The Health Effects Institute
Synopsis
Research Report Number 80
Exploration of a Link Between Exposure to
Carbon Monoxide and Atherosclerosis
Stephen R. Thom and Harry Ischiropoulos
University of Pennsylvania Medical Center, Philadelphia, PA
BACKGROUND
Carbon monoxide, an indoor and outdoor air pollutant, is produced by the incomplete combustion of fossil fuels. People are exposed to carbon monoxide from sources such as automobile emissions, industrial processes, sidestream or mainstream cigarette smoke, and poorly vented space heaters and gas stoves. Exposure to high levels of carbon monoxide (500 parts per million [ppm]) can lead to respiratory failure and death. Even low-level exposures can cause adverse health effects (for example, chest pain) in some people with coronary artery disease when they exercise. For this reason, the U.S. Environmental Protection Agency has set National Ambient Air Quality Standards for carbon monoxide of 9 ppm averaged over eight hours and 35 ppm averaged over one hour.
Some researchers speculate that exposure to carbon monoxide may be related to the development of atherosclerosis, a progressive disease characterized by deposits of fat, cholesterol, cells, and connective tissue in blood vessels. These deposits, called plaques, can block blood flow. That carbon monoxide may have a role in atherosclerosis is a public health concern because this disease is the leading contributor to deaths by heart attack and stroke in the United States.
One possible mechanism by which carbon monoxide might influence the development of atherosclerotic plaques involves nitric oxide, a gas once regarded primarily as an air pollutant from combustion sources. However, we now know that nitric oxide is also produced in the body. It is involved in regulating many physiological reactions, including those in the cardiovascular, immune, and nervous systems. Nitric oxide also can be converted to toxic metabolites such as peroxynitrite, which converts cholesterol-carrying low-density lipoproteins to a form that contributes to atherosclerotic plaque formation. An earlier study by Dr. Stephen Thom and coworkers demonstrated that blood platelets isolated from rats exposed to high levels of carbon monoxide (1,000 or 3,000 ppm) released nitric oxide. This finding suggested a possible link between carbon monoxide exposure, peroxynitrite formation from nitric oxide, and atherosclerosis. The Health Effects Institute funded this pilot study to examine the effects of low concentrations of carbon monoxide on platelets and cells isolated from blood vessels.
APPROACH
Thom and Ischiropoulos exposed blood platelets (taken from rats) and endothelial cells (isolated from bovine blood vessels) to varying concentrations of carbon monoxide and measured how much nitric oxide was released. To determine if exposure to carbon monoxide causes endothelial cells to produce peroxynitrite, the investigators looked for markers of its presence in the culture medium and in the cells. They also exposed rats to carbon monoxide by inhalation, isolated platelets from the rats' blood, and measured how much nitric oxide was released.
RESULTS AND IMPLICATIONS
The investigators reported that platelets and endothelial cells exposed to carbon monoxide released nitric oxide; this was observed after exposure to carbon monoxide levels of 10 or 20 ppm, which are environmentally relevant concentrations. In addition, they found that platelets isolated from the blood of rats exposed to 20 to 1000 ppm carbon monoxide also released nitric oxide. However, concentrations of 50 or 100 ppm carbon monoxide were required to produce markers of peroxynitrite formation in endothelial cells. These concentrations approach levels of carbon monoxide that are toxic to humans; therefore, how relevant these results are to ambient exposures is uncertain.
These findings, which support the investigators' hypothesis of a possible connection between exposure to carbon monoxide and the release of nitric oxide by platelets and endothelial cells, warrant further investigation to determine their functional significance. The suggestive link to peroxynitrite formation, however, is based on experiments conducted in a cell culture system. Because such experiments do not include cellular constituents that are present and normally react with nitric oxide in vivo, thus reducing its availability for further reaction, additional work is required with experimental systems that more accurately reflect the natural cellular milieu to confirm the link.
TABLE OF CONTENTS
Mechanism of Oxidative Stress from Low Levels of Carbon Monoxide
Investigators' Report
Stephen R. Thom and Harry Ischiropoulos
* Abstract
* Introduction
* Specific Aims
* Methods and Study Design
o Animals and Reagents
o Preparation of Carbon Monoxide Gas Supply
o Platelet Isolation
o Release of NO by Platelets
o Vascular Endothelial Cell Culture Procedures
o Nitrite plus Nitrate Production
o L-Arginine Transport by Bovine Pulmonary Artery Endothelial Cells
o Nitric Oxide Synthase Activity
o Oxidation of p-Hydroxyphenylacetic Acid
o Assessment of the Effect of Illumination on Carbon Monoxide-Mediated Oxidant Production
o Nitrotyrosine in Bovine Pulmonary ArteryEndothelial Cells
o Detection of Reduced Sulfhydryls
o Dihydrorhodamine 123 Oxidation by Bovine Pulmonary Artery Endothelial Cells
o Chromium Release by Bovine Pulmonary Artery Endothelial Cells
o Ethidium Homodimer-1 Uptake
o Statistical Methods
* Results
o Specific Aim 1: Platelet Studies
+ Platelets Release NO in Response to CarbonMonoxide Exposure
+ Exposure to Carbon Monoxide Does Not Enhance the Activity of Nitric Oxide Synthase
+ Carbon Monoxide Can Displace NO from Endogenous Binding Sites in Platelets
+ Nitrite plus Nitrate Production by Platelets Exposed to Carbon Monoxide
+ NO Release by Platelets Obtained from Rats Exposed to Carbon Monoxide
o Specific Aim 2: Vascular Endothelial Cell Studies
+ Nitrite plus Nitrate as an Index of NO Production by Endothelial Cells
+ Assessment of the Effect of Illumination on Carbon Monoxide-Mediated Nitrite plus Nitrate Production
+ Enhanced NO Production Does Not Involve Alteration in L-Arginine Transport by Vascular Endothelial Cells
+ Carbon Monoxide Exposure Does Not Alter Nitric Oxide Synthase Activity
+ Exposure to Carbon Monoxide Increases Production of Agents Capable of Oxidizing p-Hydroxyphenylacetic Acid
+ Nitrotyrosine in Cells
+ Cellular Content of Sulfhydryls
+ Dihydrorhodamine 123 Oxidation
+ Cytotoxicity of Carbon Monoxide Exposure
+ Vital Staining with Ethidium Homodimer-1
* Discussion
* Implications
* References
Commentary
Health Review Committee
* Introduction
* Scientific Background
o The Development of Atherosclerosis
+ Association Between Carbon Monoxide Exposure and Atherosclerosis
+ Nitric Oxide, Peroxynitrite, and Atherosclerotic Plaque Formation
* Justification for the Study
* Technical Evaluation of the Study
o Attainment of Objectives
o Methods and Results
+ Platelets
+ Endothelial Cells
* Physiological Significance
* Implications for Future Research
* Conclusions |
