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Blood, 15 March 2008, Vol. 111, No. 6, pp. 3024-3033. Prepublished online as a Blood First Edition Paper on January 8, 2008; DOI 10.1182/blood-2007-06-098657. This Article Full Text (PDF) Supplemental Figure All Versions of this Article: blood-2007-06-098657v1 111/6/3024    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Garnacho, C. Articles by Muro, S. Search for Related Content PubMed PubMed Citation Articles by Garnacho, C. Articles by Muro, S. Social Bookmarking                   What's this?  Previous Article  |  Next Article  Submitted June 29, 2007 Accepted December 18, 2007 RhoA activation and actin reorganization involved in endothelial CAM-mediated endocytosis of anti-PECAM carriers: critical role for tyrosine 686 in the cytoplasmic tail of PECAM-1 Carmen Garnacho, Vladimir Shuvaev, Anu Thomas, Lindsey McKenna, Jing Sun, Michael Koval, Steven Albelda, Vladimir Muzykantov, and Silvia Muro* Department of Pharmacology, School of Medicine, University of Pennsylvania, Philadelphia, PA, United States Institute for Environmental Medicine, School of Medicine, University of Pennsylvania, Philadelphia, PA, United States Pulmonary Critical Care Division of the Dept. of Medicine, School of Medicine, University of Pennsylvania, Philadelphia, PA, United States Division of Pulmonary, Allergy & Critical Care Medicine, Dept. of Medicine, Emory University School of Medicine, Atlanta, GA, United States Targeted Therapeutics Program of the Institute for Translational Medicine & Therapeutics, School of Medicine, University of Pennsylvania, Philadelphia, PA, United States * Corresponding author; email: silvia{at}mail.med.upenn.edu. Platelet-Endothelial Cell Adhesion Molecule-1 (PECAM-1), a transmembrane glycoprotein involved in leukocyte transmigration, represents a good target for endothelial drug delivery, e.g., using antibody-directed nanocarriers (anti-PECAM/NCs). Although endothelial cells do not internalize PECAM antibodies, PECAM-1 engagement by multivalent anti-PECAM conjugates and nanocarriers causes endocytosis via non-classical CAM-mediated pathway. We found that endothelial uptake of multivalent anti-PECAM complexes is associated with PECAM-1 phosphorylation. Using model REN cells expressing a series of PECAM-1 deletion and point mutants, we found that PECAM-1 cytoplasmic domain and, more precisely, PECAM-1 tyrosine 686, is critical in mediating RhoA activation and recruitment of EGFP-RhoA to anti-PECAM/NC binding sites at the plasmalemma, actin polymerization into phalloidin-positive stress fibers, and finally CAM-endocytosis of anti-PECAM/NCs. Endothelial targeting and endocytosis of anti-PECAM/NCs was markedly efficient and did not compromise endothelial barrier function in vitro (determined by immunostaining of VE-cadherin and 125I-albumin transport across endothelial monolayers) or in vivo (determined by electron microscopy imaging of pulmonary capillaries and 125I-albumin transport from the blood into the lung tissue after intravenous injection of anti-PECAM/NCs in mice). These results reveal PECAM-1 signaling and interactions with cytoskeleton required for CAM-endocytosis, potentially useful for safe intra-endothelial drug delivery by anti-PECAM/NCs. CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

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