(Received for publication, May 26, 1995; and in revised form, July 3, 1995)
From the
Rabbit smooth muscle cells (SMC) express types I and II
scavenger receptors (ScR) that are up-regulated by platelet secretion
products. In the current studies we investigated the effect of growth
factors secreted by platelets on ScR activity in rabbit and human SMC.
Platelet-derived growth factor (PDGF BB) and transforming growth factor
(TGF-
) at 10 ng/ml increased ScR
activity in rabbit SMC (by approximately 4- and 2-fold, respectively)
but not in human SMC. Epidermal growth factor (EGF) or insulin-like
growth factor I (IGF-I) alone had little effect on SMC ScR activity.
The growth factors had synergistic effects on ScR activity and on types
I and II ScR mRNA expression. In rabbit SMC, PDGF BB, EGF, and
TGF-
together stimulated ScR activity 12-fold. In
human SMC, EGF and TGF-
, together with either IGF-I or
PDGF BB, stimulated receptor activity approximately 7-fold. Growth
factor-mediated induction of ScR activity in rabbit and human SMC was
blocked by the tyrosine kinase inhibitor tyrphostin 47, whereas the
induction of ScR activity in rabbit but not human SMC was blocked by
the protein kinase C inhibitor MDL.29,152. Studies using neutralizing
antibodies demonstrated that TGF-
is the predominant
factor in in vitro preparations of platelet secretory products
which regulates ScR activity. The growth factors that act
synergistically in regulating ScR activity in vitro are all
present in atherosclerotic lesions, where they are produced by
macrophages, endothelial cells, SMC, and platelets. The data suggest
that these growth factors may regulate ScR activity in SMC in vivo and contribute to foam cell formation.
One characteristic feature of atherosclerotic lesions is the unregulated accumulation of lipoprotein-derived cholesterol and cholesteryl esters in macrophages and smooth muscle cells of the arterial intima. Lipid is deposited in these cells as droplets that give the cells a foamy appearance when viewed by phase-contrast microscopy(1, 2, 3, 4) . The mechanism of lipid accumulation and foam cell formation is not known with certainty; however, lipid accumulation in macrophages has been postulated to result from the scavenger receptor-mediated internalization of modified lipoproteins(5, 6, 7, 8) . We have demonstrated recently that smooth muscle cells also express scavenger receptors, and we have postulated a similar mechanism for lipid accumulation in these cells(9, 10, 11, 12) .
The expression of the scavenger receptor in both macrophages and smooth muscle cells can be induced during atherogenesis. Whereas circulating monocytes do not express scavenger receptors, scavenger receptor expression is induced to high levels when the monocytes adhere to the endothelium, penetrate between the endothelial cells, and differentiate to macrophages in the subendothelial space(2, 4, 13, 14) . During atherogenesis, smooth muscle cells migrate from the media to the intima of the arterial wall, where they proliferate and accumulate lipid, becoming foam cells. Scavenger receptor activity is detected in smooth muscle cells in the intima but not in normal vascular smooth muscle cells(15, 16, 17) , demonstrating that smooth muscle cell scavenger receptor activity is up-regulated in atherosclerotic lesions. The factors contributing to the regulation of smooth muscle cell scavenger receptor activity in vivo are unknown.
We have shown that scavenger receptor activity in rabbit smooth muscle cells is induced in vitro by incubation of the cells with phorbol esters, serum, or secretion products from platelets(10) . The expression of types I and II scavenger receptors in smooth muscle cells is normally quite low; however, incubation of the cells with phorbol esters increases receptor activity up to 20-fold(10, 11, 12) . Types I and II scavenger receptor cDNAs have been cloned from a cDNA library prepared from phorbol ester-treated rabbit smooth muscle cells (18) . Incubation of rabbit smooth muscle cells with secretion products from human platelets, stimulated with thrombin or with the calcium ionophore A23187, also results in up to a 5-6-fold increase in receptor activity(10, 11, 12) . These data suggest that platelet secretory products present in serum or at sites of platelet aggregation may contribute to the regulation of smooth muscle cell scavenger receptor activity.
In the present studies we examined
the effect of several known platelet secretory products on scavenger
receptor activity in human and rabbit smooth muscle cells. The results
demonstrate that platelet-derived growth factor (PDGF) ()BB,
epidermal growth factor (EGF), insulin-like growth factor I (IGF-I),
and transforming growth factor
(TGF-
), growth factors present in
atherosclerotic
lesions(19, 20, 21, 22, 23) ,
synergistically stimulate types I and II scavenger receptor gene
expression and receptor activity in both human and rabbit smooth muscle
cells.
For lipoprotein metabolism assays, the cells
were treated with growth factors as described above. After three washes
with serum-free medium, the cells were incubated with I-labeled Ac-LDL (5 µg/ml) at 37 °C for 16 h.
Cell-associated radioactivity (i.e. bound and internalized
lipoprotein) and trichloroacetic acid-soluble lipoprotein degradation
products in the medium were quantitated as described(30) .
Nonspecific degradation (i.e. the amount of degradation
obtained in the presence of a 100-fold excess of unlabeled Ac-LDL) has
been subtracted from all data.
To neutralize their activity, platelet secretory
products (40 µg/ml) were preincubated for 1 h at 37 °C with
blocking antibodies to the growth factors (concentrations indicated in
the legend to Fig. 4) before addition to the cells. The
concentrations of anti-PDGF and anti-TGF- antibodies
used were sufficient to block the effect of recombinant PDGF BB (50
ng/ml) and TGF-
(20 ng/ml) on scavenger receptor
activity in rabbit smooth muscle cells, respectively. The
concentrations of anti-IGF-I and anti-EGF antibodies used were
sufficient to block the effect of IGF-I (20 ng/ml) and EGF (10 ng/ml)
on smooth muscle cell proliferation measured by assaying the
mitochondrial dehydrogenase activity as described (Boehringer
Mannheim).
Figure 4:
Ability of blocking antibodies to
neutralize the stimulatory effect of platelet secretory products on
scavenger receptor activity. Smooth muscle cells (SMC) were
preincubated with 40 µg of platelet secretory products in the
absence or presence of blocking antibodies against PDGF (33 µg/ml),
EGF (13 µg/ml), IGF-I (10 µg/ml), and TGF- (8
µg/ml) for 16 h at 37 °C and then assayed for their ability to
internalize DiI-labeled Ac-LDL. The data are reported as the mean
± the range for two independent
experiments.
Our previous studies have shown that types I and II scavenger
receptors are expressed by rabbit smooth muscle cells and that
incubation of the cells with platelet secretory products up-regulates
receptor activity(10) . Upon activation, platelets have been
reported to secrete a number of products, including fibronectin,
fibrinogen, serotonin, and the growth factors PDGF, EGF,
TGF-, and IGF-I(32) . To determine whether
these secretory products stimulate scavenger receptor activity in
smooth muscle cells, we preincubated rabbit or human smooth muscle
cells with them for 16 h at 37 °C and examined the uptake of
DiI-labeled Ac-LDL by fluorescence microscopy. Fibrinogen (10-70
ng/ml), fibronectin (10-70 ng/ml), and serotonin (10-70
ng/ml) had little effect on scavenger receptor activity, whereas
preincubation with the combination of the four growth factors
significantly enhanced the uptake of DiI-labeled Ac-LDL by both rabbit
and human smooth muscle cells (data not shown).
To obtain a
quantitative estimate of the increase in receptor activity induced by
the growth factors and to assess the effect of each growth factor
individually, additional experiments were performed. The cells were
preincubated with the recombinant growth factors either alone or in
combination, and the uptake of DiI-labeled Ac-LDL was assessed by FACS
analysis. When DiI-labeled lipoproteins are internalized by cells and
degraded, the DiI is quantitatively retained in the lysosomes. The
amount of DiI in the cells is therefore directly proportional to the
amount of lipoprotein metabolized(28) . In rabbit smooth muscle
cells (Fig. 1A), PDGF BB (10 ng/ml) stimulated the
internalization of DiI-labeled Ac-LDL by 3.7-fold, whereas neither PDGF
AA (10-70 ng/ml) nor PDGF AB (10-70 ng/ml) had any effect
on the uptake of DiI-labeled Ac-LDL (data not shown). TGF- (10 ng/ml) up-regulated scavenger receptor activity
2-fold.
The growth factors IGF-I (20 ng/ml) or EGF (10 ng/ml) had little effect
on receptor activity when added to cells alone at the concentrations
shown here or at concentrations up to 100 ng/ml (data not shown).
However, when these growth factors were added in combination,
synergistic effects were observed. The four combinations, IGF-I and
TGF-
, EGF and PDGF BB, TGF-
and PDGF
BB, and EGF and TGF-
, stimulated receptor activity to
a significantly greater extent than that observed with any growth
factor alone. The greatest synergistic effect with two growth factors
was with EGF and TGF-
, in which receptor activity was
increased 7.5-fold. An even greater increase in receptor activity
(12-fold) was obtained with a combination of the three growth factors
EGF, TGF-
, and PDGF BB. Addition of IGF-I with the
three growth factors had little, if any, additional effect on receptor
activity.
Figure 1:
Fluorescence-activated cell sorter
analysis showing the effect of growth factors on the uptake of
DiI-labeled Ac-LDL by rabbit and human smooth muscle cells (SMC). Cells were preincubated in serum-free medium for 16 h
at 37 °C with or without EGF (10 ng/ml), PDGF BB (10 ng/ml),
TGF- (10 ng/ml), and IGF-I (20 ng/ml) either alone or
in combination. The cells were then incubated with DiI-labeled Ac-LDL
(5 µg/ml) for 8 h at 37 °C. After incubation the cells were
subjected to FACS analysis as described under ``Experimental
Procedures.'' The relative fluorescence intensity, which is
proportional to the uptake of DiI-labeled Ac-LDL, was determined for
each group of cells. The data are reported as the fold increase in the
uptake of DiI-labeled Ac-LDL, which represents the average fluorescent
intensity of each group normalized to the fluorescent intensity of the
control smooth muscle cells incubated in the absence of growth factors.
The data are the mean ± standard deviation (n =
3). In this experiment platelet secretory products (40 µg/ml)
stimulated scavenger receptor activity
4-fold (data not shown). Panels A and B represent the effect of the growth
factors on scavenger receptor activity in rabbit and human smooth
muscle cells, respectively.
In human smooth muscle cells the growth factors also had
synergistic effects on scavenger receptor activity (Fig. 1B). Whereas IGF-I, EGF, TGF-,
and PDGF BB alone had little effect on receptor activity, EGF and
TGF-
together had a substantial effect, up-regulating
receptor activity approximately 3.4-fold. The addition of either IGF-I
or PDGF BB, together with these two growth factors, increased receptor
activity up to 7-fold. The addition of a fourth growth factor had no
additional stimulatory effect. A similar effect of the combination of
growth factors was obtained with a second human aortic smooth muscle
cell line (data not shown). The growth factors, therefore, had similar
effects on human and rabbit smooth muscle cell scavenger receptor
activity, except that PDGF was more potent in the rabbit cells, and
IGF-I had more of an effect in the human cells.
The effect of the
recombinant growth factors on the internalization of both Ac-LDL and
native LDL by rabbit and human smooth muscle cells was examined to
determine if the regulation of receptor activity was specific for the
scavenger receptor or if LDL receptor activity was also increased. In
rabbit smooth muscle cells scavenger receptor activity was stimulated
by the mixture of the four growth factors (PDGF BB, EGF,
TGF-, and IGF-I), whereas LDL receptor activity was
not increased (Fig. 2A). In contrast, in human smooth
muscle cells, the uptake of both Ac-LDL and native LDL was up-regulated
by the combination of the four growth factors (Fig. 2B).
Figure 2:
Comparison of the effect of growth factors
on the internalization of DiI-labeled Ac-LDL and DiI-labeled LDL by
rabbit and human smooth muscle cells (SMC). Rabbit (panel
A) or human (panel B) smooth muscle cells were
preincubated with medium alone or with medium containing a combination
of EGF, PDGF BB, TGF-, and IGF-I for 16 h at 37 °C
before assessing their ability to internalize the DiI-labeled
lipoproteins (as described in the legend to Fig. 1). Values are
the fold increase in the uptake of DiI-labeled Ac-LDL or LDL relative
to the internalization observed in cells grown in the absence of growth
factors. The data are reported as the mean ± the range for two
independent experiments.
Whereas the internalization of
DiI-labeled Ac-LDL has been shown to be proportional to the amount of
Ac-LDL internalized and degraded in other systems, it was important to
determine directly whether the enhanced uptake of DiI-labeled Ac-LDL
induced by the growth factors also led to increased lipoprotein
metabolism in these studies. For this purpose, the smooth muscle cells
were pretreated with the combination of the four growth factors, and
the effect on the metabolism of I-Ac-LDL was determined.
The degradation of
I-labeled Ac-LDL was enhanced 3.8-fold
in human smooth muscle cells incubated with the growth factors at 37
°C for 16 h, and the cellular association of
I-Ac-LDL, which represents bound and internalized
I-Ac-LDL, was enhanced 2.9-fold (data not shown). These
data therefore confirm the results of the FACS analysis (Fig. 1B) and demonstrate growth factor-mediated
enhanced internalization and degradation of Ac-LDL by human smooth
muscle cells. Similar results were obtained in rabbit smooth muscle
cells (data not shown).
Previously we have shown that expression of types I and II scavenger receptor was increased by phorbol ester treatment of rabbit smooth muscle cells(11, 12) . To determine whether the enhancement of scavenger receptor activity by these growth factors is due to an induction of types I and II scavenger receptor expression, we performed RNase protection assays to examine scavenger receptor mRNA levels using a probe specific for both the types I and II isoforms. In this particular study scavenger receptor mRNA was not detected in rabbit smooth muscle cells under basal tissue culture conditions (Fig. 3). However, treatment of the smooth muscle cells with the combination of the four growth factors for 16 h increased the expression of both types I and II scavenger receptor mRNA. Similar induction of scavenger receptor mRNA expression was observed in human smooth muscle cells (data not shown).
Figure 3:
RNase protection assay of total RNA from
control and growth factor-treated rabbit smooth muscle cells. Total RNA
(10 µg) isolated from the control smooth muscle cells and from
smooth muscle cells treated with a mixture of EGF, PDGF BB,
TGF-, and IGF-I for 16 h at 37 °C (as described in
the legend to Fig. 1) was subjected to RNase protection assay
under the conditions described under ``Experimental
Procedures.'' bp, base pairs.
To determine the mechanism of growth factor stimulation, we first tested the effect of a tyrosine kinase inhibitor on the stimulatory effect of the growth factors on scavenger receptor activity. Incubation of rabbit smooth muscle cells with growth factors led to a 12-fold increase in scavenger receptor activity that was essentially abolished by tyrphostin 47, a tyrosine kinase inhibitor (Table 1). Tyrphostin 1, an inactive analog of tyrphostin 47, had little effect. In human smooth muscle cells the growth factor-induced increase in receptor expression was also blocked by tyrphostin 47 (Table 1). These data demonstrate that the growth factor-mediated induction of scavenger receptor activity in both rabbit and human smooth muscle cells requires tyrosine kinase activity.
In some cases tyrosine kinase-mediated signal transduction is associated with protein kinase C activation, and we have shown that protein kinase C is involved in the up-regulation of scavenger receptor activity in rabbit smooth muscle cells(10) . For these reasons we examined the effect of protein kinase C inhibitors on growth factor-induced up-regulation of scavenger receptor activity. Coincubation of rabbit smooth muscle cells with growth factors together with MDL.29,152 (50 or 100 µM), a protein kinase C inhibitor(24) , abolished the stimulatory effect of the growth factors on scavenger receptor activity (Table 1). This result suggests that protein kinase C functions in the signal transduction pathway leading to receptor up-regulation in rabbit smooth muscle cells. In contrast, MDL.29,152 (at concentrations up to 150 µM) did not block stimulation of scavenger receptor activity by growth factors in human smooth muscle cells (Table 1). The MDL.29,152 (50 µM) did, however, block the phorbol ester-induced stimulation of scavenger receptor activity in human smooth muscle cells and blocked the phorbol ester-induced differentiation (adhesion) of the human monocyte cell line THP-1, demonstrating that the inhibitor was active in human cells (data not shown). The data suggest differences between the mechanism of regulation of scavenger receptor activity by growth factors in human and rabbit smooth muscle cells.
Our data clearly demonstrate that
PDGF BB, EGF, IGF-I, and TGF- in combination
synergistically stimulate scavenger receptor activity in smooth muscle
cells. We next performed experiments to determine whether these growth
factors are the components in our preparation of platelet secretory
products responsible for stimulation of scavenger receptor activity. We
first estimated the amount of the four growth factors present in
platelet secretory products using Western blot analysis. The platelet
secretory products contained approximately 0.35 ng of
TGF-
, 0.11 ng of PDGF, and less than 0.03 ng of EGF
and IGF-I/µg (Table 2). Therefore, 40 µg of platelet
secretory products, the level that gave maximum stimulation of
scavenger receptor activity, contained
4.4 ng of PDGF,
14 ng
of TGF-
, and less than 1 ng each of EGF and IGF-I. We
next performed neutralization experiments to determine whether these
growth factors play any functional role in mediating the stimulatory
effect of platelet secretory products on scavenger receptor activity.
As shown in Fig. 4, an anti-PDGF antibody, used at a
concentration sufficient to block the effect of recombinant PDGF BB,
did not block the stimulatory effect of platelet secretory products on
scavenger receptor activity. Antibodies specific for EGF and IGF-I also
had little effect. Anti-TGF-
antibody, on the other
hand, blocked the stimulatory effect of platelet secretory products by
70%, suggesting that TGF-
is one of the active
components in platelet secretory products. However, approximately 30%
of the stimulatory activity remained, and the addition of anti-PDGF and
anti-TGF-
together or the addition of a mixture of the
four antibodies did not result in a greater inhibition of the
stimulatory effect than that observed with anti-TGF-
alone. These data indicate that TGF-
is
responsible for 70% of the stimulatory effect of platelet secretory
products on scavenger receptor activity and that the remaining 30% of
the activity is not mediated by the growth factors tested.
Two
additional experiments support the conclusion that PDGF is not the
active component in platelet secretory products. First, when 4.4 ng of
recombinant PDGF BB (the amount of PDGF estimated to be present in 40
µg of platelet secretory products, which gives maximum stimulation
of receptor activity) was used to treat the rabbit smooth muscle cells,
it failed to stimulate scavenger receptor activity, indicating that the
amount of PDGF in 40 µg of platelet secretory products is too low
to be responsible for the effect of platelet secretory products in our in vitro studies (see Fig. 5). Second, an antagonist to
PDGF (Trapidil, 100 µg/ml) which blocked the effect of recombinant
PDGF (50 ng/ml) (33) failed to block the stimulatory effect of
platelet secretory products on scavenger receptor activity (data not
shown). We also tested the ability of recombinant TGF- (14 ng) and PDGF (4.4 ng) (the amounts present in the maximally
active concentration of platelet secretory products) alone and together
to up-regulate scavenger receptor activity (Fig. 5). Neither
TGF-
nor PDGF alone or in combination increased
receptor activity to the level obtained with the platelet secretory
products, again suggesting that there are other factors besides
TGF-
in platelet secretory products which can increase
receptor activity either alone or in combination with
TGF-
.
Figure 5:
Effect of platelet secretory products,
PDGF BB, and TGF- alone or in combination on scavenger
receptor activity in rabbit smooth muscle cells (SMC). Smooth
muscle cells were pretreated with 40 µg/ml of platelet secretory
products (PSP) or with either recombinant PDGF BB or
TGF-
alone or together, at the concentrations
indicated, for 16 h at 37 °C and then assayed as described in Fig. 1.
We have shown previously that types I and II scavenger
receptor activity in rabbit smooth muscle cells can be up-regulated by
phorbol esters, platelet secretory products in serum, and secretion
products from activated
platelets(10, 11, 12) . In addition, it has
been reported that tumor necrosis factor (TNF-
) and
interferon
(IFN-
) increase scavenger receptor activity in
rabbit smooth muscle cells (17) . In the current studies we
demonstrated that growth factors stimulate scavenger receptor activity
in both human and rabbit smooth muscle cells. PDGF BB and
TGF-
increased scavenger receptor ac-tivity
approximately 4- and 2-fold, respectively, in rabbit smooth muscle
cells but not in human smooth muscle cells. EGF or IGF-I, when used
alone, had little effect on scavenger receptor activity in either human
or rabbit smooth muscle cells. However, when incubated with the cells
in combination, these growth factors showed synergistic effects on
scavenger receptor activity. In rabbit smooth muscle cells PDGF BB and
either EGF or TGF-
, and EGF together with
TGF-
, had synergistic effects in raising scavenger
receptor activity, and the addition of all three growth factors
together increased receptor activity 12-fold. In human smooth muscle
cells EGF and TGF-
clearly had a synergistic effect on
the induction of scavenger receptor activity, and the addition of
either IGF-I or PDGF BB together with these two growth factors
increased receptor activity up to 7-fold. We demonstrated that the
increase in scavenger receptor activity in both rabbit and human smooth
muscle cells by the growth factors is correlated with an increase in
type I and II scavenger receptor mRNA expression. We have shown
previously that the scavenger receptor expressed by the smooth muscle
cells had properties essentially identical to those of the bovine
macrophage types I and II scavenger
receptors(11, 12) .
In rabbit smooth muscle cells
the growth factors specifically increased scavenger receptor activity
with no effect on LDL receptor activity. In striking contrast to this,
in human smooth muscle cells both scavenger receptor and LDL receptor
activities were increased. Other investigators have demonstrated
previously that growth factors (such as TGF- and PDGF) stimulate
LDL receptor activity in human smooth muscle
cells(34, 35) . The reasons for the difference in
regulation of LDL receptor activity in the rabbit and human smooth
muscle cells remain to be determined; however, they could be related to
differences in the signal transduction pathways in the rabbit and human
cells or to differences in transcription factor binding sites in the
human and rabbit LDL receptor genes.
Our results concerning regulation of scavenger receptor activity by growth factors differ slightly from those of Inaba et al.(36, 37) , in which they observed a stimulatory effect of PDGF BB or EGF alone on scavenger receptor activity in human smooth muscle cells. We noted a slight stimulatory effect of PDGF BB alone but did not see an effect of EGF alone in two lines of human smooth muscle cells. The difference in these results may be related to the difference in experimental conditions or may simply reflect differences in the properties of various lines of human smooth muscle cells.
Whereas scavenger receptor activity in smooth muscle cells is
regulated over a wide range, in differentiated macrophages scavenger
receptor activity can be regulated in a narrow range by various growth
factors and cytokines. Incubation of macrophages with macrophage
colony-stimulating factor results in an increase in scavenger receptor
activity(38) , whereas incubation of macrophages with
TGF-(39) ,
IFN-
(40, 41, 42) , and TNF-
(43) reduces receptor activity. The reason for the differential
effects of IFN-
, TNF-
, and TGF-
on scavenger
receptor activity in smooth muscle cells and macrophages is unknown.
The regulation of scavenger receptor activity in smooth muscle cells
and macrophages, however, is fundamentally different. In macrophages
scavenger receptor expression is constitutively stimulated, and they
express a high level of receptor activity. In smooth muscle cells
scavenger receptor expression is low in the absence of stimulation.
As we have shown, platelet secretory products regulate scavenger receptor activity in smooth muscle cells. Platelet secretory products can also affect scavenger receptor activity in macrophages. It has been reported that activated platelets secrete a protein-like factor that stimulates scavenger receptor activity in macrophages (44) and that platelets secrete a ligand for the scavenger receptor which competitively inhibits the binding of modified LDL to the scavenger receptor(45, 46) . Several platelet secretory products, including serotonin, fibrinogen, fibronectin, and PDGF, inhibit scavenger receptor activity in human monocyte-derived macrophages(47) . The current data demonstrating that growth factors present in platelet secretory products stimulate scavenger receptor activity in smooth muscle cells suggest that platelets in atherosclerotic lesions could modulate lipid accumulation in both smooth muscle cells and macrophages.
The recent studies of Li et
al.(17) demonstrate that after balloon injury, the aorta
smooth muscle cells in the neointima of hypercholesterolemic rabbits
express scavenger receptors, whereas smooth muscle cells in the media
do not. These data clearly demonstrate that scavenger receptor
expression is up-regulated in smooth muscle cells in atherosclerotic
lesions. The factors leading to scavenger receptor regulation in smooth
muscle cells in vivo are unknown. However, following
deendothelialization induced by balloon injury, extensive platelet
deposition would occur, making growth factors secreted by activated
platelets available to regulate scavenger receptor
activity(48, 49, 50) . In fact, platelets are
not the only source of growth factors in the atherosclerotic lesion.
Injured endothelial cells, activated macrophages, and smooth muscle
cells themselves all secret growth factors that we have demonstrated to
stimulate scavenger receptor activity in smooth muscle
cells(22) . It has been shown that the expression of PDGF,
IGF-I, TGF-, and EGF is increased in atherosclerotic
lesions(19, 20, 21, 22, 23, 51, 52) ,
suggesting that they could play a role in regulation of scavenger
receptor activity in smooth muscle cells. Although platelets are not an
early component of atherosclerotic lesions, in the absence of
deendothelialization, macrophages are. In fact, macrophages are the
primary component of early fatty streak lesions, present initially in
vast excess to the migrating and proliferating smooth muscle cells.
These macrophages are known to secret PDGF, EGF, IGF-I, and
TGF-
, as well as IFN-
and TNF-
(22) ,
all of which regulate scavenger receptor activity in smooth muscle
cells in vitro. We propose that these growth factors and
cytokines are responsible for the regulation of scavenger receptor
activity in atherosclerotic lesions.
Thus far, little is known about the mechanisms of stimulation of scavenger receptor activity in macrophages and smooth muscle cells. Wu et al.(53) have shown that scavenger receptor expression in human monocyte-derived macrophages is regulated via a signal transduction pathway involving ras, Ap1 (c-jun and junB), and ets domain proteins. Our preliminary data show that stimulation of scavenger receptor activity by growth factors requires cellular tyrosine kinase activity in both human and rabbit smooth muscle cells. It has been shown that the binding of EGF, IGF-I, and PDGF to their corresponding receptors activates receptor-associated tyrosine kinase activity followed by a host of intracellular signal events including activation of the ras-mitogen-activated protein kinase pathway, which in turn can activate jun family proteins(54) . It will be important to determine whether or not the ras-mitogen-activated protein kinase pathway and jun proteins are involved in the growth factor-induced stimulation of scavenger receptor activity in smooth muscle cells.
It has been proposed that modified LDL is the atherogenic ligand for types I and II scavenger receptor on macrophages which causes the massive accumulation of lipid and foam cell formation(5, 6, 7, 8, 55, 56, 57, 58) . We have demonstrated that smooth muscle cells express types I and II scavenger receptors that bind, internalize, and degrade modified LDL(10, 11, 12) , suggesting that the scavenger receptor-mediated uptake of modified LDL by smooth muscle cells may also lead to lipid accumulation and foam cell formation in vivo as proposed in macrophages. The data in the current study suggest that growth factors secreted by cells in developing atherosclerotic lesions could stimulate scavenger receptor activity in smooth muscle cells and contribute to lipid accumulation and foam cell formation.