Cell injury and death
Isolated normal LDL is not toxic to vascular cells even at concentrations
equivalent to levels found in hypercholesterolemic subjects. By contrast,
oxidized LDL, particularly LDL oxidized in vitro, but also LDL- isolated
from human plasma, is highly toxic. Treatment of endothelial cells with
20 ug/ml LDL- produces acute injury and death within 24 hours. This concentration
of LDL- is comparable than that found in many hypercholesterolemic patients,
and although equivalency between exposures to cultured cells and effects
to cells in vivo are not to be expected, these finding serve as
an index for assessing the potential to elicit cellular responses related
to injury and to atherogenesis. In vivo modified forms of LDL (ie.
LDL-) are generally less oxidized than in vitro oxidized LDL and
are also less cytotoxic. Indeed, extensive oxidation of LDL is known to
cause rapid clearance from the circulation, hence highly oxidized LDL is
not encountered in human plasma. Minimally oxidized forms of LDL are, however
found in most subject in variable amounts (usually making up about 1 %
of the total LDL. These minimally oxidized LDL are toxic only at high concentrations,
but at lower levels (as encountered in plasma) induce several cell responses
indicative of early atherosclerosis.
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Induction of inflammatory factors and cytokines
A key response is the recruitment of inflammatory cells, their margination
into the vessel wall and subsequent activation. These early events are
elicited through cytokines that are released from cells interacting with
the modified LDL. A well studied example is the formation of the chemoattractant
peptide, MCP-1, by endothelial cells exposed to minimally oxidized LDL.
Figure
7 (39 Kb) shows a scheme for the events associated with
MCP-1 formation and its stimulation of macrophage chemotaxis. The interaction
of the minimally modified LDL (mmLDL) with endothelial cells induces a
number of gene products. The peroxidized phospholipids appear to be responsible
for induction of MCP-1. This occurs by interaction of with platelet-activating
factor (PAF) receptors or related receptors as the oxidized phospholipids
are structurally analogous to PAF. The activated receptor signals activation
of nuclear transcription factor(s) that bind to the gene promoter of the
MCP-1 gene inducing transcription of the message for MCP-1 protein. Thus,
by two hours of exposure, the endothelial cells synthesize and release
MCP-1 which on encountering circulating monocyte/macrophages induces them
to chemotax to the endothelial cells. This process occur in concert with
other signaling events that are provoked by other oxidized lipids found
in mmLDL. For example, lipid hydroperoxides are reported to stimulate expression
of adhesion molecules for inflammatory cells through signaling of transcription
factors in a manner analogous to that described for MCP-1.
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Mechanisms for stimulated cell division
In addition to signaling cytokine/adhesion molecules, oxidized LDL appears
to stimulate genes associated with antioxidant responses and with transcription
factors interacting with genes that regulate the cell cycle and cell proliferation.
Figure
8 (nfkbtrn.jpg 29 Kb) depicts the events involved in activation
of transcription factors (such as NFkB, although a similar process applies
to factors such as AP-1 which consists of oncogenic proteins c-fos and
c-jun). According to this scheme, peroxidation products and related oxidants
activate protein kinases (such as protein kinase C - PKC) which phosphorylate
the inhibitory peptide, IkB, associated with NFkB. The release and degradation
of IkB permits NFkB to bind to promoter sequences in DNA that regulate
transcription of genes such as those expressing adhesion molecules and
MCP-1. Events that suppress kinase-mediated phosphorylation or prevent
formation of peroxides, such as antioxidants (notably vitamin E), prevent
aberrant transcriptional activation of these genes as well as genes controlling
cell proliferation. This inhibition has been shown to involve direct interaction
of vitamin with PKC. The level of PKC-mediated phosphorylation of the c-fos/c-jun
components of AP-1 is inhibited leading to diminished binding and transcriptional
activation of genes regulating cell proliferation.
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