Inhibition of vascular smooth muscle cell proliferation

Inhibition of vascular smooth muscle
cell proliferation
Vascular smooth muscle cell proliferation represents
a signifi cant central event in the formation of the fi -
brous atherosclerotic plaque. α-Tocopherol at physiological
concentrations specifi cally inhibits mitogeninduced
proliferation of vascular smooth muscle cells
and certain other cell types in a parallel manner to
the inhibition of PKC activity (Boscoboinik et al.,
1991a,b; Chatelain et al, 1993). The degree of inhibition
of cell proliferation depends on the mitogen
responsible for stimulating growth. Proliferation induced
by platelet-derived growth factor (PDGF), endothelin
and unmodifi ed LDL was almost completely
inhibited by α-tocopherol, whereas proliferation
produced by other mitogens, such as bombesin and
lysophosphatidic acid, was only moderately or slightly
inhibited (Azzi et al., 1993). In cultured smooth mus muscle
cells, α-tocopherol activated the cellular release of
the growth-inhibiting transforming growth factor-β
(TGF-β) (Özer et al., 1995). Calphostin C, a specifi c
PKC inhibitor, also inhibits smooth muscle cell proliferation,
supporting the notion that the antiproliferative
effect of α-tocopherol is mediated through the
inhibition of PKC activity (Tasinato et al., 1995).

9.7.6 Protection of prostacyclin generation
in arteries
Prostacyclin (PGI2), a member of the prostaglandin
family, is a product of arachidonic acid metabolism
(see Fig. 5.3). Prostacyclin is a potent stimulator of
adenylyl cyclase, the enzyme which converts ATP to
cyclic AMP. Because platelet aggregation is inhibited
by cyclic AMP, prostacyclin acts as a platelet anti-aggregating
agent. This effect upon platelets is opposed
by another product of arachidonic metabolism, thromboxane A2, which inhibits adenylyl cyclase.
Prostacyclin is also a strong vasodilator; moreover,
it inhibits polymorphonuclear leucocyte adhesion to
endothelial cells in vitro (Boxer et al., 1980).
In experimental atherosclerosis the capacity of
arterial endothelial cells to generate prostacyclin at
the site of plaque formation is considerably impaired
(Gryglewski et al., 1978). This decrease in prostacyclin
production may be caused by oxidative stress.
Prostacyclin synthesis in cultured aortic endothelial
cells was depressed by the presence of high glucose
concentration in the medium and synthesis could be
restored by the simultaneous addition of vitamin E
(Kunisaki et al., 1992). Vitamin E probably acted as
an antioxidant by preventing the build-up of lipid
hydroperoxides which are inhibitory to prostacyclin
synthesis. The results suggested that vitamin E may
restore depressed prostacyclin production by the vascular
wall in hyperglycaemic conditions such as those
seen in patients with diabetes mellitus. Szczeklik et
al. (1985) reported that feeding an atherogenic diet
to rabbits for a week resulted in elevation of plasma
lipid hydroperoxides and a 90% decrease in arterial
generation of prostacyclin. Enrichment of the atherogenic
diet with 100 mg of vitamin E daily prevented
the increase in lipid hydroperoxides and protected the
prostacyclin generating system in arteries.
Chan & Leith (1981) showed that prostacyclin
synthesis in dissected rabbit aorta is decreased when
the tissue is depleted of vitamin E. Enrichment of cultured
human endothelial cells with vitamin E caused
an increase in arachidonic acid release and spontaneous
prostacyclin synthesis (Tran & Chan, 1988, 1990).
A potentiating effect of vitamin E on arachidonic acid
release in megakaryocytes was attributed to an upregulation
of phospholipase A2 (Chan et al., 1998).
The above fi ndings suggest that a defect in the local
production of prostacyclin may be an underlying factor
in the pathogenesis of atherosclerosis in terms of
platelet aggregation and monocyte adhesion to endothelial
cells. An increased dietary intake of vitamin
E can overcome this defect.
9.7.7 Preservation of nitric oxide-mediated
arterial relaxation
Background information can be found in Section 4.5.9.
Nitric oxide, an important relaxant of arterial
smooth muscle, is released from vascular endothelial
cells in response to acetylcholine (Furchgott &
Zawadzki, 1980) and other agents. Exposure of the
vascular wall to oxidized LDL (oxLDL) inhibits the
release of nitric oxide, thereby preventing relaxation
(Kugiyama et al., 1990). This dysfunction has, in part,
been attributed to PKC stimulation (Ohgushi et al.,
1993).
Keaney et al. (1996) fed rabbits diets defi cient in or
supplemented with α-tocopherol and examined the
effects of the vitamin on oxLDL-induced endothelial
dysfunction. Exposure of thoracic aorta segments
from vitamin-defi cient animals to ox-LDL produced
dose-dependent inhibition of acetylcholine-mediated
relaxation, while similarly treated segments from
animals consuming α-tocopherol showed no such inhibition.
Vessel resistance to endothelial dysfunction
in the vitamin-replete animals was strongly correlated
with the vascular content of α-tocopherol. Incorporation
of α-tocopherol into the vasculature limited
endothelial dysfunction induced by phorbol ester, a
direct activator of PKC. Using cultured human aortic
endothelial cells, Keaney et al. (1996) confi rmed that
oxLDL stimulates endothelial PKC and that cellular
incorporation of α-tocopherol inhibits this stimulation.
Desrumaux et al. (1999) showed that plasma
phospholipid transfer protein, by supplying vascular
endothelial cells with α-tocopherol, plays a distinct
role in the prevention of endothelial dysfunction.
9.7.8 Inhibition of platelet aggregation
α-Tocopherol has long been known to inhibit platelet
aggregation in vitro, an effect that was initially attributed
to the inhibition of lipid peroxidation. However,
α-tocopheryl quinone (which is not an antioxidant)
also inhibits platelet aggregation (Cox et al., 1980),
making this theory unlikely. Freedman et al. (1996)
found that inhibition of platelet aggregation by α-tocopherol
was closely linked to its incorporation into
platelets. Incorporation of α-tocopherol inhibited
phorbol ester-induced stimulation of platelet PKC,
thus implicating this enzyme in the inhibition of aggregation.
Platelet-rich plasma from healthy individuals receiving
varying doses of supplemental vitamin E showed a
marked decrease in platelet adhesion compared to presupplementation
adhesiveness. A daily supplement of
400 IU was near the optimum to reduce platelet adhesivity
(Jandak et al., 1988). Vitamin E supplementation
246 Vitamins: their role in the human body
(727 mg per day) of healthy humans with low antioxidant
status over a 5-week period showed no effects on
the capacity of platelets to aggregate in vitro (Stampfer
et al., 1988). Over a 5-month period, supplementation
of human diets with a combination of 300 mg vitamin
E, 600 mg ascorbic acid, 27 mg β-carotene and 75 μg
selenium in yeast signifi cantly reduced platelet aggregability
(Salonen et al., 1991).
9.7.9 Protection against oxLDL-mediated
cytotoxicity
Marchant et al. (1995) incubated LDL with copper
ions for varying periods in the presence or absence of
α-tocopherol. They then incubated these LDL preparations
with human monocyte-macrophages and
measured the toxicity produced in the cells. Toxicity
increased with increasing duration of copper-catalysed
oxidation in the absence of α-tocopherol. The
presence of α-tocopherol protected the LDL from
oxidation and the cells from toxicity. Martin et al.
(1998) reported that enrichment of human aortic
endothelial cells with α-tocopherol in vitro dosedependently
increased their resistance to cytotoxic
injury from oxLDL.
9.7.10 Effects of vitamin E supplementation
on experimentally induced atherosclerosis
Verlangieri & Bush (1992) studied the effects of
α-tocopherol supplementation on experimentally
induced atherosclerosis in monkeys using duplex
ultrasound scanning and B-mode imaging to monitor
lesion formation and progression. The animals
were randomly assigned to one of four groups and
atherosclerosis was monitored over a 36-month period.
One group was fed a basal diet, while three other
groups consumed an atherogenic diet (basal diet plus
0.4% cholesterol, w/w). Two of the latter groups also
received α-tocopherol, one at the onset of the study
(prevention group) and the other after atherosclerosis
was established (regression group). The dosage
of α-tocopherol was approximately eight times the
daily requirement for primates. A steady initial rise
in percent stenosis was detected in the three groups
fed an atherogenic diet. However, stenosis in the
unsupplemented animals progressed more rapidly
and to greater extent than stenosis in the prevention
group. In the regression group, stenosis reached a
plateau at 4 months post-supplementation, while
that of the unsupplemented group continued to rise.
The data indicated that, while α-tocopherol does not
totally prevent atherosclerosis, it appears to lessen the
severity and reduce the rate of disease progression.
Moreover, supplemental vitamin E may regress wellestablished
lesions.
9.7.11 Epidemiological studies and clinical
trials
Epidemiological studies have shown increased protection
against coronary artery disease in subjects
consuming vitamin E in daily doses >100 mg taken
for more than 2 years (Rimm et al., 1993; Stampfer et
al., 1993).
The CHAOS clinical trial (Stephens et al., 1996)
was designed to study the effects of α-tocopherol at
doses of 400 IU or 800 IU daily on the risk of cardiovascular
death and nonfatal myocardial infarction in
patients with established coronary atherosclerosis at
recruitment. The results showed that α-tocopherol,
compared to placebo, reduced the risk of non-fatal
myocardial infarction by 77%; there was no difference
in deaths due to cardiovascular causes. The results of
more recent clinical trials have not agreed with the
results of the CHAOS trial. HOPE Study Investigators
(2000) reported that 400 IU of vitamin E administered
daily for 4 to 6 years had no benefi cial effects on
cardiovascular outcomes in a high-risk population of
patients who were 55 years or older. In an Italian trial
(GISSI-Prevenzione Investigators, 1999), the number
of patients with nonfatal myocardial infarction was
slightly higher in patients receiving 300 IU of vitamin
E per day than those receiving placebo, and the
number of deaths from coronary heart disease was
slightly smaller. Neither difference was statistically
signifi cant.
The lack of agreement between CHAOS and GSSI
has been discussed by Pryor (2000) in his extensive review
of the effects of vitamin E on heart disease, Pryor
concluded that in view of the diffi culty in obtaining
more than about 30 IU per day from a balanced diet,
vitamin E supplementation (100 to 400 IU per day)
should be part of a general programme of hearthealthy
behaviour that includes a fruit- and vegetable-
rich diet and regular exercise.