摘要
目的：在体外分离、诱导培养和鉴定大鼠骨髓来源的EPCs，5-BrdU进行标记。
方法：采用Percoll密度梯度离心法和差速贴壁培养法，分离并抽取SD大鼠股骨和胫骨骨髓单个核细胞(MNCs)，在给予碱性成纤维细胞生长因子(Basic fibroblastgrowth factor，bFGF)等生长因子的条件下诱导培养14天。通过普通光学显微镜观察细胞的形态学特征。免疫细胞化学染色以鉴定其表面抗原表达情况。用双荧光染色检测Dil标记的乙酰化低密度脂蛋白(Acetyl low density lipoprotein，acLDL)和FITC标记的荆豆凝集素l(Ulex europaeus agglutinin，UEA-1)进行鉴定。分离培养的EPCs用不同浓度的5-溴脱氧尿嘧啶（5-Brdu）进行标记，MTT观察5-Brdu对其增殖活性的影响。5-BrdU标记EPCs备用。
结果：贴壁细胞起初呈铺路石样，随后呈现梭形样排列生长。CD34、CDl33、Flk-1(VEGFR)和vWF(Ⅷ因子)在贴壁细胞中呈不同时段的阳性表达。EPCs摄取Dil-acLDL，且结合FITC-UEA-1。5-BrdU可以标记EPCs，且不会影响细胞增殖活性。
结论：采用Percoll密度梯度离心法和差速贴壁结合法，同时给予VEGF、bFGF及EGF诱导，可以获得较高纯度的EPCs，且该细胞具备了成熟的ECs的部分功能学特性，5-Brdu标记EPCs可行。
目的：通过化学偶联法制备抗肌凝蛋白轻链/CD34双特异性抗体（BsAb），并装配在内皮祖细胞（EPCs）表面。
方法：抽取、分离和培养自体EPCs。通过化学偶联法构建BsAb以及FITC标记的BsAb。分别将10ng、25ng、50ng和100ng的FITC标记的BsAb与1×10~6个EPCs混合，在荧光显微镜及流式细胞仪下观察抗体与细胞的结合情况，以筛选出抗体装配的最佳浓度。使用上述实验所筛选出的条件，装配BsAb和EPCs备用。
结果：通过化学偶联法成功制备抗肌凝蛋白轻链/CD34双特异性抗体（BsAb），并能够装配在内皮祖细胞（EPCs）表面。BsAb在EPCs表面装配的最佳浓度为50ng/106细胞。结论：BsAb构建成功，并在EPCs表面成功装配BsAb。
目的：本部分我们探讨了超声破坏微泡对促进BsAb标记的EPCs向缺血心肌定向移植的可行性和治疗效果。
方法：使用前两部分中所制备的BsAb标记的EPCs。选取42只6周龄健康的SD大鼠制备大鼠急性心肌梗死模型，划分为①正常组（Normal），②心梗组（MI），③单纯双抗组（BsAb），④单纯超声+微泡组（US+MB），⑤单纯细胞组（EPCs）；⑥细胞+抗体组（EPCs+BsAb），⑦细胞+超声微泡+抗体组（EPCs+BsAb+US+MB）。建模8小时后，给予超声微泡和细胞移植处理，最后一组经尾静脉注射1ml经BsAb装配的EPCs，并注射脂质超声微泡溶液1ml，超声波辐照大鼠心前区。移植30天后，取大鼠心肌组织，切片进行免疫组织化学染色5-BrdU以检测EPCs的迁移和分布情况。利用超声心动图检测各组大鼠心功能变化情况，计算左心室射血分数(LVEF)及左心室短轴缩短率(FS)等以评价心功能。采用免疫组织化学法检测大鼠在细胞移植后30天后心脏中Ⅷ因子的表达情况。利用Real time PCR和Westernblot检测细胞移植后各组大鼠VEGF及SDF-1基因的mRNA表达和蛋白表达水平。
结果： EPCs+BsAb+US+MB组大鼠心肌5-BrdU免疫染色阳性细胞明显高于MI组，P<0.01。超声心电图检测大鼠左心室射血分数(LVEF)及左心室短轴缩短率(FS)等，EPCs+BsAb+US+MB组左心室收缩功能明显优于MI组，基本恢复至Normal组水平。Ⅷ因子免疫组化染色显示，EPCs+BsAb+US+MB组毛细血管数较其他各组多。RT-PCR和Western blot检测VEGF及SDF-1的表达水平，EPCs+BsAb+US+MB组VEGF和SDF-1的mRNA水平和蛋白表达水平均高于MI组，P<0.01，与Normal组没有显著差异。结论：BsAb与超声破坏微泡技术联合使用，可以促进BsAb标记的EPCs归巢至大鼠缺血心肌，促进血管新生，为干细胞移植治疗心肌梗死提供了新的思路。
目的：探讨FTO基因多态性与癌症风险。
方法：收集Pubmed和Embase数据库的文献。使用Meta分析，计算固定效应模型95%置信区间(CI)的合并比值比(OR)。13个研究中心的16277名患者以及31153名对照人员被纳入研究。
结果：除了胰腺癌外(OR=1.10,95%CI1.03–1.19)，FTO多态性与癌症风险增加相关性无显著性差异。(OR=1.01,95%CI0.98–1.04)。研究中没有文献偏差(Begg'stest: P=0.760; Egger’s test: P=0.553)。
结论：Meta分析表明尽管FTO基因多态性和胰腺癌有微弱关联，但是FTO基因多态性和癌症风险增加没有明显关联。但是这个结论仍然需要谨慎，因为这些研究中都没有把BMI/肥胖考虑进去。
Objective: To extract, culture and identify rat bone marrow-derived EPCs in vitro,labeled cells with5-BrdU.
Methods: The SD rat femur and tibia bone marrow mononuclear cells (MNCs) wereisolated and extracted by Percoll density gradient centrifugation and speed differenceadhesion culture. MNCs were cultured with administration of basic fibroblast growthfactor (bFGF), vascular endothelialcell growth factor (VEGF) and epidermal growthfactor (EGF) for14days. The morphological features were further observed withoptical microscope. Immunocytochemistry was used to identify their surface antigenexpression. Double fluorescence staining was used to detect Di-labeled Acetyl lowdensity lipoprotein (Di-acLDL) and FITC-labeled Ulex europaeus agglutinin (UEA-1).Identified EPCs were incubated with5-BrdU. EPCs were incubated with5-BrdU atdifferent concentrations, cells proliferation was assayed by MTT. EPCs Labeled with5-BrdU were for subsequent use.
Results: Adherent cells grew showing an initial cobblestone, and then a spindle-like arrangement style. CD34, CDl33, Flk-1(VEGFR) and vWF (VIII of factor) werepositively expressed at different stages in the adherent cells. The EPCs uptookDi-acLDL and bound FITC-UEA-1.5-BrdU can be used to lable EPCs，did not affectcells proliferation.
Conclusions: Percoll density gradient centrifugation and speed difference cultivationcombined withVEGF, bFGF, and EGF could obtain highly purified EPCs, and thesecells showed some of the features of mature ECs.5-BrdU can be used to lable EPCs.
Objective: To prepare anti-myosin light chain/CD34bispecific antibody (BsAb) bychemical heteroconjugation, and to assemble them on the surface of EPCs.
Methods: EPCs were extracted, isolated and cultured as described above. BsAb andFITC-BsAb were generated by chemical heteroconjugation. EPCs were mixed withFITC-BsAb at10ng,20ng,50ng and100ng, the binding activity was detected byfluorescence and flow cytomety to screen out the optimal concentration of the antibody.EPCs were assembled with these BsAbs for further purpose.
Results: BsAb was successfully generated and assembled on the surface of EPCs. Themost optimal concentration was50ng/106cells.
Conclusions: BsAb was successfully generated and further assembled on the surfaceof EPCs.
Objective: To study the feasibility of ultrasound microbubble destruction promotingBsAb-labeled EPCs to transplant to the ischemic myocardium.
Methods: BsAb-labeled EPCs were obtained as previously discribed.426-week,healthy SD rats were used as acute myocardial infarction model. They were divided asfollowing:1. Normal;2. MI;3. Antibody;4. US and MB;5. EPCs;6EPCs andantibody;7. EPCs, US, MB and antibody. Rats were treated with ultrasound andtransplanation8hours after modeling. The last group was injected1ml BsAb-labeled ofEPCs, and was further injected with lipid ultrasound microbubble solution1ml. Theprecordium of the rats were irradiation. After30days, the myocardial tissues werestained with5-BrdU to detect the migration and distribution of EPCs.Echocardiography was used to detect the heart function. The function were detected byleft ventricular ejection fraction (LVEF) and left ventricular fractional shortening (FS).VIIIwas detected by immunochemistry after30days. Real time PCR and western blotwere equipped to detect the expression of VEGF and SDF-1.
Results: Group7(EPCs, UM and antibody) was stained with more5-BrdUsignificantly. Group7showed high LVEF and FS compared with MI groups. Group7had more positive capillaries. The mRNA and protein level of VEGF and SDF-1ingroup7rats increased compared with MI groups through real time PCR and Westernblot. There was no difference between group7and Normal group of the mRNA andprotein level of VEGF and SDF-1through real time PCR and Western blot.
Conclusions: Combined with UM, BsAb could promote the target-directed migrationof EPCs and capillaries genesis. The work provided new strategy for the treatment of myocardial infarction.
Objective: to clarify the association between FTO polymorphism and cancer risk.
Methods: Published literature from PubMed and Embase databases had been retrieved.Pooled odds ratio (OR) with95%confidence interval (CI) was calculated byfixed-effects model.13studies involving16,277cases and31,153controls had beenidentified.
Results: FTO polymorphism was not significantly associated with the increased riskof cancer (OR=1.01,95%CI0.98–1.04), except that a statistically significantassociation with pancreatic cancer (OR=1.10,95%CI1.03–1.19). No publication biaswas detected (Begg's test: P=0.760; Egger’s test: P=0.553).
Conclusions: Our meta-analysis implicated that no association between FTOpolymorphism and the increased risk of cancer existed, although it was marginallyassociated with pancreatic cancer. However, the conclusion should be made withcaution for most included studies had not take BMI/obesity into account.
引文
[1] Barile L, Chimenti I, Gaetani R, et al.Cardiac stem cells: isolation, expansion andexperimental use for myocardial regeneration [J]. Nat Clin Pract Cardiovasc Med.2007,4Suppl1:S9-S14.
[2]Memon I A, Sawa Y, Miyagawa S, et al.Combined autologous cellularcardiomyoplasty with skeletal myoblasts and bone marrow cells in canine hearts forischemic cardiomyopathy [J]. J Thorac Cardiovasc Surg.2005,130(3):646-653.
[3]Segers V F, Van Riet I, Andries L J, et al.Mesenchymal stem cell adhesion tocardiac microvascular endothelium: activators and mechanisms [J]. Am J PhysiolHeart Circ Physiol.2006,290(4):H1370-H1377.
[4]Kawamoto A, Gwon H C, Iwaguro H, et al.Therapeutic potential of ex vivoexpanded endothelial progenitor cells for myocardial ischemia [J]. Circulation.2001,103(5):634-637.
[5]Li Y P, Paczesny S, Lauret E, et al.Human mesenchymal stem cells license adultCD34+hemopoietic progenitor cells to differentiate into regulatory dendritic cellsthrough activation of the Notch pathway [J]. J Immunol.2008,180(3):1598-1608.
[6]Zen K, Okigaki M, Hosokawa Y, et al.Myocardium-targeted delivery of endothelialprogenitor cells by ultrasound-mediated microbubble destruction improves cardiacfunction via an angiogenic response [J]. J Mol Cell Cardiol.2006,40(6):799-809.
[7]Dong F, Ha X Q.Effect of endothelial progenitor cells in neovascularization andtheir application in tumor therapy [J]. Chin Med J (Engl).2010,123(17):2454-2460.
[8]D' A I, Nargi E, Mastrangelo F, et al.Vascular endothelial growth factor enhancesin vitro proliferation and osteogenic differentiation of human dental pulp stem cells[J]. J Biol Regul Homeost Agents.2011,25(1):57-69.
[9]Liang F, Wang Y F, Nan X, et al.In vitro differentiation of human bonemarrow-derived mesenchymal stem cells into blood vessel endothelial cells [J].Zhongguo Yi Xue Ke Xue Yuan Xue Bao.2005,27(6):665-669.
[10]Asahara T, Murohara T, Sullivan A, et al.Isolation of putative progenitorendothelial cells for angiogenesis [J]. Science.1997,275(5302):964-967.
[11]Kuliczkowski W, Derzhko R, Prajs I, et al.Endothelial progenitor cells and leftventricle function in patients with acute myocardial infarction: potential therapeuticconsidertions [J]. Am J Ther.2012,19(1):44-50.
[12]Zen K, Okigaki M, Hosokawa Y, et al.Myocardium-targeted delivery of endothelialprogenitor cells by ultrasound-mediated microbubble destruction improves cardiacfunction via an angiogenic response [J]. J Mol Cell Cardiol.2006,40(6):799-809.
[13]Hu C H, Li Z M, Du ZM, et al.Expanded human cord blood-derived endothelialprogenitor cells salvage infarcted myocardium in rats with acute myocardialinfarction [J]. Clin Exp Pharmacol Physiol.2010,37(5-6):551-556.
[14]Stein A, Knodler M, Makowski M, et al.Local erythropoietin and endothelialprogenitor cells improve regional cardiac function in acute myocardial infarction [J].BMC Cardiovasc Disord.2010,10:43.
[15]Scheubel R J, Holtz J, Friedrich I, et al.Paracrine effects of CD34progenitor cellson angiogenic endothelial sprouting [J]. Int J Cardiol.2010,139(2):134-141.
[16]Wang Z, Liao Y, Dong J, et al.Clinical significance and pathogenic role ofanti-cardiac myosin autoantibody in dilated cardiomyopathy [J]. Chin Med J(Engl).2003,116(4):499-502.
[17]Zhao T C, Tseng A, Yano N, et al.Targeting human CD34+hematopoietic stemcells with anti-CD45x anti-myosin light-chain bispecific antibody preservescardiac function in myocardial infarction [J]. J Appl Physiol.2008,104(6):1793-1800.
[18]邓玮.BMSC在微泡及BiAb辅助下修复心肌纤维化的实验研究及PⅢNP对冠心病合并代谢综合征患者的预测价值[D].重庆医科大学,2011.
[19]Taniyama Y, Tachibana K, Hiraoka K, et al.Development of safe and efficientnovel nonviral gene transfer using ultrasound: enhancement of transfectionefficiency of naked plasmid DNA in skeletal muscle [J]. Gene Ther.2002,9(6):372-380.
[20]李晓东.超声波空化效应的生物学机制[J].临床超声医学杂志.2004,12(6):40-41.
[21]李巧.超声微泡联合Ad-EGFP/HIF-1α介导EPCs移植治疗大鼠心肌梗死的实验研究[D].重庆医科大学,2010.
[22]李兴升，王志刚，李雪霖等,超声破坏微泡造影剂促进心肌梗死大鼠血管新生的实验研究[J].中华超声影像学杂志.2007,16(21):1071-1073.
[23]Yoshida J, Ohmori K, Takeuchi H, et al.Treatment of ischemic limbs based on localrecruitment of vascular endothelial growth factor-producing inflammatory cellswith ultrasonic microbubble destruction [J]. J Am Coll Cardiol.2005,46(5):899-905.
[24]徐亚丽,诊断超声联合微泡促进MSCs归巢缺血心肌及改善心功能的实验研究[D],第三军医大学.2009.
[25]Li X, Wang Z, Ran H, et al.Experimental research on therapeutic angiogenesisinduced by hepatocyte growth factor directed by ultrasound-targeted microbubbledestruction in rats [J]. J Ultrasound Med.2008,27(3):453-460.
[1] Dong F, Ha X Q.Effect of endothelial progenitor cells in neovascularization andtheir application in tumor therapy [J]. Chin Med J (Engl).2010,123(17):2454-2460.
[2] Janic B, Arbab A S.The role and therapeutic potential of endothelial progenitorcells in tumor neovascularizatio [J]n. ScientificWorldJournal.2010,10:1088-1099.
[3]李桂琼,陈庆伟.内皮祖细胞移植策略研究进展[J].心血管病学进展.2011(2).
[4] Kajstura J, Leri A, Bolli R, et al.Endothelial progenitor cells: neovascularizationor more [J]? J Mol Cell Cardiol.2006,40(1):1-8.
[5] Jujo K, Ii M, Losordo D W.Endothelial progenitor cells in neovascularization ofinfarcted myocardium [J]. J Mol Cell Cardiol.2008,45(4):530-544.
[6]李巧.超声微泡联合Ad-EGFP/HIF-1α介导EPCs移植治疗大鼠心肌梗死的实验研究[D].重庆医科大学,2010.
[7] Gluhovschi C,Gluhovschi G,Potencz E, et al. The endothelial cell markers vonWillebrand Factor (vWF), CD31and CD34are lost in glomerulonephritis andno longer correlate with the morphological indices of glomerular sclerosis,interstitial fibrosis, activity and chronicity [J]. Folia Histochem Cytobiol.2012,2(48):230-236.
[8] Fujita Y, Asahra T.Evaluation of circulating endothelial progenitor cell incardiovascular risk. Circ J.2011,75(11):2541-2542.
[9] Nuri M M,Hafeez S. Autologous bone marrow stem cell transplant in acutemyocardial infarction [J]. J Pak Med Assoc.2012,1(62):2-6.
[10] Samper E,Diez-Juan A,Montero J A, et al. Cardiac Cell Therapy: BoostingMesenchymal Stem Cells Effects [J]. Stem Cell Rev.2012.
[11] AtesokK,Matsumoto T,Karlsson J, et al. An emerging cell-based strategy inorthopaedics: endothelial progenitor cells[J]. Knee Surg Sports TraumatolArthrosc.2012.
[12] Camacho Villa AY,Reyes Maldonado E,Montiel Cervantes LA, et al. CD133+CD34+and CD133+CD38+blood progenitor cells as predictors of plateletengraftment in patients undergoing autologous peripheral blood stem celltransplantation[J]. Transfus Apher Sci.2012,139(2):133-141.
[13]崔磊,于海莹,尹烁等.体外诱导大鼠骨髓内皮祖细胞(EPCs)成内皮细胞的实验研究[J].中华创伤骨科杂志,2007(6):1154-1159.
[14] Burger D,Touyz R M. Cellular biomarkers of endothelial health: microparticles,endothelial progenitor cells, and circulating endothelial cells[J]. J Am SocHypertens.2012,2(6):85-99.
[15] Daub K, Langer H, Seizer P, et al.Platelets induce differentiation of humanCD34+progenitor cells into foam cells and endothelial cells [J]. FASEB J.2006,20(14):2559-2561.
[16]赵婷,黎健.内皮祖细胞与氧化应激[J].医学分子生物学杂志,2008(5):344-347.
[17] Birk D M, Barbato J, Mureebe L, et al.Current insight on the biology and clinicalaspects of VEGF regulation [J]. Vasc Endovascular Surg.2008,42(6):517-530.
[18] He S,Xia T,Wang H, et al. Multiple releases of polyplexes of plasmids VEGF andbFGF from electrospun fibrous scaffolds towards regeneration of mature bloodvessels[J]. Acta Biomater2012.
[19] Dardik R, Livnat T, Seligsohn U.Variable effect of alpha v suppression onVEGFR-2expression in endothelial cell of different vascular beds [J]. ThrombHaemost.2009,102(5):975-982.
[20] Kim I, Moon S O, Kim S H, et al.Vascular endothelial growth factor expression ofintercellular adhesion molecule1(ICAM-1), vascular cells adhesion molecule1(VCAM-1), and E-selectin through nuclear factor-kappa B activation inendothelial cell [J]. J Biol Chem.2001,276(10):7614-7620.
[21] Yin L, Ohanyan V, Pung Y F, et al. Induction of vascular progenitor cells fromendothelial cells stimulates coronary collateral growth[J]. Circ Res.2012,2(110):241-252.
[22] Yang J, Ii M, Kamei N, et al.CD34+cells represent highly functional endothelialprogenitor cells in murine bone marrow [J]. PLoS One.2011,6(5):e20219.
[23] Lippross S, Loibl M, Hoppe S, et al.Platelet released growth factors boostexpansion of bone marrow derived CD34(+) and CD133(+) endothelial progenitorcells for autologous grafting [J]. Platelets.2011,22(6):422-432.
[24] Yao W, Firth A L, Sacks R S, et al.Identification of putative endothelial progenitorcells (CD34+CD133+Flk-1+) in the endarterectomized tissue of patients withchronic thromboembolic pulmonary hypertensions [J]. Am J Physiol Lung CellMol Physiol.2009,296(6):L870-L878.
[25] Yang X, Cepko C L.Flk-1, a receptor for vascular endothelial growth factor(VEGF), is expressed by the retinal progenitor cells [J]. J Neurosci.1996,16(19):6089-6099.
[26] Kim M, Kim J H, Kim J H, et al.Differential expression of the stem cell markersand vascular endothelial growth factors in human retinoblastoma tissue [J].Korean J Ophthalmol.2010,24(1):35-39.
[27] Ueno H,Koyama H,Mima Y, et al. Comparison of the effect of cilostazol withaspirin on circulating endothelial progenitor cells and small-dense LDLcholesterol in diabetic patients with cerebral ischemia: a randomized controlledpilot trial[J]. J Atheroscler Thromb.2011,10(18):883-890.
[1]Kajstura J, Leri A, Bolli R, et al.Endothelial progenitor cells: neovascularization ormore [J]? J Mol Cell Cardiol.2006,40(1):1-8.
[2]Awad O, Jiao C, Ma N, et al.Obese diabetic mouse environment differentiallyaffects the primitive and monocytic endothelial cells progenitors [J]. Stem Cells.2005,23(4):575-583.
[3] Wang Z, Liao X, Dong J, et al.The clinical significance and pathogenic roles ofanti-cardiac myosin autoantibody in dilated cardiomyopathy [J]. Chin Med J.2003,116(04):498-502.
[4] Stamova S,Feldmann A,Cartellieri M, et al. Generation of single-chain bispecificgreen fluorescent protein fusion antibodies for imaging of antibody-induced T cellsynapses[J]. Anal Biochem.2012,2(423):261-268.
[5] Molhoj M, Hoffmann P, Baeuerle P A, et al. CD19-/CD3-bispecific antibody of theBiTE class is far superior to tandem diabody with respect to redirected tumor celllysis [J]. Mol Immunol.2007,8(44):1935-1944.
[6] Sarkar S,Tang X L,Das D, et al. A bispecific antibody based assay shows potentialfor detecting tuberculosis in resource constrained laboratory settings [J]. PLoS One.2012,2(7):e32340.
[7]Lee P J, Fang Q, Davol R A, et al.Antibody targeting of stem cells to infarctedmyocardium [J]. Stem Cells.2007,25(03):713-717.
[8]Kim M, Kim J H, Kim J H, et al.Differential expression of stem cell markers andvascular endothelial growth factor in human retinoblastoma tissue [J]. Korean JOphthalmol.2010,24(1):35-39.
[9] Cioffi M,Dorado J,Baeuerle, P A, et al. EpCAM/CD3-Bispecific T-cell engagingantibody MT110eliminates primary human pancreatic cancer stem cells[J]. ClinCancer Res.2012,2(18):465-474.
[10]Kawamoto A, Gwon H C, Iwaguro H, et al.Therapeutic potential of ex vivoexpanded the endothelial progenitor cell for myocardial ischemia [J]. Circulation.2001,103(5):634-637.
[11]杨亚荣,曾秋棠,郎明健等.内皮祖细胞移植对大鼠心肌梗死后缺血心肌血管再生及心功能的影响[J].中国组织工程研究，2008(12):4815-4818.
[12]Samper E,Diez-Juan A,Montero J A, et al. Cardiac Cell Therapy: BoostingMesenchymal Stem Cells Effects [J]. Stem Cell Rev.2012.
[13]Dong F, Ha X Q.Effect of endothelial progenitor cells in neovascularization andtheir application in tumor therapy [J]. Chin Med J (Engl).2010,123(17):2454-2460.
[14]Davol P A, Smith J A, Kouttab N, et al.Anti-CD3x anti-HER2bispecific antibodyeffectively redirects armed T cells to inhibit tumor development and growth inhormone-refractory prostate cancer-bearing severe combined immunodeficientbeige mice [J]. Clin Prostate Cancer.2004,3(2):112-121.
[15]Kiewe P, Hasmuller S, Kahlert S, et al.Phase I trial of the trifunctional anti-HER2x anti-CD3antibody ertumaxomab in metastatic breast cancer [J]. Clin Cancer Res.2006,12(10):3085-3091.
[16] Baeuerle P A,Itin C. Clinical Experience with Gene Therapy and BispecificAntibodies for T Cell-based Therapy of Cancer[J]. Curr Pharm Biotechnol.2012.
[17]Asahara T, Murohara T, Sullivan A, et al. Isolation of the putative progenitorendothelial cells for angiogenesis [J]. Science.1997,275(5302):964-967.
[18]Lum L G, Fok H, Sievers R, et al.Targeting of Lin-Sca+hematopoietic stem cellswith bispecific antibodies to injured myocardium [J]. Blood Cells Mol Dis.2004,32(1):82-87.
[19]Lippross S, Loibl M, Hoppe S, et al.Platelet released growth factors boostexpansion of bone marrow derived CD34(+) and CD133(+) endothelial progenitorcells for autologous grafting [J]. Platelets.2011,22(6):422-432.
[20]邓玮.BMSC在微泡及BiAb辅助下修复心肌纤维化的实验研究及PⅢNP对冠心病合并代谢综合征患者的预测价值[D].重庆医科大学,2011.
[21]Scheubel R J, Holtz J, Friedrich I, et al.Paracrine effects of CD34progenitor cellson angiogenic endothelial sprouting [J]. Int J Cardiol.2010,139(2):134-141.
[22]Nuri M M,Hafeez S. Autologous bone marrow stem cell transplant in acutemyocardial infarction [J]. J Pak Med Assoc.2012,1(62):2-6.
[23]Wassmann S, Werner N, Czech T, et al.Improvement of endothelial functions bysystemic transfusion of vascular progenitor cell [J]. Circ Res.2006,99(8):e74-e83.
[1] A. Kawamoto, H. C. Gwon, H. Iwaguro,et al. Therapeutic potential of ex vivoexpanded endothelial progenitor cells for myocardial ischemia[J].Circulation,2001,103(5):634-637.
[2] O. Awad, C. Jiao, N. Ma,et al. Obese diabetic mouse environment differentiallyaffects primitive and monocytic endothelial cell progenitors[J]. StemCells,2005,23(4):575-583.
[3] A. Stein, M. Knodler, M. Makowski,et al. Local erythropoietin and endothelialprogenitor cells improve regional cardiac function in acute myocardialinfarction[J]. BMC Cardiovasc Disord,2010,10:43.
[4]杨亚荣,曾秋棠,郎明健等.内皮祖细胞移植对大鼠心肌梗死后缺血心肌血管再生及心功能的影响[J].中国组织工程研究，2008(12):4815-4818.
[5] W. Kuliczkowski, R. Derzhko, I. Prajs, et al. Endothelial progenitor cells and leftventricle function in patients with acute myocardial infarction: potentialtherapeutic considertions[J]. Am J Ther,2012,19(1):44-50.
[6]李晓东.超声波空化效应的生物学机制[J].临床超声医学杂志.2004,12(6):40-41.
[7]李巧.超声微泡联合Ad-EGFP/HIF-1α介导EPCs移植治疗大鼠心肌梗死的实验研究[D].重庆医科大学,2010.
[8]徐亚丽,诊断超声联合微泡促进MSCs归巢缺血心肌及改善心功能的实验研究[D],第三军医大学.2009.
[9] B. H. Annex, M. Simons. Growth factor-induced therapeutic angiogenesis in theheart: protein therapy[J]. Cardiovasc Res,2005,65(3):649-655.
[10] U. F. Florenzano. Acute myocardial infarction mortality in Chile: Thrombolysis orangioplasty[J]. Rev Med Chil,2011,139(11):1393-1395.
[11] V. F. Segers, I. Van Riet, L. J. Andries,et al. Mesenchymal stem cell adhesion tocardiac microvascular endothelium: activators and mechanisms[J]. Am J PhysiolHeart Circ Physiol,2006,290(4): H1370-H1377.
[12] M. Shabbir, A. M. Kayani, O. Qureshi,et al. Predictors of fatal outcome in acutemyocardial infarction[J]. J Ayub Med Coll Abbottabad,2008,20(3):14-16.
[13] A. Mullasari. Strategy of in ambulance thrombolysis followed by routine PCI inacute myocardial infarction[J]. Indian Heart J,2009,61(5):448-453.
[14] W. Xu, X. Zhang, H. Qian,et al. Mesenchymal stem cells from adult human bonemarrow differentiate into a cardiomyocyte phenotype in vitro[J]. Exp Biol Med(Maywood),2004,229(7):623-631.
[15] L. Barile, I. Chimenti, R. Gaetani,et al. Cardiac stem cells: isolation, expansion andexperimental use for myocardial regeneration[J]. Nat Clin Pract CardiovascMed,2007,4Suppl1: S9-S14.
[16] S. Miyagawa, G. Matsumiya, T. Funatsu,et al. Combined autologous cellularcardiomyoplasty using skeletal myoblasts and bone marrow cells for humanischemic cardiomyopathy with left ventricular assist system implantation: reportof a case[J]. Surg Today,2009,39(2):133-136.
[17] Y. P. Li, S. Paczesny, E. Lauret,et al. Human mesenchymal stem cells license adultCD34+hemopoietic progenitor cells to differentiate into regulatory dendriticcells through activation of the Notch pathway[J]. J Immunol,2008,180(3):1598-1608.
[18] Y. L. Xu, Y. H. Gao, Z. Liu,et al. Myocardium-targeted transplantation ofmesenchymal stem cells by diagnostic ultrasound-mediated microbubbledestruction improves cardiac function in myocardial infarction of New Zealandrabbits[J]. Int J Cardiol,2010,138(2):182-195.
[19] C. Toma, M. F. Pittenger, K. S. Cahill,et al. Human mesenchymal stem cellsdifferentiate to a cardiomyocyte phenotype in the adult murine heart[J].Circulation,2002,105(1):93-98.
[20] K. Zen, M. Okigaki, Y. Hosokawa,et al. Myocardium-targeted delivery ofendothelial progenitor cells by ultrasound-mediated microbubble destructionimproves cardiac function via an angiogenic response[J]. J Mol CellCardiol,2006,40(6):799-809.
[21] T. Asahara, T. Murohara, A. Sullivan,et al. Isolation of putative progenitorendothelial cells for angiogenesis[J]. Science,1997,275(5302):964-967.
[22] J. M. Hare, S. V. Chaparro. Cardiac regeneration and stem cell therapy[J]. CurrOpin Organ Transplant,2008,13(5):536-542.
[23] C. H. Hu, Z. M. Li, Du ZM,et al. Expanded human cord blood-derived endothelialprogenitor cells salvage infarcted myocardium in rats with acute myocardialinfarction[J]. Clin Exp Pharmacol Physiol,2010,37(5-6):551-556.
[24]李兴升,王志刚,李雪霖等,超声破坏微泡造影剂促进心肌梗死大鼠血管新生的实验研究[J].中华超声影像学杂志.2007,16(21):1071-1073.
[25] P. A. Davol, J. A. Smith, N. Kouttab,et al. Anti-CD3x anti-HER2bispecificantibody effectively redirects armed T cells to inhibit tumor development andgrowth in hormone-refractory prostate cancer-bearing severe combinedimmunodeficient beige mice[J]. Clin Prostate Cancer,2004,3(2):112-121.
[26] P. Kiewe, S. Hasmuller, S. Kahlert,et al. Phase I trial of the trifunctional anti-HER2x anti-CD3antibody ertumaxomab in metastatic breast cancer[J]. Clin CancerRes,2006,12(10):3085-3091.
[27] T. C. Zhao, A. Tseng, N. Yano,et al. Targeting human CD34+hematopoietic stemcells with anti-CD45x anti-myosin light-chain bispecific antibody preservescardiac function in myocardial infarction[J]. J Appl Physiol,2008,104(6):1793-1800.
[28] W. Yao, A. L. Firth, R. S. Sacks,et al. Identification of putative endothelialprogenitor cells (CD34+CD133+Flk-1+) in endarterectomized tissue of patientswith chronic thromboembolic pulmonary hypertension[J]. Am J Physiol Lung CellMol Physiol,2009,296(6): L870-L878.
[29] R. Senior, S. Bhattacharya, P. Manspeaker,et al.99mTc-antimyosin antibodyimaging for the detection of acute myocardial infarction in human beings[J]. AmHeart J,1993,126(3Pt1):536-542.
[30] R. J. Lee, Q. Fang, P. A. Davol,et al. Antibody targeting of stem cells to infarctedmyocardium[J]. Stem Cells,2007,25(3):712-717.
[31] C. Herder, T. Tonn, R. Oostendorp,et al. Sustained expansion and transgeneexpression of coagulation factor VIII-transduced cord blood-derived endothelialprogenitor cells[J]. Arterioscler Thromb Vasc Biol,2003,23(12):2266-2272.
[32] G. Chodorowska, J. Chodorowski, A. Wysokinski. Vascular endothelial growthfactor (VEGF) in physiological and pathological conditions[J]. Ann Univ MariaeCurie Sklodowska Med,2004,59(2):8-14.
[33] D. M. Birk, J. Barbato, L. Mureebe,et al. Current insights on the biology andclinical aspects of VEGF regulation[J]. Vasc Endovascular Surg,2008,42(6):517-530.
[34] X. Yang, C. L. Cepko. Flk-1, a receptor for vascular endothelial growth factor(VEGF), is expressed by retinal progenitor cells[J]. J Neurosci,1996,16(19):6089-6099.
[35]董静.血管内皮生长因子的血管新生疗法治疗缺血性心血管疾病[J].中国分子心脏病学杂志,2005(2):507-510.
[36]沈帆霞,陈生弟.血管内皮细胞生长因子的血管新生、神经新生及神经保护作用[J].中国现代神经疾病杂志,2006(2):143-146.
[37]吕菁君,林国生,李庚山.骨髓单个核细胞白体移植促进心脏血管再生[J].中华心胸血管外科杂志,2004(2):102-104.
[38] I. Kim, S. O. Moon, S. H. Kim,et al. Vascular endothelial growth factor expressionof intercellular adhesion molecule1(ICAM-1), vascular cell adhesion molecule1(VCAM-1), and E-selectin through nuclear factor-kappa B activation inendothelial cells[J]. J Biol Chem,2001,276(10):7614-7620.
[39]余跃,干细胞基础与临床.中国科学技术大学出版社,2008:102.
[40] D. J. Ceradini, A. R. Kulkarni, M. J. Callaghan,et al. Progenitor cell trafficking isregulated by hypoxic gradients through HIF-1induction of SDF-1[J]. NatMed,2004,10(8):858-864.
[41] A. T. Askari, S. Unzek, Z. B. Popovic,et al. Effect of stromal-cell-derived factor1on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy[J].Lancet,2003,362(9385):697-703.
[42] G. S. Lin, J. J. Lu, X. J. Jiang,et al. Autologous transplantation of bone marrowmononuclear cells improved heart function after myocardial infarction[J]. ActaPharmacol Sin,2004,25(7):876-886.
[43]赵婷,黎健.内皮祖细胞与氧化应激[J].医学分子生物学杂志,2008(5):344-347.
[1] Wang D, Zheng W, Wang SM, et al. Estimation of cancer incidence and mortalityattributable to overweight, obesity, and physical inactivity in china. Nutr Cancer.2012;64:48-56.
[2] Frayling TM, Timpson NJ, Weedon MN, Zeggini E, Fret al. A common variant in theFTO gene is associated with body mass index and predisposes to childhood andadult obesity. Science.2007;316:889–94.
[3] Xi B, Mi J.FTO polymorphisms are associated with obesity but not with diabetes inEast Asian populations: a meta-analysis. Biomed Environ Sci.2009;22:449-57.
[4] Adeyemo A, Chen G, Zhou J, et al. FTO genetic variation and association withobesity in West Africans and African Americans. Diabetes.2010;59:1549-54.
[5] Wang H, Dong S, Xu H, et al. Genetic variants in FTO associated with metabolicsyndrome: a meta-and gene-based analysis. Mol Biol Rep.2011; doi:10.1007/s11033-011-1377-y.
[6] Cecil JE, Tavendale R, Watt P, et al. An obesity-associated FTO gene variant andincreased energy intake in children. N Engl J Med.2008;359:2558-66.
[7] Kaklamani V, Yi N, Sadim M, et al. The role of the fat mass and obesity associatedgene (FTO) in breast cancer risk. BMC Med Genet.2011;12:52.
[8] Kusinska R, Górniak P, Pastorczak A, et al. Influence of genomic variation in FTOat16q12.2, MC4R at18q22and NRXN3at14q31genes on breast cancer risk.Mol Biol Rep.2012;39:2915–9.
[9] Lurie G, Gaudet MM, Spurdle AB, et al. The obesity-associated polymorphismsFTO rs9939609and MC4R rs17782313and endometrial cancer risk innon-Hispanic white women. PLoS One.2011;6:e16756.
[10] Delahanty RJ, Beeghly-Fadiel A, Xiang YB, et al. Association of obesity-relatedgenetic variants with endometrial cancer risk: a report from the ShanghaiEndometrial Cancer Genetics Study. Am J Epidemiol.2011;174:1115–26.
[11] Gaudet MM, Yang HP, Bosquet JG, et al. No association between FTO or HHEXand endometrial cancer risk. Cancer Epidemiol Biomarkers Prev.2010;19:2106–9.
[12] Tang H, Dong X, Hassan M,et al.Body mass index and obesity-anddiabetes-associated genotypes and risk for pancreatic cancer. Cancer EpidemiolBiomarkers Prev.2011;20:779–92.
[13] Pierce BL, Austin MA, Ahsan H. Association study of type2diabetes geneticsusceptibility variants and risk of pancreatic cancer: an analysis of PanScan-I data.Cancer Causes Control.2011;22:877–83.
[14] Meyer TE, Boerwinkle E, Morrison AC, et al. Diabetes genes and prostate cancerin the Atherosclerosis Risk in Communities study. Cancer Epidemiol BiomarkersPrev.2010;19:558–65.
[15] Lewis SJ, Murad A, Chen L, et al. Associations between an obesity related geneticvariant (FTO rs9939609) and prostate cancer risk. PLoS One.2010;5:e13485.
[16] Tarabra E, Actis GC, Fadda M, et al. The obesity gene and colorectal cancer risk: apopulation study in Northern Italy. Eur J Intern Med.2012;23:65–9.
[17] Brennan P, McKay J, Moore L, et al. Obesity and cancer: Mendelian randomizationapproach utilizing the FTO genotype. Int J Epidemiol.2009;38:971–5.
[18] DerSimonian R, Laird N. Meta-analysis in clinical trials. Control Clin Trials.1986;7:177–88.
[19] Mantel N, Haenszel W. Statistical aspects of the analysis of data from retrospectivestudies of disease. J Natl Cancer Inst.1959;22:719–48.
[20] Begg CB, Mazumdar M. Operating characteristics of a rank correlation test forpublication bias. Biometrics.1994;50:1088–101.
[21] Egger M, Davey Smith G, Schneider M, et al.Bias in meta-analysis detected by asimple, graphical test. BMJ.1997;315:629–34.
[22] Nock NL, Plummer SJ, Thompson CL, et al.FTO polymorphisms are associatedwith adult body mass index (BMI) and colorectal adenomas in African-Americans.Carcinogenesis.2011;32:748–56.
[23] Fischer J, Koch L, Emmerling C, et al. Inactivation of the Fto gene protects fromobesity. Nature.2009;458:894–8.
[24] Zimmermann E, Kring SI, Berentzen TL, et al. Fatness-associated FTO genevariant increases mortality independent of fatness--in cohorts of Danish men.PLoS One.2009;4:e4428.
[25] Stratigopoulos G, Padilla SL, LeDuc CA, et al. Regulation of Fto/Ftm geneexpression in mice and humans. Am J Physiol Regul Integr Comp Physiol.2008;294:R1185–96.
[26] Jowett JB, Curran JE, Johnson MP, et al. Genetic variation at the FTO locusinfluences RBL2gene expression. Diabetes.2010;59:726–32.
[1] Asahara T, Murohara T, Sullivan A, et al. Isolation of putative progenitorendothelial cells for angiogenesis[J]. Science,1997,275(5302):964–967.
[2] Wassmann S, Werner N, Czech T, et al.Improvement of endothelial function bysystemic transfusion of vascular progenitor cells[J]. Circ Res,2006,99(8): e74–e83.
[3] Aicher A, Rentsch M, Sasaki K, et al.Nonbone marrow-derived circulatingprogenitor cells contribute to postnatal neovascularization following tissue ischemia[J].Circ Res,2007,100(4):581–589.
[4] Sengenes C, Miranville A, Maumus M, et al.Chemotaxis and differentiation ofhuman adipose tissue CD34+CD31-progenitor cells: role of stromal derived factor-1released by adipose tissue capillary endothelial cells[J]. Stem Cells,2007,25(9):2269–2276.
[5] Ingram DA，Mead LE，Moore DB, et a1.Vessel wal1-Derived endothelial cellsrapidly proliferate because they contain a complete hierarchy of endothelial progenitorcells[J]. Blood,2005,105(7):2783-2786．
[6] Kawamoto A, Losordo DW, et al.Endothelial progenitor cells for cardiovascularregeneration[J]. Trends Cardiovasc Med,2008,18:33–37.
[7] Kawamoto A, Asahara T. Role of progenitor endothelial cells in cardiovasculardisease and upcoming therapies[J]. Catheter Cardiovasc Interv,2007,70(4):477–484.
[8] Shiba Y, Takahashi M, Yoshioka T, et al.M-CSF accelerates neointimal formationin the early phase after vascular injury in mice: the critical role of the SDF-1-CXCR4system[J]. Arterioscler Thromb Vasc Biol,2007,27(2):283–289.
[9] Ozuyaman B, Ebner P, Niesler U, et al.Nitric oxide differentially regulatesproliferation and mobilization of endothelial progenitor cells but not of hematopoieticstem cells[J]. Tromb Haemost,2005,94(4):770-772.
[10] Faouzia Zemani, Jean-Sébastien Silvestre, Fran oise Fauvel-Lafeve, et al. Ex VivoPriming of Endothelial Progenitor Cells With SDF-1BeforeTransplantation CouldIncrease Their Proangiogenic Potential[J]. Arterioscler Thromb Vasc Biol,2008,28(4):644-650.
[11] Pitchford SC, Furze RC, Jones CP, et al.Differential mobilization of subsets ofprogenitor cells from the bone marrow[J]. Stem Cell,2009,4(1):62-72.
[12] Aicher A, Zeiher AM, Dimmeler S. Mobilizing endothelial progenitor cells[J].Hypertension,2005,45(3):321–325.
[13] Qin G, Ii M, Silver M, et al.Functional disruption of alpha4integrin mobilizes bonemarrow-derived endothelial progenitors and augments ischemic neovascularization[J]. JExp Med,2006,203(1):153–163.
[14] Iwaguro H, Yamaguchi J, Kalka C, et al.Endothelial progenitor cell vascularendothelial growth factor gene transfer for vascular regeneration[J]. Circulation,2002,105(6):732–8.
[15] Jiang M, Wang C, He B, et al.Angiogenesis by transplantation of HIF-1a modifiedEPCs into ischemic limbs[J]. J Cell Biochem,2008,103(1):321-334.
[16] Ming-Bao Song，Xue-Jun Yu，Guang-Xu Zhu, et al.Transfection of HGF geneenhances endothelial progenitor cell (EPC) function and improves EPC transplantefficiency for balloon-induced arterial injury in hypercholesterolemic rats[J]. VascPharmacol,2009,51(2):205-213.
[17] Shantsila E, Watson T, Tse HF, et al. New insights on endothelial progenitor cellsubpopulations and their angiogenic properties[J]. J Am Coll Cardiol,2008,51(6):669-671.
[18] Case J, Mead LE, Bessler WK, et al.Human CD34+AC133+VEGFR-2+cells arenot endothelial progenitor cells but distinct, primitive hematopoietic progenitors[J]. ExpHematol,2007,35(7):1109-1118.
[19] Yoder MC, Mead LE, Prater D, et al.Redefining endothelial progenitor cells viaclonal analysis and hematopoietic stem/progenitor cell principals[J]. Blood,2007,109(5):1801–1809.
[20] Wang XX, Zhang FR, Shang YP, et al. Transplantation of Autologous EndothelialProgenitor Cells May Be Beneficial in Patients With Idiopathic Pulmonary ArterialHypertension: A Pilot Randomized Controlled Trial[J]. J Am Coll Cardiol,2007,49(10):1566-1571.
[21] Nakamura T, Torimura T, Sakamoto M, et al. S ignificance and therapeuticpotential of endothelial progenitor cell transplantation in a cirrhotic liver rat model[J].Gastroenterology,2007,133(1):91-107.
[22] Liu F, Liu ZD, Wu N, et al.Transplanted endothelial progenitor cells amelioratecarbon tetrachloride-induced liver cirrhosis in rats[J]. Liver Transpl,2009,15(9):1092-2100.
[23] Beeres SL, Bax JJ, Dibbets-Schneider P, et al.Intramyocardial injection ofautologous bone marrow mononuclear cells in patients with chronic myocardialinfarction and severe left ventricular dysfunction[J]. Am J Cardiol,2007,100(7):1094-1098.
[24] George J, Afek A, Abashidze A, et al.Transfer of endothelial progenitor and bonemarrow cells influences atherosclerotic plaque size and composition in apolipoprotein Eknockout mice[J]. Arterioscl Throm Vasc biol,2005,25(12):2636–2641.
[25] Tateno K, Minamino T, Toko H, et al. Critical roles of muscle-secreted angiogenicfactors in therapeutic neovascularization[J]. Circ Res2006,98(9):1194–1202.
[26] Kawamoto A, Iwasaki H, Kusano K, et al.CD34-positive cells exhibit increasedpotency and safety for therapeutic neovascularization after myocardial infarctioncompared with total mononuclear cells[J]. Circulation,2006,114(20):2163–2169.
[27] Hofmann M, Wollert KC, Meyer GP, et al.Monitoring of bone marrow cell hominginto the infarcted human myocardium[J]. Circulation,2005,111(17):2198–2202.
[28] Schuh A, Liehn EA, Sasse A, et al.Transplantation of endothelial progenitor cellsimproves neovascularization and left ventricular function after myocardial infarction ina rat model[J]. Basic Res Cardiol,2008,103(1):69-77.
[29] Li M, Takenaka H, Asai J, et al. Endothelial progenitor thrombospondin-1mediatesdiabetes-induced delay in reendothelialization following arterial injury[J]. Circ Res,2006,98(5):697–704.
[30]Abou-Saleh H, Yacoub D, Théorêt JF, et al.Endothelial progenitor cells bind andinhibit platelet function and thrombus formation[J]. Circulation,2009,120(22):2230-2239.30970826国家自然科学基金资助项目《免疫双特异抗体联合超声微泡介导内皮祖细胞移植促进血管新生》