(Circulation. 2000;102:225.)
© 2000 American Heart Association, Inc.
Basic Science Reports |
Shear Stress Downregulation of Platelet-Derived Growth Factor Receptor-ß and Matrix Metalloprotease-2 Is Associated With Inhibition of Smooth Muscle Cell Invasion and Migration
From the Laboratorio di Patologia Vascolare, Istituto Dermopatico dell’Immacolata, Istituto di Ricovero e Cura a Carattere Scientifico, Rome (R.P., C.G., G.M., E.T., M.C.C.), and the Istituto di Ricerche Farmacologiche Mario Negri, Bergamo (A.R.), Italy.
Correspondence to Dr Maurizio C. Capogrossi, Laboratorio di Patologia Vascolare, Istituto Dermopatico dell’Immacolata, Via dei Monti di Creta, 104, 00167 Rome, Italy. E-mail capogrossi{at}idi.it
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Abstract |
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Background—After endovascular injury, smooth muscle cells (SMCs) may be exposed to hemodynamic shear stress (SS), and these forces modulate neointima accumulation. The effect of SS on SMC migration and invasion is unknown, and it was examined in the present study.
Methods and Results—Bovine aortic SMCs were exposed to laminar SS of 12 dyne/cm2 for 3 (SS3) or 15 (SS15) hours; control (C3 and C15) SMCs were kept under static conditions. Platelet-derived growth factor (PDGF)-BB–directed SMC migration and invasion were evaluated by a modified Boyden chamber assay with filters coated with either gelatin or reconstituted basement membrane proteins (Matrigel), respectively. SS15 inhibited both SMC migration and invasion (P<0.0001). There was no significant difference between SS3 and C3 cells. Media conditioned with SS15 cells exhibited a reduction in matrix metalloprotease-2 (MMP-2) by zymography and Western analysis. Northern blot analysis revealed no effect of SS15 on MMP-2 mRNA. In contrast, SS15 decreased MMP-2 activator and membrane-type MMP (MT-MMP or MMP-14) mRNA and protein. Furthermore, SS15 decreased PDGF receptor-ß (PDGF-Rß) mRNA and protein (P<0.05), and the SS-dependent decrease in PDGF-BB–directed cell migration was rescued by overexpressing PDGF-Rß.
Conclusions—SS inhibits SMC migration and invasion via diminished PDGF-Rß expression. This effect of SS is associated with decreased MMP-2 secretion and MT-MMP downregulation.
Key Words: stress • muscle, smooth • metalloproteinases • platelet-derived factors • restenosis
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Introduction |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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Under physiological conditions, smooth muscle cells (SMCs) are localized predominantly in the medial layer of blood vessels and are not in contact with the bloodstream. After vascular injury, however, SMCs may be exposed to hemodynamic shear stress, and in balloon-injured carotid arteries, low flow enhanced but high flow inhibited neointima accumulation.1 2 Furthermore, intimal thickening in injured carotids exposed to low flow was associated with increased matrix metalloprotease 2 (MMP-2),2 a member of a family of enzymes that digest extracellular matrix (ECM) components3 and play a key role in neointima accumulation after vascular injury.4 5 6 MMP-2 is secreted in a latent form, and membrane-type MMP (MT-MMP or MMP-14), which is an integral membrane protein rather than a secreted protein, is responsible for the activation of pro-MMP-2 on the cell surface.7 Furthermore, MMP-2 binding to the integrin
vß3 on the cell
surface leads to MMP-2 activation.8 All active MMPs are
inhibited by a family of natural inhibitors called tissue
inhibitors of matrix metalloprotease (TIMPs), which form a
1-to-1 complex with MMPs.3
In addition to MMPs, vascular injury modulates SMC expression of
platelet-derived growth factor (PDGF) and of its
receptors9 ; these effects are time-dependent, differ
between SMCs from the neointima and the media, and are
associated with functional differences in SMC responses to
PDGF.10 11 12 13 PDGF ligands consist of homodimers or
heterodimers of A and B chains, and there are 3 PDGF isoforms: AA, AB,
and BB. These PDGF isoforms bind PDGF- or -ß receptor (PDGF-R)
subunits with different affinities and induce their dimerization.
PDGF-R can bind either the A or B chain, whereas PDGF-Rß binds
only B chain with high affinity.9 The key role of PDGF in
neointima accumulation after vascular injury has been shown
by in vivo experiments. Exogenous PDGF-BB enhanced intimal
thickening,14 whereas antibodies to PDGF inhibited SMC
accumulation after angioplasty,15 16 and similar results
were obtained with an antisense oligonucleotide against
the PDGF-Rß subunit17 and a specific receptor tyrosine
kinase inhibitor.18 Furthermore, it has been
shown that endothelial cell PDGF expression was
modulated by shear stress both in vitro19 and in
vivo.20 In light of the importance of shear stress in
vascular wall remodeling after injury, the objective of the present
study was to determine in vitro whether, and eventually by what
mechanism, shear stress modulates SMC migration and invasion. The
results show that laminar shear stress inhibited both SMC migration and
invasion through a reconstituted ECM layer. These responses were
associated with diminished MMP-2 secretion, as well as reduced MT-MMP
and PDGF-Rß mRNA and protein. The shear-dependent decrease in cell
migration was abolished in SMCs forced to overexpress PDGF-Rß.
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Methods |
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SMC Isolation
Bovine aorta SMCs were isolated as described.21 Cells between passages 3 and 12 were used in all experiments, and positive staining for anti-SMC
-actin (Dako A/S) demonstrated >95%
SMC population purity.
Shear Stress Apparatus
Near-confluent SMC monolayers in DMEM with 10% FCS were exposed
to uniform fluid shear stress of 12 dyne/cm2 for
either 3 (SS3) or 15 (SS15) hours in a cone-and-plate
apparatus22 maintained at 37°C in humidified
air with 5% CO2. Control SMCs (C3 and C15) were
kept under static conditions. Cells were detached with trypsin and used
in migration/invasion assays; alternatively, cells were lysed and used
for either Northern or Western blots.
Invasion and Migration Assay
Previous studies have validated the use of a modified Boyden
chamber assay to differentiate between cell migration and cell
invasion, demonstrating the key role of MMPs in these
processes.23 24 Polycarbonate filters (8-µm pores;
Nucleopore Costar Scientific Corp) were coated with either 5 mg/L
gelatin (Sigma Chemical Co) or reconstituted basement membrane proteins
(Matrigel; Collaborative Research) for migration and invasion assays,
respectively.
PDGF-BB, 1 to 10 ng/mL (Collaborative Research), was the chemoattractant. After 4 hours of incubation, cells on the filter were fixed with ethanol and stained with toluidine. Cells from 5 randomly chosen high-power (x400) fields on the lower side of the filter were counted, and all experiments were run in triplicate.
Gelatin Zymography
SDS polyacrylamide 10% gels containing gelatin (1
mg/mL) were used to identify proteins with
gelatinolytic activity.24 Latent
collagenases were not activated with aminophenyl
mercuric acetate, and by gelatin zymography, only zymogen forms of MMPs
were identified. In the presence of SDS, otherwise inactive forms can
lyse the substrate in the gel because of detergent-induced
conformational change of the enzyme.
ELISA Test
TIMP-2 in cell culture supernatant was measured by ELISA
according to the manufacturer’s instructions (Amersham).
Northern Blot Analysis
Total RNA was isolated from SMCs and analyzed by gel
electrophoresis.5 The cDNA fragment of human MMP-2, MMP-9
(kindly provided by W.G. Stetler-Stevenson, Laboratory of Pathology,
National Institutes of Health, Bethesda, Md), MT-MMP (kindly provided
by M.T. Crow), and PDGF-Rß (kindly provided by C.H. Heldin,
Biomedical Center, Ludwig Institute for Cancer Research, Uppasala,
Sweden) were labeled with [-32P]dCTP
by the random-primer method.
Either the housekeeping GAPDH gene or the 18S rRNA was used for normalization.
Flow Cytometry
SMCs exposed to laminar flow and control cells were
analyzed by flow cytometry
(fluorescence-activated cell sorter [FACS], Becton
Dickinson).
Briefly, 2x105 cells/mL were incubated with
primary antibodies against
vß3 and
vß5 integrins, and
1-, 2-, and
ß1-integrin subunits (Chemicon) and
FITC-conjugated secondary antibodies (Dako A/S) were fixed in 1%
paraformaldehyde and analyzed.
Western Blot Analysis
Activated MMP-2 and its zymogen form as well as
PDGF-Rß and MT-MMP were detected in SMC lysates by Western blot
analysis. Conditioned media (CM) were examined for the
presence of MMP-2. Proteins (100 µg) were separated by 6% SDS-PAGE
gel under reducing conditions and blotted. The membranes were blocked
in PBS/5% nonfat dry milk, washed, and incubated with the following
primary antibodies: rabbit polyclonal anti-human PDGF-Rß IgG antibody
(Santa Cruz Inc), mouse monoclonal anti-human MT-MMP IgG antibody
(Oncogene Research Products), and mouse monoclonal anti-human MMP-2
IgG antibody (Oncogene Research Products). Horseradish
peroxidase–conjugated anti-rabbit (PDGF-Rß) and anti-mouse (MT-MMP
and MMP-2) secondary antibodies were used, and chemiluminescence was
detected according to the manufacturer’s specifications
(Amersham).
Plasmids and Transfection Methodology
These experiments were performed to examine the effect of
PDGF-Rß overexpression on SMC function. CMV.PDGF-Rß (kindly
provided by C.H. Heldin) or equal amounts of pCDNA3 (Invitrogen) empty
vector were cotransfected with either CMV.LacZ or pEGFP-N1 (Clontech)
reporter vectors (3:1 ratio). SMCs (1.7x106
cells/100-mm-diameter dish) were transfected with lipofectamine plus
reagent (Gibco BRL). After 48 hours, SMCs were exposed to shear stress
for 15 hours and studied in migration and invasion assays. Cells were
also either assayed for ß-gal activity25 26 or examined
for GFP fluorescence by UV light microscopy.27 By
this approach, it was possible to normalize results for ß-gal units
or to examine the effect of shear stress only on GFP-positive cells.
Cotransfection with 2 independent vectors results in the
internalization of both plasmids by the same cell28 ;
therefore, these experiments made it possible to determine the effect
of shear stress on PDGF-Rß–transfected cells, overcoming the
potential limitations of low transfection efficiency, ie, transfection
of 5% to 10% of the total population.
Statistical Analysis
Continuous variables were analyzed by Student’s
t test and 1-way ANOVA. Post hoc tests according to the
Student-Newman-Keuls method were used when the ANOVA P value
indicated a statistically significant difference among groups. Data are
expressed as mean±SD. A value of P
0.05 was deemed
statistically significant.
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Results |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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Effect of Shear Stress on SMC Migration and Invasion
Laminar shear stress of 12 dyne/cm2 for 3 hours did not affect SMC migration and invasion (Figure 1
). In contrast, 15 hours of exposure to
shear stress inhibited both migration (23±1 versus 33±3 cells/field
for SS15 and C15 cells, respectively; 31±3% inhibition;
P<0.0001) and invasion (10±2 versus 22±3 cells/field for
SS15 and C15 cells, respectively; 54±9.9% inhibition;
P<0.0001) (Figure 1); the inhibitory
effect was more pronounced on invasion than on migration
(P<0.05). In the absence of PDGF-BB as a chemoattractant,
there was no effect of shear stress on SMC migration (cells/field: SS3
14±3, C3 16±5, SS15 7±1, C15 8±1) and invasion (cells/field: SS3
27±9, C3 27±10, SS15 6.7±1, C15 7.5±2; n=4). This result suggests
that the inhibitory effect of SS15 was limited to
PDGF-BB–directed migration and invasion and was not due to nonspecific
inhibition of cell motility. It is noteworthy that both SMC migration
and invasion were lower in cells kept in either control or shear stress
conditions for 15 hours versus those maintained for 3 hours. Because
all experiments were started when cells had reached near-confluence and
cells continued proliferating throughout the course of the experiment,
it is possible that more marked contact inhibition of SMC motility may
account for this difference between the 15- and 3-hour experiments.
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Effect of Shear Stress on MMPs and TIMP-2
Because MMPs are required for SMCs to migrate across an ECM layer,
the effect of laminar flow on MMPs was evaluated. CM were examined for
their gelatinolytic activity by zymography, and
bands of gelatinase activity with molecular weights of 72 and 92 kDa,
which correspond to MMP-2 and MMP-9, respectively, in their secreted
forms, were identified (Figure 2). The
decrease in MMP-2 in SS3 versus C3 media was not statistically
significant, whereas a significant 40±7% decrease in the 72-kDa band
density was present in CM from SS15 versus C15 (P<0.05)
(Figure 2). SS3 and SS15 did not modulate MMP-9 gelatinase
(Figure 2).
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In these experiments, latent collagenase was not activated with aminophenyl mercuric acetate; therefore, only the 72-kDa gelatinolytic band was identified by gelatin zymography.
This band is considered to be the zymogen, and not the active MMP-2
protein (66 kDa). The same CM as used for gelatin zymography were
examined by Western analysis, and SS15 was found to decrease
both 72- and 66-kDa MMP-2 (Figure 2
).
Because MMP-2 activity is inhibited by TIMP-2 and to a lesser extent by
TIMP-1,3 29 the effect of shear stress on TIMP-2 in CM was
evaluated by ELISA, but no differences among groups were identified (C3
9±1 ng/mL versus SS3 8±0.3 ng/mL; C15 10±3.3 ng/mL versus SS15
8±0.1 ng/mL; n= 3). In additional experiments, we determined whether
SS15 induced changes in MMP-2 mRNA or MT-MMP mRNA and protein that
could account for the decrease in MMP-2 reported above. Neither MMP-2
mRNA (Figure 3) nor MMP-9 mRNA (data not
shown) levels were modulated by shear stress. In contrast, after 15
hours of shear stress, MT-MMP mRNA exhibited a 33±1% reduction versus
C15 (P<0.001; n=3), and this effect was associated with a
diminution in MT-MMP protein by Western analysis (Figure 3). These results suggest that the decrease in 66-kDa MMP-2 may
be related to the effect of shear stress to diminish MT-MMP expression
rather than to a direct effect on MMP-2 expression.
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Effect of Shear Stress on SMC Integrin Expression
Integrins expressed on the cell surface modulate migration, and it
has also recently been shown that MMP-2 colocalizes with the integrin
vß3 and that by this
mechanism, the cell may achieve directional migration.8
Therefore, we examined the distribution of
vß3 as well as of
vß5 integrin and of
1-, 2-, and
ß1-integrin subunits on SMCs after shear
stress. Flow cytometry showed that SMCs express these integrins on
their surface but that no significant changes were induced by shear
stress for 3 and 15 hours. This experiment was repeated 3 times with
similar results (data not shown).
Effect of Shear Stress on PDGF-Rß
The results presented so far suggest that a decrease in
MMP-2 may be responsible for the effect of shear stress to inhibit SMC
invasion across a layer of reconstituted basement membrane proteins but
do not explain the diminished SMC migration in response to PDGF-BB.
Therefore, both PDGF-Rß mRNA and protein levels were evaluated; after
15 hours of exposure to shear stress, PDGF-Rß mRNA levels decreased
by 35±8% versus control (Figure 4A),
and PDGF-Rß protein expression decreased by 70±3%
(P<0.05) (Figure 4B).
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The functional role of the shear stress–mediated decrease in PDGF-Rß
was further characterized in SMCs transfected with an expression vector
carrying the cDNA for PDGF-Rß. These cells overexpressed PDGF-Rß,
as shown by Western analysis (Figure 5A, inset), and after SS15, they
exhibited no decrease in migration in response to 5 and 10 ng/mL
PDGF-BB (Figure 5A). A similar result was obtained in invasion
assays in response to 10 ng/mL PDGF-BB (not shown). In contrast, cells
transfected with a control plasmid responded to SS15 with a
decrease in migration and invasion comparable to that described in
untransfected cells (Figure 1). To avoid the potential
limitations posed by a background of untransfected SMCs, additional
experiments were performed in which PDGF-Rß was cotransfected with a
green fluorescent protein (GFP) expression vector. After
migration, only GFP-positive cells, which are expected to have been
transformed also with PDGF-Rß or the control expression
vector,28 were counted. Figure 5B shows that
PDGF-BB–directed migration of SMCs transfected with PDGF-Rß was
enhanced versus control both under static conditions and after SS15.
Furthermore, shear stress–mediated inhibition of SMC migration was
abolished. Taken together, these results confirm that shear stress
downregulation of PDGF-Rß plays a key role in the inhibition of
PDGF-BB–directed SMC migration.
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Discussion |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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The present study shows that SMC exposure to laminar flow, at arterial levels of shear stress, reduced PDGF-BB–directed cell migration and invasion. A major mechanism for this effect was the decrease in PDGF-Rß mRNA expression and protein levels, because shear-dependent inhibition of SMC migration was abolished in cells forced to overexpress PDGF-Rß. It is of interest that diminished PDGF binding10 and mitogenic response to PDGF10 13 have also been observed in SMCs derived from the intima of injured carotid arteries, suggesting the possibility that SMC exposure to hemodynamic shear stress in vivo may be a mechanism for these effects.
A second mechanism that may contribute to the effect of shear
stress to diminish SMC invasiveness across Matrigel is the decrease in
MMP-2. MMPs are required for SMCs to invade a barrier constituted by
ECM components,3 24 and lower MMP-2 levels without a
change in TIMP-2, the main MMP-2 inhibitor,29
may account, at least in part, for the results shown in Figure 1B. The decrease in MMP-2 occurred without an apparent
concomitant decrease in MMP-2 mRNA levels but was associated with a
significantly decreased expression of MT-MMP, a membrane
activator of MMP-2.7 In addition, some
integrins modulate SMC migration in vitro30 and in
vivo.31 We examined the effect of shear stress on
vß3 and
vß5 integrins as well
as on 1-, 2-, and
ß1-integrin subunits; all integrins tested were
expressed in SMCs, but none were modulated by shear stress.
Although the possibility that cell selection may occur during experimental shear stress cannot be excluded, our results suggest that shear stress–mediated inhibition of SMC migration may act in synergy with the previously shown effect of shear stress to decrease SMC proliferation21 32 to prevent intima thickening after vascular injury.
The physiological significance of our results is related to the ability of shear stress to modulate neointima accumulation after vascular injury and in vascular grafts. It has been shown that high blood flow inhibits neointima accumulation in the balloon-injured rat1 and rabbit2 carotid artery, and MMP-2 activity was lower in the blood vessels exposed to high flow versus the low-flow group.2 Other studies have shown that shear stress inhibits neointima accumulation in vascular grafts.33 34 An important role for blood flow in neointima accumulation is also supported by clinical observations that restenosis after balloon angioplasty is less likely to occur in lesions that have been widely dilated and in which the intima has been dissected,35 ie, under conditions in which SMCs are exposed to the blood stream and shear stress is more likely to be laminar than in a stenotic blood vessel. The issue could be raised that SMC migration from the media to the intima occurs in response to endovascular injury and that if only SMCs exposed to shear stress were inhibited in their migratory ability, this would be irrelevant, because these cells are on the luminal surface of vascular wall. However, because MMP-2 is secreted, it is likely that it may have functional effects not only on cells that secrete it but on all cells that are exposed to it. Thus, inhibition of MMP-2 secretion by cells exposed to shear stress may diminish migration of SMCs not directly in contact with the blood stream and may affect ECM volume and composition in neointima.
In conclusion, our results suggest that SMC exposure to shear stress may minimize the ability of SMCs to migrate from the media toward the lumen of the vessel and lead to the accumulation of neointima because of diminished SMC responsiveness to both PDGF-BB and MMP-2 activity.
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Acknowledgments |
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This work was supported in part by Biomed 2 grants BMH4-CT95-1160 and BMH4-CT97-2270. The authors thank Gabriella Ricci and Cinzia Carloni for excellent secretarial assistance.
Received September 29, 1999; revision received December 31, 1999; accepted February 7, 2000.
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 V. Gupta and K. J. Grande-Allen Effects of static and cyclic loading in regulating extracellular matrix synthesis by cardiovascular cells Cardiovasc Res, December 1, 2006; 72(3): 375 - 383. [Abstract] [Full Text] [PDF]
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J. S. Garanich, M. Pahakis, and J. M. Tarbell Shear stress inhibits smooth muscle cell migration via nitric oxide-mediated downregulation of matrix metalloproteinase-2 activity Am J Physiol Heart Circ Physiol, May 1, 2005; 288(5): H2244 - H2252. [Abstract] [Full Text] [PDF]
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P. Zahradka, G. Harding, B. Litchie, S. Thomas, J. P. Werner, D. P. Wilson, and N. Yurkova Activation of MMP-2 in response to vascular injury is mediated by phosphatidylinositol 3-kinase-dependent expression of MT1-MMP Am J Physiol Heart Circ Physiol, December 1, 2004; 287(6): H2861 - H2870. [Abstract] [Full Text] [PDF]
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 M. B. Dancu, D. E. Berardi, J. P. Vanden Heuvel, and J. M. Tarbell Asynchronous Shear Stress and Circumferential Strain Reduces Endothelial NO Synthase and Cyclooxygenase-2 but Induces Endothelin-1 Gene Expression in Endothelial Cells Arterioscler Thromb Vasc Biol, November 1, 2004; 24(11): 2088 - 2094. [Abstract] [Full Text] [PDF]
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 N. Auge, F. Maupas-Schwalm, M. Elbaz, J.-C. Thiers, A. Waysbort, S. Itohara, H.-W. Krell, R. Salvayre, and A. Negre-Salvayre Role for Matrix Metalloproteinase-2 in Oxidized Low-Density Lipoprotein-Induced Activation of the Sphingomyelin/Ceramide Pathway and Smooth Muscle Cell Proliferation Circulation, August 3, 2004; 110(5): 571 - 578. [Abstract] [Full Text] [PDF]
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 J. F. LaDisa Jr., L. E. Olson, I. Guler, D. A. Hettrick, S. H. Audi, J. R. Kersten, D. C. Warltier, and P. S. Pagel Stent design properties and deployment ratio influence indexes of wall shear stress: a three-dimensional computational fluid dynamics investigation within a normal artery J Appl Physiol, July 1, 2004; 97(1): 424 - 430. [Abstract] [Full Text] [PDF]
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 B.A. Kelly, B.C. Bond, and L. Poston Aortic adaptation to pregnancy: elevated expression of matrix metalloproteinases-2 and -3 in rat gestation Mol. Hum. Reprod., May 1, 2004; 10(5): 331 - 337. [Abstract] [Full Text] [PDF]
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S. Mattiussi, P. Turrini, L. Testolin, F. Martelli, G. Zaccagnini, A. Mangoni, L. M Barlucchi, A. Antonini, B. Illi, C. Cirielli, et al. p21Waf1/Cip1/Sdi1 mediates shear stress-dependent antiapoptotic function Cardiovasc Res, March 1, 2004; 61(4): 693 - 704. [Abstract] [Full Text] [PDF]
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S. Q. Liu, D. Tang, C. Tieche, and P. K. Alkema Pattern formation of vascular smooth muscle cells subject to nonuniform fluid shear stress: mediation by gradient of cell density Am J Physiol Heart Circ Physiol, August 7, 2003; 285(3): H1072 - H1080. [Abstract] [Full Text] [PDF]
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S. Q. Liu, C. Tieche, D. Tang, and P. Alkema Pattern formation of vascular smooth muscle cells subject to nonuniform fluid shear stress: role of PDGF-{beta} receptor and Src Am J Physiol Heart Circ Physiol, August 7, 2003; 285(3): H1081 - H1090. [Abstract] [Full Text] [PDF]
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L. Wang, M. Andersson, L. Karlsson, M.-A. Watson, D. J. Cousens, S. Jern, and D. Erlinge Increased Mitogenic and Decreased Contractile P2 Receptors in Smooth Muscle Cells by Shear Stress in Human Vessels With Intact Endothelium Arterioscler Thromb Vasc Biol, August 1, 2003; 23(8): 1370 - 1376. [Abstract] [Full Text] [PDF]
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 A. C. Lake, A. Bialik, K. Walsh, and J. J. Castellot Jr CCN5 Is a Growth Arrest-Specific Gene That Regulates Smooth Muscle Cell Proliferation and Motility Am. J. Pathol., January 1, 2003; 162(1): 219 - 231. [Abstract] [Full Text] [PDF]
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 S. Tateshima, Y. Murayama, J. P. Villablanca, T. Morino, K. Nomura, K. Tanishita, and F. Vinuela In Vitro Measurement of Fluid-Induced Wall Shear Stress in Unruptured Cerebral Aneurysms Harboring Blebs Stroke, January 1, 2003; 34(1): 187 - 192. [Abstract] [Full Text] [PDF]
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 S. Yun, A. Dardik, M. Haga, A. Yamashita, S. Yamaguchi, Y. Koh, J. A. Madri, and B. E. Sumpio Transcription Factor Sp1 Phosphorylation Induced by Shear Stress Inhibits Membrane Type 1-Matrix Metalloproteinase Expression in Endothelium J. Biol. Chem., September 13, 2002; 277(38): 34808 - 34814. [Abstract] [Full Text] [PDF]
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 B. H. Rauch, E. Bretschneider, M. Braun, and K. Schror Factor Xa Releases Matrix Metalloproteinase-2 (MMP-2) From Human Vascular Smooth Muscle Cells and Stimulates the Conversion of Pro-MMP-2 to MMP-2: Role of MMP-2 in Factor Xa-Induced DNA Synthesis and Matrix Invasion Circ. Res., May 31, 2002; 90(10): 1122 - 1127. [Abstract] [Full Text] [PDF]
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R. Palumbo, C. Gaetano, A. Antonini, G. Pompilio, E. Bracco, L. Ronnstrand, C.-H. Heldin, and M. C. Capogrossi Different Effects of High and Low Shear Stress on Platelet-Derived Growth Factor Isoform Release by Endothelial Cells: Consequences for Smooth Muscle Cell Migration Arterioscler Thromb Vasc Biol, March 1, 2002; 22(3): 405 - 411. [Abstract] [Full Text] [PDF]
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