(Circulation. 1997;95:2441-2447.)
© 1997 American Heart Association, Inc.
Articles |
In Vivo Gene Transfection of Human Endothelial Cell Nitric Oxide Synthase in Cardiomyocytes Causes Apoptosis-Like Cell Death
Identification Using Sendai Virus–Coated Liposomes
From the Second Department of Internal Medicine, Health Service Center, University of Tokyo, Tokyo, and the Institute for Molecular and Cellular Biology (Y.K.), Osaka University, Osaka, Japan.
Correspondence and reprint requests to Professor T. Toyo-oka, The Second Department of Internal Medicine, Tokyo University Hospital, University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113, Japan.
Background Nitric oxide (NO) has various actions on the cardiovascular system, although its pathophysiological significance in myocardial cells remains obscure. The aim of the present study was to identify direct NO actions on cardiomyocytes by gene transfection in vivo using a newly developed vector under physiological conditions.
Methods and Results Liposomes containing the
ß-galactosidase (ß-gal) gene alone or with the human
endothelial cell nitric oxide synthase (ecNOS) gene
were coated with UV-inactivated Sendai virus and injected
into the left ventricular wall of rat heart in vivo.
Histological examination confirmed that the
transfection efficiency was comparable to adenovirus-mediated
transfection and that the new vector per se caused no inflammation.
ß-Gal expression was confined to cardiomyocytes between
two intercalated discs, suggesting that the transfected gene did not
permeate the discs. An immunohistochemical study showed that
cotransfection of the ecNOS gene induced massive myocardial cell
shrinkage in both transfected cells and the adjacent myocytes in a
time- and dose-dependent manner. Histochemical findings in shrunk cells
coincided with apoptosis as identified by terminal
deoxynucleotidyl transferase–mediated dUTP-biotin
nick-end labeling. Electron microscopy of the lesion revealed
myofibrillar degradation and accumulation of mitochondria but no
apoptotic bodies. Pretreatment with the NOS
inhibitor N
-nitro-L-arginine
methyl ester abolished these morphological alterations.
Conclusions The efficient expression of the human ecNOS gene in vivo suggests that NO or its toxic metabolite caused myocardial degradation, a part of which was compatible with apoptosis of the transfected cardiomyocytes themselves and the adjacent cells as a paracrine effect. These morphological features mimicked acute myocarditis or ischemic injury.
Key Words: genes • endothelium-derived factors • immunohistochemistry • myocardium • molecular biology
This article has been cited by other articles:
 |
|
 |
|
  J. D. Hunter and M. Doddi Sepsis and the heart Br. J. Anaesth., November 25, 2009; (2009) aep339v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Davis, M. V. Westfall, D. Townsend, M. Blankinship, T. J. Herron, G. Guerrero-Serna, W. Wang, E. Devaney, and J. M. Metzger Designing Heart Performance by Gene Transfer Physiol Rev, October 1, 2008; 88(4): 1567 - 1651. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Ly, Y. Kawase, R. Yoneyama, and R. J. Hajjar Gene Therapy in the Treatment of Heart Failure Physiology, April 1, 2007; 22(2): 81 - 96. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E Belcher, J Mitchell, and T Evans Myocardial dysfunction in sepsis: no role for NO? Heart, June 1, 2002; 87(6): 507 - 509. [Full Text] [PDF]
|
|
|
|
 |
|
 T. Kawada, M. Nakazawa, S. Nakauchi, K. Yamazaki, R. Shimamoto, M. Urabe, J. Nakata, C. Hemmi, F. Masui, T. Nakajima, et al. Rescue of hereditary form of dilated cardiomyopathy by rAAV-mediated somatic gene therapy: Amelioration of morphological findings, sarcolemmal permeability, cardiac performances, and the prognosis of TO-2 hamsters PNAS, January 22, 2002; 99(2): 901 - 906. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Iwata, S. Sai, Y. Nitta, M. Chen, R. de Fries-Hallstrand, J. Dalesandro, R. Thomas, and M. D. Allen Liposome-Mediated Gene Transfection of Endothelial Nitric Oxide Synthase Reduces Endothelial Activation and Leukocyte Infiltration in Transplanted Hearts Circulation, June 5, 2001; 103(22): 2753 - 2759. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. J. Szabolcs, N. Ma, E. Athan, J. Zhong, M. Ming, R. R. Sciacca, J. Husemann, A. Albala, and P. J. Cannon Acute Cardiac Allograft Rejection in Nitric Oxide Synthase-2-/- and Nitric Oxide Synthase-2+/+ Mice : Effects of Cellular Chimeras on Myocardial Inflammation and Cardiomyocyte Damage and Apoptosis Circulation, May 22, 2001; 103(20): 2514 - 2520. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. E. McDonald, M. N. Grinman, C. M. Carthy, and K. R. Walley Endotoxin infusion in rats induces apoptotic and survival pathways in hearts Am J Physiol Heart Circ Physiol, November 1, 2000; 279(5): H2053 - H2061. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Hikiji, W. S. Shin, T. Koizumi, T. Takato, T. Susami, Y. Koizumi, Y. Okai-Matsuo, and T. Toyo-Oka Peroxynitrite production by TNF-alpha and IL-1beta : implication for suppression of osteoblastic differentiation Am J Physiol Endocrinol Metab, June 1, 2000; 278(6): E1031 - E1037. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 U. Weiland, J. Haendeler, C. Ihling, U. Albus, W. Scholz, H. Ruetten, A. M. Zeiher, and S. Dimmeler Inhibition of endogenous nitric oxide synthase potentiates ischemia-reperfusion-induced myocardial apoptosis via a caspase-3 dependent pathway Cardiovasc Res, February 1, 2000; 45(3): 671 - 678. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Mortensen, J. Skouv, D. M. Hougaard, and L.-I. Larsson Endogenous Endothelial Cell Nitric-oxide Synthase Modulates Apoptosis in Cultured Breast Cancer Cells and Is Transcriptionally Regulated by p53 J. Biol. Chem., December 31, 1999; 274(53): 37679 - 37684. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Veit, J.-P. Boissel, M. Buerke, T. Grosser, J. Meyer, and H. Darius Highly efficient liposome-mediated gene transfer of inducible nitric oxide synthase in vivo and in vitro in vascular smooth muscle cells Cardiovasc Res, August 15, 1999; 43(3): 808 - 822. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Koglin, D. J. Granville, T. Glysing-Jensen, J. S. Mudgett, C. M. Carthy, B. M. McManus, and M. E. Russell Attenuated Acute Cardiac Rejection in NOS2 -/- Recipients Correlates With Reduced Apoptosis Circulation, February 16, 1999; 99(6): 836 - 842. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. J. Ing, J. Zang, V. J. Dzau, K. A. Webster, and N. H. Bishopric Modulation of Cytokine-Induced Cardiac Myocyte Apoptosis by Nitric Oxide, Bak, and Bcl-x Circ. Res., January 22, 1999; 84(1): 21 - 33. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Stein, T. Eschenhagen, J. Rudiger, H. Scholz, U. Forstermann, and I. Gath Increased expression of constitutive nitric oxide synthase III, but not inducible nitric oxide synthase II, in human heart failure J. Am. Coll. Cardiol., November 1, 1998; 32(5): 1179 - 1186. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Koglin, T. Glysing-Jensen, J. S. Mudgett, and M. E. Russell NOS2 Mediates Opposing Effects in Models of Acute and Chronic Cardiac Rejection : Insights from NOS2-Knockout Mice Am. J. Pathol., November 1, 1998; 153(5): 1371 - 1376. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gawaz, F.-J. Neumann, T. Dickfeld, W. Koch, K.-L. Laugwitz, H. Adelsberger, K. Langenbrink, S. Page, D. Neumeier, A. Schomig, et al. Activated Platelets Induce Monocyte Chemotactic Protein-1 Secretion and Surface Expression of Intercellular Adhesion Molecule-1 on Endothelial Cells Circulation, September 22, 1998; 98(12): 1164 - 1171. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Jeppsson, C. Pellegrini, T. O'Brien, V. M. Miller, H. D. Tazelaar, and C. G.A. McGregor Transbronchial gene transfer of endothelial nitric oxide synthase to transplanted lungs Ann. Thorac. Surg., August 1, 1998; 66(2): 318 - 324. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. A Doggrell and L. Brown Rat models of hypertension, cardiac hypertrophy and failure Cardiovasc Res, July 1, 1998; 39(1): 89 - 105. [Full Text] [PDF]
|
|
|
|
|
|
J. Chen, Y. Wang, Y. Wang, T. Nakajima, K. Iwasawa, H. Hikiji, M. Sunamoto, D.-K. Choi, Y. Yoshida, Y. Sakaki, et al. Autocrine Action and Its Underlying Mechanism of Nitric Oxide on Intracellular Ca2+ Homeostasis in Vascular Endothelial Cells J. Biol. Chem., September 8, 2000; 275(37): 28739 - 28749. [Abstract] [Full Text] [PDF]
|
|