Dynamic Regulation of Vascular Myosin Light Chain (MYL9) with Injury and Aging Lina A. Shehadeh1,2,3*, Keith A. Webster2,3,4, Joshua M. Hare1,3, Roberto I. Vazquez-Padron2,3,5* 1 Department of Medicine, Division of Cardiology, University of Miami Leonard M. Miller School of Medicine, Miami, Florida, United States of America, 2 Vascular Biology Institute, University of Miami Leonard M. Miller School of Medicine, Miami, Florida, United States of America, 3 Interdisciplinary Stem Cell Institute, University of Miami Leonard M. Miller School of Medicine, Miami, Florida, United States of America, 4 Department of Pharmacology, University of Miami Leonard M. Miller School of Medicine, Miami, Florida, United States of America, 5 Department of Surgery, University of Miami Leonard M. Miller School of Medicine, Miami, Florida, United States of America Abstract Background: Aging-associated changes in the cardiovascular system increase the risk for disease development and lead to profound alterations in vascular reactivity and stiffness. Elucidating the molecular response of arteries to injury and age will help understand the exaggerated remodeling of aging vessels. Methodology/Principal Findings: We studied the gene expression profile in a model of mechanical vascular injury in the iliac artery of aging (22 months old) and young rats (4 months old). We investigated aging-related variations in gene expression at 30 min, 3 d and 7 d post injury. We found that the Myosin Light Chain gene (MYL9) was the only gene differentially expressed in the aged versus young injured arteries at all time points studied, peaking at day 3 after injury (4.6 fold upregulation (p,0.05) in the smooth muscle cell layers. We confirmed this finding on an aging aortic microarray experiment available through NCBI’s GEO database. We found that Myl9 was consistently upregulated with age in healthy rat aortas. To determine the arterial localization of Myl9 with age and injury, we performed immunohistochemistry for Myl9 in rat iliac arteries and found that in healthy and injured (30 days post injury) arteries, Myl9 expression increased with age in the endothelial layers. Conclusions/Significance: The consistent upregulation of the myosin light chain protein (Myl9) with age and injury in arterial tissue draws attention to the increased vascular permeability and to the age-caused predisposition to arterial constriction after balloon angioplasty. Citation: Shehadeh LA, Webster KA, Hare JM, Vazquez-Padron RI (2011) Dynamic Regulation of Vascular Myosin Light Chain (MYL9) with Injury and Aging. PLoS ONE 6(10): e25855. doi:10.1371/journal.pone.0025855 Editor: Marcello Rota, Brigham & Women’s Hospital - Harvard Medical School, United States of America Received May 27, 2011; Accepted September 12, 2011; Published October 7, 2011 Copyright: 2011 Shehadeh et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by awards from the American Heart Association (Scientist Development Grant 0930169N) and American Federation for Aging Research (M1001096) to LAS; and by the American Heart Association (Scientist Development Award 0535167B) and NIH-NHLBI [1K01HL096413-01] to RIVP. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: lshehadeh@med.miami.edu (LAS); RVazquez@med.miami.edu (RIV-P) Introduction Vascular diseases remain the most common cause of death in the world [1]. Aging increases the risk for hypertension, coronary artery disease, and heart failure. The aging-associated changes in the cardiovascular system lead to profound alterations in vascular reactivity and stiffness [2,3]. Aging also increases the risk for vascular proliferative diseases such atherosclerosis [4] and restenosis after percutaneous coronary interventions. In fact, experimental data suggesting that age increases the risk for vascular diseases date back to 1982. Using a balloon injury model Stemerman et al. [5] showed that VSMCs of aged rats proliferate more in response to injury than those of young animals. These results were reproduced in rabbits and primates [4,6] and extended to a model of vascular injury in mice [7]. We previously demonstrated that aged mice develop more neointima following arterial injury than their younger counterparts [7]. Based on the aforementioned observations, the aim of this study was to investigate the effects of aging on the early vascular response to injury in a model of arterial stenosis. Therefore, we studied the global gene expression profiles in healthy and injured arteries of aged and young rats at multiple time points. Moreover, we attempted to verify our results by analysis of a public dataset on gene expression of aged and young healthy aortas spanning four different ages [8]. Finally, we studied the localization of the identified contractility gene, Myl9, within the arterial wall of aged and young, healthy and injured iliac arteries. Results Myl9 is the only gene differentially expressed in all 3 different time-points after wire injury in aged versus young rats We can start to comprehend the dynamics of vascular injury only if we examine vascular remodeling with time. Therefore, we elected to study global gene expression of vascular remodeling in a 3-point time series. Looking at the global gene expression in the iliac arteries of old versus young rats 30 minutes, 3 days, or 7 days after balloon injury, we found that Myosin Light Chain, Myl9, was the only gene that overlapped as differentially expressed in all the PLoS ONE | www.plosone.org 1 October 2011 | Volume 6 | Issue 10 | e25855