doi: 10.1161/CIRCULATIONAHA.105.575688
(Circulation. 2005;112:e359-e360.)
© 2005 American Heart Association, Inc.
Correspondence |
Letter Regarding Article by Rotmans et al, "In Vivo Cell Seeding With Anti-CD34 Antibodies Successfully Accelerates Endothelialization but Stimulates Intimal Hyperplasia in Porcine Arteriovenous Expanded Polytetrafluoroethylene Grafts"
Division of Cardiovascular Diseases, Molecular Medicine Program, Mayo Clinic, Rochester, Minn
Rapid (re)endothelialization of autologous and synthetic vascular grafts is regarded as a key element in reducing the risk of acute thrombosis and, by limiting adverse vessel wall remodeling, which includes intimal hyperplasia, preserving long-term patency. Rotmans et al1 (and the accompanying editorial2) challenge this tenet of vascular biology in their report of human anti-CD34 antibody–coated polytetrafluoroethylene (PTFE) grafts in a porcine AV fistula model. Rapid "endothelialization" of grafts is described, which, rather than improving functional outcome, was associated with significantly increased intimal hyperplasia at the venous anastomosis. The authors conclude that future studies will need to address a putative deficiency in the functional capability of captured endothelial progenitor cells (EPCs), highlighting approaches based on the use of regulatory proteins or alternative cell combinations.
In our opinion, the data do not support these conclusions. The graft endoluminal cells were inadequately characterized with lectin (a surface antigen shared by several blood-borne cells, including monocytes) as the major discriminant of an endothelial cell. For instance, endothelial nitric oxide synthase would be much more specific for endothelial cell identification. Moreover, the fundamental premise of the study—that human anti-CD34 antibody has high affinity and specificity for circulating porcine EPCs—is unsound. The non–peer-reviewed citation supporting the claim of cross-reactivity does not characterize this phenomenon in any detail, either quantitatively or qualitatively.3 Extensive testing of a number of anti-CD34 antibodies in the pig by our group and others4,5 has been disappointing, with nonspecific binding and unequivocal failure to identify a subset of circulating porcine EPCs with this detection approach. Indeed, it is acknowledged in the field that a CD34 antibody that specifically identifies porcine EPCs currently does not exist. The effects observed in this study are therefore not at all surprising, with enhanced inflammatory cell recruitment to the graft most likely accounting for the significant increase in intimal hyperplasia. In this context, an isotype-matched IgG antibody–seeded graft, essential to exclude nonspecific binding of circulating cells in this model, may have been a more valuable control than the bare PTFE conduit used. Implanted grafts preseeded ex vivo with confirmed EPCs may have provided further insight into the true nature of the unexpected findings.
In summary, these data appear insufficient to provide a basis for the reevaluation of the role of endothelialization in modulating intimal hyperplasia and should not deter further study targeting progenitor-driven endothelialization of vascular devices as a worthy clinical end point.
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1. Rotmans JI, Heyligers JM, Verhagen HJ, Velema E, Nagtegaal MM, de Kleijn DP, de Groot FG, Stroes ES, Pasterkamp G. In vivo cell seeding with anti-CD34 antibodies successfully accelerates endothelialization but stimulates intimal hyperplasia in porcine arteriovenous expanded polytetrafluoroethylene grafts. Circulation. 2005; 112: 12–18.
2. Roy-Chaudhury P. Endothelial progenitor cells, neointimal hyperplasia, and hemodialysis vascular access dysfunction: novel therapies for a recalcitrant clinical problem. Circulation. 2005; 112: 3–5.
3. Kutryk MJ, Kuliszewski MA. In vivo endothelial progenitor cell seeding of stented arterial segments and vascular grafts. Circulation. 2003; 108 (suppl): IV-573. Abstract.
4. Kawamoto A, Asahara T, Losordo DW. Transplantation of endothelial progenitor cells for therapeutic neovascularization. Cardiovasc Radiat Med. 2002; 3: 221–225.[CrossRef][Medline] [Order article via Infotrieve]
5. Sun J, Sinkora J, Wertz N, Moravkova A, Butler JE. Characterization of porcine CD19 and anti-CD19 monoclonal antibodies. Mol Immunol. 2004; 41: 929–938.[CrossRef][Medline] [Order article via Infotrieve]
Laboratory of Experimental Cardiology, University Medical Center, Utrecht, The Netherlands, Department of Vascular Medicine, Academic Medical Center, Amsterdam, The Netherlands
Departments of Vascular Surgery and Hematology, University Medical Center, Utrecht, The Netherlands
Department of Vascular Surgery, University Medical Center, Utrecht, The Netherlands
Laboratory of Experimental Cardiology, University Medical Center, Utrecht, The Netherlands
Department of Vascular Medicine, Academic Medical Center, Amsterdam, The Netherlands
Department of Hematology, University Medical Center, Utrecht, The Netherlands
We thank Drs Doyle and Caplice for their interest in our article.1 As they emphasized, immunohistochemical characterization of the adhered cells in a pig model is hampered by the lack of specific antibodies for endothelial antigens in pigs. Accordingly, we discussed the scenario that we cannot completely rule out entrapment of CD34(–) cells on the anti-CD34 antibody–covered graft. However, several observations argue against inflammatory cell recruitment to the graft as the major cause of the increased intimal hyperplastic response. First, flow cytometric analysis of our anti-human CD34 antibody showed consistent cross-reactivity with porcine mononuclear cells that significantly exceeded modest nonspecific binding to an isotype-matched antibody. Second, staining with MAC387, a selective marker for granulocytes and macrophages of both human and porcine origin, did not reveal any positive cells at the luminal side of the anti-CD34–coated ePTFE graft. In this respect, it is worthwhile to reappraise the optimism with respect to the role of bone-marrow–derived cells. Recent observations have underlined that EPCs can also have adverse effects on vascular integrity. First, in the MAGIC trial, infusion of progenitor cells in coronary arteries on stenting resulted in a significant increase of in-stent restenosis.2 More recently, coating of a graft with vascular endothelial growth factor in rats resulted in increased intimal hyperplasia in the seeded grafts, despite achieving impressive graft endothelialization.3 Both studies clearly emphasize potentially adverse effects of endothelial (progenitor) cells on neointimal formation under suboptimal conditions.
We agree with Doyle and Caplice that additional experiments with implanted grafts preseeded ex vivo with carefully defined EPCs would have further strengthened our observations. In fact, these experiments have been performed previously by Griese et al.4 Ex vivo seeding with extensively characterized EPCs in rabbits resulted in a significant increase in ePTFE graft endothelialization, whereas the authors did not show a beneficial effect on intimal hyperplasia on the seeded grafts.
In summary, we concur with Doyle and Caplice that the exact mechanism of the adverse effects of anti-CD34–coated ePTFE grafts warrants further investigation. We hope that our findings provide a further stimulus, rather than an impediment, for further research on the therapeutic application of EPCs.
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This work was supported by Dutch Kidney Foundation grant CSP 6001.
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1. Rotmans JI, Heyligers JM, Verhagen HJ, Velema E, Nagtegaal MM, de Kleijn DP, de Groot FG, Stroes ES, Pasterkamp G. In vivo cell seeding with anti-CD34 antibodies successfully accelerates endothelialization but stimulates intimal hyperplasia in porcine arteriovenous expanded polytetrafluoroethylene grafts. Circulation. 2005; 112: 12–18.
2. Kang HJ, Kim HS, Zhang SY, Park KW, Cho HJ, Koo BK, Kim YJ, Soo LD, Sohn DW, Han KS, Oh BH, Lee MM, Park YB. Effects of intracoronary infusion of peripheral blood stem-cells mobilised with granulocyte-colony stimulating factor on left ventricular systolic function and restenosis after coronary stenting in myocardial infarction: the MAGIC cell randomised clinical trial. Lancet. 2004; 363: 751–756.[CrossRef][Medline] [Order article via Infotrieve]
3. Randone B, Cavallaro G, Polistena A, Cucina A, Coluccia P, Graziano P, Cavallaro A. Dual role of VEGF in pretreated experimental ePTFE arterial grafts. J Surg Res. 2005; 127: 70–79.[CrossRef][Medline] [Order article via Infotrieve]
4. Griese DP, Ehsan A, Melo LG, Kong D, Zhang L, Mann MJ, Pratt RE, Mulligan RC, Dzau VJ. Isolation and transplantation of autologous circulating endothelial cells into denuded vessels and prosthetic grafts: implications for cell-based vascular therapy. Circulation. 2003; 108: 2710–2715.
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