent vascularprotection. Science signaling.2009;2:ra81.
[25] Zhang Q, Kandic I, Kutryk MJ. Dysregulation of angiogenesis-related microRNAs inendothelial progenitor cells from patients with coronary artery disease. Biochemicaland biophysical research communications.2011;405:42-6.
[26] Zhang J, Zhang Z, Zhang DY, Zhu J, Zhang T, Wang C. microRNA126inhibits thetransition of endothelial progenitor cells to mesenchymal cells via thePIK3R2-PI3K/Akt signalling pathway. PloS one.2013;8:e83294.
[27] Meng S, Cao JT, Zhang B, Zhou Q, Shen CX, Wang CQ. Downregulation ofmicroRNA-126in endothelial progenitor cells from diabetes patients, impairs theirfunctional properties, via target gene Spred-1. Journal of molecular and cellularcardiology.2012;53:64-72.
[28] Yan T, Liu Y, Cui K, Hu B, Wang F, Zou L. MicroRNA-126regulates EPCs function:implications for a role of miR-126in preeclampsia. Journal of cellular biochemistry.2013;114:2148-59.
[29] Mocharla P, Briand S, Giannotti G, Dorries C, Jakob P, Paneni F, et al. AngiomiR-126expression and secretion from circulating CD34(+) and CD14(+) PBMCs: role forproangiogenic effects and alterations in type2diabetics. Blood.2013;121:226-36.
[30] Fruman DA, Cantley LC, Carpenter CL. Structural organization and alternativesplicing of the murine phosphoinositide3-kinase p85alpha gene. Genomics.1996;37:113-21.
[31] Ueki K, Fruman DA, Yballe CM, Fasshauer M, Klein J, Asano T, et al. Positive andnegative roles of p85alpha and p85beta regulatory subunits of phosphoinositide3-kinase in insulin signaling. The Journal of biological chemistry.2003;278:48453-66.
[32] Geering B, Cutillas PR, Nock G, Gharbi SI, Vanhaesebroeck B. Class IAphosphoinositide3-kinases are obligate p85-p110heterodimers. Proceedings of theNational Academy of Sciences of the United States of America.2007;104:7809-14.
[33] Alcazar I, Cortes I, Zaballos A, Hernandez C, Fruman DA, Barber DF, et al. p85betaphosphoinositide3-kinase regulates CD28coreceptor function. Blood.2009;113:3198-208.
[34] Fruman DA, Snapper SB, Yballe CM, Davidson L, Yu JY, Alt FW, et al. Impaired Bcell development and proliferation in absence of phosphoinositide3-kinase p85alpha.Science.1999;283:393-7.
[1] T. Asahara，T. Murohara，A. Sullivan，M. Silver，R. van der Zee，T. Li，et al．Isolationof putative progenitor endothelial cells for angiogenesis[J]．Science，1997，275（5302）：964-967.
[2] Y. Matsuo，T. Imanishi，Y. Hayashi，Y. Tomobuchi，T. Kubo，T. Hano，et al．Theeffect of endothelial progenitor cells on the development of collateral formation inpatients with coronary artery disease[J]．Intern Med，2008，47（3）：127-134.
[3] N. Rodriguez-Losada， J. M. Garcia-Pinilla， M. F. Jimenez-Navarro， F. J.Gonzalez．Endothelial progenitor cells in cell-based therapy for cardiovasculardisease[J]．Cell Mol Biol (Noisy-le-grand)，2008，54（1）：11-23.
[4]陈书艳，许勤，王飞，周卿，颜雪芸，舒红，等．人外周血内皮祖细胞移植促裸鼠缺血组织血管新生的初步研究[J]．中国微循环，2005，9（1）：28-29.
[5] P. D. Lambiase，R. J. Edwards，P. Anthopoulos，S. Rahman，Y. G. Meng，C. A.Bucknall，et al．Circulating humoral factors and endothelial progenitor cells inpatients with differing coronary collateral support[J]．Circulation，2004，109（24）：2986-2992.
[6] V. Jodon de Villeroche，J. Avouac，A. Ponceau，B. Ruiz，A. Kahan，C. Boileau，et al．Enhanced late-outgrowth circulating endothelial progenitor cell levels inrheumatoid arthritis and correlation with disease activity[J]．Arthritis Res Ther，2010，12（1）：R27.
[7]陈超，杨述华，冯勇，陈东，禹虔，王小红．激素性股骨头坏死患者中两种内皮祖细胞功能的改变[J]．中华实验外科杂志，2012，29（1）：108-111.
[8] M. C. Yoder，L. E. Mead，D. Prater，T. R. Krier，K. N. Mroueh，F. Li，etal．Redefining endothelial progenitor cells via clonal analysis and hematopoieticstem/progenitor cell principals[J]．Blood，2007，109（5）：1801-1809.
[9] D. A. Ingram，L. E. Mead，H. Tanaka，V. Meade，A. Fenoglio，K. Mortell，etal．Identification of a novel hierarchy of endothelial progenitor cells using humanperipheral and umbilical cord blood[J]．Blood，2004，104（9）：2752-2760.
[10] H. Naito，H. Kidoya，S. Sakimoto，T. Wakabayashi，N. Takakura．Identificationand characterization of a resident vascular stem/progenitor cell population inpreexisting blood vessels[J]．EMBO J，2012，31（4）：842-855.
[11] M. Vasa，S. Fichtlscherer，K. Adler，A. Aicher，H. Martin，A. M. Zeiher，etal．Increase in circulating endothelial progenitor cells by statin therapy in patientswith stable coronary artery disease[J]．Circulation，2001，103（24）：2885-2890.
[12] C. P. Khoo，P. Pozzilli，M. R. Alison．Endothelial progenitor cells and their potentialtherapeutic applications[J]．Regen Med，2008，3（6）：863-876.
[13] H. Bompais，J. Chagraoui，X. Canron，M. Crisan，X. H. Liu，A. Anjo，et al．Humanendothelial cells derived from circulating progenitors display specific functionalproperties compared with mature vessel wall endothelial cells[J]．Blood，2004，103（7）：2577-2584.
[14] G. P. Fadini，C. Agostini，A. Avogaro．Autologous stem cell therapy for peripheralarterial disease meta-analysis and systematic review of theliterature[J]．Atherosclerosis，2010，209（1）：10-17.
[15] S. Dimmeler，J. Burchfield，A. M. Zeiher．Cell-based therapy of myocardialinfarction[J]．Arterioscler Thromb Vasc Biol，2008，28（2）：208-216.
[16] M. J. Lipinski，G. G. Biondi-Zoccai，A. Abbate，R. Khianey，I. Sheiban，J. Bartunek，et al．Impact of intracoronary cell therapy on left ventricular function in the setting ofacute myocardial infarction: a collaborative systematic review and meta-analysis ofcontrolled clinical trials[J]．J Am Coll Cardiol，2007，50（18）：1761-1767.
[17] T. H. Luo，Y. Wang，Z. M. Lu，H. Zhou，X. C. Xue，J. W. Bi，et al．The changeand effect of endothelial progenitor cells in pig with multiple organ dysfunctionsyndromes[J]．Crit Care，2009，13（4）：R118.
[18] D. A. Ingram，N. M. Caplice，M. C. Yoder．Unresolved questions, changingdefinitions, and novel paradigms for defining endothelial progenitor cells[J]．Blood，2005，106（5）：1525-1531.
[19]邱明，肖明朝，苟欣，杨钰兴．体外分离培养大鼠骨髓内皮祖细胞及其生物学特性鉴定[J]．中国组织工程研究与临床康复，2009，13（36）：7083-7086.
[20] F. Timmermans，J. Plum，M. C. Yoder，D. A. Ingram，B. Vandekerckhove，J.Case．Endothelial progenitor cells: identity defined?[J]．J Cell Mol Med，2009，13（1）：87-102.
[21]郭素芬，杨向红．bFGF和PDGF-BB对内皮祖细胞分化的内皮样细胞和平滑肌样细胞增殖和迁移的影响[J]．中国动脉硬化杂志，2011，19（9）：735-740.
[22]方叶青，谢秀梅，何晋，陈晓彬．冠心病患者外周血内皮祖细胞的数量和活性[J]．中国动脉硬化杂志，2008，16（5）：381-384.
[23]赵忠海，李洪秋，阿良，陈宾，汪少伯．自体富血小板血浆对人骨髓来源内皮祖细胞功能活性的影响[J]．中国组织工程研究与临床康复，2010，14（14）：2607-2611.
[24] R. J. Medina，C. L. O'Neill，M. Sweeney，J. Guduric-Fuchs，T. A. Gardiner，D. A.Simpson，et al．Molecular analysis of endothelial progenitor cell (EPC) subtypesreveals two distinct cell populations with different identities[J]． BMC MedGenomics，2010，3：18.
[25] G. Matsumura，S. Miyagawa-Tomita，T. Shin'oka，Y. Ikada，H. Kurosawa．Firstevidence that bone marrow cells contribute to the construction of tissue-engineeredvascular autografts in vivo[J]．Circulation，2003，108（14）：1729-1734.
[26] Y. Lin，D. J. Weisdorf，A. Solovey，R. P. Hebbel．Origins of circulating endothelialcells and endothelial outgrowth from blood[J]．J Clin Invest，2000，105（1）：71-77.
[27] M. Reyes， A. Dudek， B. Jahagirdar， L. Koodie， P. H. Marker， C. M.Verfaillie．Origin of endothelial progenitors in human postnatal bone marrow[J]．JClin Invest，2002，109（3）：337-346.
[28] M. Peichev，A. J. Naiyer，D. Pereira，Z. Zhu，W. J. Lane，M. Williams，etal．Expression of VEGFR-2and AC133by circulating human CD34(+) cells identifiesa population of functional endothelial precursors[J]．Blood，2000，95（3）：952-958.
[29] M. Navarro-Sobrino，M. Hernandez-Guillamon，I. Fernandez-Cadenas，M. Ribo，I. A. Romero，P. O. Couraud，et al．The angiogenic gene profile of circulatingendothelial progenitor cells from ischemic stroke patients[J]．Vasc Cell，2013，5（1）：3.
[30] T. Murayama，O. M. Tepper，M. Silver，H. Ma，D. W. Losordo，J. M. Isner，etal．Determination of bone marrow-derived endothelial progenitor cell significance inangiogenic growth factor-induced neovascularization in vivo[J]．Exp Hematol，2002，30（8）：967-972.
[31] C. H. Yoon，J. Hur，K. W. Park，J. H. Kim，C. S. Lee，I. Y. Oh，et al．Synergisticneovascularization by mixed transplantation of early endothelial progenitor cells andlate outgrowth endothelial cells: the role of angiogenic cytokines and matrixmetalloproteinases[J]．Circulation，2005，112（11）：1618-1627.
[32] A. Casamassimi，M. L. Balestrieri，C. Fiorito，C. Schiano，C. Maione，R. Rossiello，et al．Comparison between total endothelial progenitor cell isolation versus enrichedCd133+culture[J]．J Biochem，2007，141（4）：503-511.
[33] T. Takahashi，C. Kalka，H. Masuda，D. Chen，M. Silver，M. Kearney，etal．Ischemia-and cytokine-induced mobilization of bone marrow-derived endothelialprogenitor cells for neovascularization[J]．Nat Med，1999，5（4）：434-438.
[34] L. E. Mead，D. Prater，M. C. Yoder，D. A. Ingram．Isolation and characterizationof endothelial progenitor cells from human blood[J]．Curr Protoc Stem Cell Biol，2008，Chapter2：Unit2C1.
[35] O. Tura，E. M. Skinner，G. R. Barclay，K. Samuel，R. C. Gallagher，M. Brittan，et al．Late outgrowth endothelial cells resemble mature endothelial cells and are notderived from bone marrow[J]．Stem Cells，2013，31（2）：338-348.
[36] J. Chen，M. Song，S. Yu，P. Gao，Y. Yu，H. Wang，et al．Advanced glycationendproducts alter functions and promote apoptosis in endothelial progenitor cellsthrough receptor for advanced glycation endproducts mediate overpression of celloxidant stress[J]．Mol Cell Biochem，2010，335（1-2）：137-146.
[37] H. Uchimura，T. Marumo，O. Takase，H. Kawachi，F. Shimizu，M. Hayashi，etal．Intrarenal injection of bone marrow-derived angiogenic cells reduces endothelialinjury and mesangial cell activation in experimental glomerulonephritis[J]．J Am SocNephrol，2005，16（4）：997-1004.
[38] M. Ii． Bone marrow-derived endothelial progenitor cells: isolation andcharacterization for myocardial repair[J]．Methods Mol Biol，2010，660：9-27.
[1] Wakefield TW, Myers DD, Henke PK. Mechanisms of venous thrombosis andresolution. Arteriosclerosis, thrombosis, and vascular biology.2008;28:387-91.
[2] Hillen F, Griffioen AW. Tumour vascularization: sprouting angiogenesis and beyond.Cancer metastasis reviews.2007;26:489-502.
[3] Carmeliet P. Mechanisms of angiogenesis and arteriogenesis. Nature medicine.2000;6:389-95.
[4] Xu Q. The impact of progenitor cells in atherosclerosis. Nature clinical practiceCardiovascular medicine.2006;3:94-101.
[5] Rafii S, Lyden D. Therapeutic stem and progenitor cell transplantation for organvascularization and regeneration. Nature medicine.2003;9:702-12.
[6] Asahara T, Murohara T, Sullivan A, Silver M, van der Zee R, Li T, et al. Isolation ofputative progenitor endothelial cells for angiogenesis. Science.1997;275:964-7.
[7] Martin K, Stanchina M, Kouttab N, Harrington EO, Rounds S. Circulating endothelialcells and endothelial progenitor cells in obstructive sleep apnea. Lung.2008;186:145-50.
[8] Shintani S, Murohara T, Ikeda H, Ueno T, Honma T, Katoh A, et al. Mobilization ofendothelial progenitor cells in patients with acute myocardial infarction. Circulation.2001;103:2776-9.
[9] Kalka C, Masuda H, Takahashi T, Gordon R, Tepper O, Gravereaux E, et al. Vascularendothelial growth factor(165) gene transfer augments circulating endothelialprogenitor cells in human subjects. Circulation research.2000;86:1198-202.
[10] Junhui Z, Xingxiang W, Guosheng F, Yunpeng S, Furong Z, Junzhu C. Reducednumber and activity of circulating endothelial progenitor cells in patients withidiopathic pulmonary arterial hypertension. Respiratory medicine.2008;102:1073-9.
[11] Smadja DM, Bieche I, Helley D, Laurendeau I, Simonin G, Muller L, et al. IncreasedVEGFR2expression during human late endothelial progenitor cells expansionenhances in vitro angiogenesis with up-regulation of integrin alpha(6). Journal ofcellular and molecular medicine.2007;11:1149-61.
[12] Murohara T, Ikeda H, Duan J, Shintani S, Sasaki K, Eguchi H, et al. Transplanted cordblood-derived endothelial precursor cells augment postnatal neovascularization. TheJournal of clinical investigation.2000;105:1527-36.
[13] Kalka C, Tehrani H, Laudenberg B, Vale PR, Isner JM, Asahara T, et al. VEGF genetransfer mobilizes endothelial progenitor cells in patients with inoperable coronarydisease. The Annals of thoracic surgery.2000;70:829-34.
[14] Asahara T. Cell therapy and gene therapy using endothelial progenitor cells forvascular regeneration. Handbook of experimental pharmacology.2007:181-94.
[15] Kawamoto A, Gwon HC, Iwaguro H, Yamaguchi JI, Uchida S, Masuda H, et al.Therapeutic potential of ex vivo expanded endothelial progenitor cells for myocardialischemia. Circulation.2001;103:634-7.
[16] Lin Y, Weisdorf DJ, Solovey A, Hebbel RP. Origins of circulating endothelial cellsand endothelial outgrowth from blood. The Journal of clinical investigation.2000;105:71-7.
[17] Ott I, Keller U, Knoedler M, Gotze KS, Doss K, Fischer P, et al. Endothelial-like cellsexpanded from CD34+blood cells improve left ventricular function afterexperimental myocardial infarction. FASEB journal: official publication of theFederation of American Societies for Experimental Biology.2005;19:992-4.
[18] Zhang B, Pan X, Anderson TA. MicroRNA: a new player in stem cells. Journal ofcellular physiology.2006;209:266-9.
[19] Georgantas RW,3rd, Hildreth R, Morisot S, Alder J, Liu CG, Heimfeld S, et al.CD34+hematopoietic stem-progenitor cell microRNA expression and function: acircuit diagram of differentiation control. Proceedings of the National Academy ofSciences of the United States of America.2007;104:2750-5.
[20] Lee RC, Feinbaum RL, Ambros V. The C. elegans heterochronic gene lin-4encodessmall RNAs with antisense complementarity to lin-14. Cell.1993;75:843-54.
[21] Ambros V, Lee RC, Lavanway A, Williams PT, Jewell D. MicroRNAs and other tinyendogenous RNAs in C. elegans. Current biology: CB.2003;13:807-18.
[22] Pasquinelli AE, Reinhart BJ, Slack F, Martindale MQ, Kuroda MI, Maller B, et al.Conservation of the sequence and temporal expression of let-7heterochronicregulatory RNA. Nature.2000;408:86-9.
[23] Lau AO, Sacci JB, Jr., Azad AF. Detection of Plasmodium yoelii stage mRNA inBALB/c mice. The Journal of parasitology.2001;87:19-23.
[24] Lagos-Quintana M, Rauhut R, Yalcin A, Meyer J, Lendeckel W, Tuschl T.Identification of tissue-specific microRNAs from mouse. Current biology: CB.2002;12:735-9.
[25] Han J, Lee Y, Yeom KH, Kim YK, Jin H, Kim VN. The Drosha-DGCR8complex inprimary microRNA processing. Genes&development.2004;18:3016-27.
[26] Choudhuri S. Small noncoding RNAs: biogenesis, function, and emergingsignificance in toxicology. Journal of biochemical and molecular toxicology.2010;24:195-216.
[27] He JF, Luo YM, Wan XH, Jiang D. Biogenesis of MiRNA-195and its role inbiogenesis, the cell cycle, and apoptosis. Journal of biochemical and moleculartoxicology.2011;25:404-8.
[28] Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell.2004;116:281-97.
[29] Peters L, Meister G. Argonaute proteins: mediators of RNA silencing. Molecular cell.2007;26:611-23.
[30] Khvorova A, Reynolds A, Jayasena SD. Functional siRNAs and miRNAs exhibitstrand bias. Cell.2003;115:209-16.
[31] Schwarz DS, Hutvagner G, Du T, Xu Z, Aronin N, Zamore PD. Asymmetry in theassembly of the RNAi enzyme complex. Cell.2003;115:199-208.
[32] Liu J, Valencia-Sanchez MA, Hannon GJ, Parker R. MicroRNA-dependentlocalization of targeted mRNAs to mammalian P-bodies. Nature cell biology.2005;7:719-23.
[33] Brennecke J, Stark A, Russell RB, Cohen SM. Principles of microRNA-targetrecognition. PLoS biology.2005;3:e85.
[34] Pillai RS, Bhattacharyya SN, Filipowicz W. Repression of protein synthesis bymiRNAs: how many mechanisms? Trends in cell biology.2007;17:118-26.
[35] Zhang MX, Ou H, Shen YH, Wang J, Wang J, Coselli J, et al. Regulation ofendothelial nitric oxide synthase by small RNA. Proceedings of the NationalAcademy of Sciences of the United States of America.2005;102:16967-72.
[36] Valencia-Sanchez MA, Liu J, Hannon GJ, Parker R. Control of translation and mRNAdegradation by miRNAs and siRNAs. Genes&development.2006;20:515-24.
[37] Ruvkun G. Molecular biology. Glimpses of a tiny RNA world. Science.2001;294:797-9.
[38] Hunter MP, Ismail N, Zhang X, Aguda BD, Lee EJ, Yu L, et al. Detection ofmicroRNA expression in human peripheral blood microvesicles. PloS one.2008;3:e3694.
[39] Vatolin S, Navaratne K, Weil RJ. A novel method to detect functional microRNAtargets. Journal of molecular biology.2006;358:983-96.
[40] Kim SK, Nam JW, Rhee JK, Lee WJ, Zhang BT. miTarget: microRNA target geneprediction using a support vector machine. BMC bioinformatics.2006;7:411.
[41] Hatfield S, Ruohola-Baker H. microRNA and stem cell function. Cell and tissueresearch.2008;331:57-66.
[42] Matsubara H, Takeuchi T, Nishikawa E, Yanagisawa K, Hayashita Y, Ebi H, et al.Apoptosis induction by antisense oligonucleotides against miR-17-5p and miR-20a inlung cancers overexpressing miR-17-92. Oncogene.2007;26:6099-105.
[43] Carleton M, Cleary MA, Linsley PS. MicroRNAs and cell cycle regulation. Cell cycle.2007;6:2127-32.
[44] Poy MN, Eliasson L, Krutzfeldt J, Kuwajima S, Ma X, Macdonald PE, et al. Apancreatic islet-specific microRNA regulates insulin secretion. Nature.2004;432:226-30.
[45] Gauthier BR, Wollheim CB. MicroRNAs:'ribo-regulators' of glucose homeostasis.Nature medicine.2006;12:36-8.
[46] Yang WJ, Yang DD, Na S, Sandusky GE, Zhang Q, Zhao G. Dicer is required forembryonic angiogenesis during mouse development. The Journal of biologicalchemistry.2005;280:9330-5.
[47] Giraldez AJ, Cinalli RM, Glasner ME, Enright AJ, Thomson JM, Baskerville S, et al.MicroRNAs regulate brain morphogenesis in zebrafish. Science.2005;308:833-8.
[48] Kuehbacher A, Urbich C, Zeiher AM, Dimmeler S. Role of Dicer and Drosha forendothelial microRNA expression and angiogenesis. Circulation research.2007;101:59-68.
[49] Suarez Y, Fernandez-Hernando C, Pober JS, Sessa WC. Dicer dependent microRNAsregulate gene expression and functions in human endothelial cells. Circulationresearch.2007;100:1164-73.
[50] Ji R, Cheng Y, Yue J, Yang J, Liu X, Chen H, et al. MicroRNA expression signatureand antisense-mediated depletion reveal an essential role of MicroRNA in vascularneointimal lesion formation. Circulation research.2007;100:1579-88.
[51] Poliseno L, Tuccoli A, Mariani L, Evangelista M, Citti L, Woods K, et al. MicroRNAsmodulate the angiogenic properties of HUVECs. Blood.2006;108:3068-71.
[52] Chen Y, Gorski DH. Regulation of angiogenesis through a microRNA (miR-130a) thatdown-regulates antiangiogenic homeobox genes GAX and HOXA5. Blood.2008;111:1217-26.
[53] Pulkkinen K, Malm T, Turunen M, Koistinaho J, Yla-Herttuala S. Hypoxia inducesmicroRNA miR-210in vitro and in vivo ephrin-A3and neuronal pentraxin1arepotentially regulated by miR-210. FEBS letters.2008;582:2397-401.
[54] Fasanaro P, D'Alessandra Y, Di Stefano V, Melchionna R, Romani S, Pompilio G, et al.MicroRNA-210modulates endothelial cell response to hypoxia and inhibits thereceptor tyrosine kinase ligand Ephrin-A3. The Journal of biological chemistry.2008;283:15878-83.
[55] Lee DY, Deng Z, Wang CH, Yang BB. MicroRNA-378promotes cell survival, tumorgrowth, and angiogenesis by targeting SuFu and Fus-1expression. Proceedings of theNational Academy of Sciences of the United States of America.2007;104:20350-5.
[56] Mendell JT. miRiad roles for the miR-17-92cluster in development and disease. Cell.2008;133:217-22.
[57] Taguchi A, Yanagisawa K, Tanaka M, Cao K, Matsuyama Y, Goto H, et al.Identification of hypoxia-inducible factor-1alpha as a novel target for miR-17-92microRNA cluster. Cancer research.2008;68:5540-5.
[58] Minami Y, Satoh M, Maesawa C, Takahashi Y, Tabuchi T, Itoh T, et al. Effect ofatorvastatin on microRNA221/222expression in endothelial progenitor cellsobtained from patients with coronary artery disease. European journal of clinicalinvestigation.2009;39:359-67.
[59] Bonauer A, Carmona G, Iwasaki M, Mione M, Koyanagi M, Fischer A, et al.MicroRNA-92a controls angiogenesis and functional recovery of ischemic tissues inmice. Science.2009;324:1710-3.
[60] Wang CH, Lee DY, Deng Z, Jeyapalan Z, Lee SC, Kahai S, et al. MicroRNA miR-328regulates zonation morphogenesis by targeting CD44expression. PloS one.2008;3:e2420.
[61] Cimmino A, Calin GA, Fabbri M, Iorio MV, Ferracin M, Shimizu M, et al. miR-15and miR-16induce apoptosis by targeting BCL2. Proceedings of the NationalAcademy of Sciences of the United States of America.2005;102:13944-9.
[62] Carmeliet P. Angiogenesis in health and disease. Nature medicine.2003;9:653-60.
[63] Wienholds E, Kloosterman WP, Miska E, Alvarez-Saavedra E, Berezikov E, de BruijnE, et al. MicroRNA expression in zebrafish embryonic development. Science.2005;309:310-1.
[64] Fish JE, Santoro MM, Morton SU, Yu S, Yeh RF, Wythe JD, et al. miR-126regulatesangiogenic signaling and vascular integrity. Developmental cell.2008;15:272-84.
[65] Wang S, Aurora AB, Johnson BA, Qi X, McAnally J, Hill JA, et al. Theendothelial-specific microRNA miR-126governs vascular integrity and angiogenesis.Developmental cell.2008;15:261-71.
[66] van Solingen C, Seghers L, Bijkerk R, Duijs JM, Roeten MK, van Oeveren-RietdijkAM, et al. Antagomir-mediated silencing of endothelial cell specific microRNA-126impairs ischemia-induced angiogenesis. Journal of cellular and molecular medicine.2009;13:1577-85.
[67] Zernecke A, Bidzhekov K, Noels H, Shagdarsuren E, Gan L, Denecke B, et al.Delivery of microRNA-126by apoptotic bodies induces CXCL12-dependent vascularprotection. Science signaling.2009;2:ra81.
[68] Zhang Q, Kandic I, Kutryk MJ. Dysregulation of angiogenesis-related microRNAs inendothelial progenitor cells from patients with coronary artery disease. Biochemicaland biophysical research communications.2011;405:42-6.
[69] Zhang J, Zhang Z, Zhang DY, Zhu J, Zhang T, Wang C. microRNA126inhibits thetransition of endothelial progenitor cells to mesenchymal cells via thePIK3R2-PI3K/Akt signalling pathway. PloS one.2013;8:e83294.
[70] Meng S, Cao JT, Zhang B, Zhou Q, Shen CX, Wang CQ. Downregulation ofmicroRNA-126in endothelial progenitor cells from diabetes patients, impairs theirfunctional properties, via target gene Spred-1. Journal of molecular and cellularcardiology.2012;53:64-72.
[71] Yan T, Liu Y, Cui K, Hu B, Wang F, Zou L. MicroRNA-126regulates EPCs function:implications for a role of miR-126in preeclampsia. Journal of cellular biochemistry.2013;114:2148-59.
[72] Mocharla P, Briand S, Giannotti G, Dorries C, Jakob P, Paneni F, et al. AngiomiR-126expression and secretion from circulating CD34(+) and CD14(+) PBMCs: role forproangiogenic effects and alterations in type2diabetics. Blood.2013;121:226-36.
[73] Zeng Y, Cai X, Cullen BR. Use of RNA polymerase II to transcribe artificialmicroRNAs. Methods in enzymology.2005;392:371-80.
[74] Ebert MS, Neilson JR, Sharp PA. MicroRNA sponges: competitive inhibitors of smallRNAs in mammalian cells. Nature methods.2007;4:721-6.
[1] Cimmino A, Calin GA, Fabbri M, Iorio MV, Ferracin M, Shimizu M, et al. miR-15andmiR-16induce apoptosis by targeting BCL2. Proceedings of the National Academy ofSciences of the United States of America.2005;102:13944-9.
[2] Le Brigand K, Robbe-Sermesant K, Mari B, Barbry P. MiRonTop: mining microRNAstargets across large scale gene expression studies. Bioinformatics.2010;26:3131-2.
[3] Lu TP, Lee CY, Tsai MH, Chiu YC, Hsiao CK, Lai LC, et al. miRSystem: an integratedsystem for characterizing enriched functions and pathways of microRNA targets. PloSone.2012;7:e42390.
[4] Wang XJ, Reyes JL, Chua NH, Gaasterland T. Prediction and identification ofArabidopsis thaliana microRNAs and their mRNA targets. Genome biology.2004;5:R65.
[5] Sethupathy P, Megraw M, Hatzigeorgiou AG. A guide through present computationalapproaches for the identification of mammalian microRNA targets. Nature methods.2006;3:881-6.
[6] Kuhn DE, Martin MM, Feldman DS, Terry AV, Jr., Nuovo GJ, Elton TS. Experimentalvalidation of miRNA targets. Methods.2008;44:47-54.
[7] La Rocca G, Badin M, Shi B, Xu SQ, Deangelis T, Sepp-Lorenzinoi L, et al.Mechanism of growth inhibition by MicroRNA145: the role of the IGF-I receptorsignaling pathway. Journal of cellular physiology.2009;220:485-91.
[8] Wymann MP, Pirola L. Structure and function of phosphoinositide3-kinases.Biochimica et biophysica acta.1998;1436:127-50.
[9] Ward SG, Finan P. Isoform-specific phosphoinositide3-kinase inhibitors as therapeuticagents. Current opinion in pharmacology.2003;3:426-34.
[10] Chagpar RB, Links PH, Pastor MC, Furber LA, Hawrysh AD, Chamberlain MD, et al.Direct positive regulation of PTEN by the p85subunit of phosphatidylinositol3-kinase. Proceedings of the National Academy of Sciences of the United States ofAmerica.2010;107:5471-6.
[11] Fruman DA, Cantley LC, Carpenter CL. Structural organization and alternativesplicing of the murine phosphoinositide3-kinase p85alpha gene. Genomics.1996;37:113-21.
[12] Ueki K, Fruman DA, Brachmann SM, Tseng YH, Cantley LC, Kahn CR. Molecularbalance between the regulatory and catalytic subunits of phosphoinositide3-kinaseregulates cell signaling and survival. Molecular and cellular biology.2002;22:965-77.
[13] Geering B, Cutillas PR, Nock G, Gharbi SI, Vanhaesebroeck B. Class IAphosphoinositide3-kinases are obligate p85-p110heterodimers. Proceedings of theNational Academy of Sciences of the United States of America.2007;104:7809-14.
[14] Fruman DA, Snapper SB, Yballe CM, Davidson L, Yu JY, Alt FW, et al. Impaired Bcell development and proliferation in absence of phosphoinositide3-kinase p85alpha.Science.1999;283:393-7.
[15] Ueki K, Fruman DA, Yballe CM, Fasshauer M, Klein J, Asano T, et al. Positive andnegative roles of p85alpha and p85beta regulatory subunits of phosphoinositide3-kinase in insulin signaling. The Journal of biological chemistry.2003;278:48453-66.
[16] Li W, Wang H, Kuang CY, Zhu JK, Yu Y, Qin ZX, et al. An essential role for theId1/PI3K/Akt/NFkB/survivin signalling pathway in promoting the proliferation ofendothelial progenitor cells in vitro. Molecular and cellular biochemistry.2012;363:135-45.
[17] Yan T, Liu Y, Cui K, Hu B, Wang F, Zou L. MicroRNA-126regulates EPCs function:implications for a role of miR-126in preeclampsia. Journal of cellular biochemistry.2013;114:2148-59.
[18] Marson A, Levine SS, Cole MF, Frampton GM, Brambrink T, Johnstone S, et al.Connecting microRNA genes to the core transcriptional regulatory circuitry ofembryonic stem cells. Cell.2008;134:521-33.
[19] Wang S, Aurora AB, Johnson BA, Qi X, McAnally J, Hill JA, et al. Theendothelial-specific microRNA miR-126governs vascular integrity and angiogenesis.Developmental cell.2008;15:261-71.
[20] Harris TA, Yamakuchi M, Ferlito M, Mendell JT, Lowenstein CJ. MicroRNA-126regulates endothelial expression of vascular cell adhesion molecule1. Proceedings ofthe National Academy of Sciences of the United States of America.2008;105:1516-21.
[21] Liu B, Peng XC, Zheng XL, Wang J, Qin YW. MiR-126restoration down-regulateVEGF and inhibit the growth of lung cancer cell lines in vitro and in vivo. Lungcancer.2009;66:169-75.
[1] Naldini L, Blomer U, Gage FH, Trono D, Verma IM. Efficient transfer, integration, andsustained long-term expression of the transgene in adult rat brains injected with alentiviral vector. Proc Natl Acad Sci U S A.1996;93:11382-8.
[2] Lois C, Hong EJ, Pease S, Brown EJ, Baltimore D. Germline transmission andtissue-specific expression of transgenes delivered by lentiviral vectors. Science.2002;295:868-72.
[3] Pfeifer A, Ikawa M, Dayn Y, Verma IM. Transgenesis by lentiviral vectors: lack of genesilencing in mammalian embryonic stem cells and preimplantation embryos. ProcNatl Acad Sci U S A.2002;99:2140-5.
[4] Delenda C. Lentiviral vectors: optimization of packaging, transduction and geneexpression. J Gene Med.2004;6Suppl1:S125-38.
[5] Lever AM, Strappe PM, Zhao J. Lentiviral vectors. J Biomed Sci.2004;11:439-49.
[6] Chang LJ, Gay EE. The molecular genetics of lentiviral vectors--current and futureperspectives. Curr Gene Ther.2001;1:237-51.
[7] Cronin J, Zhang XY, Reiser J. Altering the tropism of lentiviral vectors throughpseudotyping. Curr Gene Ther.2005;5:387-98.
[8] Croyle MA, Callahan SM, Auricchio A, Schumer G, Linse KD, Wilson JM, et al.PEGylation of a vesicular stomatitis virus G pseudotyped lentivirus vector preventsinactivation in serum. J Virol.2004;78:912-21.
[9] Zufferey R, Donello JE, Trono D, Hope TJ. Woodchuck hepatitis virusposttranscriptional regulatory element enhances expression of transgenes delivered byretroviral vectors. J Virol.1999;73:2886-92.
[10] Iwakuma T, Cui Y, Chang LJ. Self-inactivating lentiviral vectors with U3and U5modifications. Virology.1999;261:120-32.
[11] Zufferey R, Dull T, Mandel RJ, Bukovsky A, Quiroz D, Naldini L, et al.Self-inactivating lentivirus vector for safe and efficient in vivo gene delivery. J Virol.1998;72:9873-80.
[12] Hematti P, Hong BK, Ferguson C, Adler R, Hanawa H, Sellers S, et al. Distinctgenomic integration of MLV and SIV vectors in primate hematopoietic stem andprogenitor cells. PLoS Biol.2004;2:e423.
[13] Loewen N, Leske DA, Cameron JD, Chen Y, Whitwam T, Simari RD, et al. Long-termretinal transgene expression with FIV versus adenoviral vectors. Mol Vis.2004;10:272-80.
[14] He Y, Zhang J, Donahue C, Falo LD, Jr. Skin-derived dendritic cells induce potentCD8(+) T cell immunity in recombinant lentivector-mediated genetic immunization.Immunity.2006;24:643-56.
[15] He Y, Falo LD. Induction of T cell immunity by cutaneous genetic immunization withrecombinant lentivector. Immunol Res.2006;36:101-17.
[16] Liu Y, Deisseroth A. Tumor vascular targeting therapy with viral vectors. Blood.2006;107:3027-33.
[1] Kahn SR, Shrier I, Julian JA, Ducruet T, Arsenault L, Miron MJ, et al. Determinantsand time course of the postthrombotic syndrome after acute deep venous thrombosis.Annals of internal medicine.2008;149:698-707.
[2] Modarai B, Burnand KG, Humphries J, Waltham M, Smith A. The role ofneovascularisation in the resolution of venous thrombus. Thrombosis and haemostasis.2005;93:801-9.
[3] Moldovan NI, Asahara T. Role of blood mononuclear cells in recanalization andvascularization of thrombi: past, present, and future. Trends in cardiovascularmedicine.2003;13:265-9.
[4] Waltham M, Burnand K, Fenske C, Modarai B, Humphries J, Smith A. Vascularendothelial growth factor naked DNA gene transfer enhances thrombus recanalizationand resolution. Journal of vascular surgery.2005;42:1183-9.
[5] Varma MR, Varga AJ, Knipp BS, Sukheepod P, Upchurch GR, Kunkel SL, et al.Neutropenia impairs venous thrombosis resolution in the rat. Journal of vascularsurgery.2003;38:1090-8.
[6] Humphries J, McGuinness CL, Smith A, Waltham M, Poston R, Burnand KG.Monocyte chemotactic protein-1(MCP-1) accelerates the organization and resolutionof venous thrombi. Journal of vascular surgery.1999;30:894-9.
[7] Henke PK, Pearce CG, Moaveni DM, Moore AJ, Lynch EM, Longo C, et al.Targeted deletion of CCR2impairs deep vein thombosis resolution in a mouse model.Journal of immunology.2006;177:3388-97.
[8] Myers DD, Jr., Henke PK, Bedard PW, Wrobleski SK, Kaila N, Shaw G, et al.Treatment with an oral small molecule inhibitor of P selectin (PSI-697) decreases veinwall injury in a rat stenosis model of venous thrombosis. Journal of vascular surgery.2006;44:625-32.
[9] Henke PK, Varma MR, Moaveni DK, Dewyer NA, Moore AJ, Lynch EM, et al.Fibrotic injury after experimental deep vein thrombosis is determined by themechanism of thrombogenesis. Thrombosis and haemostasis.2007;98:1045-55.
[10] Comerota AJ. Quality-of-life improvement using thrombolytic therapy for iliofemoraldeep venous thrombosis. Reviews in cardiovascular medicine.2002;3Suppl2:S61-7.
[11] Hockel M, Schlenger K, Doctrow S, Kissel T, Vaupel P. Therapeutic angiogenesis.Archives of surgery.1993;128:423-9.
[12] Modarai B, Burnand KG, Sawyer B, Smith A. Endothelial progenitor cells arerecruited into resolving venous thrombi. Circulation.2005;111:2645-53.
[13] M, Burnand K, Fenske C, Modarai B, Humphries J, Smith A. Vascular endothelialgrowth factor naked DNA gene transfer enhances thrombus recanalization andresolution. Journal of vascular surgery.2005;42:1183-9.
[14] Modarai B, Humphries J, Burnand KG, Gossage JA, Waltham M, Wadoodi A, et al.Adenovirus-mediated VEGF gene therapy enhances venous thrombus recanalizationand resolution. Arteriosclerosis, thrombosis, and vascular biology.2008;28:1753-9.
[15] Waltham M, Burnand KG, Collins M, McGuinness CL, Singh I, Smith A. Vascularendothelial growth factor enhances venous thrombus recanalisation and organisation.Thrombosis and haemostasis.2003;89:169-76.
[16] Corriere MA, Guzman RJ. Method of cell transplantation promoting the organizationof intraarterial thrombus. Perspectives in vascular surgery and endovascular therapy.2006;18:188-90.
[17] Moldovan NI. Tissular insemination of progenitor endothelial cells: the problem, anda suggested solution. Advances in experimental medicine and biology.2003;522:99-113.
[18] Wakefield TW, Myers DD, Henke PK. Mechanisms of venous thrombosis andresolution. Arteriosclerosis, thrombosis, and vascular biology.2008;28:387-91.
[19] Varma MR, Moaveni DM, Dewyer NA, Varga AJ, Deatrick KB, Kunkel SL, et al.Deep vein thrombosis resolution is not accelerated with increased neovascularization.Journal of vascular surgery.2004;40:536-42.
[20] Ali T, Humphries J, Burnand K, Sawyer B, Bursill C, Channon K, et al. Monocyterecruitment in venous thrombus resolution. Journal of vascular surgery.2006;43:601-8.
[21] Evans CE, Grover SP, Humphries J, Saha P, Patel AP, Patel AS, et al. Antiangiogenictherapy inhibits venous thrombus resolution. Arteriosclerosis, thrombosis, andvascular biology.2014;34:565-70.
[22] Zangari M, Fink LM, Elice F, Zhan F, Adcock DM, Tricot GJ. Thrombotic events inpatients with cancer receiving antiangiogenesis agents. Journal of clinical oncology:official journal of the American Society of Clinical Oncology.2009;27:4865-73.
[23] Drouet L.[Thromboembolic risk associated with use of angiogenesis inhibitors usedfor the treatment of cancers]. Pathologie-biologie.2008;56:195-204.
[24] Waltham M, Burnand KG, Collins M, Smith A. Vascular endothelial growth factorand basic fibroblast growth factor are found in resolving venous thrombi. Journal ofvascular surgery.2000;32:988-96.
[25] Evans CE, Humphries J, Mattock K, Waltham M, Wadoodi A, Saha P, et al. Hypoxiaand upregulation of hypoxia-inducible factor1{alpha} stimulate venous thrombusrecanalization. Arteriosclerosis, thrombosis, and vascular biology.2010;30:2443-51.
[26] Evans CE, Humphries J, Waltham M, Saha P, Mattock K, Patel A, et al. Upregulationof hypoxia-inducible factor1alpha in local vein wall is associated with enhancedvenous thrombus resolution. Thrombosis research.2011;128:346-51.
[27] Henke PK, Wakefield TW, Kadell AM, Linn MJ, Varma MR, Sarkar M, et al.Interleukin-8administration enhances venous thrombosis resolution in a rat model.The Journal of surgical research.2001;99:84-91.
[28] Asahara T, Murohara T, Sullivan A, Silver M, van der Zee R, Li T, et al. Isolation ofputative progenitor endothelial cells for angiogenesis. Science.1997;275:964-7.
[29] Rafii S, Lyden D. Therapeutic stem and progenitor cell transplantation for organvascularization and regeneration. Nature medicine.2003;9:702-12.
[30] Feigl W, Sinzinger H, Kern HG.[Thrombus organization and atherosclerosis]. DieMedizinische Welt.1978;29:1132-4.
[31] Stirling GA, Tsapogas MJ, Girolami PL. Organization of thrombi. The British journalof surgery.1966;53:232-5.
[32] Khan SS, Solomon MA, McCoy JP, Jr. Detection of circulating endothelial cells andendothelial progenitor cells by flow cytometry. Cytometry Part B, Clinical cytometry.2005;64:1-8.
[33] Dignat-George F, Sampol J, Lip G, Blann AD. Circulating endothelial cells: realitiesand promises in vascular disorders. Pathophysiology of haemostasis and thrombosis.2003;33:495-9.
[34] Singh I, Burnand KG, Collins M, Luttun A, Collen D, Boelhouwer B, et al. Failure ofthrombus to resolve in urokinase-type plasminogen activator gene-knockout mice:rescue by normal bone marrow-derived cells. Circulation.2003;107:869-75.
[35] de Boer HC, Verseyden C, Ulfman LH, Zwaginga JJ, Bot I, Biessen EA, et al. Fibrinand activated platelets cooperatively guide stem cells to a vascular injury and promotedifferentiation towards an endothelial cell phenotype. Arteriosclerosis, thrombosis,and vascular biology.2006;26:1653-9.
[36] Alessio AM, Beltrame MP, Nascimento MC, Vicente CP, de Godoy JA, Silva JC, etal. Circulating progenitor and mature endothelial cells in deep vein thrombosis.International journal of medical sciences.2013;10:1746-54.
[37] Shintani S, Murohara T, Ikeda H, Ueno T, Honma T, Katoh A, et al. Mobilization ofendothelial progenitor cells in patients with acute myocardial infarction. Circulation.2001;103:2776-9.
[38] Petrauskiene V, Falk M, Waernbaum I, Norberg M, Eriksson JW. The risk of venousthromboembolism is markedly elevated in patients with diabetes. Diabetologia.2005;48:1017-21.
[39] Bouzeghrane F, Zhang X, Gevry G, Raymond J. Deep vein thrombosis resolution isimpaired in diet-induced type2diabetic mice. Journal of vascular surgery.2008;48:1575-84.
[40] Li XQ, Meng QY, Wu HR. Effects of bone marrow-derived endothelial progenitorcell transplantation on vein microenvironment in a rat model of chronic thrombosis.Chinese medical journal.2007;120:2245-9.
[41] Santo SD, Tepper OM, von Ballmoos MW, Diehm N, Volzmann J, Baumgartner I, etal. Cell-based therapy facilitates venous thrombus resolution. Thrombosis andhaemostasis.2009;101:460-4.
[42] Meng Q, Li X, Yu X, Lei F, Jiang K, Li C. Transplantation of ex vivo expanded bonemarrow-derived endothelial progenitor cells enhances chronic venous thrombusresolution and recanalization. Clinical and applied thrombosis/hemostasis: officialjournal of the International Academy of Clinical and Applied Thrombosis/Hemostasis.2011;17:E196-201.
[43] Kalka C, Masuda H, Takahashi T, Kalka-Moll WM, Silver M, Kearney M, et al.Transplantation of ex vivo expanded endothelial progenitor cells for therapeuticneovascularization. Proceedings of the National Academy of Sciences of the UnitedStates of America.2000;97:3422-7.
[44] Urbich C, Dimmeler S. Endothelial progenitor cells: characterization and role invascular biology. Circulation research.2004;95:343-53.
[45] Meyer GP, Wollert KC, Lotz J, Steffens J, Lippolt P, Fichtner S, et al. Intracoronarybone marrow cell transfer after myocardial infarction: eighteen months' follow-updata from the randomized, controlled BOOST (BOne marrOw transfer to enhanceST-elevation infarct regeneration) trial. Circulation.2006;113:1287-94.
[46] De Palma M, Venneri MA, Roca C, Naldini L. Targeting exogenous genes to tumorangiogenesis by transplantation of genetically modified hematopoietic stem cells.Nature medicine.2003;9:789-95.
[47] Rehman J, Li J, Orschell CM, March KL. Peripheral blood "endothelial progenitorcells" are derived from monocyte/macrophages and secrete angiogenic growth factors.Circulation.2003;107:1164-9.
[48] Xia L, Fu GS, Yang JX, Zhang FR, Wang XX. Endothelial progenitor cells mayinhibit apoptosis of pulmonary microvascular endothelial cells: new insights into celltherapy for pulmonary arterial hypertension. Cytotherapy.2009;11:492-502.
[49] Urbich C, Aicher A, Heeschen C, Dernbach E, Hofmann WK, Zeiher AM, et al.Soluble factors released by endothelial progenitor cells promote migration ofendothelial cells and cardiac resident progenitor cells. Journal of molecular andcellular cardiology.2005;39:733-42.
[50] Yang Z, von Ballmoos MW, Faessler D, Voelzmann J, Ortmann J, Diehm N, et al.Paracrine factors secreted by endothelial progenitor cells prevent oxidativestress-induced apoptosis of mature endothelial cells. Atherosclerosis.2010;211:103-9.
[51] Zhang Q, Kandic I, Barfield JT, Kutryk MJ. Coculture with Late, but Not Early,Human Endothelial Progenitor Cells Up Regulates IL-1beta Expression in THP-1Monocytic Cells in a Paracrine Manner. Stem cells international.2013;2013:859643.
[52] Meng QY, Li XQ, Yu XB, Lei FR, Jiang K, Li CY. Transplantation ofVEGF165-gene-transfected endothelial progenitor cells in the treatment of chronicvenous thrombosis in rats. Chinese medical journal.2010;123:471-7.
[53] Kwaan HC, Samama MM. The significance of endothelial heterogeneity inthrombosis and hemostasis. Seminars in thrombosis and hemostasis.2010;36:286-300.
[54] Smadja DM, Laurendeau I, Avignon C, Vidaud M, Aiach M, Gaussem P. Theangiopoietin pathway is modulated by PAR-1activation on human endothelialprogenitor cells. Journal of thrombosis and haemostasis: JTH.2006;4:2051-8.
[55]杨震,陶军,涂昌,徐明国,王妍,王洁梅, et al.流体切应力调节内皮祖细胞分泌组织纤溶酶原激活物的研究.中华心血管病杂志.2005;33:840-2.
[56] Smadja DM, Bieche I, Silvestre JS, Germain S, Cornet A, Laurendeau I, et al. Bonemorphogenetic proteins2and4are selectively expressed by late outgrowthendothelial progenitor cells and promote neoangiogenesis. Arteriosclerosis,thrombosis, and vascular biology.2008;28:2137-43.
[57] Iwaguro H, Yamaguchi J, Kalka C, Murasawa S, Masuda H, Hayashi S, et al.Endothelial progenitor cell vascular endothelial growth factor gene transfer forvascular regeneration. Circulation.2002;105:732-8.
[58] Murasawa S, Llevadot J, Silver M, Isner JM, Losordo DW, Asahara T. Constitutivehuman telomerase reverse transcriptase expression enhances regenerative propertiesof endothelial progenitor cells. Circulation.2002;106:1133-9.
[59] Lei FR, Li XQ, Liu H, Zhu RD, Meng QY, Rong JJ. Rapamycin and3-methyladenineregulate apoptosis and autophagy in bone-derived endothelial progenitor cells.Chinese medical journal.2012;125:4076-82.
[60] Mitchell PS, Parkin RK, Kroh EM, Fritz BR, Wyman SK, Pogosova-Agadjanyan EL,et al. Circulating microRNAs as stable blood-based markers for cancer detection.Proceedings of the National Academy of Sciences of the United States of America.2008;105:10513-8.
[61] Latronico MV, Catalucci D, Condorelli G. Emerging role of microRNAs incardiovascular biology. Circulation research.2007;101:1225-36.
[62] Cordes KR, Srivastava D. MicroRNA regulation of cardiovascular development.Circulation research.2009;104:724-32.
[63] Suarez Y, Sessa WC. MicroRNAs as novel regulators of angiogenesis. Circulationresearch.2009;104:442-54.
[64] Bonauer A, Boon RA, Dimmeler S. Vascular microRNAs. Current drug targets.2010;11:943-9.
[65] van Solingen C, Seghers L, Bijkerk R, Duijs JM, Roeten MK, van Oeveren-RietdijkAM, et al. Antagomir-mediated silencing of endothelial cell specific microRNA-126impairs ischemia-induced angiogenesis. Journal of cellular and molecular medicine.2009;13:1577-85.
[66] Harris TA, Yamakuchi M, Kondo M, Oettgen P, Lowenstein CJ. Ets-1and Ets-2regulate the expression of microRNA-126in endothelial cells. Arteriosclerosis,thrombosis, and vascular biology.2010;30:1990-7.
[67] Zernecke A, Bidzhekov K, Noels H, Shagdarsuren E, Gan L, Denecke B, et al.Delivery of microRNA-126by apoptotic bodies induces CXCL12-dependent vascularprotection. Science signaling.2009;2:ra81.
[68] Feng R, Chen X, Yu Y, Su L, Yu B, Li J, et al. miR-126functions as a tumoursuppressor in human gastric cancer. Cancer letters.2010;298:50-63.
[69] Guo C, Sah JF, Beard L, Willson JK, Markowitz SD, Guda K. The noncoding RNA,miR-126, suppresses the growth of neoplastic cells by targeting phosphatidylinositol3-kinase signaling and is frequently lost in colon cancers. Genes, chromosomes&cancer.2008;47:939-46.
[70] Tavazoie SF, Alarcon C, Oskarsson T, Padua D, Wang Q, Bos PD, et al. Endogenoushuman microRNAs that suppress breast cancer metastasis. Nature.2008;451:147-52.
[71] Wang X, Tang S, Le SY, Lu R, Rader JS, Meyers C, et al. Aberrant expression ofoncogenic and tumor-suppressive microRNAs in cervical cancer is required for cancercell growth. PloS one.2008;3:e2557.
[72] Shen WF, Hu YL, Uttarwar L, Passegue E, Largman C. MicroRNA-126regulatesHOXA9by binding to the homeobox. Molecular and cellular biology.2008;28:4609-19.
[73] Zhang J, Du YY, Lin YF, Chen YT, Yang L, Wang HJ, et al. The cell growthsuppressor, mir-126, targets IRS-1. Biochemical and biophysical researchcommunications.2008;377:136-40.
[74] Zhang Q, Kandic I, Kutryk MJ. Dysregulation of angiogenesis-related microRNAs inendothelial progenitor cells from patients with coronary artery disease. Biochemicaland biophysical research communications.2011;405:42-6.
[75] Qiang L, Hong L, Ningfu W, Huaihong C, Jing W. Expression of miR-126andmiR-508-5p in endothelial progenitor cells is associated with the prognosis of chronicheart failure patients. International journal of cardiology.2013;168:2082-8.
[76] Meng S, Cao JT, Zhang B, Zhou Q, Shen CX, Wang CQ. Downregulation ofmicroRNA-126in endothelial progenitor cells from diabetes patients, impairs theirfunctional properties, via target gene Spred-1. Journal of molecular and cellularcardiology.2012;53:64-72.
[77] Mocharla P, Briand S, Giannotti G, Dorries C, Jakob P, Paneni F, et al.AngiomiR-126expression and secretion from circulating CD34(+) and CD14(+)PBMCs: role for proangiogenic effects and alterations in type2diabetics. Blood.2013;121:226-36.
[78] Ranghino A, Cantaluppi V, Grange C, Vitillo L, Fop F, Biancone L, et al. Endothelialprogenitor cell-derived microvesicles improve neovascularization in a murine modelof hindlimb ischemia. International journal of immunopathology and pharmacology.2012;25:75-85.
[79] Deregibus MC, Cantaluppi V, Calogero R, Lo Iacono M, Tetta C, Biancone L, et al.Endothelial progenitor cell derived microvesicles activate an angiogenic program inendothelial cells by a horizontal transfer of mRNA. Blood.2007;110:2440-8.
[80] Cantaluppi V, Biancone L, Figliolini F, Beltramo S, Medica D, Deregibus MC, et al.Microvesicles derived from endothelial progenitor cells enhance neoangiogenesis ofhuman pancreatic islets. Cell transplantation.2012;21:1305-20.
[81] Cantaluppi V, Gatti S, Medica D, Figliolini F, Bruno S, Deregibus MC, et al.Microvesicles derived from endothelial progenitor cells protect the kidney fromischemia-reperfusion injury by microRNA-dependent reprogramming of residentrenal cells. Kidney international.2012;82:412-27.
[82] Wang S, Aurora AB, Johnson BA, Qi X, McAnally J, Hill JA, et al. Theendothelial-specific microRNA miR-126governs vascular integrity and angiogenesis.Developmental cell.2008;15:261-71.
[83] Fish JE, Santoro MM, Morton SU, Yu S, Yeh RF, Wythe JD, et al. miR-126regulatesangiogenic signaling and vascular integrity. Developmental cell.2008;15:272-84.
[84] Horwich MD, Zamore PD. Design and delivery of antisense oligonucleotides to blockmicroRNA function in cultured Drosophila and human cells. Nature protocols.2008;3:1537-49.
[85] Mae M, Andaloussi SE, Lehto T, Langel U. Chemically modified cell-penetratingpeptides for the delivery of nucleic acids. Expert opinion on drug delivery.2009;6:1195-205.
[86] Fluiter K, Mook OR, Baas F. The therapeutic potential of LNA-modified siRNAs:reduction of off-target effects by chemical modification of the siRNA sequence.Methods in molecular biology.2009;487:189-203.
[87] Krutzfeldt J, Rajewsky N, Braich R, Rajeev KG, Tuschl T, Manoharan M, et al.Silencing of microRNAs in vivo with 'antagomirs'. Nature.2005;438:685-9.
1.Asahara T.Isolation of putative progenitor endothelial cells for angiogenesis.Science,1997,275(5302):964-967.
2.Shi Q.Evidence for circulating bone marrow-derived endothelial cells.Blood,1998,92(2):362-367.
3.Hur J.Characterization of two types of endothelial progenitor cells and their differentcontributions to neovasculogenesis.Arterioscler Thromb Vasc Biol,2004,24(2):288-293.
4.Yang J,Li M,Kamei N,et al.CD34+cells represent highly functional endothelual cells inmurine bone marrow.PLoS ONE,2011,6(5):e20219-20219.
5.Rae PC,Kelly RDW,Egginton S,et al.Angiogenic potential of endothelial progenitor cellsand embryonic stem cells.Vascular cell,2011,3(11):1-15.
6.Wen Y,Meng L,et al.Direct autologous bone marrow-derived stem cell transplantation forischemic heart disease: a meta-analysis.Expert Opin Biol Ther,2011,11(5):559-567.
7.Peichev M,Naiyer A J,Pereira D,et al.Expression of VEGFR-2and AC133by circulatinghuman CD34+cells identifies a population of functional endothelialprecursors.Blood.2000,95(3),952-958.
8.Navarro-Sobrino M,Hernández-Guillamon M,Fernandez-Cadenas I,et al.The angiogenicgene profile of circulating endothelial progenitor cells from ischemic strokepatients.Vascular cell,2013,5(1):1-7.
9.Murayama T,Tepper OM,Silver M,et al.Determination of bone marrow-derivedendothelial progenitor cell significance in angiogenic growth factor-inducedneovascularization in vivo.Exp Hematol,2002,30(8):967-972．
10.Yoon CH,Hur J,Park KW,et al.Synergistic neovascularization by mixed transplantationof early endothelial progenitor cells and late outgrowth endothelial Cells:the role ofangiogenic cytokines and matrix metalloproteinases.Circulation,2005,112(11):1618-1627．
11.Casamassimi A,Balestrieri M L,Fiorit o C,et al.Comparison between total endothelialprogenitor cell isolation versus enriched CD133+culture.JBiochem,2007,141(4):503-511.
12.Takahashi T,Kalka C,Masuda H,et al.Ischemia-and cytokine-induced mobilization ofbone marrow-derived endothelial progenitor cells for neovascularization.Naturemedicine.1999,5(4):434-438.
13.Mead LE, Prater D, Yoder MC, et al. Isolation and characterization of endothelialprogenitor cells from human blood.CurrProtoc Stem Cell Biol.2008;Chapter2:Unit2C.1.
14.Tura O, Skinner EM, Barclay GR, et al. Late outgrowth endothelial cells resemblemature endothelial cells and are not derived from bone marrow. Stem Cells.2013;31(2):338-348.
15.Chen J, Song M, Yu S, et at. Advanced glycation endproducts alter functions andpromote apoptos in endothelial progenitor cells through receptor for advancedglycation endproducts mediate over pression of cell oxidant stress. Mol Cell Biochem.2010;335(1-2):137-146.
16.Hideki U,Takeshi M,Osamu T,et al.Intrarenal injection of bone marrow-derivedangiogenic cells reduces endothelial injury and mesangial cell activation inexperimental glomerulonephritis.J Am Soc Nephrol,2005,16(4):997-1004.
17.li M.Bone Marrow-Derived Endothelial Progenitor Cells:Isolation and Characterizationfor Myocardial Repair.Methods Mol Biol.2010;660:9-27.
18.Jiang H，Liang C，Liu x，et a1.Palmitic acid promotes endothelialprogenitor cellsapoptosis via p38and JNK mitogen-activated protein kinase pathways.Atherosclerosis,2010,210(1):71．
19.Tura O,Crawford J,Barclay GR,et a1.Granuloeyte colony stimulating factor(G-CSF)depresses angiogenesis in vivo and in vitro：implications for sourcing cells forvascular regeneration therapy.J Thromb Haemost,2010,8(7):1614．
20.Malan D,Wenzel D,Sehmidt A,et a1.Endothelial beta1integrinsregulate sprouting andnetwork formation during vascular development.Development,2010,137(6):993．
21.Schluterman MK,Chapman SL,Korpanty G,et a1.Loss of fibulin5binding to beta1integrins inhibits tumor growth by increasing the level of ROS.Dis Model Mech,2010,3(5-6):333．
22.Kalka C,Masuda H,Takahashi T,et al.Transplantation of ex vivo expanded endothelialprogenitor cells for therapeutic neovascularization.Proc Natl Acad SciUSA,2000,97(7):3422-3427．
23.杨志明,边云飞,萧传实.血管内皮祖细胞在血管新生疗法的应用.中华老年心脑血管病杂志,2002,14(6):421-422．
24.Schatteman GC,Dunnwald M,Jiao C.Biology of bone marrow-derived endothelial cellprecursors.Am J Physiol Heart Circ Physiol,2007,292(1):11-18.
25.Timmermans F,Plum J,Yoder M C,et al.Endothelial progen it or cells:identity defined.JCell Mol Med,2009,13(1):87-102.
26.Jeon O,Song S J,Bhang S H,et al.Additive effect of endothelial progenit or cellmobilization and bone marrow mononuclear cell transplantation on angiogenesis inmouse ischemic limbs.J Biomed Sci,2007,14(3):323-330.
26.Rotmans J I,Heyliger s J M,Verhagen H J,et al.In vivo cell seeding w ith ant-i CD34antibodies successfully accelerates endothelialization but stimulates intimalhyperplasia in porcine arteriovenous expanded polytetrafluoroethylene grafts.Circulation,2005,112(1):12-18.
27.Rubio D,Garcia-Castro J,Martin M C,et al.Spontaneous human adult stem celltransformation.Cancer Res,2005,65(8):3035-3039.
28.Dobaczewski M,Bujak M,Zymek P,et al.Extracellular matrix remodeling incanine andmouse myocardial infarcts.Cell Tissue Res,2006,324(3):475-488．
29.Wei J,Blum S,Unger M,et al.Embryonic endothelial progenitor cells armed with asuicide gene target hypoxic lung metastases after intravenous delivery.Cancer Cell,2004,5(5):477-488．
30.Jiang Y,Jahagirdar BN,Reinhardt RL,et al.Pluripotency of mesenchymal stem cellsderived from adult marrow.Nature,2002,418(6893):41-49．
31.Zarnore PD,Haley B.The big world of smallRNAs.Science,2005,309:1519-1524.
32.Hutvagner G,Zamore PD.A microRNA in a multiple-turnover RNAi enzyme complex.Science,2002,297(5589):2056-2060.
33.Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell,2004,116(2):281-297.
34.Ambros V,Lee ＲC.Identification of microＲNAs and other tiny noncodingＲNAs bycDNA cloning.Methods Mol Biol,2004,265:131-158.
35.Pfeffer S,Zavolan M,Grasser F A,et al.Identification of virus-encoded micro ＲNAs.Science,2004,304(5671):734-736.
36.Chen P Y,Manninga H,Slanchev K,et al.The developmental mi Ｒ NA profiles ofzebrafish as determined by small ＲNA cloning.Genes Dev,2005,19(11):1288-1293.
37.Maniataki E,De Planell Saguer M D A,Mourelatos Z.Immunoprecipitation of microＲNAs and directional cloning of microＲNAs.Methods Mol Biol,2005,309:283-294．
38.Fu H,TieY,XuC,et al.Identification of human fetal liver miRNAs by a hovelmethod.Cancer,2005,579(17):3849-3854.
39.Pfeffer S,Sewer A,Lagos Quintana M.Identification of microＲNAs of the herpesvirusfamily.Nat Meth,2005,2(4):269-276.
40.Beuvink I,Kolb F A,Budach W,et al.A novel microarray approach reveals new tssuespecific signatures of known and predicted mammalian microＲNAs.Nucleic AcidsＲes,2007,35(7): e52.
41.Lim L P,Lau N C,Weinstein E G,et al.The microRNAs of Caenorhabditis elegans. GenesDev,2003,17:991-1008.
42.Várallyay E,Burgyán J,Havelda Z.Detection of microＲNAs by Northern blot analysesusing LNA probes.Methods,2007,43(2):140-145．
43.Várallyay E,Burgyán J,Havelda Z.MicroＲNA detection by northern blotting usinglocked nucleic acid probes.Nat Protoc,2008,3(2):190-196．
44.Liu CG,Calin GA,Meloon B,et al.An oligonucleotide microchip for genome-widemicroRNA profiling in human and mouse tissues.Proc Natl Acad Sci,2004,101(26):9740-9744.
45.Castoldi M,Schmidt S,Benes V,et al.miChip: an array-based method for microＲNAexpression profiling using locked nucleic acid capture probes.NatProtoc,2008,3(2):321-329．
46.Vorwerk S,Ganter K,Cheng Y,et al.Microfluidic-based enzymatic on-chip labeling ofmiRNAs.New Biotechnology2008,25(2-3):142-149.
47.欧阳松应,杨冬,欧阳红生,等.实时荧光定量PC Ｒ技术及其应用.生命的化学,2004,24(1):74-76．
48.Kersten S,Mandard S,Tan N S,et al.Characterizati on of the fasting-induced adiposefactor FIAF,a novel peroxisome proliferators-activated receptor target gene.J BiolChem,2000,275(37):28488-28493.
49.Mandard S,Zandbergen F,Van Straten E,et al.The fasting-induced adipose factor/angiopoietin-like protein4is physically associated with lipoproteins and governsplasma lipid levels and adiposity.J Biol Chem,2006,281(2):934-944.
50.Morin Ｒ D,O'Connor M D,Griffith M,et al.Application of massively parallelsequencing to micro Ｒ NA profiling and discovery in human embryonic stemcells.Genome Ｒes,2008,18(4):610-621．
51.Kuchenbauer F,Mah SM,Heuser M,et al.Comprehensive analysis of mammalianmiRNA*species and their role in myeloid cells.Blood,2011,118(12):3350–3358.
52.Dupuis DE,Maas S.MiRNA editing.Methods Mol Biol,2010,667:267–279.
53.Fernandez-Valverde SL,Taft RJ,Mattick JS.Dynamic isomiR regulation in Drosophiladevelopment.RNAna,2010,16(10):1881–1888.
54.Thomson DW,Bracken CP,Goodall GJ.Experimental strategies for microRNA targetidentification.Nucleic AcidsRes,2011,39(16):6845–6853.
55.Kozomara A,Griffiths-Jones S.miRBase:integrating microRNA annotation anddeep-sequencing data.Nucleic Acids Res,2011,39(Database issue):152–157.
56.Zhang B,Pan X,Anderson TA.MicroRNA：a new player in stem cells.J Cell Physiol,2006,209(2):1266-269．
57.Georgantas RW,Hildreth R,Morisot S,et al.CD34+hematopoietic stem-progenitor cellmicroRNA expression and function:a circuit diagram of differentiation control.ProcNatl Acad Sci USA,2007,104(8):2750-2755.
58.Chen CZ,Li L,Lodish HF,et a1.MicroRNAs modulate hematopoietic lineagedifferentiation,Science,2004,303(5654):83-86.
