Note: the American Heart Association takes money from PhARMA,
and has been exposed as being too PhARMA friendly. The Association president released a statement
that seemed to have been written by PhARMA in 2009, when a study exposed that
Ezetimibe taken with a statin had no positive effects upon cholesterol profile.
Arteriosclerosis, Thrombosis, and Vascular
Biology. 2003; 23: 769-775
Published online before print March 27, 2003doi:
Statins Inhibit Secretion of
Metalloproteinases-1, -2, -3, and -9 From Vascular Smooth Muscle Cells and
J. Chase*, Andrew
Objective— Production of several metalloproteinases (MMPs)
from smooth muscle cells (SMCs) and macrophages causes matrix destruction and
atherosclerotic plaque instability. Statins, which inhibit HMG-CoA reductase
and hence cholesterol and isoprenoid synthesis, stabilize plaques. We
investigated whether statins inhibit MMP secretion from SMCs and macrophages.
Methods and Results— We used human saphenous vein and rabbit
aortic SMC and foamy macrophages from cholesterol-fed rabbits. Cerivastatin (50
nmol/L) inhibited inducible MMP-1, -3, and -9 secretion from human SMC by
52±19%, 71±18%, and 73±17%, respectively (P<0.01, n=3). Similar
dose-related effects of cerivastatin (50 to 500 nmol/L), simvastatin (1 to 20
μmol/L), and lovastatin (5 to 20 μmol/L) were consistent with their relative
potencies against HMG-CoA reductase. Statins also inhibited inducible MMP-1,
-3, and -9 and constitutive MMP-2 secretion but not TIMP-1 or -2 secretion from
rabbit SMC. Statins also dose-dependently inhibited MMP-1, -3, and -9 secretion
from rabbit foam cells; cerivastatin (50 nmol/L) inhibited by 68±18%, 74±14%,
and 74±14%, respectively (P<0.01, n=4). Statins similarly decreased
collagenolytic, caseinolytic, and gelatinolytic activity. Mevalonate and
geranylgeranylpyrophosphate but not squalene reversed the effects, showing
dependence on isoprenoid, not cholesterol depletion. Statins did not affect MMP
Conclusions— Statins inhibit secretion of a several MMPs
from both SMCs and macrophages, which could therefore contribute to their
Matrix metalloproteinases (MMPs) play a major role
in atherosclerosis, restenosis after angioplasty, and vein-graft stenosis by
remodelling the extracellular matrix.1,2
Matrix remodelling by MMPs liberates the vascular smooth muscle cells (VSMCs)
from their pericellular matrix cage and permits migration during responses to
Overexpression of MMPs, including MMP-1, MMP-3, and MMP-9, has been
demonstrated in human and animal atherosclerotic plaques,7–16
where it is colocalized with morphological and mechanical determinants of
plaque rupture. MMPs together can catalyze the complete destruction of
which is the main component of fibrous caps responsible for their tensile
strength. Loss of collagen leads to structural weakness and less resistance to
the mechanical stresses imposed during systole.18
This results ultimately in plaque rupture, the key event in triggering coronary
thrombosis and hence acute coronary syndromes such as unstable angina and
Expression of MMPs-1, -3, and -9 is upregulated in
cells present in atheromas, including endothelial cells,20
Inflammatory mediators, including interleukin-1 (IL-1), CD-40 ligand, and tumor
necrosis factor-α, upregulate MMP activity in vascular cells, especially in
combination with platelet-derived growth factor (PDGF) or basic fibroblast
Tissue inhibitors of metalloproteinases (TIMPs) are a family of naturally
occurring specific inhibitors of MMPs whose activity in atherosclerotic plaques
seems to correlate with decreased MMP activity30,31
and hence reduced matrix remodelling.
Statins are a structurally related group of
hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors that are widely
used to treat hyperlipidemia. Their use is associated with significant
reduction of adverse coronary events, including myocardial infarction, and a
marginal regression of plaque size.32,33
Furthermore, recent studies, both in vitro and in vivo, have suggested that the
of statins may extend to mechanisms beyond
These pleiotropic effects of statins are mediated by their ability to block the
synthesis of isoprenoid intermediates, which serve as lipid attachments for a
variety of intracellular signaling molecules, especially Rho-family small
GTP-binding proteins, whose proper membrane localization and function are
dependent on isoprenylation.34,35,37
The pleiotropic effects of statins include improving or restoring endothelial function,
inhibiting the proliferation and migration of SMCs, decreasing vascular
inflammation, and, importantly, enhancing the stability of atherosclerotic
Recently, studies demonstrated that statins reduced MMP-9 secretion by
macrophages and MMP-1 secretion from vascular endothelial cells.14,38–40
If these effects were more general to other MMP family members and other plaque
resident cells, they might have an important role in plaque stabilization. We
therefore investigated whether statins modulate MMP-1, -2, -3, and -9
expression in cultured rabbit and human VSMCs and foam cell macrophages
elicited in cholesterol-fed rabbits.
Sheep polyclonal anti-rabbit MMP-1 and MMP-3 antibodies were a generous gift
from Dr G. Murphy, University of East Anglia, Norwich, UK. Mouse anti-human
MMP-1 antibody was purchased from Chemicon International. Sheep anti-human MMP-3
antibody was purchased from the Binding Site. Human recombinant IL-1-α and
human recombinant PDGFBB were purchased from R&D System. Cerivastatin was
from Bayer, UK, Ltd, and simvastatin was from Merck Research Laboratories. All
other reagents were purchased from Sigma Chemical Company unless otherwise
Primary cultures of human saphenous vein and rabbit aortic smooth muscle
cells were prepared by modifications of the explant technique, as previously
described in detail.21
Explants were maintained in complete medium composed of DMEM containing
penicillin-streptomycin (100 U/mL and 100 μg/mL, respectively), 8 mmol/L
L-glutamine, and 15% FBS (Advanced Protein Products). After 10 to 14 days,
cells were subcultured by trypsin/EDTA treatment. Cells between passages 1
through 3 were plated at a density of 2×105 cells/well into 6-well
culture plates for zymography and Western blotting or 1×106 cells/75
cm2 flasks for RNA studies. For all experiments, subconfluent cells
were rendered quiescent by incubation in serum-free DMEM supplemented with
0.25% (vol/vol) lactalbumin hydrolysate (Gibco BRL) for 3 days. Cultures were
then exposed to fresh serum-free medium containing the appropriate
concentration of the agent under investigation for 48 hours.
Rabbit experimental foam cells were isolated from subcutaneous granulomas of
cholesterol-fed New Zealand White rabbits, as previously described.26,29
Briefly, rabbits began a 1% cholesterol diet 2 weeks before implantation of 2
to 6 polyurethane sponges (Baxter Scientific) under the dorsal skin. Sponges
remained in place for 4 to 5 weeks to allow macrophage accumulation while the
animal remained on a 1% cholesterol diet throughout. The recovered sponges were
gently squeezed over sterile test tubes, and the exudates were layered onto a
discontinuous metrizamide gradient (bottom cushion 10 mL of 10% metrizamide
[wt/vol], top 3 to 4 mL cell suspension) and centrifuged at 1200g for
15 minutes at 10°C. Foam cells were recovered from the floating layer and
washed 3 times, and aliquots were prepared for oil red O staining to confirm
lipid content and immunocytochemistry by using the rabbit macrophage-specific
marker RAM 11. One rabbit yielded ≈2×107 foam cells. Cells were
placed at a density of 5×105 cells/well into 24-well plates,
nonadherent cells were discarded after 45 minutes, and the adherent foam cells
were then exposed to fresh macrophage serum-free medium (2 g/L bicarbonate-buffered
RPMI 1640 media supplemented with 100 U/mL penicillin, 100 μg/mL streptomycin,
2 mmol/L L-glutamine, 0.25% [vol/vol] lactalbumin) containing the appropriate
concentration of the agent under investigation for 48 hours.
Zymography for MMP-9 and Reverse Zymography for TIMP Activity
MMP-9 activity was detected in conditioned media, as previously described.21
Briefly, 15-μL aliquots of conditioned media diluted 1:1 with nonreducing
Laemmli sample buffer (2×) were electrophoresed at 4°C in 7.5%
SDS-polyacrylamide gels containing 2 mg/mL gelatin derived from calf skin
collagen. For reverse zymography, culture supernatants from rabbit SMCs were
concentrated 5-fold. Aliquots (40 μL) of nonreduced media were electrophoresed
at 4°C in 12% SDS-polyacrylamide gels containing 0.5 mg/mL gelatin and 10% baby
hamster kidney cell, serum-free, conditioned media as a source of gelatinase.
In either case after electrophoresis, SDS was removed and gelatinase activity
was revealed by overnight incubation at 37°C and staining with 0.1% Coomassie
Brilliant Blue. Zymograms were quantified in the linear range by densitometry
with a GS 690 Image Analysis software system (Bio-Rad).
Western Blotting for MMP-1 and MMP-3
Western blotting was performed on conditioned media samples concentrated
10-fold by ultrafiltration using Amicon 10 centrifugal concentrators (Amicon,
Stonehouse). Samples were separated by SDS-PAGE and blotted onto a
Hybond-nitrocellulose membrane (Amersham) with the use of a semidry blotting
apparatus. Blocking of nonspecific binding and dilutions of the primary (40
μg/mL) anti-MMP1 or anti-MMP3 and secondary antibodies (1:2000, DAKO) used 5%
skimmed milk powder/Tris-buffered saline/0.2% Tween 20. Protein was visualized
using an enhanced chemiluminescence system (Amersham). Bands were quantified by
Collagenolytic, β-Caseinolytic, and Gelatinolytic Activity Assays
Freshly isolated culture supernatants were assayed for collagenolytic,
β-caseinolytic, and gelatinolytic activity on the basis of the cleavage of
fluorescently labeled substrates by using the Type I Collagenase Assay Kit,
Stromelysin Activity Assay Kit, and Type IV Collagenase Assay Kit (Yagai Corp),
respectively, according to the manufacturer’s instructions.
Semiquantitative Analysis by Reverse Transcriptase–Polymerase Chain
Total cellular RNA was prepared from 2×106 rabbit aortic VSMCs
using an RNeasy Mini kit (Qiagen) according to the manufacturer’s instructions.
Procedures for reverse transcription (RT)-polymerase chain reaction (PCR) and
the primers used to measure rabbit MMP-1, MMP-3, MMP-9, TIMP-1, TIMP-2, and
GAPDH mRNA levels have been described in our previous work.29
Cell Viability and Proliferation Studies
After harvesting conditioned media, viable cell numbers were assessed by the
3-(4,5-dimethylthiazol-2-yl)-2,-5-diphenyltetrazolium bromide (MTT) assay
(Sigma) according to the manufacturer’s instruction. Apoptosis was assayed by
using Cell Death Detection Elisaplus (Roche), a photometric enzyme
immunoassay for the quantitative determination of cytoplasmic
histone-associated DNA fragments.
Each experiment was performed at least 3 times. Data are presented as
mean±SEM and analyzed using the Student’s t test using Bonferroni
correction for multiple comparisons. P<0.05 was considered
Statins Inhibit MMP-1, -3, and -9 Production From Human VSMCs
As previously detailed,25
secretion of MMP-1, -3, and -9 from human VSMC was increased from undetectable
levels by a combination of IL-α and PDGFBB (Figure 1A).
Secretion of all 3 MMPs was decreased 52±19%, 71±18%, and 73±17%, respectively
(P<0.01, n=3) by 50 nmol/L cerivastatin, a plasma concentration
previously shown to be associated with cholesterol-lowering effects in vivo.41,42
Lovastatin at a concentration of 5 μmol/L also inhibited IL-α and
PDGFBB-stimulated MMP-1, -3, and -9 secretion by 94±5%, 78±24%, and 74±9.8%,
respectively (P<0.01, n=3) (Figure I, available online at
http://atvb.ahajournals. org), consistent with the relative potencies of
cerivastatin and lovastatin as HMGCoA reductase inhibitors.43
Interestingly, cerivastatin (not shown) and lovastatin (Figure I) also
decreased constitutive secretion of MMP-2 by 57±2.3%.
View larger version:
Figure 1. Cerivastatin inhibited
production of MMP-1, -3, and -9 by human VSMCs. Cerivastatin (50 nmol/L) had no
effect on MMP-1, -3, and -9 production in unstimulated human VSMCs but
inhibited production in response to IL-1α (20 ng/mL) and PDGFBB (20
ng/mL) over 48 hours. Values are mean±SEM of 3 separate observations. *P<0.05,
**P<0.01 compared with cells stimulated with IL-1a and PDGFBB
Lovastatin Decreases MMP-1, -2, -3, and -9 But Not TIMP-1 or -2 Secretion
From Rabbit VSMCs
Lovastatin also concentration-dependently decreased MMP-1, -3, and -9
secretion induced by IL-α and PDGFBB and constitutive MMP-2 secretion in rabbit
SMC (Figure 2A),
which demonstrates that the effect is not species-specific. In contrast, it had
no effect on constitutive TIMP-1 or -2 secretion (Figure 2B),
which implies that statin shifted the MMP/TIMP balance toward inhibition by
View larger version:
Figure 2. Dose-dependent inhibition of
MMP-1, -2, -3, and -9 but not TIMP-1 and -2 production from rabbit VSMC by
lovastatin. Quiescent RVSMCs were incubated with IL-1α (20 ng/mL), PDGFBB
(20 ng/mL), and differing concentrations of lovastatin for 48 hours. A, MMP-1
and -3 were measured in conditioned media by Western blotting, and MMP-2 and -9
by gelatin zymography and related to the production in the absence of
lovastatin. B, TIMP-1 and -2 were measured in conditioned media by reverse
zymography. Values are mean±SEM of the number of separate observations shown.
**P<0.01 vs absence of lovastatin.
Using rabbit VSMCs, we investigated whether decreased MMPs secretion was
mediated by decreases in MMP mRNA levels by semiquantitative RT-PCR. Consistent
with our previous work,23,25
mRNA levels of MMP-1, MMP-3, and MMP-9 were upregulated by combination of IL-1
with PDGFBB (Figure II, available online at http://atvb.ahajournals.org), but
MMP-2 was constitutive (not shown). TIMP-1 and TIMP-2 mRNA levels were also
constitutive. Lovastatin had no significant effect on mRNA levels of MMPs or
TIMPs (Figure II).
Statins Decrease MMP-1, -3, and -9 Secretion From Rabbit Foam Cell
In agreement with previous work,26,29
rabbit macrophage foam cells expressed MMP-1, MMP-3, and MMP-9 without
exogenous stimuli (Figure 3);
secretion of MMP-2 was much lower and could not be quantified (results not
shown). Cerivastatin concentration-dependently inhibited spontaneous MMP-1, -3,
and -9 production from rabbit foam cell macrophages (Figure 3);
cerivastatin (50 nmol/L) inhibited by 68±18%, 74±14%, and 74±14%, respectively
(P<0.01, n=4). Maximal inhibition was also observed with
simvastatin at concentrations of 1 μmol/L and greater (Figure IIIA, available
online at http://atvb.ahajournals.org) and concentration-dependently by
lovastatin greater than 1 μmol/L (Figure IIIB). Lovastatin (10 μmol/L)
inhibited MMP-1, MMP-3, and MMP-9 secretion by 79±14%, 80±10%, and 66±17%,
respectively (P<0.01, n=3) but, similarly to VSMC, had no effect on
mRNA levels for MMP-1, -3, or -9 levels by semiquantitative PCR (results not
View larger version:
Figure 3. Cerivastatin inhibited
production of MMP-1, -3, and -9 by rabbit foam cells. Cerivastatin (5 to 500
nmol/L) inhibited production of MMP-1, -3, and -9 by foam cells over 48 hours
in a dose-dependent manner. Values are mean±SEM of 3 separate observations. *P<0.05,
**P<0.01 compared with control cells.
As in rabbit SMCs, viable cell numbers were measured by MTT assay. Neither
cerivastatin nor simvastatin had any effect on MTT activity (not shown), but
lovastatin inhibited MTT metabolism at a concentration of 1 to 20 μmol/L (see
Figure IV, available online at http://atvb.ahajournals.org). However, this
effect on MTT did not seem to be mediated by loss of cell numbers, because
total protein levels were not systematically affected (not shown). Furthermore,
when we used the Cell Death Detection Elisaplus assay as a sensitive
method to detect death of foam cells, lovastatin did not lead to cell death
even at a concentration of 20 μmol/L (P>0.05, n=5) (Figure IVB). To
correct for any variation in cell numbers, the loading volumes of samples shown
in Figure IIIB were normalized according to total protein. When these data were
additionally normalized to MTT activity (solid bars in Figure IIIB), 10 and 20
μmol/L lovastatin still significantly reduced MMP-1, -3, and -9 secretion,
which demonstrates that loss of cell viability could not be the main cause of
Effects of Mevalonate and Isoprenoids on Action of Lovastatin in Rabbit
VSMCs and Foam Cells
Incubation of cells with HMG-CoA reductase inhibitors causes mevalonate
starvation. Mevalonate metabolism yields a series of isoprenoids, including the
cholesterol precursor, squalene, and geranylgeranyl-pyrophosphate (GGPP), an
important lipid attachment for the posttranslational modification of Rho
To test by which pathway statins inhibit MMPs, we attempted to rescue MMP
secretion from rabbit VSMCs with mevalonate, squalene, or GGPP in the presence
of lovastatin. Addition of squalene did not reduce the inhibitory effect of
lovastatin on MMP-1, -2, -3, and -9 secretion (Figure 4). In
contrast, the addition of mevalonate or GGPP completely reversed the effects of
lovastatin on MMP-1 and -3 and partially reversed the effect on MMP-2 and -9
View larger version:
Figure 4. Reversal by mevalonate and
isoprenoids of the effects of lovastatin on MMP-1, -2, -3, and -9 expression in
rabbit VSMCs. Quiescent rabbit VSMCs were incubated with 100 μmol/L mevalonate,
10 μmol/L squalene, or 10 μmol/L GGPP in the presence of IL-1α, PDGFBB
(20 ng/mL), and lovastatin (5 μmol/L) for 48 hours. Production of MMP-1, -2,
-3, and -9 inhibited by lovastatin was rescued by mevalonate and GGPP but not
by squalene. Values are mean±SEM of the number of separate observations shown.
*P<0.05, **P<0.01 vs absence of lovastatin. #P<0.05
vs inhibited with lovastatin alone.
Similarly to SMCs, we incubated rabbit foam cells with mevalonate, squalene,
or GGPP in the presence of lovastatin. The addition of squalene slightly
reversed the effect of lovastatin on MMP-1 secretion but had no effects on
MMP-3 or -9 (Figure
5). The addition of mevalonate (100 μmol/L) completely reversed lovastatin
effects on MMP-3 and MMP-9 and reversed the effect on MMP-1 by 92±2% (P<0.01,
n=3). The addition of GGPP completely reversed MMP-1, -3, and -9 secretion (Figure 5).
View larger version:
Figure 5. Reversal by mevalonate and
isoprenoids of the effect of lovastatin on MMP-1, -3, and -9 production in
rabbit foam cells. Rabbit foam cells were incubated with 100 μmol/L mevalonate,
10 μmol/L squalene, or 15 μmol/L GGPP in the presence of 10 μmol/L lovastatin
for 48 hours. Inhibition of MMP-1 through -3 and -9 production was reversed by
mevanolate (mev) and GGPP but not squalene (squal). Values are mean±SEM of the
number of separate observations shown. *P<0.05, **P<0.01
vs absence of lovastatin; #P<0.05, ##P<0.01 vs
Effects of Lovastatin on MMP Activity in Conditioned Media From Rabbit
VSMCs and Foam Cells
Using fluorescently labeled substrates, collagenolytic, β-caseinolyic, and
gelatinolytic activities could not be detected in rabbit SMC-conditioned media
either in the absence or presence of IL-1 and PDGFBB (data not shown). Hence,
we could not evaluate any effect of lovastatin. However, consistent with our
conditioned media from rabbit granuloma foam cells contained measurable
proteolytic activity against fluorescently labeled type I collagen, β-casein,
and gelatin substrates, which correspond to activities of MMP-1 through -3 and
-9, respectively. Each activity was strongly inhibited by lovastatin, and the
inhibition was completely reversed by mevalonate but not squalene (Figure 6). GGPP
completely reversed collagenolytic and gelatinolytic activity and partially
reversed β-caseinolytic activity by 81±10% (P<0.05, n=3).
View larger version:
Figure 6. Reversal by mevalonate and
isoprenoids of the effect of lovastatin on metalloproteinase activity in rabbit
foam cells. Collagenolytic, β-caseinolytic, and gelatinolytic activity, which
corresponded to MMP-1, -3, and -9 activity, respectively, were measured using
fluorescently labeled substrates. Values are mean±SEM of the number of separate
observations shown. *P<0.05, **P<0.01 vs absence of
lovastatin, # P<0.05, ##P<0.01 vs lovastatin alone.
In this study, we demonstrated for the first time that incubation of rabbit
and human VSMCs with several statins in vitro reduced the secretion of MMP-1,
-2, -3, and -9, but not that of TIMP-1 and -2, by a mechanism that involved
isoprenylation and was posttranscriptional. In rabbit foam cells, in which
activation of MMP-1, -3, and -9 secretion was activated in vivo, subsequent in
vitro incubation with statins also effectively inhibited MMP-1, -3, and -9
secretion and activity by a prenylation-dependent mechanism. The effects of
cerivastatin were obtained at 50 nmol/L and above, within the range of
concentrations obtained in humans with the highest clinically used dose (0.8
They are also in the range of concentrations found to be effective in previous
studies in rabbits.42,44
The dose-related effects of the other statins were obtained at 100-fold higher
concentrations, consistent with their known potencies against HMG-CoA reductase
and the higher clinical doses 10 to 80 mg/d.43,45
The concentrations of statins used in our experiments were consistent with
those found to cause other non–lipid-lowering effects.35,46
Furthermore, we established directly that mevalonate starvation is the basis
for our observed effects on MMP expression as it is for the cholesterol
lowering of statins. Hence, although all of our experiments were conducted in
vitro, we believe that they could be of relevance to the action of statins in
MMPs are expressed by both SMCs and macrophage foam cells in human
atherosclerotic plaques, as demonstrated by immunocytochemistry and in situ
Our activity measurements showed that part of MMP-1, -3, and -9 secreted by
foam cells was in an active form, consistent with in situ zymography data on
human and rabbit atherosclerotic plaques.31,48
In contrast, MMP-1, -2, -3, and -9 secreted from isolated SMC cultures in vitro
were either in a latent form or there was an excess of TIMPs so that no
proteolytic activity could be detected. Presumably, in atherosclerotic plaques,
MMP proenzymes secreted from SMCs can become activated, either through the
action of oxidative species49
or MMPs secreted from macrophages, plasmin, or other serine proteases, for
example from mast cells.50
Statins decreased secretion of a broad spectrum of MMPs from both SMC and
foamy macrophages, which implies a beneficial effect on plaque stability.
Indeed, statins have been shown previously to reduce MMP protein expression and
activities when administered to hyperlipidemic rabbits,13,14
and this is accompanied by change in plaque morphology consistent with
Moreover, the plaque-stabilizing effects of statins in animal models can
apparently be obtained even independently of cholesterol lowering,36,52
which implies a direct effect on mechanisms leading to plaque instability.
However, in previous in vivo studies,13,14,51,52
reduction of macrophage foam cell numbers and MMP activity occurred together,
and, except in the case of MMP-9, an effect on MMP secretion per se was not
demonstrated. Our experiments conducted ex vivo on foam cells produced in vivo
demonstrate that there is indeed a direct effect of statins on MMP-1 and -3
production from macrophages as well as confirming the reported effect on MMP-9.14,38
By contrast, statins did not affect the production of TIMP-1 and -2, which
potentially inhibit all of the MMP studies here and implies that statins
shifted the MMP/TIMP balance toward inactive enzymes. Consistent with this, we
showed directly in foam cells that lovastatin decreased the collagenolytic,
β-caseinolytic, and gelatinolytic activities, which are predominantly
associated with MMP-1, -3, and -9, respectively.
Investigating the mechanism underlying inhibition of MMP secretion, we first
showed rescue of MMP-1, -3, and -9 secretion by mevalonate, consistent with
bypass of the blockade of HMG-CoA reductase. The potency series
cerivastatin>>simvastatin>lovastatin is also consistent with their
known potencies against the enzyme.43,45
Treatment with lovastatin may cause mevalonate starvation inside VSMCs and foam
cells. Mevalonate metabolism yields squalene, the precursor of cholesterol and
GGPP, which is important in prenylation of proteins. For example, translocation
of Rho GTPase family members from the cytoplasm to the plasma membrane is
dependent on geranylgeranylation.35
Rescue of MMP secretion by GGPP implies that inhibition of prenylation is the
mechanism for the inhibitory effect of statins on MMP secretion. Our RT-PCR
result clearly demonstrated that lovastatin had no significant effects on mRNA
level of MMP-1, -3, and -9, even when we titrated the number of cycles of PCR
to avoid saturation. Interestingly, the previously reported inhibitory effect
of fluvastatin on MMP-9 secretion from human macrophages was accompanied by a
doubling of steady-state mRNA levels for MMP-9.38
Thus, both studies agree that statins inhibit for MMP secretion by a
posttranslational mechanism. Such a mechanism helps to explain inhibition of
secretion by statins of MMPs with widely differing transcriptional regulation.
For example MMP-1, -3, and -9 secretion in VSMC and MMP-1 and -3 secretion in
foam cells is regulated and depends on the transcription factor nuclear
factor-κB, but MMP-2 secretion in VSMC and MMP-9 secretion in macrophages is
constitutive and independent of nuclear factor-κB.29
Our results showing that secretion of TIMPs is unaffected by statins implies
that the posttranslational mechanism is selective for MMPs, not merely an
overall inhibition of protein synthesis.
Additional studies, beyond the scope of this article, will be required to
fully understand the mechanisms of how the drug affects posttranscriptional
processes for MMPs. Previous studies demonstrated that the inhibition of
geranylgeranyl transferase with L-839,867 and the inhibition of Rho by C3
exoenzyme significantly decreased production of MMPs.39,40
Rho are small GTP-binding proteins that cycle between the inactive GDP-bound
state and active GTP-bound state; they play crucial roles in diverse cellular
events such as cytoskeleton organization, membrane trafficking, secretion,
transcriptional regulation, cell growth control, and development.35
In summary, we demonstrate for the first time that statins inhibit the
secretion of a broad spectrum of MMPs from both SMC and foam cell macrophages.
The effect is mediated by inhibition of prenylation and seems to be mainly
posttranslational. Inhibition of MMP secretion could contribute to the
plaque-stabilizing potential of statins.
Previous SectionNext Section
Supported by a Fellowship to Z. Luan from the China Scholarship Council and
a Fellowship to A.J. Chase from the British Heart Foundation. We thank Dr Ray
Bush for expert husbandry of the cholesterol-fed rabbits.
authors contributed equally to this study.
- Received February 26, 2003.
- Accepted March 10, 2003.
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