Elevated Levels of VE-Cadherin-Positive Endothelial Microparticles in Patients With Type 2 Diabetes Mellitus and Coronary Artery Disease
Hidenobu Koga, MD*,
Seigo Sugiyama, MD, PhD*,*,
Kiyotaka Kugiyama, MD, PhD ,
Keisuke Watanabe, MD*,
Hironobu Fukushima, MD*,
Tomoko Tanaka, MD*,
Tomohiro Sakamoto, MD, PhD*,
Michihiro Yoshimura, MD, PhD*,
Hideaki Jinnouchi, MD, PhD and
Hisao Ogawa, MD, PhD*
* Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan
2nd Department of Internal Medicine, Interdisciplinary Graduate School of Medicine and Engineering University of Yamanashi, Yamanashi, Japan
Department of Jinnouchi Diabetes Center, Kumamoto, Japan
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Figure 1 (A) An immunofluorescent photomicrograph showing CD144-expression in human coronary artery endothelial cells (HCAEC). The green signal indicates fluorescein isothiocyanate (FITC)-CD144 while the blue signal indicates the nucleus (n = 4). (B) A histogram analysis of CD144-expression in HCAEC detected by FACScalibur. The thin line represents control immunoglobulin G (IgG) and the thick line represents CD144 (n = 5). (C to H) Histogram analyses of CD144-expression in microparticles derived from a variety of cells involved in atherosclerosis detected using FACScalibur. (C) HCAEC; (D) HAoSMC (human aorta smooth muscle cell); (E) platelets; (F) lymphocytes; (G) monocytes; (H) macrophages. The thin lines represent control IgG and the thick lines represent CD144 (n = 5). (I to J) Histogram analyses of CD31-expression in microparticles derived from HCAEC and platelets. (K) Representative dot-plot graphs of microparticles expressing CD31-CD42b derived from human platelets. The lower right section of graph shows CD31-positive and CD42b-negative microparticles derived from isolated human platelets.
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Figure 2 (A) A bar graph showing CD144-EMP (endothelium-derived microparticles) levels in cultured medium of HCAEC and the number of dead HCAEC after exposure for 2 h to various concentrations of H2O2. The solid bars represent CD144-EMP levels and open bars represent the number of dead HCAEC (n = 4) (B) A linear-regression graph demonstrating the significant correlation between the number of dead HCAECs and CD144-EMP levels in cultured medium after exposure to various concentrations of H2O2 (r = 0.791, p = 0.002). Abbreviations as in Figure 1.
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Figure 3 (A) Forward scatter (FSC) and side scatter (SSC) analysis of microparticles in human plasma. The section containing microparticles was gated using a size <1.5 µm in FSC-SSC scatter using standard beads (bead size = 4.2µm). (B) A histogram analysis of CD144-expression in the gated microparticles in human plasma. The thin line represents control-IgG and the thick line represents CD144. (n = 5) (C to D) Representative dot-plot graphs of microparticles expressing CD144-CD42b in human plasma in patients with or without DM. The lower right section of each graph contains EMP as CD144-positive and CD42b-negative microparticles. DM = diabetes mellitus; other abbreviations as in Figures 1 and 2.
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Figure 4 (A to D) Linear regression analysis showing the relationship between plasma levels of CD144-EMP or soluble intercellular adhesion molecule-1 (sICAM-1) and percentage change in coronary artery luminal diameter at the proximal segment of LAD in response to intracoronary acetylcholine (10 µg/min, 50 µg/min at left main trunk coronary artery [LMT]) infusion. Open circles are nondiabetic control patients and filled circles are patients with DM. (E to F) Correlation between plasma levels of CD144-EMP or sICAM-1 and percent increase from baseline values of coronary blood flow in response to intracoronary infusion of acetylcholine (10 µg/min at LMT). Open circles are nondiabetic control patients and filled circles are patients with DM. Abbreviations as in Figures 1 to 3.
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Figure 5 (A) Box-and-whisker plot showing plasma CD144-EMP values in patients with and without DM. (B) Box-and-whisker plot showing plasma CD144-EMP values in control patients, DM patients with coronary artery disease (CAD), and DM patients without CAD. In these plots, lines within boxes represent median values; the upper and lower lines of the boxes represent the 25th and 75th percentiles, respectively; and the upper and lower bars outside the boxes represent the 90th and 10th percentiles, respectively. Abbreviations as in Figures 1 to 3.
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