© 1999 American Heart Association, Inc.
Cellular Biology |
Human Activin-A Is Expressed in the Atherosclerotic Lesion and Promotes the Contractile Phenotype of Smooth Muscle Cells
From the Academic Medical Center (M.A.E., J.M.N., T.A.E.v.A., B.E.v.A., H.P., C.J.M.d.V.), University of Amsterdam, Department of Biochemistry, Amsterdam and Erasmus University (R.H.N.v.S.), Department of Endocrinology and Reproduction, Rotterdam, The Netherlands.
Correspondence to Carlie J.M. de Vries, PhD, Department of Biochemistry, Academic Medical Center K1-163, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands. E-mail c.j.devries{at}amc.uva.nl
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Abstract—Activin is a member of the transforming growth factor-ß superfamily, and it modulates the proliferation and differentiation of various target cells. In this study, we investigated the role of activin in the initiation and progression of human atherosclerosis. The expression of activin, its physiological inhibitor follistatin, and activin receptors were assayed in human vascular tissue specimens that represented various stages of atherogenesis. In situ hybridization experiments revealed activin mRNA in endothelial cells and macrophages and a strong induction of activin expression in neointimal smooth muscle cells from the early onset of atherogenesis. We developed an "in situ free-activin binding assay" by using biotinylated follistatin, which allowed us to detect bioactive activin at specific sites in atherosclerotic lesions. The mRNAs encoding the activin receptors are expressed similarly in normal and atherosclerotic tissue, which indicates that activin-A signaling in atherogenesis is most likely dependent on changes in growth factor concentrations rather than on receptor levels. In vitro, activin induces the contractile, nonproliferative phenotype in cultured smooth muscle cells, as is reflected by increased expression of smooth muscle-specific markers (SM
-actin and SM22). Our data provide evidence that
activin induces redifferentiation of neointimal smooth
muscle cells, and we hypothesize that activin is involved in
plaque stabilization.
Key Words: atherosclerosis • smooth muscle cell • follistatin • transforming growth factor-ß
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Introduction |
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Activin belongs to the transforming growth factor-ß (TGF-ß) superfamily of growth factors, and it was initially identified as a protein that induces the release of follicle-stimulating hormone by pituitary cells.1 2 Subsequently, it was demonstrated that activin is involved in the growth and differentiation of various cell types.3 4 5 6 7 8 The glycoprotein activin consists of ßA and ßB chains, which are held in homodimeric or heterodimeric configurations by disulfide bonds. The distantly related inhibin-
subunit can dimerize
with either of the activin-ß subunits, resulting in inhibin A or
inhibin B, which inhibit follicle-stimulating hormone release
from pituitary cells.9 Activin signaling is mediated by
cell-surface type I and type II activin receptors (ActR). Two type I
receptors have been described, ActR-I (ALK-2) and ActR-IB (ALK-4), as
have 2 type II receptors, ActR-II and ActR-IIB.10 The
activity of activin is regulated by follistatin, a 34-kDa
glycoprotein, which binds activin with high affinity in
equimolar complexes that are unable to bind and activate the
activin receptors.11 12 A potential role for activin in atherogenesis has been proposed on the basis of the observation that this factor modulates the proliferation and differentiation of several cell types that are involved in atherogenesis, notably endothelial cells (ECs), macrophages, and smooth muscle cells (SMCs). Activin inhibits the propagation of human ECs,13 and it enhances the differentiation of monocytic cells into macrophages.14 Most remarkably, with respect to activin function in atherogenesis, foam cell formation of THP-1–derived macrophages is inhibited.15 The effect of activin on cultured SMCs is controversial; activin has induced DNA synthesis in rat SMCs in some studies,16 17 whereas others reported that activin did not affect rat SMC growth.18 In separate studies, activin19 and follistatin20 were expressed in atherosclerotic lesions of hyperlipidemic rabbits, and enhanced activin expression was also observed in the rat carotid artery after balloon injury.17
In the present study, we investigated the role of activin in human atherogenesis in detail. In situ hybridization of human tissue specimens demonstrated the expression of activin receptors in SMCs, ECs, and macrophages, as well as the distinct expression of activin and follistatin in atherosclerotic lesions. Immunohistochemical examinations revealed partial colocalization of activin and follistatin protein; this observation prompted us to design a method to detect free, bioactive activin in atherosclerotic lesions. We took advantage of the knowledge that free activin (not bound in complexes) can associate with follistatin, and we developed a new, highly specific "in situ free-activin binding assay" to show the presence of significant amounts of bioactive activin in advanced atherosclerotic lesions. To reveal the function of activin in human atherogenesis, we evaluated the expression of activin, activin receptors, and follistatin in cultured SMCs. Finally, we demonstrated that activin mediates the differentiation of proliferating SMCs toward a contractile phenotype, as was illustrated by the increased expression of smooth muscle (SM)–specific genes.
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Materials and Methods |
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Tissue Specimens
Human tissue samples were obtained from organ donors or from patients undergoing vascular surgery with the informed consent of relatives or the patients, according to protocols approved by the Medical Ethical Committee of the Academic Medical Center of the University of Amsterdam. The specimens were fixed in 3.8% vol/vol formaldehyde in PBS and embedded in paraffin or snap-frozen in liquid nitrogen within 5 minutes after resection. After sectioning (5 µm) and mounting on 3-aminopropyl-triethoxysilane–coated glass slides, the specimens were subjected to in situ hybridization, immunohistochemistry, or the in situ free-activin binding assay. Table 1
describes the specimens.
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Immunohistochemistry
Antibody 1A4 (DAKO) recognizes SM -actin and was used
to detect SMCs, whereas antibody HAM56 (DAKO) was used to recognize
macrophages in the sections. Activin was detected with
monoclonal antibody E4 (Serotec), and follistatin with an anti-peptide
antibody (provided by Prof S. Werner, ETH, Zurich, Switzerland).
Immunohistochemical examinations were performed on 5-µm paraffin
sections (1A4, HAM56) or acetone-fixed cryosections (E4,
anti-follistatin) of tissue specimens. For pretreatment, the sections
were rehydrated, incubated with 0.3% hydrogen peroxide to block
endogenous peroxidase activity, and blocked with 10%
vol/vol preimmune goat serum (DAKO) in Tris-buffered saline (TBS;
10 mmol/L Tris [pH 8.0] and 150 mmol/L NaCl). Subsequently,
the sections were incubated with specific antibodies, followed by
incubation with biotinylated secondary antibodies, which were detected
with streptavidin-horseradish peroxidase conjugates (DAKO). ECs were
specifically recognized with Ulex europaeus lectin, which
was detected with an anti–Ulex lectin-horseradish peroxidase
conjugate. Peroxidase activity was visualized with aminoethylcarbazole
and hydrogen peroxide. After counterstaining with hematoxylin, the
sections were embedded in glycergel (Sigma).
In Situ Hybridization/RNase Protection Assay
In vitro transcription of linearized plasmid DNA was performed
to obtain radiolabeled anti-sense or sense riboprobes
([35S]-UTP for in situ hybridization and
[32P]-UTP for RNase protection [Amersham]);
see Table 2 for details on probes. The in
situ hybridization assays and RNase protection assays were performed as
described previously.22 As a control for the specificity
of the anti-sense riboprobes, matching sense riboprobes were assayed
for each gene; the sense probes gave neither background nor an
aspecific signal.
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In Situ Free-Activin Binding Assay
Ten micrograms of purified follistatin (see Growth Factor
Incubation and Total RNA Isolation) was biotinylated for 2 hours at
0°C with 3.25 µg Sulfo-N-hydroxysuccinimide-long
chain (5 µg/µL in dimethylsulphoxide; Pierce) in PBS at a
total volume of 20 µL. The reaction was stopped with 1 µL 1 mol/L
Tris-HCl (pH 8.0). The biotinylated follistatin was purified on a
Sephadex G25 column (Pharmacia). Subsequently, the preparation was
incubated on cryosections that were pretreated as described in
Immunohistochemistry. After 2 hours of incubation at room temperature
in 250 nmol/L biotinylated follistatin and TBS containing 1%
bovine serum albumin, the sections were washed 3 times for
2 minutes with TBS and incubated with avidin-horseradish
peroxidase conjugate, which was visualized with aminoethylcarbazole
(see Immunohistochemistry).
Cell Culture
SMCs were derived from human vessel explants and originated from
the iliac artery and aorta of organ donors. The cultured SMCs were
characterized by immunofluorescence with a murine
monoclonal antibody directed against SM -actin (1A4, DAKO), which
was detected with a Cy3-conjugated goat anti-mouse antibody (Jackson
Laboratories). With this method, the cells show uniform fibrillar
staining. The cells were used at passage 5 to 7. Cell cultures were
performed at 37°C in a humidified 5% CO2
chamber in 40% Medium-199 with L-glutamine/L-amino acids, 40% RPMI
1640 with HEPES buffer/L-glutamine, and 20% vol/vol human serum
supplemented with penicillin, streptomycin, and Fungizone (GIBCO
BRL).
Growth Factor Incubation and Total RNA Isolation
Purified, recombinant human activin-A [lot 15365-36(1)]
and follistatin (lot B3904) were obtained from Dr Pawson through the
National Hormone and Pituitary Program, the National Institute of
Diabetes and Digestive and Kidney Disease, the National Institute of
Child Health and Human Development, and the US Department of
Agriculture (Bethesda, Md). For growth factor incubations, SMCs
were made quiescent for 72 hours in serum-free standard medium
containing 10 mg/L bovine insulin, 5.5 mg/L human transferrin, and 6.7
µg/L sodium selenite (GIBCO BRL). Subsequently, recombinant
human follistatin, activin-A, or human serum was added to the culture
medium during the periods and at the concentrations indicated. After
incubation, the cells were subjected to total RNA isolation with Trizol
reagent (GIBCO BRL). After reprecipitation, the RNA samples were
subjected to RNase protection analyses and Northern blotting
procedures.
Northern Blotting
Northern blots were made as described previously23
using Hybond N nylon membranes (Amersham). SM -actin mRNA was
detected by hybridization with the oligonucleotide
5'-AGTGCTGTCCTCT-TCTTCACACATA-3' and radiolabeled with
[32P]-
ATP (Amersham). After the removal of
free nucleotides on a spin column, the probe was hybridized
overnight at 50°C in 10 mmol/L sodium phosphate (pH 6.8), 6xSSC
(20xSSC is 0.3 mol/L sodium citrate [pH 7.0] and 3 mol/L NaCl),
5xDenhardt’s (50xDenhardt’s contains 1% wt/vol Ficoll, 1% wt/vol
polyvinylpyrrolidone, and 1% wt/vol bovine serum albumin),
1 mmol/L EDTA, 0.5% wt/vol SDS, and 0.1 mg/mL salmon sperm DNA.
The blot was washed for 15 minutes at 65°C, increasing the stringency
to 2xSSC and 0.1% wt/vol SDS. SM22 mRNA was detected with a
1027-bp, full-length rat cDNA (provided by Dr Shanahan, Cambridge, UK),
radiolabeled with [32P]-ATP (Amersham) using
a random oligolabeling kit (Gibco BRL), and purified on a
oligonucleotide-removal column (Qiagen). Probe
hybridization in formamide was performed as described
previously.23 The blot was washed most stringently at
65°C with 0.1xSSC and 0.1% wt/vol SDS.
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Expression of mRNA Encoding Activin, Follistatin, and Activin Receptors in Human Vascular Tissue
We determined the expression of activin, follistatin, and activin receptor mRNA in human vascular tissue at different stages of atherosclerosis. The vascular specimens were characterized by immunohistochemistry on consecutive sections with specific antibodies to detect macrophages, SMCs, and ECs (Figures 1a
and 1d, 1b, and 1c,
respectively). To illustrate the expression of the studied genes in
lesion macrophages, we show the data from a radioactive in situ
hybridization performed in an aorta lesion containing a substantial
fibrotic core and, at the luminal side, a large infiltrate of
macrophages in Figures 1a and 1d. Both activin-ßA
(Figure 1e) and follistatin (Figure 1f) are expressed by
macrophages (Figure 1d); variations in expression levels
between individual cells are observed.
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, macrophage; and
NI, neointima (double-headed arrow). Original
magnification x25 (a) or x100 (b through j).
Only a few macrophages expressed small amounts of ActR-I mRNA
(Figure 1g
), whereas ActR-IB (Figure 1h) was expressed
more homogeneously. ActR-II and ActR-IIB were expressed by
all macrophages present in the vessel wall (Figures 1i and 1j). The type I receptors are usually expressed at
relatively low levels, which emphasizes the need to check for the
absence of background signals with corresponding sense riboprobes. Each
of the riboprobes applied in these hybridization experiments was
designed to delimit nonspecific hybridization with complementary strand
probes (data not shown).
In Figure 2, an example of the mRNA
expression of activin-ßA, follistatin, and activin receptors in
neointimal SMCs and ECs is shown. Activin-ßA
(Figure 2e) and follistatin (Figure 2f) showed
variable expression in the neointimal SMCs (Figure 2b): SMCs with high and low expression of these genes were
randomly distributed throughout the early lesion. ActR-I and ActR-IB
mRNAs (Figure 2g and 2h) were detected at relatively low levels
in subsets of SMCs, and both type II receptors (Figure 2i and 2j) were expressed at high levels in all neointimal SMCs
and in the medial SMCs. In addition, activin-ßB and inhibin mRNA
expression were not detected in macrophages, SMCs, or ECs (data
not shown).
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Because our studies revealed that the activin receptors were not
regulated in atherogenesis, we examined in more detail the expression
of activin-ßA and follistatin mRNAs, which encode soluble factors. In
Figures 3a through 3d, cross-sections of
an early aortic fibrotic lesion are shown. Follistatin mRNA (Figure 3c) was expressed at similar levels in the medial and in the
neointimal SMCs (Figure 3a). Activin-ßA mRNA
expression (Figure 3d) was, however, higher in the
neointimal SMCs than the medial SMCs. The lesion shown in
Figures 3e through 3h represents an advanced aortic
lesion with extensive neointima formation and a large
macrophage infiltrate (Figure 3f). Activin-ßA mRNA
(Figure 3h) was detected in the media, and its expression was
considerably higher in both neointimal cell types, ie, in
neointimal SMCs and in macrophages (Figures 3e and 3f). Follistatin mRNA was readily detected in medial SMCs
and, in contrast to activin-ßA expression, was not increased in the
neointima (Figures 3g and 3h). On the basis of these
data, which showed increased mRNA expression of activin in the
neointima and similar expression of follistatin in media
and neointima, we hypothesized that free activin will only
be available in the atherosclerotic lesion.
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Immunohistochemistry and In Situ Free-Activin Binding
Assay
In addition to the in situ hybridization experiments, the
expression of activin and follistatin protein was assayed with specific
antibodies. In agreement with the data obtained by the in situ
hybridization experiments, activin and follistatin protein colocalized
to some extent, as was shown in different specimens (Figures 4c, 4d, 4h, 4i, 4m, and 4n). To evaluate
the presence of free activin, which is not inactivated in
complex formation with follistatin, or other presently unknown
activin-inhibiting proteins, we developed an in situ free-activin
binding assay. This assay is based on the fact that only free,
bioactive activin can bind exogenously added follistatin. We applied
biotinylated follistatin in these experiments and, subsequently, bound
molecules were detected with streptavidin-conjugates.
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Dose-response experiments demonstrated that the signal of the
free-activin binding assay was linear and saturable. Competition
experiments, in which an incubation of the sections with excess
unbiotinylated follistatin was performed, resulting in complete
competition of the signal, confirmed the specificity of the assay (data
not shown). In early lesions (Figures 4a through 4e),
follistatin and activin were detected both in the media and the
neointima (Figures 4c and 4d, respectively), and the
in situ free-activin binding assay showed that no excess free activin
was present (Figure 4e). In advanced lesions, however,
activin and follistatin were relatively highly expressed in areas with
high densities of both macrophages and neointimal
SMCs, as is shown in Figures 4f through 4i and Figures 4k
through 4n. Free activin (Figures 4j and 4o) is detected as a
fraction of total activin protein (Figures 4i and 4n),
specifically at those sites where SMCs and macrophages are in
close proximity (Figures 4f and 4g and 4k and 4l, respectively).
In conclusion, the immunohistochemical data, in combination with the
data obtained by the in situ free-activin binding assay, show that
bioactive activin is present in the neointimal region
of the atherosclerotic vessel wall.
Expression of Activin, Follistatin, and Activin Receptor mRNA in In
Vitro–Activated SMCs
Our data obtained from the analysis of vascular specimens
demonstrated that the induction of activin in atherogenesis was most
prominent in neointimal SMCs and macrophages.
Medial SMCs present a contractile, fully differentiated
phenotype, whereas the activated, atherosclerotic,
neointimal SMCs are proliferating and/or migrating
entities. To mimic this process in atherosclerosis in
vitro, SMCs were subjected to growth stimulation, and the expression
levels of activin, follistatin, and activin receptors were determined.
SMCs were obtained as explant cultures derived either from the iliac
artery (a muscular artery) or from the elastic aorta. Because of the
different phenotypic properties of SMCs, we chose to culture the SMCs
for 3 days in serum-free medium to induce a quiescent, contractile
phenotype. Subsequently, SMC proliferation was stimulated by
the addition of 20% human serum to the culture. Then, total RNA was
isolated from the (un)stimulated SMCs and the RNA was subjected to
RNase protection analyses (Figure 5). In addition, RNase protection
analyses were performed using placenta RNA and tRNA as the
positive and negative controls, respectively.
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For each riboprobe, the hybridizations were performed
simultaneously with a GAPDH riboprobe to verify the loading
of equal amounts of RNA (Figures 5b, 5d, 5f, 5h, and 5k).
Activin-ßA mRNA expression (Figure 5a) in SMCs and serum
stimulation had no effect on activin-ßA mRNA levels in iliac artery
SMCs, although in aortic SMCs, activin mRNA was induced 2.5-fold.
Notably, we found that follistatin mRNA (Figure 5c) was
downregulated 5-fold in serum-stimulated iliac artery SMCs. This
observation was also made in aortic SMCs, although at relatively lower
expression levels. Activin-ßB and inhibin- expression were
assayed, but they could not be detected by RNase protection
analysis. It is conceivable that on atherogenic stimulation of
SMCs, the balance of follistatin and activin levels is shifted toward
an excess of free, bioavailable activin.
The expression level of ActR-I mRNA (Figure 5e) was higher in
iliac artery SMCs than in aortic SMCs; it was decreased in
serum-stimulated, iliac artery SMCs, but in aortic SMCs, serum had no
effect. RNase protection analyses for ActR-IB (Figure 5g) showed an expression pattern similar to that of ActR-I,
although overall ActR-IB mRNA expression was less pronounced. ActR-II
mRNA (Figure 5i) was downregulated 5-fold in serum-stimulated,
iliac artery SMCs; aortic SMCs displayed less abundant ActR-II mRNA
expression, which is not affected by the stimulation of these cells.
The expression of ActR-IIB mRNA (Figure 5j) was relatively low
and not substantially influenced by the different culture conditions.
These RNase protection analyses demonstrate that SMCs express
at least one type I receptor and one type II receptor.
Activin Induces the Expression of SM -Actin and SM22 mRNA
in SMCs
Arrested proliferation and the induction of differentiation in
SMCs coincides with a change from the activated and
proliferative phenotype into the nonactivated,
contractile phenotype of these cells. SMCs with a
nonproliferative, contractile phenotype express both the
SM-specific variant of -actin (SM -actin) and SM22 at high
levels.24 25 26 27 To assess the effect of activin on SMCs, we
added purified activin-A for 0, 3, 6, 12, and 24 hours to cultured SMCs
derived from the human aorta and iliac artery. In addition, the SMCs
were incubated with follistatin to block the activity of
endogenously synthesized activin. Subsequently, the
expression of the differentiation markers SM -actin and SM22 was
assayed by Northern blot analysis (Figure 6). Follistatin had no effect on the
expression of these markers in iliac artery SMCs, but the expression of
SM22 was reduced 2-fold in aortic SMCs (Figure 6d),
indicating that endogenous activin contributes to the
induction of SM22 expression in these cells. Clearly, activin-A
induces the expression of SMC-specific markers. After 24 hours of
activin treatment, a 2-fold increase in SM -actin expression was
observed in aortic SMCs (Figure 6b), whereas SM22 expression
did not change (Figure 6d). The activin treatment of iliac
artery SMCs had more pronounced effects: after 24 hours of incubation,
SM -actin and SM22 mRNA levels increased 10-fold and 5-fold,
respectively (Figures 6a and 6c). Collectively, these
observations demonstrate that activin promotes the differentiation of
proliferative SMCs into SMCs with a nonproliferative, contractile
phenotype.
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Discussion |
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In this study, we determined, for the first time, the mRNA and protein expression of both activin and follistatin in human vascular tissue at different stages of atherosclerosis. These experiments revealed the presence of activin-A in the normal media, with a higher level of expression in the lesion; follistatin was expressed at similar levels in the normal vessel wall and in the atherosclerotic plaque. Activin and follistatin colocalized to a large extent and, because follistatin can bind and neutralize the biological activity of this growth factor, it was essential to assess the presence of free, bioactive activin. A newly developed, in situ free-activin binding assay allowed us to selectively detect the presence of free activin in the advanced atherosclerotic lesion, especially in areas rich in both macrophages and neointimal SMCs. These results are consistent with data obtained by in vitro studies, showing that monocytic cells, upon differentiation into macrophages, and growth-stimulated aortic SMCs produce more activin-ßA mRNA than nonactivated cells.3 28 At this point, it should be added that neither in situ hybridization nor RNase protection analyses revealed any expression of activin-ßB and inhibin-
transcripts, which
excludes the possible presence of appreciable amounts of activin-AB,
activin-B, and inhibin in the vessel wall or in cultured SMCs. The
exclusive expression of the ßA subunit in the human vasculature is
reminiscent of the situation in rat vessels, where also only activin-A
expression has been observed.17 29 We developed a free-activin binding assay to demonstrate the presence of bioactive activin in situ. This assay is based on the high-affinity interaction between follistatin and free activin, which results in almost irreversible complex formation.30 Potential interactions of follistatin and bone morphogenetic proteins can be excluded because these interactions are highly reversible, and such complexes will dissociate during the procedure, which involves vigorous washing steps.31 The mere presence of bioactive activin, especially in the advanced atherosclerotic lesion, suggests a role for this factor in the progression of the disease. A prerequisite for functional involvement of activin in atherogenesis is, however, the presence of type I and type II activin receptors, as well as downstream signaling components. Our RNase protection analyses on cultured SMCs and in situ hybridization studies on vascular specimens demonstrated that SMCs, ECs, and macrophages express at least one type I and one type II receptor, which indicates that these cell types can bind activin and probably also respond to activin. Because the expression levels of activin receptors are similar in the media and in the neointima, in contrast to a neointimal increase in bioactive activin, activin function in atherogenesis seems to be regulated by changes in expression levels of the soluble factor, rather than by altered expression levels of its receptors.
Limited information is available on the signaling pathways involved downstream of the activin receptors, except for the binding and phosphorylation of the transcription factors Smad2 and Smad3 by the activin-receptor complex. A complex of these phosphorylated Smads and Smad4 is subsequently translocated to the nucleus of the activin-stimulated cell to directly modulate the expression of, at present, mostly unknown genes.10 Although the presence in vivo of activin signal transduction pathway components, including the Smads, needs to be confirmed, the demonstration of a physiologically relevant response of in vitro cultured cells on activin treatment supports the in vivo significance of activin in atherogenesis. Therefore, we will discuss the effect of activin on the most relevant cell types involved in atherogenesis: ECs, SMCs, and macrophages.
First, in human ECs, activin and TGF-ß inhibit DNA
synthesis,13 and we confirmed these observations (data not
shown). Second, in human SMCs derived from various vascular
origins, we showed that activin does not affect DNA synthesis, whereas
TGF-ß does (data not shown). Also, in rat SMCs, one study
showed that activin does not influence DNA
synthesis,18 although these data contrast with
results obtained by other groups, which show increased DNA synthesis by
activin.16 17 To establish the
physiological importance of activin receptors on
human SMCs and the presence of functional signaling pathways, the
effect of activin on SMC differentiation was analyzed. In vitro
cultured SMCs exhibited a significant increase in mRNA expression of
the SM-specific markers SM22 and SM -actin in response to
activin. The augmented expression of these genes is indicative of SMC
differentiation toward a nonactivated, fully differentiated
phenotype, which is commonly associated with SMCs residing in
the normal media or in the fully differentiated fibrous cap of advanced
lesions.24 25 26 27 In the present study, we observed that
iliac artery SMCs were more responsive to activin than aortic SMCs,
which may be attributed to the relatively higher expression levels of
ActR-I, ActR-IB, and ActR-II in the iliac artery SMCs (Figure 5).
To our knowledge, this is the first study showing that human SMCs originating from either the elastic aorta or the muscular iliac artery differ in their response to a growth factor. In this respect, it is of interest to note that chicken vascular SMCs reportedly exhibit opposite growth responses to TGF-ß, depending on their embryonic origin (ectodermal or mesodermal).32 These differences correlated with the extent of glycosylation of the TGF-ß type II receptors. At present, it is not known whether activin receptors on human SMCs of distinct vascular origin would be differentially glycosylated, which could result in a different response toward activin in elastic and muscular SMCs. Finally, in human THP-1–derived macrophages, activin and TGF-ß inhibit scavenger receptor expression, resulting in reduced lipid accumulation in these cells.15 33 This would suggest that activin and TGF-ß reduce atheroma foam cell formation during atherogenesis.
The distinct responses of these 3 cell types to activin illustrate the pleiotropic effects of this growth and differentiation factor and may give an indication of the functional involvement of activin in atherogenesis. It has been hypothesized that plaque stability is inversely related to macrophage and lipid content and is directly related to the number of SMCs present in the lesion.34 By contrast, intimal SMCs are thought to suppress plaque rupture and subsequent local thrombosis, thereby preventing acute clinical problems. We propose, on the basis of our knowledge that activin inhibits foam cell formation in combination with our observations that it induces the redifferentiation of vascular SMCs, that activin, like TGF-ß, may be beneficial for plaque stability.
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Acknowledgments |
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This work was supported by the Netherlands Heart Foundation (grant 95-153).
Received May 17, 1999; accepted September 8, 1999.
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References |
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