Endothelial Progenitor Cells at Work
http://www.100md.com
《动脉硬化血栓血管生物学》
From Molecular Cardiology, Department of Internal Medicine IV, University of Frankfurt, Germany.
Tissue replacement in the adult organism by cell-specific differentiation of autologous stem/progenitor cells has evolved as a fascinating concept in stem cell biology. After organ damage, bone marrow–derived circulating or tissue-resident progenitor cells are thought to differentiate toward the type of cell needed for repair. According to this concept, maturation of these cells would be expected to result at best in a perfect morphological and functional replacement of the injured tissue. However, in this issue of Arteriosclerosis, Thrombosis, and Vascular Biology, He et al show that endothelial progenitor cells (EPCs) are more than just as capable of in vitro angiogenic tube formation as mature endothelial cells (ECs), but are truly advantageous when it comes to stress tolerance.1
See page 2021
EPCs were originally characterized as cells that are mobilized from the bone marrow and circulate in the peripheral blood and express certain surface membrane markers including the vascular endothelial growth factor receptor (VEGF-R2) KDR and the hematopoietic progenitor cell markers CD34 and CD133.2–4 During ex vivo expansion, these cells develop morphological and functional characteristics typical for ECs, including formation of vascular-like structures in matrigel and other in vitro angiogenesis assays. Most importantly, however, transplanted EPCs exhibit an extraordinary potent in vivo capacity to improve the neovascularization of ischemic tissue in the adult organism.5 In this regard, EPCs were shown to be more effective than mature ECs in animal models of hind limb ischemia,6–8 although mature ECs are well established to exert a potent in vitro angiogenic activity. Thus, the angiogenic capacity of various cell types in ischemic tissue differs from their vascular-like structure forming activity in in vitro angiogenesis assays.
But what enables EPCs to promote postischemic vessel growth that much? To understand the advantageous profile of EPCs for fulfilling the task of neovascularization after ischemia, one has to consider the changes in the tissue environment after ischemia. After anaerobic metabolism during ischemia, the tissue pH decreases, and proteolytic enzymes are released from damaged lysosomes of necrotic cell remnants. At infarct border zones, reperfusion results in the accumulation of reactive oxygen species (ROS) and, subsequently, of oxidized metabolites. Eventually, invading neutrophils and macrophages contribute additional oxygen radicals (Figure). Thus, for the purpose of recreating a perfusion network in a postischemic environment, angiogenic precursor cells first of all have to withstand oxidative stress. Such reflections may have been made by He et al to ask the question whether the superiority of EPCs to perform regenerative neovascularization in a temporarily nonperfused tissue could be caused by an enhanced cytotoxic resistance. To test this hypothesis, they compare viability and function of umbilical vein and mature coronary ECs versus EPCs after exposure toward the naturally occurring ROS-generating cytokine tumor necrosis factor (TNF) and a synthetic superoxide generator. The results are astonishing: EPCs display a unique robustness against ROS-driven cytotoxicity. Not only is cell viability preserved, but also the angiogenic capacities of EPCs are fully protected against the ROS assault. When tracking down the pathways of antioxidant resistance signaling, He et al come across the enzyme family of superoxide dismutases (SOD). Among those, they identify manganese SOD (MnSOD) as the mediator of survival, which is expressed at highest levels in EPCs compared with mature ECs. The protective relevance of this enzyme is demonstrated by the capacity of adenoviral overexpression of MnSOD to largely enhance the resistance of mature ECs toward superoxide generation close to the levels of EPCs.
Endothelial progenitor cells (EPCs) contribute to postischemic neovascularization in the adult organism. Ischemic tissue and ischemia/reperfusion–related injury provide cytotoxic stresses (left), which may compromise EPC function and survival. To preserve the neovascularizing capacities of EPCs in this environment, EPCs are equipped with a protective set of genes including MnSOD (right).
These data give new insight into the functional capacities of EPCs to protect themselves against the rough conditions under which they have to work to recreate a capillary network (Figure). Indeed, expression of MnSOD at high levels may maintain EPC survival and thus explain their extraordinary capacities for postischemic neovascularization. However, 2 recent studies additionally suggested that an increased resistance against stress could exemplify a rather general stem cell feature beyond the protection against apoptosis. Transcriptional profiling of mouse embryonic as well as adult neural and hematopoietic stem cells revealed a characteristic pattern of so-called "stemness genes," the expression of which is typical for these cells and distinguishes them from mature cells.9,10 Among other gene families, a set of stress response genes involved in the redox balance were identified to be a characteristic feature of stem cells, proposing that one essential attribute of "stemness" includes the high resistance to stress.9,10 These descriptive studies, however, did not address what is functionally controlled by redox regulatory enzymes in stem cells. Observations in neural progenitor cells have shown that factors promoting self-renewal cause these cells to enter a more reduced redox state, whereas exposure to signaling molecules that promote differentiation leads to an excess of oxidation in these progenitors.11 Thus, ROS may regulate the balance between self-renewal and differentiation. At the same time, maturation processes coincide with changes in the redox balance,11 suggesting that antioxidant enzymes could play a major role for the preservation of "stemness." According to this hypothesis, one may speculate that the increased expression of MnSOD in EPCs compared with mature ECs reported by He et al actively maintains EPCs in an undifferentiated, self-renewing state to allow for progenitor cell expansion at sites of repair. Additionally, other functional capacities of EPCs such as migration, which determines the functional improvement of patients after cell transplantation,12 might be critically influenced by the redox equilibrium.13
Although He et al elegantly demonstrate that MnSOD expression is sufficient for EC protection from cell death, it remains to be shown whether EPC survival actually depends on the presence of MnSOD, or whether there are redundant tools against ROS available. Interestingly, while the homozygous MnSOD-deficient mice die from dilated cardiomyopathy at postnatal day 10,14 even the heterozygous MnSOD-deficient mice experience increased oxidative stress in various tissues.15 In line with the importance of MnSOD in the in vivo gene deficiency model, Dernbach et al demonstrated that survival as well as the migratory capacity of EPCs, an important functional property of EPCs required for capillary sprouting, is reduced after genetic knock-down by transient transfection of RNA interference oligonucleotides against MnSOD.13 However, the results of Dernbach et al also show that only the combined inhibition of catalase, MnSOD, and glutathione peroxidase significantly decreases EPC survival and migration in the presence of hydrogen peroxide.13 From these data, it appears likely that redundant antioxidant defense strategies warrant maintenance of the equilibrium and EPC viability.
Thus, the expression of MnSOD is essential for EPC function. But are all EPCs equally equipped with enough MnSOD to protect themselves against untimely differentiation and cell death? He et al made use of EPC preparations from healthy human donors. However, it is exactly the functional defectiveness of EPCs that could underlie insufficient postischemic regeneration of ischemic tissue in patients with coronary artery disease.16–19 Therefore, based on the data of He et al, it will be mandatory to compare the expression of MnSOD between EPCs derived from healthy donors versus patients with increased cardiovascular risk, manifest coronary artery disease, and/or ischemic cardiomyopathy. Preliminary data indeed indicate that the expression of MnSOD is reduced in EPCs of patients with coronary artery disease (C.U., unpublished data, 2004). If the regenerative potential of EPCs in these patients essentially depends on the expression and activity of MnSOD, one could envision to overexpress MnSOD therapeutically in these cells to improve survival and enhance their regenerative potential for autologous transplantation.
References
He T, Peterson TE, Holmuhamedov EL, Terzic A, Caplice NM, Oberley LW, Katusic ZS. Human endothelial progenitor cells tolerate oxidative stress caused by intrinsically high expression of manganese superoxide dismutase. Arterioscler Thromb Vasc Biol. 2004; 24: 2021–2027.
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.
Peichev M, Naiyer AJ, Pereira D, Zhu Z, Lane WJ, Williams M, Oz MC, Hicklin DJ, Witte L, Moore MA, Rafii S. Expression of VEGFR-2 and AC133 by circulating human CD34(+) cells identifies a population of functional endothelial precursors. Blood. 2000; 95: 952–958.
Urbich C, Dimmeler S. Endothelial progenitor cells: characterization and role in vascular biology. Circ Res. 2004; 95: 343–353.
Murohara T, Ikeda H, Duan J, Shintani S, Sasaki K, Eguchi H, Onitsuka I, Matsui K, Imaizumi T. Transplanted cord blood-derived endothelial precursor cells augment postnatal neovascularization. J Clin Invest. 2000; 105: 1527–1536.
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.
Kocher AA, Schuster MD, Szabolcs MJ, Takuma S, Burkhoff D, Wang J, Homma S, Edwards NM, Itescu S. Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function. Nat Med. 2001; 7: 430–436.
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.
Ramalho-Santos M, Yoon S, Matsuzaki Y, Mulligan RC, Melton DA. "Stemness: " transcriptional profiling of embryonic and adult stem cells. Science. 2002; 298: 597–600.
Ivanova NB, Dimos JT, Schaniel C, Hackney JA, Moore KA, Lemischka IR. A stem cell molecular signature. Science. 2002; 298: 601–604.
Noble M, Smith J, Power J, Mayer-Proschel M. Redox state as a central modulator of precursor cell function. Ann N Y Acad Sci. 2003; 991: 251–271.
Britten MB, Abolmaali ND, Assmus B, Lehmann R, Honold J, Schmitt J, Vogl TJ, Martin H, Schachinger V, Dimmeler S, Zeiher AM. Infarct remodeling after intracoronary progenitor cell treatment in patients with acute myocardial infarction (TOPCARE-AMI): mechanistic insights from serial contrast-enhanced magnetic resonance imaging. Circulation. 2003; 108: 2212–2218.
Dernbach E, Urbich C, Brandes RP, Hofmann WK, Zeiher AM, Dimmeler S. Anti-oxidative stress–associated genes in circulating progenitor cells: evidence for enhanced resistance against oxidative stress. Blood. 2004;May 25 .
Li Y, Huang TT, Carlson EJ, Melov S, Ursell PC, Olson JL, Noble LJ, Yoshimura MP, Berger C, Chan PH, et al. Dilated cardiomyopathy and neonatal lethality in mutant mice lacking manganese superoxide dismutase. Nat Genet. 1995; 11: 376–381.
Van Remmen H, Salvador C, Yang H, Huang TT, Epstein CJ, Richardson A. Characterization of the antioxidant status of the heterozygous manganese superoxide dismutase knockout mouse. Arch Biochem Biophys. 1999; 363: 91–97.
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.
Hill JM, Zalos G, Halcox JP, Schenke WH, Waclawiw MA, Quyyumi AA, Finkel T. Circulating endothelial progenitor cells, vascular function, and cardiovascular risk. N Engl J Med. 2003; 348: 593–600.
Rauscher FM, Goldschmidt-Clermont PJ, Davis BH, Wang T, Gregg D, Ramaswami P, Pippen AM, Annex BH, Dong C, Taylor DA. Aging, progenitor cell exhaustion, and atherosclerosis. Circulation. 2003; 108: 457–463.
Heeschen C, Lehmann R, Honold J, Assmus B, Aicher A, Walter DH, Martin H, Zeiher AM, Dimmeler S. Profoundly reduced neovascularization capacity of bone marrow mononuclear cells derived from patients with chronic ischemic heart disease. Circulation. 2004; 109: 1615–1622.(Lothar R?ssig; Carmen Urb)
Tissue replacement in the adult organism by cell-specific differentiation of autologous stem/progenitor cells has evolved as a fascinating concept in stem cell biology. After organ damage, bone marrow–derived circulating or tissue-resident progenitor cells are thought to differentiate toward the type of cell needed for repair. According to this concept, maturation of these cells would be expected to result at best in a perfect morphological and functional replacement of the injured tissue. However, in this issue of Arteriosclerosis, Thrombosis, and Vascular Biology, He et al show that endothelial progenitor cells (EPCs) are more than just as capable of in vitro angiogenic tube formation as mature endothelial cells (ECs), but are truly advantageous when it comes to stress tolerance.1
See page 2021
EPCs were originally characterized as cells that are mobilized from the bone marrow and circulate in the peripheral blood and express certain surface membrane markers including the vascular endothelial growth factor receptor (VEGF-R2) KDR and the hematopoietic progenitor cell markers CD34 and CD133.2–4 During ex vivo expansion, these cells develop morphological and functional characteristics typical for ECs, including formation of vascular-like structures in matrigel and other in vitro angiogenesis assays. Most importantly, however, transplanted EPCs exhibit an extraordinary potent in vivo capacity to improve the neovascularization of ischemic tissue in the adult organism.5 In this regard, EPCs were shown to be more effective than mature ECs in animal models of hind limb ischemia,6–8 although mature ECs are well established to exert a potent in vitro angiogenic activity. Thus, the angiogenic capacity of various cell types in ischemic tissue differs from their vascular-like structure forming activity in in vitro angiogenesis assays.
But what enables EPCs to promote postischemic vessel growth that much? To understand the advantageous profile of EPCs for fulfilling the task of neovascularization after ischemia, one has to consider the changes in the tissue environment after ischemia. After anaerobic metabolism during ischemia, the tissue pH decreases, and proteolytic enzymes are released from damaged lysosomes of necrotic cell remnants. At infarct border zones, reperfusion results in the accumulation of reactive oxygen species (ROS) and, subsequently, of oxidized metabolites. Eventually, invading neutrophils and macrophages contribute additional oxygen radicals (Figure). Thus, for the purpose of recreating a perfusion network in a postischemic environment, angiogenic precursor cells first of all have to withstand oxidative stress. Such reflections may have been made by He et al to ask the question whether the superiority of EPCs to perform regenerative neovascularization in a temporarily nonperfused tissue could be caused by an enhanced cytotoxic resistance. To test this hypothesis, they compare viability and function of umbilical vein and mature coronary ECs versus EPCs after exposure toward the naturally occurring ROS-generating cytokine tumor necrosis factor (TNF) and a synthetic superoxide generator. The results are astonishing: EPCs display a unique robustness against ROS-driven cytotoxicity. Not only is cell viability preserved, but also the angiogenic capacities of EPCs are fully protected against the ROS assault. When tracking down the pathways of antioxidant resistance signaling, He et al come across the enzyme family of superoxide dismutases (SOD). Among those, they identify manganese SOD (MnSOD) as the mediator of survival, which is expressed at highest levels in EPCs compared with mature ECs. The protective relevance of this enzyme is demonstrated by the capacity of adenoviral overexpression of MnSOD to largely enhance the resistance of mature ECs toward superoxide generation close to the levels of EPCs.
Endothelial progenitor cells (EPCs) contribute to postischemic neovascularization in the adult organism. Ischemic tissue and ischemia/reperfusion–related injury provide cytotoxic stresses (left), which may compromise EPC function and survival. To preserve the neovascularizing capacities of EPCs in this environment, EPCs are equipped with a protective set of genes including MnSOD (right).
These data give new insight into the functional capacities of EPCs to protect themselves against the rough conditions under which they have to work to recreate a capillary network (Figure). Indeed, expression of MnSOD at high levels may maintain EPC survival and thus explain their extraordinary capacities for postischemic neovascularization. However, 2 recent studies additionally suggested that an increased resistance against stress could exemplify a rather general stem cell feature beyond the protection against apoptosis. Transcriptional profiling of mouse embryonic as well as adult neural and hematopoietic stem cells revealed a characteristic pattern of so-called "stemness genes," the expression of which is typical for these cells and distinguishes them from mature cells.9,10 Among other gene families, a set of stress response genes involved in the redox balance were identified to be a characteristic feature of stem cells, proposing that one essential attribute of "stemness" includes the high resistance to stress.9,10 These descriptive studies, however, did not address what is functionally controlled by redox regulatory enzymes in stem cells. Observations in neural progenitor cells have shown that factors promoting self-renewal cause these cells to enter a more reduced redox state, whereas exposure to signaling molecules that promote differentiation leads to an excess of oxidation in these progenitors.11 Thus, ROS may regulate the balance between self-renewal and differentiation. At the same time, maturation processes coincide with changes in the redox balance,11 suggesting that antioxidant enzymes could play a major role for the preservation of "stemness." According to this hypothesis, one may speculate that the increased expression of MnSOD in EPCs compared with mature ECs reported by He et al actively maintains EPCs in an undifferentiated, self-renewing state to allow for progenitor cell expansion at sites of repair. Additionally, other functional capacities of EPCs such as migration, which determines the functional improvement of patients after cell transplantation,12 might be critically influenced by the redox equilibrium.13
Although He et al elegantly demonstrate that MnSOD expression is sufficient for EC protection from cell death, it remains to be shown whether EPC survival actually depends on the presence of MnSOD, or whether there are redundant tools against ROS available. Interestingly, while the homozygous MnSOD-deficient mice die from dilated cardiomyopathy at postnatal day 10,14 even the heterozygous MnSOD-deficient mice experience increased oxidative stress in various tissues.15 In line with the importance of MnSOD in the in vivo gene deficiency model, Dernbach et al demonstrated that survival as well as the migratory capacity of EPCs, an important functional property of EPCs required for capillary sprouting, is reduced after genetic knock-down by transient transfection of RNA interference oligonucleotides against MnSOD.13 However, the results of Dernbach et al also show that only the combined inhibition of catalase, MnSOD, and glutathione peroxidase significantly decreases EPC survival and migration in the presence of hydrogen peroxide.13 From these data, it appears likely that redundant antioxidant defense strategies warrant maintenance of the equilibrium and EPC viability.
Thus, the expression of MnSOD is essential for EPC function. But are all EPCs equally equipped with enough MnSOD to protect themselves against untimely differentiation and cell death? He et al made use of EPC preparations from healthy human donors. However, it is exactly the functional defectiveness of EPCs that could underlie insufficient postischemic regeneration of ischemic tissue in patients with coronary artery disease.16–19 Therefore, based on the data of He et al, it will be mandatory to compare the expression of MnSOD between EPCs derived from healthy donors versus patients with increased cardiovascular risk, manifest coronary artery disease, and/or ischemic cardiomyopathy. Preliminary data indeed indicate that the expression of MnSOD is reduced in EPCs of patients with coronary artery disease (C.U., unpublished data, 2004). If the regenerative potential of EPCs in these patients essentially depends on the expression and activity of MnSOD, one could envision to overexpress MnSOD therapeutically in these cells to improve survival and enhance their regenerative potential for autologous transplantation.
References
He T, Peterson TE, Holmuhamedov EL, Terzic A, Caplice NM, Oberley LW, Katusic ZS. Human endothelial progenitor cells tolerate oxidative stress caused by intrinsically high expression of manganese superoxide dismutase. Arterioscler Thromb Vasc Biol. 2004; 24: 2021–2027.
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.
Peichev M, Naiyer AJ, Pereira D, Zhu Z, Lane WJ, Williams M, Oz MC, Hicklin DJ, Witte L, Moore MA, Rafii S. Expression of VEGFR-2 and AC133 by circulating human CD34(+) cells identifies a population of functional endothelial precursors. Blood. 2000; 95: 952–958.
Urbich C, Dimmeler S. Endothelial progenitor cells: characterization and role in vascular biology. Circ Res. 2004; 95: 343–353.
Murohara T, Ikeda H, Duan J, Shintani S, Sasaki K, Eguchi H, Onitsuka I, Matsui K, Imaizumi T. Transplanted cord blood-derived endothelial precursor cells augment postnatal neovascularization. J Clin Invest. 2000; 105: 1527–1536.
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.
Kocher AA, Schuster MD, Szabolcs MJ, Takuma S, Burkhoff D, Wang J, Homma S, Edwards NM, Itescu S. Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function. Nat Med. 2001; 7: 430–436.
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.
Ramalho-Santos M, Yoon S, Matsuzaki Y, Mulligan RC, Melton DA. "Stemness: " transcriptional profiling of embryonic and adult stem cells. Science. 2002; 298: 597–600.
Ivanova NB, Dimos JT, Schaniel C, Hackney JA, Moore KA, Lemischka IR. A stem cell molecular signature. Science. 2002; 298: 601–604.
Noble M, Smith J, Power J, Mayer-Proschel M. Redox state as a central modulator of precursor cell function. Ann N Y Acad Sci. 2003; 991: 251–271.
Britten MB, Abolmaali ND, Assmus B, Lehmann R, Honold J, Schmitt J, Vogl TJ, Martin H, Schachinger V, Dimmeler S, Zeiher AM. Infarct remodeling after intracoronary progenitor cell treatment in patients with acute myocardial infarction (TOPCARE-AMI): mechanistic insights from serial contrast-enhanced magnetic resonance imaging. Circulation. 2003; 108: 2212–2218.
Dernbach E, Urbich C, Brandes RP, Hofmann WK, Zeiher AM, Dimmeler S. Anti-oxidative stress–associated genes in circulating progenitor cells: evidence for enhanced resistance against oxidative stress. Blood. 2004;May 25 .
Li Y, Huang TT, Carlson EJ, Melov S, Ursell PC, Olson JL, Noble LJ, Yoshimura MP, Berger C, Chan PH, et al. Dilated cardiomyopathy and neonatal lethality in mutant mice lacking manganese superoxide dismutase. Nat Genet. 1995; 11: 376–381.
Van Remmen H, Salvador C, Yang H, Huang TT, Epstein CJ, Richardson A. Characterization of the antioxidant status of the heterozygous manganese superoxide dismutase knockout mouse. Arch Biochem Biophys. 1999; 363: 91–97.
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.
Hill JM, Zalos G, Halcox JP, Schenke WH, Waclawiw MA, Quyyumi AA, Finkel T. Circulating endothelial progenitor cells, vascular function, and cardiovascular risk. N Engl J Med. 2003; 348: 593–600.
Rauscher FM, Goldschmidt-Clermont PJ, Davis BH, Wang T, Gregg D, Ramaswami P, Pippen AM, Annex BH, Dong C, Taylor DA. Aging, progenitor cell exhaustion, and atherosclerosis. Circulation. 2003; 108: 457–463.
Heeschen C, Lehmann R, Honold J, Assmus B, Aicher A, Walter DH, Martin H, Zeiher AM, Dimmeler S. Profoundly reduced neovascularization capacity of bone marrow mononuclear cells derived from patients with chronic ischemic heart disease. Circulation. 2004; 109: 1615–1622.(Lothar R?ssig; Carmen Urb)