ABSTRACT
Brief Report
The aim of the present study was to assess whether a family history of cardiovascular disease and higher than normal serum levels of asymmetric dimethylarginine could be correlated with an inhibition of nitric oxide synthase. A new method, an Enzyme Linked Immunosorbant Assay (ELISA) for asymmetric dimethylarginine, was used to assess the serum levels of asymmetric dimethylarginine. The analysis of volunteers with a family history of cardiovascular disease and the elderly showed that individuals with serum levels of asymmetric dimethylarginine greater than 0.4 mM may be predisposed to endothelium dysfunction.
Endothelial dysfunction is suspected as a major cause of vascular disease. Endothelial derived nitric oxide has inhibitory effects on cardiovascular disorders including vascular disease associated with hypertension, diabetes, renal insufficiency, hypercholesterolemia, hyperhomocysteinemia, vascular aging and estrogen deficiency. Nitric oxide is formed from L-arginine by oxidation of the guanidino-nitrogen terminal. The enzymes that synthesize nitric oxide exist in several isoforms in endothelium, vascular smooth muscle, macrophages and neurons (1). These enzymes are inhibited by asymmetric dimethylarginine. Asymmetric dimethylarginine is synthesized by methylation of arginine residues in proteins and is a product of protein hydrolysis. The concentration of asymmetric dimethylarginine is elevated in patients with vascular disease (2,3). Additionally, asymmetric dimethylarginine is degraded by the enzyme dimethylarginine dimethylaminohydrolase. This enzyme is found in the same tissues as nitric oxide synthase, and disruption in the synthesis of nitric oxide by asymmetric dimethylarginine is thought to contribute to vascular disease (4). There is some controversy as to the absolute amount of asymmetric dimethylarginine in serum; however, our study determined that an ELISA assay with an antibody specific to asymmetric dimethylarginine can be used to accurately measure this analyte in serum. To control for confounding effects of hypertension, diabetes, renal dysfunction and smoking on concentrations of asymmetric dimethylarginine, the study excluded volunteers with these conditions.
Results:
In the present study, we measured asymmetric dimethylarginine (ADMA) levels in the serum of 33 volunteers; their ages ranged from 16 to 83 years old.
These were volunteers selected without our knowledge of their family history or predisposition to cardiovascular disease. The additional criteria for the volunteers included no medications or food and drink other than water eight hours prior to blood collection. All subjects provided written informed consent prior to blood donation.
The (ADMA) levels in the serum of volunteers were classified as low risk or high risk for vascular disease based on the ADMA concentration needed to inhibit at least 50% of endothelial nitric oxide synthase (5). The nitric oxide synthase assay has been used to show that patients with chronic renal failure have plasma asymmetric dimethylarginine levels that inhibit production of nitric oxide. We concluded that the ability to inhibit nitric oxide synthase should be considered since this is the target of ADMA. The designations LR and HR were assigned for convenience to identify samples after measurement of serum ADMA concentrations. The age range of the volunteers in the low risk group was 16 to 67. The mean age of male volunteers was 33.5 years, and the mean age of female volunteers was 33.1 years (Table I).
Most high risk group members had a family history of cardiovascular disease and/or other risk factors, although this trend was not universal. For example, HR02 was an 80 year white female with no previous family history of vascular disease. HR02�s son, HR08, was 58 years old, and her 83 year-old husband, HR13, suffered from Alzheimer's disease. At age 67, HR13 had coronary by-pass surgery. All other members of the high risk group, aged 44 to 70 years, had fathers or grandfathers who died of complications of heart disease between the ages of 47 to 60. However, these volunteers did not exhibit any symptoms of cardiovascular disease. The volunteer listed as HR06, was a white female; her father died from a heart attack at age 59, and HR06�s brother had a heart attack at age 52 (the same age their father was diagnosed with heart disease). When HR06 was evaluated for traditional risk factors, she had a fasting cholesterol level of 270 mg/dL. This individual was not taking medication at the time the study was completed. She was one of three individuals in the study that had a higher than normal level of a traditional cardiovascular disease risk factor. The volunteers, indicated by HR04 and HR07 are black males, age 44 and 49 respectively. Neither is taking medication; however, both individuals have a cholesterol reading of 180 mg/dL. Their father is healthy while their mother and her sister have diabetes and hypertension. Their paternal grandfather lived to be 103, however their maternal grandfather died of complications of heart disease at age 47.
Since our data suggest that those with a family history of heart disease may have ADMA levels above 0.4mM, we decided to measure the ADMA levels in serum of those patients known to be at risk for cardiovascular disease. The data in Table II indicate that 22 of 24 individuals predisposed to developing cardiovascular disease have ADMA amounts greater than 0.4 mM, as determined with the antibody test.
Discussion:
The results from the study suggest that individuals
with a family history for cardiovascular disease and the elderly may have
higher than normal serum levels of ADMA, and be predisposed to endothelial
dysfunction and vascular disease. The level of cholesterol in volunteers
younger than 40 did not correlate with high ADMA amounts, however, the
individuals in the high risk group with a family history of heart disease
and high cholesterol do have high serum ADMA levels. These data suggest
there are overlapping pathways in cholesterol metabolism and asymmetric
dimethylarginine. The results of these studies indicate that it is now
feasible to complete multiple measurements of ADMA amounts on the same
patient several times during clinical studies; and determine whether there
exists a pathophysiological role for ADMA in cardiovascular disease.
Table II: Patients at
risk for cardiovascular
disease have high ADMA levels
The ADMA concentrations from Healthy Volunteers
The linear part of the standard curve is presented because the ADMA concentrations were obtained from this region.
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