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Analytical Cellular Pathology
Volume 2015, Article ID 326385, 6 pages
Research Article

A Simple Method to Assess In Vivo Proliferation in Lung Vasculature with EdU: The Case of MMC-Induced PVOD in Rat

1Faculté de Médecine, Université Paris-Sud, Le Kremlin-Bicêtre, France
2AP-HP, Centre de Référence de l’Hypertension Pulmonaire Sévère, Département Hospitalo-Universitaire (DHU) Thorax Innovation, Service de Pneumologie et Réanimation Respiratoire, Hôpital de Bicêtre, Le Kremlin-Bicêtre, France
3UMRS 999, INSERM and Université Paris-Sud, Laboratoire d’Excellence (LabEx) en Recherche sur le Médicament et l’Innovation Thérapeutique (LERMIT), Centre Chirurgical Marie Lannelongue, Le Plessis-Robinson, France
4Groupe de Recherche en Hypertension Pulmonaire, Centre de Recherche de l’Institut Universitaire de Cardiologie et de Pneumologie de Québec, QC, Canada

Received 26 May 2015; Revised 26 July 2015; Accepted 28 July 2015

Academic Editor: Matthias Stope

Copyright © 2015 Antigny Fabrice et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


5-Ethynyl-2′-deoxyuridine (EdU) incorporation is becoming the gold standard method for in vitro and in vivo visualization of proliferating cells. The small size of the fluorescent azides used for detection results in a high degree of specimen penetration. It can be used to easily detect DNA replication in large tissue samples or organ explants with low proliferation and turnover of cells formerly believed to be in a “terminal” state of differentiation. Here we describe a protocol for the localization and identification of proliferating cells in quiescent or injured pulmonary vasculature, in a model of pulmonary veno-occlusive disease (PVOD). PVOD is an uncommon form of pulmonary hypertension characterized by progressive obstruction of small pulmonary veins. We previously reported that mitomycin-C (MMC) therapy is associated with PVOD in human. We demonstrated that MMC can induce PVOD in rats, which currently represents the sole animal model that recapitulates human PVOD lesions. Using the EdU assay, we demonstrated that MMC-exposed lungs displayed areas of exuberant microvascular endothelial cell proliferation which mimics pulmonary capillary hemangiomatosis, one of the pathologic hallmarks of human PVOD. In vivo pulmonary cell proliferation measurement represents an interesting methodology to investigate the potential efficacy of therapies aimed at normalizing pathologic angioproliferation.