Published online before print October 16, 2008, doi: 10.1161/CIRCRESAHA.108.180463
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© 2008 American Heart Association, Inc.
Integrative Physiology |
Sustained Persistence of Transplanted Proangiogenic Cells Contributes to Neovascularization and Cardiac Function After Ischemia
From Molecular Cardiology (T.Z., C.-H.Y., T.T., M.M.-R., G.C., C.U., A.M.Z., S.D.), Department of Internal Medicine III, University Frankfurt; Max Planck Institute (A.W., T.B.), Bad Nauheim; Junior Group Molecular Imaging (F.K.), German Cancer Research Center, Heidelberg; Angewandte Virologie und Gentherapie (S.S., M.G.), Georg-Speyer-Haus, Frankfurt; and Gemeinschaftspraxis für Pathologie (C.I.), Frankfurt, Germany.
Correspondence to Prof Dr Stefanie Dimmeler Molecular Cardiology, Department of Internal Medicine III University of Frankfurt, Theodor-Stern-Kai 7 60590 Frankfurt am Main, Germany. E-mail Dimmeler{at}em.uni-frankfurt.de
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Abstract |
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Circulating blood–derived vasculogenic cells improve neovascularization of ischemic tissue by a broad repertoire of potential therapeutic actions. Whereas initial studies documented that the cells incorporate and differentiate to cardiovascular cells, other studies suggested that short-time paracrine mechanisms mediate the beneficial effects. The question remains to what extent a physical incorporation is contributing to the beneficial effects of cell therapy. By using the inducible suicide gene thymidine kinase to deplete transplanted cells, we determined the contribution of physical incorporation in 3 animal models. After acute myocardial infarction, depletion of cells 14 days after infusion resulted in a reduction of capillary density and a substantial deterioration of heart function. Likewise, neovascularization of Matrigel plugs and ischemic limbs was significantly suppressed when infused cells were depleted 7 days after infusion. Induction of cell death in the previously transplanted cells reduced perfusion and led to vascular leakage as evidenced by Evans blue extravasation. These results indicate that physical incorporation and persistence of cells contribute to cell-mediated improvement of neovascularization and cardiac function. Long-term paracrine activities and/or cell intrinsic mechanisms may have contributed to the maintenance of functional improvement.
Key Words: progenitor cells • neovascularization • cell therapy
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Introduction |
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Cell therapy is a promising option for treating ischemic diseases. Adult stem and progenitor cells from various sources have experimentally been shown to augment the functional recovery after ischemia. Clinical trials confirmed that autologous cell therapy using bone marrow–derived or circulating blood-derived progenitor cells is safe and provides beneficial effects1–3 (see also recent metaanalysis4). Endothelial progenitor cells (EPCs) were initially isolated from bone marrow or peripheral blood and were characterized by the expression of the hematopoietic stem cell marker CD34 or CD133 and endothelial markers such as vascular endothelial growth factor receptor 2 (KDR).5,6 Isolated CD34+ cells differentiate to endothelial cells in vitro and incorporate into newly formed vessels during angiogenesis in vivo.5 These pioneering studies suggested that bone marrow–derived cells can participate in vascular repair. Indeed, a single hematopoietic stem cell was shown to give rise to both blood cells and vascular endothelium.7–9 The contribution of circulating cells to the endothelium was further supported by sex-mismatched heart transplantation in humans.10,11 Other investigators used culture assays to enrich for EPC-yielding cells, which coexpress endothelial and myeloid marker ("early EPCs," proangiogenic cells), or to show a more mature endothelial cell phenotype on further culture and expansion ("late or outgrowing EPCs").12,13 Although the origin and the phenotype of the cultured EPCs is not entirely clear and may vary depending on the defined specific culture conditions,14 the therapeutic potential and improved neovascularization mediated by the cultured cells has been documented extensively in ischemic models.12,15,16
Progenitor cell–mediated improved neovascularization of ischemic tissue might be attributable to various therapeutic actions including a physical incorporation of the infused cells in the endothelium leading to the formation of new capillaries.16–18 Transplanted or infused hematopoietic or endothelial progenitor cells also were shown to differentiate to cardiac muscle cells.5,19,20 However, the capacity of hematopoietic or endothelial progenitor cells to acquire a cardiac phenotype is not well established.21 Bone marrow–derived cells also were detected in the perivascular region, where they were proposed to provide paracrine factors contributing to vessel growth.22 Indeed, cultured endothelial progenitor cells release proangiogenic and antiapoptotic cytokines, which may contribute to enhanced angiogenesis and endogenous repair.23,24 Based on these findings, it was speculated that short-term paracrine mechanisms account for the observed therapeutic benefits of cell therapy.25–27
Thus, despite ample data providing evidence for functional improvements after cell therapy, the mechanisms by which cells augment the functional recovery after ischemia are not well defined, and it is not known to what extent the physical incorporation and persistence of transplanted cells contributes to the beneficial effects of cell therapy.
To determine the role of cell incorporation and persistence after cell therapy, we used the suicide gene thymidine kinase (TK), which activates the prodrug gancyclovir (GV),28 to induce cell death in infused cells at specific time points after infusion and assessed the influence on neovascularization and cardiac function after induction of ischemia. Our results demonstrate that depletion of cells weeks after infusion reduces vascularization and causes a deterioration of function in 3 different animal models.
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Materials and Methods |
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Culture of Proangiogenic Cells
Human peripheral blood mononuclear cells (MNCs) were isolated by density gradient centrifugation with Biocoll (Biochrom AG) from peripheral venous blood according to Vasa et al.29 Immediately following isolation, 8x106 MNCs per milliliter of medium were plated on culture flask coated with human fibronectin (Sigma, Taufkirchen, Germany) and maintained in endothelial basal medium (EBM; LONZA, Verviers, Belgium) supplemented with EGM SingleQuots and 20% FBS. After 3 days in culture, nonadherent cells were removed by thorough washing with PBS, and adherent cells were incubated in fresh medium.
Cell Culture
Pooled human umbilical vein endothelial cells (HUVECs) were purchased from LONZA (Verviers, Belgium) and were cultured in EBM medium supplemented with EGM SingleQuots and 10% FBS. HEK 293 cells were maintained in DMEM containing 10% FBS and penicillin–streptomycin.
Preparation of Lentiviral Stocks
Self-inactivating lentiviral vectors containing the enhanced green fluorescent protein (GFP) gene or the viral TK gene of the herpes simplex type 2 virus and a WPRE (woodchuck posttranscriptional regulatory element) under the control of a spleen focus-forming virus promoter were generated by transient transfection in 293T cells using pCMV
R8.91 as packaging plasmid and pMD2.G for vesicular stomatitis virus–G protein (VSV-G) pseudotyping as described.30,31 After 8 hours, the medium was replaced by DMEM/10 mmol/L sodium butyrate for 20 hours. Lentiviral particles were collected in EBM supplemented with EGM SingleQuots and 20% FBS every 24 hours for 2 days, pooled (200 to 250 mL), and filtered through 0.22-µm filters.
Lentiviral Transduction
For lentiviral transduction, EPCs were transduced on days 3 and 4 after isolation. Transduction was carried out by adding viral supernatant to the EBM supplemented with EGM SingleQuots and 20% FBS. After 8 hours, medium was changed and EPCs were transduced a second time. Transduced EPCs were used on days 5 or 6 after isolation for the following experiments.
Matrigel Plug Model
All animal experiments were approved by the Refierungspracsidium Darmstadt, Germany. A total of 500 µL of Matrigel Basement Membrane Matrix (BD Biosciences) without supplements was injected subcutaneously into 6- to 8-week-old female athymic nude mice (Harlan) along the abdominal midline. After 7 or 14 days, blood vessel infiltration in Matrigel plugs was quantified by analysis of lectin and smooth muscle actin-stained sections using microscopy.
Hindlimb Ischemia Model
Eight-week-old male Balb/c nude mice (Charles River) were anesthetized with isoflurane. Right femoral artery and vein were coagulated and then cut out to induce critical ischemia. One day after operation, EPCs (1 million) were administrated via the tail vein. Laser Doppler perfusion image was taken as indicated and perfusion of each limb was calculated from mean value multiplied by the number of pixel of the region below the inguinal ligament. Data were represented as ratios of the ischemic limb to the nonischemic limb. Evans blue (30 mg/kg) was injected intravenously at day 9. Mice were euthanized 24 hours after dye injection and cardioperfused with normal saline. Tissue was harvested and frozen. Evans blue was extracted from tissue using trichloroacetic acid and ethanol. Absorbance was measured at 620 nm.
Acute Myocardial Infarction Model
Left coronary artery occlusion was performed in female nu/nu mice (8 to 9 weeks; Harlan) under mechanical ventilation and anesthesia. Acute myocardial ischemia was induced by ligating the left coronary artery. Cells (2 million) were intravenously infused 24 hours after operation.
Physiological Assessment of Left Ventricular Function
In the myocardial infarction study, transthoracic echocardiography was performed to evaluate left ventricular function by measuring ejection fraction 2, 3, and 4 weeks after myocardial ischemia.
MRI Procedure
Cardiac MRI was performed under volatile isoflurane (1.5% to 2.0%) anesthesia with a Bruker Pharmascan 7.0 T, a custom-built circularly polarized birdcage resonator and use of the Early Access Package for Self-gated Cardiac Imaging (Intragate).32 This measurement is based on the gradient echo method (repetition time=44.4 ms; echo time=6.0 ms; field of view=2.20x2.20 cm; slice thickness=1.0 mm; matrix=128x128; repetitions=100). The imaging plane was localized using scout images showing the 4- and 2-chamber view of the heart, followed by acquisition in short axis view, orthogonal on the septum in both scouts. Multiple contiguous short-axis slices consisting of 6 to 8 slices were acquired for complete coverage of the left ventricle. All MRI data were analyzed using Qmass digital imaging software (Medis, Leiden, The Netherlands).
Immunostainings
Sections were deparaffinized and were incubated with biotinylated Isolectin B4 (Vector B1201) followed by streptavidin Alexa Fluor 488. TUNEL assays (Roche) was performed according to the instructions of the manufacturer. Confocal microscopic analysis was performed using a Zeiss LSM 510 Meta.
Statistical Analysis
Data are expressed as means±SEM. Two treatment groups were compared by Mann–Whitney test, 3 or more treatment groups were compared by 1-way ANOVA, followed by post hoc analysis adjusted with a least significant difference correction for multiple comparisons (SPSS). Results were considered statistically significant when P<0.05.
An expanded Materials and Methods section is available in the online data supplement at http://circres.ahajournals.org.
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Results |
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Induction of Cell Death by Virally Transduced Suicide Gene
To investigate the physical contribution of proangiogenic cells, peripheral blood–derived mononuclear cells were cultivated on fibronectin-coated dishes under conditions favoring endothelial commitment as previously described.15,16 Adherent cells at day 4 express endothelial markers (KDR, von Willebrand factor, endothelial nitric oxide synthase), but also hematopoietic/myeloid markers such as CD31, CD45, and CD14 (see previous publications for cell characteristics13,33). To determine the role of cell incorporation and persistence, cells were transduced using a lentiviral vector encoding TK. The transfection efficiency of the GFP control vector was >80%, and control experiments confirmed that the viral transduction did not affect cell function (viability, migration, colony forming activity) (data not shown). Addition of GV induced toxicity in the TK-transduced cells but not in the GFP control vector–expressing cells (Figure 1a). To determine potential indirect effects of GV-induced cytotoxicity, culture supernatants of the GV treated TK-expressing cells were added to HUVECs or cardiomyocytes. However, conditioned medium did not induce cell death (Figure 1b and data not shown). Moreover, TK-expressing cells were cocultured with GFP-transfected HUVECs before adding GV to induce cell death in the TK-expressing cells. The viability of GFP-transfected HUVECs was not reduced by inducing cell death in the TK-expressing cells (99.9% of control), indicating that the treatment with GV selectively kills the transduced cells.
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Depletion of Infused Cells Impairs Functional Recovery After Acute Myocardial Infarction
Next, we infused TK-expressing cells in nude mice after acute myocardial infarction to determine the effect of cell ablation on heart function in vivo (Figure 2a). Myocardial infarction was induced by permanent ligation of the left coronary artery, and TK- or GFP-transduced cells were injected 1 day later. After 4 weeks, 1.55±0.2% of cells were positive for human specific Alu sequence in the border zone of the infarcts. Human cells were preferentially localized in or around vessels, whereas only a small number of myocytes (<0.1%) were detected (Figures I and II in the online data supplement). Two weeks after acute myocardial infarction, left ventricular ejection fraction was significantly improved in both cell-treated groups compared with PBS control mice (Figure 2b). In GFP cell-treated mice, the ejection fraction was not affected by GV treatment, which was initiated 2 weeks after cell transplantation, and the beneficial effect of cell therapy was maintained during the 4-week observation period. In contrast, left ventricular ejection fraction significantly deteriorated after injection of GV in TK cell–treated mice, indicating that ablation of the cells 2 weeks after infusion impaired heart function. To confirm these results, MRI was used as an additional means to corroborate our results. Again, left ventricular ejection fraction was significantly improved in mice treated with TK cells in the absence of GV (Figure 2c and 2e). Injection of GV after 2 weeks in TK cell–treated mice resulted in a reduced left ventricular ejection fraction at 4 weeks compared with TK cell–treated mice not receiving GV (Figure 2c). Consistent with these results, end-systolic volume was significantly reduced in mice injected with TK cells but significantly increased by additional GV treatment (Figure 2d and 2e).
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Because injected cells preferentially incorporated in or around capillaries, we investigated whether ablation of the cells affected capillary density. We detected a significant reduction of the increased capillary density area in TK cell–treated mice 4 weeks after myocardial infarction, when GV was injected at 2 weeks (Figure 2f; for higher magnification, see supplemental Figure IV). To obtain a more detailed view of the acute effects induced by GV treatment in TK cell–treated mice, TUNEL stainings were performed. As shown in Figure 2g and supplemental Figure III, GV treatment induced cell death in TK cell–treated mice 3 days after onset of GV treatment (1.7±0.28-fold compared with mice, which had received TK cells without GV treatment, P<0.05).
Depletion of Infused Cells Reduces Neovascularization
Having demonstrated that depletion of transplanted cells reduced capillary density, we investigated the relevance of cell incorporation in 2 models of vascularization. First, we infused TK-expressing cells in nude mice implanted with a Matrigel plug to determine the effect of cell ablation on angiogenesis. Infusion of GFP- or TK-expressing cells increased the number of invading vessel-like structures in a time-dependent manner (Figure 3a and 3b). To determine whether persistence of the cells contributes to angiogenesis 7 days after infusion of the cells, we started GV infusion at day 7 and documented vessel growth at day 14 (Figure 3a and 3b). As shown in Figure 3b, GV treatment at day 7 significantly reduced Matrigel plaque vascularization in mice treated with TK cells, but not in mice receiving GFP cells. Similarly, in TK cell–infused mice, the number of lectin-positive cells was decreased by GV treatment to 38.9±1.4% (P<0.05). However, only a partial inhibition by GV-induced cell depletion was observed and the vascularization of plaques from mice receiving TK cells and GV was significantly higher compared with the PBS controls group (Figure 3b), indicating that in this angiogenesis model vascularization is mediated by both immediate paracrine effects on host-derived endothelial cells and physical incorporation of the infused cells.
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To additionally determine the role of physical incorporation and persistence of infused cells in blood flow recovery after ischemia, we used a hindlimb ischemia model (Figure 4a and 4b). Infusion of TK-transduced cells time-dependently improved blood flow recovery after ischemia (Figure 4b). At day 7 after infusion of TK cells, mice were randomized to receive either GV or PBS and laser Doppler-derived blood flow was analyzed at 14 and 21 days (Figure 4b). The TK cell–treated mice receiving PBS showed a further increase in blood flow recovery (Figure 4b, blue line). However, in the TK cell–treated mice receiving GV, perfusion did not further increase and remained at the levels detected at day 7 (Figure 4b, red line). Consistently, capillary density was significantly reduced by GV treatment in TK cell–treated mice (Figure 4c and 4d). Moreover, 3 days after injection of GV, a reduced perfusion was detected by MRI in TK cell–treated mice compared with TK cell–treated mice, which received PBS (supplemental Figure V). However, the extension of the observation period revealed a recovery of perfusion at day 21 in the GV-treated group, indicating that ablation of cells does not irreversibly affect the recovery after ischemia. As a control, we infused TK-expressing macrophages. Intravenous infusion of macrophages did not augment laser Doppler–derived blood flow compared with PBS and was not affected by GV treatment (data not shown).
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Depletion of Infused Cells Induce Vascular Leakage
Previous studies documented the incorporation of transplanted human EPCs in 10 to 20% of the capillaries in the ischemic hindlimb muscle.16,18 Therefore, we investigated whether ablation of the TK-transduced cells might induce capillary damage and leakage leading to edema formation and transient perfusion defects in the hindlimb ischemia model. Injections of Evans blue in TK cell–treated mice 3 days after onset of GV treatment (Figure 4e; GV treatment started at day 7) revealed multiple spots of Evans blue staining in the thigh muscle of GV-injected mice (305±78% increase quantified by dye extraction of muscle tissue compared with PBS controls, P<0.05), whereas no staining was evident in PBS-injected mice (Figure 4e), suggesting that ablation of the previously administered cells by GV injection induces vascular damage and increases permeability.
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Discussion |
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Our data demonstrate that ablation of transplanted cells 1 week after infusion reduced vessel growth and perfusion recovery in 2 different models of neovascularization. Moreover, ablation of infused cells 2 weeks after administration induced a rapid decline of ejection fraction and capillary density in a model of acute myocardial infarction. These data demonstrate that infused cells are physically incorporated into host tissue and persist weeks after administration. Thus, it appears unlikely that the beneficial effects of cell therapy using EPCs can be solely explained by short-time paracrine effects. However, the present study does not exclude that the incorporated cells may provide long term paracrine effects. Of note, the functional consequences of cell depletion were different in the animal models used. In the angiogenesis Matrigel plug assay, vessel growth was only partially inhibited, indicating that a paracrine mechanism promoting host-derived angiogenesis within the first days after infusion significantly contributed to the vascularization of the plugs. After hindlimb ischemia, a profound increase in vascular leakage and decreased perfusion was detected after cell ablation. However, this defect was transient and mice treated with GV partially recovered at day 21, indicating that host-derived endogenous repair compensated for the loss of cells at later time points. The extent to which early paracrine mechanisms may have contributed to the rapid recovery detected later is difficult to evaluate.
In contrast to the partial inhibitory or transient effects detected in the Matrigel and hindlimb ischemia model, depletion of the cells after acute myocardial infarction induced a profound and sustained deterioration of cardiac function and capillary density was still significantly reduced in the GV treated group at 4 weeks. These findings might be explained by the crucial role of angiogenesis in the heart. Two recently published experimental models demonstrated that disruption of angiogenesis in the heart induced a decline in heart function,34 resulting in heart failure during pressure overload or induction of hypertrophy.35 In addition, the induction of cell death in cardiac myocytes derived from the injected cells might have contributed to a reduction of heart function. However, although we detected human cardiac myocytes, the number was below 0.1%, suggesting that this is unlikely to account for the deterioration of function. However, we cannot exclude that the activated prodrug GV can be transported by gap junctions to physically connected neighboring cells,36 although we excluded unspecific cytotoxicity of GV in vivo (supplemental Figure VI) and "kiss of death" effects imposed by TK-transduced cells on neighboring endothelial cells in culture in vitro. Because cultivated vascular progenitor cells were previously shown to connect to cardiac myocytes via gap junctions in vitro,20 and transplanted c-kit+ bone marrow–derived cells were shown to form interactions with host cells via gap junctions,37 such bystander effects might have amplified the detrimental effects caused by depletion of TK-expressing cells on cardiac function in the myocardial infarction model. Nevertheless, even if bystander effects on physically connected neighboring cells might have contributed to the observed abrogation of neovascularization recovery and deterioration of cardiac function following ablation of the administered cells, such mechanisms would require physical incorporation of the cells into the host tissue. Thus, sustained persistence of the administered cells in the host tissue contributes to the beneficial effects of cell therapy on neovascularization and recovery of cardiac function. The contribution of specific cell lineages to the maintenance of functional improvement warrants further investigations.
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Acknowledgments |
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We thank Tino Roexe and Ariane Fischer for expert technical assistance.
Sources of Funding
This study was supported by Deutsche Forschungsgemeinschaft grants Di600/6-3 (to S.D.) and TR-SFB23 (to F.K.) and Excellence Cluster Cardiopulmonary Systems grant Exc 147/1.
Disclosures
A.M.Z. and S.D. are cofounders and advisors to t2cure GmbH.
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Footnotes |
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*Both authors contributed equally to this work.
Original received May 31, 2008; revision received September 30, 2008; accepted October 6, 2008.
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References |
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1. Schachinger V, Erbs S, Elsasser A, Haberbosch W, Hambrecht R, Holschermann H, Yu J, Corti R, Mathey DG, Hamm CW, Suselbeck T, Assmus B, Tonn T, Dimmeler S, Zeiher AM. Intracoronary bone marrow-derived progenitor cells in acute myocardial infarction. N Engl J Med. 2006; 355: 1210–1221.
2. Janssens S, Dubois C, Bogaert J, Theunissen K, Deroose C, Desmet W, Kalantzi M, Herbots L, Sinnaeve P, Dens J, Maertens J, Rademakers F, Dymarkowski S, Gheysens O, Van Cleemput J, Bormans G, Nuyts J, Belmans A, Mortelmans L, Boogaerts M, Van de Werf F. Autologous bone marrow-derived stem-cell transfer in patients with ST-segment elevation myocardial infarction: double-blind, randomised controlled trial. Lancet. 2006; 367: 113–121.[CrossRef][Medline] [Order article via Infotrieve]
3. Wollert KC, Meyer GP, Lotz J, Ringes-Lichtenberg S, Lippolt P, Breidenbach C, Fichtner S, Korte T, Hornig B, Messinger D, Arseniev L, Hertenstein B, Ganser A, Drexler H. Intracoronary autologous bone-marrow cell transfer after myocardial infarction: the BOOST randomised controlled clinical trial. Lancet. 2004; 364: 141–148.[CrossRef][Medline] [Order article via Infotrieve]
4. Lipinski MJ, Biondi-Zoccai GG, Abbate A, Khianey R, Sheiban I, Bartunek J, Vanderheyden M, Kim HS, Kang HJ, Strauer BE, Vetrovec GW. Impact of intracoronary cell therapy on left ventricular function in the setting of acute myocardial infarction: a collaborative systematic review and meta-analysis of controlled clinical trials. J Am Coll Cardiol. 2007; 50: 1761–1767.
5. Asahara T, Murohara T, Sullivan A, Silver M, van der Zee R, Li T, Witzenbichler B, Schatteman G, Isner JM. Isolation of putative progenitor endothelial cells for angiogenesis. Science. 1997; 275: 964–967.
6. Shi Q, Rafii S, Wu MH, Wijelath ES, Yu C, Ishida A, Fujita Y, Kothari S, Mohle R, Sauvage LR, Moore MA, Storb RF, Hammond WP. Evidence for circulating bone marrow-derived endothelial cells. Blood. 1998; 92: 362–367.
7. Bailey AS, Jiang S, Afentoulis M, Baumann CI, Schroeder DA, Olson SB, Wong MH, Fleming WH. Transplanted adult hematopoietic stems cells differentiate into functional endothelial cells. Blood. 2004; 103: 13–19.
8. Grant MB, May WS, Caballero S, Brown GA, Guthrie SM, Mames RN, Byrne BJ, Vaught T, Spoerri PE, Peck AB, Scott EW. Adult hematopoietic stem cells provide functional hemangioblast activity during retinal neovascularization. Nat Med. 2002; 8: 607–612.[CrossRef][Medline] [Order article via Infotrieve]
9. Pelosi E, Valtieri M, Coppola S, Botta R, Gabbianelli M, Lulli V, Marziali G, Masella B, Muller R, Sgadari C, Testa U, Bonanno G, Peschle C. Identification of the hemangioblast in postnatal life. Blood. 2002; 100: 3203–3208.
10. Thiele J, Varus E, Wickenhauser C, Kvasnicka HM, Lorenzen J, Gramley F, Metz KA, Rivero F, Beelen DW. Mixed chimerism of cardiomyocytes and vessels after allogeneic bone marrow and stem-cell transplantation in comparison with cardiac allografts. Transplantation. 2004; 77: 1902–1905.[CrossRef][Medline] [Order article via Infotrieve]
11. Minami E, Laflamme MA, Saffitz JE, Murry CE. Extracardiac progenitor cells repopulate most major cell types in the transplanted human heart. Circulation. 2005; 112: 2951–2958.
12. Hur J, Yoon CH, Kim HS, Choi JH, Kang HJ, Hwang KK, Oh BH, Lee MM, Park YB. Characterization of two types of endothelial progenitor cells and their different contributions to neovasculogenesis. Arterioscler Thromb Vasc Biol. 2004; 24: 288–293.
13. Urbich C, Heeschen C, Aicher A, Dernbach E, Zeiher AM, Dimmeler S. Relevance of monocytic features for neovascularization capacity of circulating endothelial progenitor cells. Circulation. 2003; 108: 2511–2516.
14. Yoder MC, Mead LE, Prater D, Krier TR, Mroueh KN, Li F, Krasich R, Temm CJ, Prchal JT, Ingram DA. Re-defining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals. Blood. 2007; 109: 1801–1809.
15. Kalka C, Masuda H, Takahashi T, Kalka-Moll WM, Silver M, Kearney M, Li T, Isner JM, Asahara T. Transplantation of ex vivo expanded endothelial progenitor cells for therapeutic neovascularization. Proc Natl Acad Sci U S A. 2000; 97: 3422–3427.
16. Urbich C, Heeschen C, Aicher A, Sasaki K, Bruhl T, Farhadi MR, Vajkoczy P, Hofmann WK, Peters C, Pennacchio LA, Abolmaali ND, Chavakis E, Reinheckel T, Zeiher AM, Dimmeler S. Cathepsin L is required for endothelial progenitor cellinduced neovascularization. Nat Med. 2005; 11: 206–213.[CrossRef][Medline] [Order article via Infotrieve]
17. Kawamoto A, Tkebuchava T, Yamaguchi J, Nishimura H, Yoon YS, Milliken C, Uchida S, Masuo O, Iwaguro H, Ma H, Hanley A, Silver M, Kearney M, Losordo DW, Isner JM, Asahara T. Intramyocardial transplantation of autologous endothelial progenitor cells for therapeutic neovascularization of myocardial ischemia. Circulation. 2003; 107: 461–468.
18. Aicher A, Heeschen C, Mildner-Rihm C, Urbich C, Ihling C, Technau-Ihling K, Zeiher AM, Dimmeler S. Essential role of endothelial nitric oxide synthase for mobilization of stem and progenitor cells. Nat Med. 2003; 9: 1370–1376.[CrossRef][Medline] [Order article via Infotrieve]
19. Orlic D, Kajstura J, Chimenti S, Jakoniuk I, Anderson SM, Li B, Pickel J, McKay R, Nadal-Ginard B, Bodine DM, Leri A, Anversa P. Bone marrow cells regenerate infarcted myocardium. Nature. 2001; 410: 701–705.[CrossRef][Medline] [Order article via Infotrieve]
20. Badorff C, Brandes RP, Popp R, Rupp S, Urbich C, Aicher A, Fleming I, Busse R, Zeiher AM, Dimmeler S. Transdifferentiation of blood-derived human adult endothelial progenitor cells into functionally active cardiomyocytes. Circulation. 2003; 107: 1024–1032.
21. Murry CE, Soonpaa MH, Reinecke H, Nakajima H, Nakajima HO, Rubart M, Pasumarthi KB, Virag JI, Bartelmez SH, Poppa V, Bradford G, Dowell JD, Williams DA, Field LJ. Haematopoietic stem cells do not transdifferentiate into cardiac myocytes in myocardial infarcts. Nature. 2004; 428: 664–668.[CrossRef][Medline] [Order article via Infotrieve]
22. Grunewald M, Avraham I, Dor Y, Bachar-Lustig E, Itin A, Yung S, Chimenti S, Landsman L, Abramovitch R, Keshet E. VEGF-induced adult neovascularization: recruitment, retention, and role of accessory cells. Cell. 2006; 124: 175–189.[CrossRef][Medline] [Order article via Infotrieve]
23. Rehman J, Li J, Orschell CM, March KL. Peripheral blood "endothelial progenitor cells" are derived from monocyte/macrophages and secrete angiogenic growth factors. Circulation. 2003; 107: 1164–1169.
24. Urbich C, Aicher A, Heeschen C, Dernbach E, Hofmann WK, Zeiher AM, Dimmeler S. Soluble factors released by endothelial progenitor cells promote migration of endothelial cells and cardiac resident progenitor cells. J Mol Cell Cardiol. 2005; 39: 733–742.[CrossRef][Medline] [Order article via Infotrieve]
25. Heil M, Ziegelhoeffer T, Mees B, Schaper W. A different outlook on the role of bone marrow stem cells in vascular growth: bone marrow delivers software not hardware. Circ Res. 2004; 94: 573–574.
26. Gnecchi M, He H, Liang OD, Melo LG, Morello F, Mu H, Noiseux N, Zhang L, Pratt RE, Ingwall JS, Dzau VJ. Paracrine action accounts for marked protection of ischemic heart by Akt-modified mesenchymal stem cells. Nat Med. 2005; 11: 367–368.[CrossRef][Medline] [Order article via Infotrieve]
27. Cho HJ, Lee N, Lee JY, Choi YJ, Ii M, Wecker A, Jeong JO, Curry C, Qin G, Yoon YS. Role of host tissues for sustained humoral effects after endothelial progenitor cell transplantation into the ischemic heart. J Exp Med. 2007; 204: 3257–3269.
28. De Palma M, Venneri MA, Roca C, Naldini L. Targeting exogenous genes to tumor angiogenesis by transplantation of genetically modified hematopoietic stem cells. Nat Med. 2003; 9: 789–795.[CrossRef][Medline] [Order article via Infotrieve]
29. Vasa M, Fichtlscherer S, Adler K, Aicher A, Martin H, Zeiher AM, Dimmeler S. Increase in circulating endothelial progenitor cells by statin therapy in patients with stable coronary artery disease. Circulation. 2001; 103: 2885–2890.
30. Scherr M, Battmer K, Blomer U, Schiedlmeier B, Ganser A, Grez M, Eder M. Lentiviral gene transfer into peripheral blood-derived CD34+ NOD/SCIDrepopulating cells. Blood. 2002; 99: 709–712.
31. Zufferey R, Nagy D, Mandel RJ, Naldini L, Trono D. Multiply attenuated lentiviral vector achieves efficient gene delivery in vivo. Nat Biotechnol. 1997; 15: 871–875[CrossRef][Medline] [Order article via Infotrieve]
32. Larson AC, White RD, Laub G, McVeigh ER, Li D, Simonetti OP. Self-gated cardiac cine MRI. Magn Reson Med. 2004; 51: 93–102.[CrossRef][Medline] [Order article via Infotrieve]
33. Vasa M, Fichtlscherer S, Aicher A, Adler K, Urbich C, Martin H, Zeiher AM, Dimmeler S. Number and migratory activity of circulating endothelial progenitor cells inversely correlate with risk factors for coronary artery disease. Circ Res. 2001; 89: e1–e7.[CrossRef][Medline] [Order article via Infotrieve]
34. Shiojima I, Sato K, Izumiya Y, Schiekofer S, Ito M, Liao R, Colucci WS, Walsh K. Disruption of coordinated cardiac hypertrophy and angiogenesis contributes to the transition to heart failure. J Clin Invest. 2005; 115: 2108–2118.[CrossRef][Medline] [Order article via Infotrieve]
35. Sano M, Minamino T, Toko H, Miyauchi H, Orimo M, Qin Y, Akazawa H, Tateno K, Kayama Y, Harada M, Shimizu I, Asahara T, Hamada H, Tomita S, Molkentin JD, Zou Y, Komuro I. p53-induced inhibition of Hif-1 causes cardiac dysfunction during pressure overload. Nature. 2007; 446: 444–448.[CrossRef][Medline] [Order article via Infotrieve]
36. Fick J, Barker FG, II, Dazin P, Westphale EM, Beyer EC, Israel MA. The extent of heterocellular communication mediated by gap junctions is predictive of bystander tumor cytotoxicity in vitro. Proc Natl Acad Sci U S A. 1995; 92: 11071–11075.
37. Rota M, Kajstura J, Hosoda T, Bearzi C, Vitale S, Esposito G, Iaffaldano G, Padin Iruegas ME, Gonzalez A, Rizzi R, Small N, Muraski J, Alvarez R, Chen X, Urbanek K, Bolli R, Houser SR, Leri A, Sussman MA, Anversa P. Bone marrow cells adopt the cardiomyogenic fate in vivo. Proc Natl Acad Sci U S A. 2007; 104: 17783–17788.
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