Modified lipoproteins in the vessel wall
Figure
5 (click to load 15Kb) illustrates the formation of oxidatively
modified lipoproteins and their effects on vascular cell function.
Most studies on atherosclerosis have focused on LDL and have examined its
oxidative modification either by inducing oxidation in vitro, or
isolating oxidatively modified forms of LDL from the total LDL recovered
from plasma (in vivo oxidation). Oxidation of LDL under either circumstance
produces more electronegative particles (LDL-, also referred to as oxidatively
modified LDL). These particles are a heterogeneous population that vary
depending on the extent of oxidation and can be isolated using high pressure
liquid chromatography as well as by electrophoresis. The isolated particles
are enriched in lipid peroxidation products, including important products
such as fatty acid hydroperoxides, aldehydic products formed by decomposition
of the hydroperoxides, cholesterol oxidation products and oxidized apoprotein
B-100 (the major protein of LDL).
Depicted
in Figure 6 (36 Kb) is the process by which
oxidative modification of LDL produces alterations in recognition and binding
to LDL receptors. Modification of the apoprotein B-100 is thought to occur
by reaction of lipid peroxidation products (notably reactive aldehydes)
with amino acids comprising the domain of the protein involved in binding
to the LDL receptor. Lysine residues are important components of this domain
and readily react with aldehydes. As the lysine residues contribute the
positive charge of the protein, reaction with aldehydes cancels these positive
charges and renders a net negative charge to the protein since negatively
charged amino acids remain in excess to the positive charged amino acids
in these domains. This gives the particle a net electrogenative character.
Elimination of approximately 20% of these lysine groups is sufficient to
alter the binding domain such that it no longer recognizes the LDL receptor.
The particle then can be bound by an alternate, or scavenger, receptor
which has a high affinity to negatively charged molecules and particles.
Binding to the scavenger receptors is accompanied by rapid endocytosis
and assimilation with components of the particles being delivered to lysosomes
where degradation takes place. This process is accompanied by specific
signaling events that evoke oxidative activity (enhanced reactive oxygen
generation) and induce a number acute response genes including genes controlling
proliferation and formation of cytokines. Moreover, assimilation of LDL
through the scavenger receptor appears not to be subject to feedback regulation,
as is the case for uptake via the normal receptor. Thus, accumulation of
cholesterol continues without appropriate down regulation of cell cholesterol
synthesis and of receptor content and activity.
Oxidized apoprotein B-100 is largely responsible for the net electronegativity
of LDL- and also determines binding of LDL to the LDL receptor of cells.
Its modification is produced by reaction of specific amino acid moieties
(notably lysine residues) with lipid peroxidation products. This modification
is a principal reason for the inability of modified LDL to bind to LDL
receptors, being instead assimilated by scavenger receptors - the so-called
"unregulated" uptake process referred to above which accounts
for foam cell formation. It is widely held that normal LDL is not toxic
to cells and evoke little if any atherogenic responses. On the other hand,
LDL-, and other modified forms of LDL, are toxic, and toxicity is attributed
to the content of lipid peroxidation products. Rapid and unregulated uptake
of the modified LDL also delivers the toxic lipid peroxidation products
at greater rates to target cells eliciting the various responses noted
in Figure 5, ie. inflammation, cytotoxicity and cholesteryl ester accumulation.
Return to Index
Modified lipoproteins in the plasma
Modified forms of LDL, especially with characteristic of LDL- have been
isolated from human plasma and from the plasma of animals which have had
experimentally-induced atherosclerosis. The formation of oxidatively modified
LDL is thought to take place in the vessel wall, within the connective
tissue matrix underlying the endothelium. Although the exact source for
modification remain uncertain, prime candidates include inflammatory cells
(resident macrophages) and endothelial cells. As described further below,
cell-induced oxidation appears to involve formation of reactive oxygen
species via aberrant stimulation of these cells and a number of enzymes
appear able to produce a flux of oxidizing agents. The oxidizing species
may include some or all of the following: hydrogen peroxide/superoxide,
transition metals free or as protein complexes (eg. heme), hypochlorous
acid and lipoxygenase (directly forming lipid hydroperoxides). Evidence
for formation in the matrix of the vessel wall is based on immunochemical
staining of modified LDL using a variety of antibodies raised against oxidatively
modified forms of LDL and co-localization with atherosclerotic lesions.
Return to Index
Relationships between the levels of modified lipoproteins
and disease progression
Studies describing a relationship between LDL modification and atherosclerosis
progression are only at the earliest stages of research, however, limited
scale experimental trials and large scale epidemiological studies (designed
to look at factors other than lipoprotein oxidation as a primary variable)
indicate that a strong correlation is likely. Using antibodies to oxidized
LDL, a number of reports show that modified forms of LDL are present in
the plasma of subjects with a history of heart disease or with diseases
that place them at risk to heart disease, eg. diabetes. The levels of modified
LDL also change dramatically following myocardial infarction and appear
to decrease when subjects take antioxidant supplements (notably vitamin
E).
Return to Index
Relationships between the levels of antioxidant vitamins
and disease progression
A recent study in the Journal of the American Medical Association (Hodis,
et al., 1995) showed that progression of atherosclerosis, measured
by ultrasonic imaging of the internal carotid arteries, was significantly
reduced when subjects consumed more than 100 IU of vitamin E per day. Although
the levels of modified LDL in these subjects is still under investigation,
other reports show that antioxidant supplementation at this level reduces
the levels of oxidized plasma lipoproteins. Based on the above data, there
has been a surge of investigations aimed at examining the relationship
between antioxidant intake and atherogenesis. To date, the preponderance
of epidemiological data shows that supplementation with vitamin E strongly
reduces the incidence of coronary artery disease attributable to atherosclerosis.
Controlled clinical trials in progress also show preliminarily, that early
markers of disease (intimal-medial thickening of carotid vessels) are reduced
in vitamin E supplemented individuals. These studies follow from animal
experiments that show that the amount of oxidized LDL detected in the vessels
using immunihistochemisrty are substantially reduced following supplementation
with vitamin E.
Return to Index
Figure
5 (click to load 15Kb) illustrates the formation of oxidatively
modified lipoproteins and their effects on vascular cell function.
Most studies on atherosclerosis have focused on LDL and have examined its
oxidative modification either by inducing oxidation in vitro, or
isolating oxidatively modified forms of LDL from the total LDL recovered
from plasma (in vivo oxidation). Oxidation of LDL under either circumstance
produces more electronegative particles (LDL-, also referred to as oxidatively
modified LDL). These particles are a heterogeneous population that vary
depending on the extent of oxidation and can be isolated using high pressure
liquid chromatography as well as by electrophoresis. The isolated particles
are enriched in lipid peroxidation products, including important products
such as fatty acid hydroperoxides, aldehydic products formed by decomposition
of the hydroperoxides, cholesterol oxidation products and oxidized apoprotein
B-100 (the major protein of LDL). 
Depicted
in Figure 6 (36 Kb) is the process by which
oxidative modification of LDL produces alterations in recognition and binding
to LDL receptors. Modification of the apoprotein B-100 is thought to occur
by reaction of lipid peroxidation products (notably reactive aldehydes)
with amino acids comprising the domain of the protein involved in binding
to the LDL receptor. Lysine residues are important components of this domain
and readily react with aldehydes. As the lysine residues contribute the
positive charge of the protein, reaction with aldehydes cancels these positive
charges and renders a net negative charge to the protein since negatively
charged amino acids remain in excess to the positive charged amino acids
in these domains. This gives the particle a net electrogenative character.
Elimination of approximately 20% of these lysine groups is sufficient to
alter the binding domain such that it no longer recognizes the LDL receptor.
The particle then can be bound by an alternate, or scavenger, receptor
which has a high affinity to negatively charged molecules and particles.
Binding to the scavenger receptors is accompanied by rapid endocytosis
and assimilation with components of the particles being delivered to lysosomes
where degradation takes place. This process is accompanied by specific
signaling events that evoke oxidative activity (enhanced reactive oxygen
generation) and induce a number acute response genes including genes controlling
proliferation and formation of cytokines. Moreover, assimilation of LDL
through the scavenger receptor appears not to be subject to feedback regulation,
as is the case for uptake via the normal receptor. Thus, accumulation of
cholesterol continues without appropriate down regulation of cell cholesterol
synthesis and of receptor content and activity.