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Atherosclerosis, calcium, and calcium antagonists
PHILIP D. HENRY, M.D.
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ALTHOUGH it has been generally recognized that
atherosclerosis is associated with a deposition of calcium in the arterial wall, the pathogenic significance of
this calcification remains unclear. In recent years, a
number of calcium-chelating agents, lanthanum chloride, and organic calcium antagonists have been shown
to exert antiatherogenic effects in animals with dietary
hypercholesterolemia. This brief review discusses possible interrelations between arterial calcification and
atherogenesis, and summarizes some of the data in the
literature on the effects of chelating and calcium antagonistic agents.
Deposition of calcium salts in the walls of human
arteries begins in childhood and progresses with age.
Electron microscopic studies of human atherosclerotic
arteries have shown that calcification in the media and
intima is associated with extracellular membranous
vesicles structurally resembling matrix vesicles in
bone and other calcifying tissues.1 These vesicles contain noncristalline particles and needle-shaped crystals
that have the diffraction patterns of apatite. The membranous structures occur in the vicinity of degenerating
cells that enclose similar vesicular formations. This
suggests that calcifying vesicles represent the debris of
mitochondria and other organelles from degenerating
medial and intimal cells. The biochemical composition
of the membranes has not yet been determined, but it
appears likely that phospholipids constitute an important component. It is of interest that atheromatous lesions contain a vitamin K-dependent (y-carboxyglutamic acid-containing) calcium-binding protein that
appears to be functionally analogous but structurally
dissimilar to osteocalcin.' Like y-carboxyglutamic
acid-containing proteins of the blood clotting system,
this protein may bind calcium and phospholipid and
play a role in regulating the calcification of atherosclerotic plaques. Results of the studies cited above are
From the Section of Cardiology, Baylor College of Medicine, Houston.
Supported in part by grants HL3 1487 and HL31488 from the National
Institutes of Health, Bethesda.
Address for correspondence: Philip D. Henry, M.D., Section of
Cardiology, The Methodist Hospital, Suite F-905, Baylor College of
Medicine, 6535 Fannin St., Houston, TX 77030.
456
based on analysis of material obtained at autopsy and
prepared by conventional histologic techniques, a procedure that does not satisfactorily preserve the physical
state and localization of electrolytes in soft tissues.
Additional investigations in which rapidly frozen (frozen-hydrated) samples and electron-probe microanalysis are used are needed, since with these techniques it
would be possible to determine if calcium in degenerating macrophages and smooth muscle cells accumulates in the mitochondria, as has been demonstrated in
other syndromes associated with cell necrosis.
Blumenthal et al. were among the first to point out
that in the human aorta medial calcification (or other
processes altering vascular elastance, such as syphylitic aortitis) is a structural alteration always preceding
the development of intimal plaques. A similar sequence of events is observed in rabbits and swine fed
vitamin D in doses inducing little or no hypercalcemia.4 In these normolipidemic animals atherosclerotic
lesions appear only months after fibromuscular
changes, cell necrosis, and calcifications have developed. Recent findings with regard to the influence of
mechanical factors on atherogenesis suggest that arterial stiffness may play a role in the localization and
development of atheromas. The relationship between
the physical properties of arteries, fluid mechanics,
and atherogenesis will require further investigative attention. In addition, the possibility must be considered
that crystalline deposits alter the deformability of atheromatous lesions. In coronary arteries, vessels subjected to continual bending movements, crystalline
bodies could be traumatic to surrounding soft tissues
and cause acute structural changes such as plaque hemorrhage or rupture. It has been recently reemphasized
that the dense neovascularization of atheromas may
render the artery susceptible to intramural hemorrhage, a possible mechanism of coronary thrombosis.6
Recently, evidence has accumulated indicating that
calcium antagonists may protect muscle and nonmuscle cells by a variety of pathophysiologic processes.
Protective effects have been reported in ischemic, cytotoxic, and degenerative syndromes involving skeletal muscle, myocardium, brain, liver, kidney, intesCIRCULATION
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tine, and the eye.7 The fact that calcium antagonists
have a protective effect supports the notion that excessive calcium uptake, in particular mitochondrial calcium overload, plays a role in mediating cell death
after various insults. An important question is whether
there are membrane physiologic alterations in atherosclerosis that might favor an excessive uptake of calcium. It is well established that modest changes in
membrane lipids may alter the functional characteristics of membrane proteins such as ion channels, ion
transporters, and receptors. The importance of the
abundance of membrane cholesterol in regulating the
functional characteristics of membranes has been well
demonstrated in red blood cells and platelets. Certain
membrane enzymes, such as ATPase transporters, appear to be particularly sensitive to changes in membrane cholesterol.'8 9 Recently Locher et al.9 have demonstrated in human red blood cells that increases in
membrane cholesterol augment the entry of calcium
through a channel that can be blocked with a dihydropyridine calcium antagonist. Inspired by experiments
relating changes in membrane lipids to calcium uptake
we performed experiments with isolated canine coronary arteries incubated in a high-cholesterol environment.8 Arteries exposed to low concentrations of cholesterol in aqueous solution (10- 2M) or to human
low-density lipoprotein underwent slow contractions
and preincubation of arteries with cholesterol sensitized the arteries to the constrictor effects of calcium
over a wide range of external calcium concentrations.
The uptake of radioactive cholesterol associated with
these changes in arterial reactivity was very low. The
fact that the constrictor effects of cholesterol were
blocked by calcium antagonists and that sterols structurally closely related to cholesterol were ineffective
suggested to us that cholesterol did not act by a simple
detergent effect. Our findings support the hypothesis
that physiologic changes in membranes caused by a
high-cholesterol environment may favor calcium uptake by arterial smooth muscle.
Interrelations between membrane lipids, calcium
uptake, and cell necrosis may be relevant to atherogenesis. Necrosis of arterial cells may play a pivotal role in
the initiation, progression, and regression of atherosclerotic lesions. '0 Results of studies of coronary arteries of children and young adults support the view that
cell death and accumulation of cellular debris in the
intima are important features distinguishing early fibrotic plaques from the pure foam cell lesions (fatty
streaks). 1" Cell death also results in the release of cytotoxic substances such as polar lipids and lytic enzymes, a process that may amplify cell necrosis and
Vol. 72, No. 3, September 1985
contribute to the stimulation of a fibroproliferative response.
If cell necrosis is an important process in the formation of atheroma and if cell death is mediated in part by
changes in membranes that facilitate the intracellular
accumulation of calcium, drugs capable of reducing
calcium uptake might exert antiatherogenic effects.
Almost 20 years ago, Wartman et al. 12 treated cholesterol-fed rabbits with subcutaneous injections of magnesium EDTA and concluded that this chelating agent
suppressed the deposition of collagen and elastin in the
arteries. Unfortunately, Wartman's experiments have
to my knowledge never been repeated, but other chelating agents were subsequently tested for their antiatherogenic effects. Kramsch et al. 1' administered diphosphonic acid and thiophene derivatives to rabbits fed a
high-cholesterol diet. The treatments did not influence
serum cholesterol or triglycerides, but inhibited calcification, lipid accumulation, and plaque formation in
arteries. In addition, these investigators demonstrated
that LaCL3 exerted antiatherogenic effects in rabbits
and monkeys.'3
In view of our considerations concerning altered
calcium uptake in atherosclerosis and the therapeutic
results obtained with chelating agents, we evaluated
the effects of nifedipine, a calcium antagonist, on atherogenesis in rabbits with diet-induced atheroscleroSiS. 14 Rabbits were fed a 2% cholesterol diet for 8
weeks with or without treatment with oral nifedipine
(40 mg/rabbit/day). High-dose oral nifedipine was
well tolerated and produced modest transient hypotensive effects. Compared with rabbits receiving placebo,
treated animals exhibited highly significant reductions
in sudanophilic lesions, aortic total cholesterol, and
aortic calcium. As with the chelating agents, the drug
did not influence the hyperlipidemic response to the
diet. Our findings were subsequently confirmed by
Willis et al. ,' 'who demonstrated that high-dose nicardipine and nifedipine produced substantial reductions
in aortic lesions, aortic total cholesterol, and aortic
triglyceride in cholesterol-fed rabbits. In another
study, low-dose nifedipine (2 mg/rabbit/day) was reported to exert antiatherogenic effects in rabbits, although the number of animals studied was small.'6
Subsequently, calcium antagonists structurally unrelated to nifedipine, such as verapamil, diltiazem, and
flunarizine, were demonstrated to exert antiatherogenic effects in cholesterol-fed rabbits.17-'9
Two groups of investigators have failed to demonstrate that calcium antagonists exert antiatherogenic
effects in cholesterol-fed rabbits. In the first study,20
however, high-dose nifedipine was administered by
457
HENRY
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the sublingual route, which may evoke severe hypotensive episodes and trigger a sympathetic discharge.
The reflex release of catecholamines, agents that stimulate the uptake of calcium, may have promoted the
intracellular accumulation of calcium. Such an effect
would oppose the therapeutic action of calcium antagonists. There is evidence that antiadrenergic drugs
such as reserpine, guanethidine, and propranolol suppress atherogenesis in cholesterol-fed rabbits.21 In the
second study,22 in which calcium antagonists were
found ineffective in suppressing atherogenesis in
cholesterol-fed rabbits given antiadrenergic drugs,
plaques of the whole aorta were approximately onetenth to one-fifth of those reported in the literature.13 9-9
Moreover, in two control groups receiving the same
cholesterol diet, plaque areas expressed as a percent of
the intimal area varied by more than 300% (4.4% vs
13.9%), a variation that would make the detection of
significant effects in drug-treated groups difficult if not
impossible.
The mechanism of action of chelating agents and
calcium antagonists remains unclear. Calcium antagonists might exert antiatherogenic effects by reducing
arterial pressure. In conscious normotensive rabbits,
however, nifedipine administered by the oral route
exerts only modest transient hypotensive effects. 14
Blumlein et al.'9 concluded that the antiatherogenic
action of verapamil was unrelated to lowering of arterial pressure. Nakao et al.23 have reported that the calcium antagonist nicardipine inhibits the migration of
cultured vascular smooth muscle from rat aorta, an
effect that might play a role in inhibiting atherogenesis. Recently Stein et al.24 demonstrated that high concentrations of verapamil enhanced the receptor-mediated uptake and disposal of low-density lipoprotein
(LDL) by aortic cells in culture. They proposed that
verapamil-induced stimulation of receptors might aid
in the removal of LDL cholesteryl esters from the
aortic interstitium. An action of calcium antagonists
via LDL receptors might explain why the Watanabe
rabbit, an animal that lacks LDL receptors, does not
respond favorably to nifedipine.25 However, studies in
Watanabe rabbits should be repeated in larger groups
of animals receiving drug under controlled conditions.
Recently, Etingin and Hajjar26 demonstrated that nifedipine produced a 50% loss of cholesterol and cholesteryl esters from lipid-laden rabbit aortic smooth
muscle in culture. They concluded that nifedipine decreased cholesterol and cholesteryl ester accumulation
in arterial smooth muscle by increasing cholesteryl
ester hydrolysis.
In summary, current evidence indicates that cal458
cium-chelating (anticalcifying) agents and calcium antagonists (calcium-channel blockers) have antiatherogenic effects in rabbits and monkeys. The drugs exert
their effects without reducing dietary hypercholesterolemia. Therefore, the mechanism of action differs
from that of hypolipidemic interventions.
Clinical implications. For many years it has been
claimed by some that the chelating agent EDTA exerts
beneficial effects in patients suffering from coronary
and peripheral artery disease. Unfortunately, treatment
with EDTA has never been subjected to a controlled
trial and no statement can be made regarding the efficacy of this controversial therapy. On the other hand,
several controlled studies with calcium blockers have
recently been initiated in North America and Europe.
To my knowledge, the ongoing trials are all based on
the evaluation of sequential coronary arteriograms of
patients undergoing catheterization for symptomatic
coronary disease. For such trials to be successful, patients in placebo groups will have to be kept off calcium antagonists, and patient groups will have to be
matched with respect to antiadrenergic manipulations
(/l-adrenergic blockade), which may influence the progression of atherosclerosis.
The excellent assistance of Patricia Farrell in the preparation
of this manuscript is gratefully acknowledged.
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Atherosclerosis, calcium, and calcium antagonists.
P D Henry
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Circulation. 1985;72:456-459
doi: 10.1161/01.CIR.72.3.456
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