(Hypertension. 2000;36:319.)
© 2000 American Heart Association, Inc.
Scientific Contributions |
Mechanical Strain–Induced Extracellular Matrix Production by Human Vascular Smooth Muscle Cells
Role of TGF-ß1
From the Cardiovascular Research Institute, University of Leicester, UK (B.W.); and the Department of Clinical Pharmacology, Austin and Repatriation Medical Centre, Heidleberg, Australia (C.J.O.).
Correspondence to B. Williams, MD, FRCP, Professor of Medicine and Director, Cardiovascular Research Institute, University of Leicester Clinical Sciences Bldg, Leicester Royal Infirmary, PO Box 65, Leicester LE2 7LX, UK. E-mail bw17{at}le.ac.uk
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Abstract |
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Abstract—Elevated blood pressure imposes increased mechanical stress on the vascular wall, and mechanical strain is a mitogenic stimulus for vascular smooth muscle (VSM) cells. The role of mechanical forces in regulating the production of noncellular material by VSM cells for VSM cells of human origin remains undefined. We thus investigated the effects of chronic cyclical mechanical strain on extracellular matrix (ECM) protein production by cultured human VSM cells. To simulate a blood pressure of 120/80 mm Hg, human VSM cells were repetitively stretched and relaxed by 10% to 16% of their original length with the Flexercell apparatus. Fibronectin and collagen protein concentrations, matrix metalloproteinase (MMP) activity, and transforming growth factor-ß1 (TGF-ß1) mRNA expression by human VSM cells were measured in response to mechanical strain. Exposing human VSM cells to 5 days of chronic cyclical mechanical strain increased fibronectin (+48%, P<0.01) and collagen (+50%, P<0.001) concentrations when compared with cells grown in static conditions. Mechanical strain also increased MMP-2 activity, the predominant matrix-degrading isoform (+11%, P<0.05) in human VSM cells, thus strain-induced ECM accumulation was not due to inhibition of ECM protein degradation. Strain also increased TGF-ß1 mRNA expression and the production of a soluble factor that increased ECM protein production. Moreover, a TGF-ß–blocking antibody inhibited the effect of strain-conditioned media on matrix production by human VSM cells. These results suggest that chronic cyclical mechanical strain can directly modulate the fibrogenic activity of human VSM cells by inducing ECM protein synthesis and MMP activity. This occurs, at least in part, through mechanical strain–induced TGF-ß1 production, a mechanism that could explain the increased vascular ECM deposition in hypertension.
Key Words: muscle, smooth, vascular • extracellular matrix • transforming growth factors
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Introduction |
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The vascular wall in hypertension is modified by progressive accumulation of noncellular material, for example, extracellular matrix (ECM),1 2 and the vascular smooth muscle (VSM) cell is the major source of ECM protein production within the vessel wall.3 4 In addition to affecting cross-sectional area available to flow, the accumulation of rigid ECM proteins such as collagen can affect arterial wall stiffness5 and therefore arterial compliance, pulse wave propagation, and pulse pressure.6 7 These matrix-dependent changes in vascular hemodynamics are important because they ultimately affect cardiovascular morbidity and mortality rates.7
Numerous factors have been implicated in cardiovascular matrix accumulation, notably neurohumoral factors and the vascular wall stress resulting from the increase in blood pressure per se.8 9 10 The arterial wall is exposed to significant mechanical strain during the cardiac cycle, much of which is experienced by the VSM cells.11 12 Mechanical strain has been shown to profoundly influence VSM cell growth, phenotype, and function.13 14 15 16 17 18 19 With regard to ECM production, some studies have shown increased matrix synthesis in response to mechanical strain by using VSM or glomerular mesangial cells in vitro or perfused rabbit aorta ex vivo. However, other studies have yielded conflicting results.14 20 21 22 23 24 25 Various explanations for this inconsistency have been cited, including different breeding strains, species, and ages of tissues from experimental animals. It seems likely, therefore, that the cellular response to mechanical strain may not be generic across species and that studies of human vascular cells are essential. Remarkably, to date, the effects of chronic cyclical mechanical strain on ECM protein production by human VSM cells have not been reported. This is addressed by the present study.
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Methods |
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Human VSM Cell Culture
VSM cells were obtained from human aortic or venous tissue that was obtained during surgical procedures as previously described.26 The donors were normotensive. Ethics committee approval and relatives’ consent were obtained. After removal of the endothelial layer by debridement, the VSM cell layer was diced into sections of <1 mm3. The cell suspension was then suspended in media [Ham’s F12 medium (Sigma Chemical Co), 0.5% chick embryo extract (Gibco, Life Technology), 2 mmol/L L-glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin] supplemented with 20% fetal bovine serum (FBS) (Gibco, Life Technology), and cultured in 100-mm tissue culture plates in humidified air supplemented with 5% CO2. The cell monolayers were extensively characterized to be well-differentiated VSM cells on the basis of light and electron microscopic morphology and immunohistochemistry. For experiments, human VSM cells from the second to the sixth passage were used.
Chronic Cyclical Mechanical Strain
Human VSM cells were stretched with the use of the Flexercell
Stress Unit (Flexcell Corp). The cells were grown on specialized
culture plates that contain an elastomer flexible base (Flexcell Corp).
These plates are fixed into a manifold that is placed in the
CO2 incubator as described above. A
computer-controlled vacuum is applied to the manifold, which stretches
the flexible bases of the culture plates and thereby stretches the
adherent cells. The rate, duration, and magnitude of the
stretch:relaxation cycle can be programmed to mimic the pattern of
stretch:relaxation that would be experienced by VSM cells within the
vascular wall in vivo. In these experiments, a blood pressure of
120/80 mm Hg was simulated by cyclically stretching human VSM
cells from between 10% to 16% of their length at a rate of 60
cycles/min.27
Northern Analysis
Northern analysis was performed as previously described,
with the use of a single-stranded oligonucleotide DNA
probe for human transforming growth factor
(TGF)-ß1 (Ingenius, R&D Systems) and stripped
and rehybridized with a cDNA fragment for human GAPDH (No. 9805/1;
Clontech). The resulting autoradiographs were subjected to
densitometric analysis (LKB Gelscan) to quantify the ratio of
TGF-ß1: GAPDH mRNA.
Extracellular Matrix Proteins
The rate of collagen synthesis was measured with the use of the
method of Peterkofsky and Diegelmann.28 At the end of the
experiments, media-containing collagen labeled with
3H-thymidine was removed from the culture wells
and added (vol/vol) to cold buffer containing 0.65 mol/L NaCl, 0.1
mol/L Tris (pH 7.4), 4.7 mmol/L CaCl2, and
2.5 mg/mL n-ethylmaleimide. BSA (100 µg/mL) was added as a
carrier. TCA (10%) was added to an aliquot of the mixture, and
material was allowed to flocculate for 30 minutes at 4°C. The
TCA-precipitated material was pelleted, washed twice with 5% TCA,
washed twice with cold ethanol, dried, and then dissolved in 0.1N NaOH.
The sample was then added to 4 mL of scintillant and counted (Packard
2200 CA scintillation counter).
Simultaneously, a second aliquot was digested with a highly specific collagenase (Collagenase Form III, Advance Biofactures Corp; 10 U/mL media) for 90 minutes at 37°C and then treated identically to the nondigested aliquot. The relative rate of collagen synthesis was determined, assuming that the ratio of proline residues in collagen relative to noncollagen protein is 5.4.28
Fibronectin concentrations in the cell supernatant were measured with the use of a specific ELISA with a rabbit anti-human fibronectin antibody (Sigma) in the coating buffer, a mouse monoclonal anti-fibronectin secondary antibody (Sigma, F7387), and an HRP-conjugated rabbit anti-mouse IgG (DAKO P260).
Zymography
ECM-degrading activity was detected in conditioned media as
previously described.29 Briefly, equal amounts of sample
protein were mixed with an equal volume of nonreducing Laemmeli sample
buffer and electrophoresed at 4°C in SDS/7.5% polyacrylamide
gels containing either 2 mg/mL gelatin derived from calf skin (collagen
type III; Sigma) or 1 mg/mL casein. After electrophoresis, the gels
were cleared of SDS by incubating for 1 hour with 2 changes of 2.5%
(vol/vol) Triton X-100. Gels were then incubated overnight in substrate
buffer (50 mmol/L Tris, pH 8.0, 50 mmol/L NaCl, 10
mmol/L CaCl2, and 0.05% Brij 35) at 37°C. The
gels were then stained with 0.1% Coomassie brilliant blue, and
gelatinolytic bands were size-calibrated with a
high-molecular-mass standard mixture of proteins (Sigma). As an
additional control, 5 µL of conditioned media from human HT-1080
fibrosarcoma cells (European Cell & Animal Cultures) that had been
treated with phorbol 12-myristate 13-acetate was loaded on each
gel. Zymograms were subjected to densitometric analysis with a
Sharp JX-330 densitometer.
Statistics
All results are expressed as mean±SEM. All experiments were
conducted at least in triplicate. Comparisons between the stretched and
control (static) cells were made by means of the independent-samples
t test, except where measurements were performed over time,
in which case pairwise comparisons (with Fisher’s least-squares
difference) between stretched and control cells were made if a
repeated-measures ANOVA demonstrated a value of P<0.05. All
comparisons were made with the use of a computerized statistical
package (Systat 5.0, SPSS Inc).
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Results |
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Effect of Mechanical Strain on ECM Protein Production and Cell Growth
Confluent human VSM cells in 6-well plates were rendered quiescent by serum depletion (1% FBS) for 48 hours and thereafter subjected to repetitive mechanical strain for the 24, 48, or 96 hours. For experiments lasting 96 hours, the media was refreshed after 48 hours. At the end of the experimental period, the media was exchanged for fresh media containing ascorbic acid (50 µg/mL) and 3H-proline (1 µCi/mL). After a further 24 hours of mechanical strain, the media were harvested for analysis of fibronectin or collagen concentrations, and cell numbers were counted. Control cells grown on similar flexible culture plates were treated identically but not exposed to repetitive strain.
Mechanical strain induced a time-dependent increase in production of ECM proteins by human VSM cells when compared with control (static) VSM cells (Figures 1 and 2). Moreover, increasing the magnitude of strain from 10% to 16% increased the production of fibronectin by a further 32±7% (P<0.05) and collagen by a further 27±9% (P<0.05), indicating the strain dependency of the response. The strain-induced increase in collagen production occurred irrespective of whether the human VSM cells were maintained in culture media supplemented with low or high concentrations of FBS, but the response was more marked in the presence of higher serum concentrations. The increase in ECM protein production represented an absolute increase per cell, as there was only a small strain-induced increase in human VSM cell number (stretched human VSM cells: 7.1±1.7x105 cells/mL versus control cells: 6.9±0.7x105 cells/mL).
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Effect of Stretch on Matrix Metalloproteinase Enzymes
Matrix metalloproteinase (MMP) enzymes degrade ECM proteins; thus,
the effect of mechanical strain on MMP activity was also examined.
Human VSM cells were stretched for up to 96 hours as described above.
Before the supernatant was harvested, the cells were rinsed twice
before incubating for a further 24 hours in 1.0 mL/well of serum-free
media. Consistent with previous reports,29 we
could not detect MMP enzyme activity that degraded casein, nor could we
detect MMP-1 (interstitial collagenase)
activity in cultured human VSM cells. However, both static and
stretched cells produced gelatinase-degrading enzymes that localized at
72 kDa and 96 kDa (Figure 3). The 72-kDa
band, which colocalized with MMP-2 in the fibrosarcoma cell serum, was
the predominant gelatinolytic enzyme. The size of
the 72-kDa band was significantly increased by chronic cyclical
mechanical strain (Figure 3), suggesting that mechanical strain
increases MMP-2 activity in human VSM cells.
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Human VSM Cells Produce a Soluble Factor That Increases Matrix
Production
Given that the production of collagen and fibronectin are
under the control of cytokines, we examined whether cyclical
mechanical strain might promote the release of a soluble factor with
the biological potential to increase ECM protein production.
The culture media overlying human VSM cells after their exposure to
chronic cyclical mechanical strain was removed every 48 hours and
transferred to human VSM cells grown on static plates. When compared
with conditioned media from unstretched cells, the conditioned media
from stretched cells caused a significant increase in the
production of both fibronectin and collagen proteins (Figure 4).
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Effect of Cyclical Mechanical Strain on TGF-ß mRNA
Expression
TGF-ß1 is a potent fibrogenic
cytokine that has been strongly implicated in the regulation of
ECM protein production by cardiovascular
tissues.30 To examine whether human VSM cells have the
potential to produce TGF-ß1, the expression of
TGF-ß1 mRNA was examined. Confluent human VSM
cells were rendered quiescent by serum depletion (1% FBS) for 48 hours
before being subjected to cyclical mechanical strain for up to 6 hours.
Compared with baseline, mechanical strain rapidly increased
TGF-ß1 mRNA expression, which was maximal after
120 minutes (Figure 5). The increase in
TGF-ß1 mRNA expression was strain dependent
because increasing strain from 10% to 16% further increased
TGF-ß1 mRNA expression by 32%.
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Effect of Anti-TGF Antibody on Collagen Production by Human
VSM Cells
A pan-specific antibody against TGF-ß1 was
used to determine if mechanical strain–induced collagen
production by human VSM cells was mediated by
TGF-ß1. Anti-TGF-ß1 (5
µg/mL) or equal amounts of a control antibody (
-globulin) were
added to conditioned media from static and stretched cells. In the
presence of anti-TGF-ß1 antibody, the
fibrogenic effect of stretch-conditioned media was markedly attenuated
(Figure 6). The data suggests that the
fibrogenic activity associated with the soluble factor released by
human VSM cells in response to mechanical strain was accounted for by
TGF-ß1.
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Discussion |
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Hypertension is associated with increased cardiovascular deposition and increased systemic turnover of ECM proteins.1 2 3 31 The present study demonstrates the powerful independent effects of chronic cyclical mechanical strain on ECM protein biology in human VSM cells. The study of VSM cells is highly appropriate in this context because this is the main cell type exposed to chronic cyclical stretch within the vessel wall and the cell type most responsible for the vascular deposition of ECM proteins.3 4 There was no difference in pattern or magnitude of response irrespective of whether the human VSM cells were of arterial or venous origin.
Our results also suggest that TGF-ß1 plays a key role in the regulation of mechanical strain–induced matrix synthesis by human VSM cells. This is consistent with several lines of evidence suggesting that TGF-ß1 may also play an important role in the development of hypertension-induced cardiovascular fibrosis.32 33 34 For example, the expression of TGF-ß1 mRNA is elevated in the aortas of hypertensive rats and VSM cells cultured from the spontaneously hypertensive rat.32 33 34 These previous studies do not, however, define the stimulus for increased TGF-ß1 mRNA expression in experimental hypertension. The present study demonstrates that human VSM cells express TGF-ß1 mRNA and that chronic cyclical mechanical strain is a powerful stimulus for TGF-ß1 mRNA expression by these human cells. Our observations are consistent with recent reports of strain-induced TGF-ß1 mRNA expression in nonhuman cell types, for example, cultured rabbit aortic cells35 and rat glomerular mesangial cells,36 and confirm the strain-magnitude dependency of this response. However, our study goes beyond the study of TGF-ß1 mRNA expression and uniquely demonstrates that mechanical strain promotes the release of a soluble factor that acts as an autocrine stimulus for ECM protein production. That this factor is likely to be TGF-ß1 is supported by our observations that (1) TGF-ß1 mRNA expression is increased by mechanical strain and that (2) a specific anti–TGF-ß1 antibody blocked the fibrogenic activity of the stretch-conditioned culture media.
Similar to other noncellular material, ECM protein metabolism is in dynamic equilibrium and undergoes constant turnover. ECM protein accumulation can thus arise from increased synthesis and/or decreased degradation. Matrix proteins are degraded by a family of MMP enzymes, which, depending on the subtype, can degrade collagen or elastin.29 Our studies show that human VSM cells secrete 72-kDa and 96-kDa gelatinases (MMP-2 and MMP-9) but do not secrete elastases. Rather than being inhibited by mechanical strain, the activity of MMP-2, the most abundant of the gelatinases, was significantly augmented by mechanical strain. TGF-ß1 has been implicated in the regulation of MMP production, most notably by inhibition of MMP-1 mRNA expression. Thus, we might have expected to observe a decrease in the activity of some MMPs with mechanical strain. Our studies were hindered by the acknowledged difficulty in reliably detecting the full repertoire of MMP activities produced by cultured cells, in particular, casein-degrading activity and MMP-1 activity (interstitial collagenase). We were therefore unable to reliably measure the activity of the MMPs (MMP-1) most likely to be inhibited by TGF-ß1. Nevertheless, the aforementioned results suggest that the mechanical strain–induced increase in ECM protein production by human VSM cells is accompanied by an increased turnover of some ECM proteins. The net effect of mechanical strain is a strain "dose-dependent" increase in vascular ECM protein synthesis. Although this effect is not large, it is a consistent finding, which, if maintained in vivo, would lead to extensive remodeling and accelerated deposition of vascular ECM matrix proteins in hypertensive subjects.
Quantitative changes in the rates of ECM accumulation in hypertension are important with regard to their impact on the structure and the hemodynamic performance and compliance of the vessel wall.7 However, qualitative shifts in vascular matrix composition may have equally profound effects on the overall response of the vessel wall to multiple stimuli. Matrix proteins bind to specific membrane associated integrin receptors and thereby initiate intracellular signaling.13 37 38 These matrix:integrin receptor complexes play a key role in transducing mechanical forces into a biological response.13 16 19 37 39 Specific patterns of matrix:integrin interactions appear to activate specific signaling pathways and ultimately influence the phenotype and proliferative response of VSM cells to external stimuli.40 Thus, in addition to defining the quantitative effects of mechanical strain on matrix production by human VSM cells, further studies are needed to define the qualitative shifts in matrix composition that may occur in human VSM cells in response to mechanical strain. Such qualitative shifts in matrix composition may ultimately be more important than simple accumulation with regard to the biology of the vascular wall in hypertension.
In summary, we report the first study to define the impact of chronic cyclical mechanical strain on ECM protein synthesis and degradation in human VSM cells. The results suggest that the increased vascular wall stress that occurs in hypertensive individuals would be sufficient to promote a strain "dose-dependent" increase in TGF-ß1 production by VSM cells and an associated increase in matrix accumulation. We propose that this most likely represents the main biological mechanism whereby hypertension promotes cardiovascular matrix accumulation.
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Acknowledgments |
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This work was conducted when Dr O’Callaghan was a National Heart Foundation of Australia Overseas Research Fellow in the Cardiovascular Research Institute, Faculty of Medicine and Biological Sciences, University of Leicester, UK. This work was also supported by grants from the British Heart Foundation (B. Williams).
Received October 15, 1999; first decision November 2, 1999; accepted March 23, 2000.
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 S. W. Watts, C. Rondelli, K. Thakali, X. Li, B. Uhal, M. H. Pervaiz, R. E. Watson, and G. D. Fink Morphological and biochemical characterization of remodeling in aorta and vena cava of DOCA-salt hypertensive rats Am J Physiol Heart Circ Physiol, May 1, 2007; 292(5): H2438 - H2448. [Abstract] [Full Text] [PDF]
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B. C. Isenberg, C. Williams, and R. T. Tranquillo Small-Diameter Artificial Arteries Engineered In Vitro Circ. Res., January 6, 2006; 98(1): 25 - 35. [Abstract] [Full Text] [PDF]
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 E. Mata-Greenwood, A. Grobe, S. Kumar, Y. Noskina, and S. M. Black Cyclic stretch increases VEGF expression in pulmonary arterial smooth muscle cells via TGF-{beta}1 and reactive oxygen species: a requirement for NAD(P)H oxidase Am J Physiol Lung Cell Mol Physiol, August 1, 2005; 289(2): L288 - L289. [Abstract] [Full Text] [PDF]
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 S. Lehoux and A. Tedgui Making Up and Breaking Up: The Tortuous Ways of the Vascular Wall Arterioscler Thromb Vasc Biol, May 1, 2005; 25(5): 892 - 894. [Full Text] [PDF]
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J. Herrmann, S. Samee, A. Chade, M. R. Porcel, L. O. Lerman, and A. Lerman Differential Effect of Experimental Hypertension and Hypercholesterolemia on Adventitial Remodeling Arterioscler Thromb Vasc Biol, February 1, 2005; 25(2): 447 - 453. [Abstract] [Full Text] [PDF]
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 L. Wang, H.-W. Liu, K. D. McNeill, G. Stelmack, J. E. Scott, and A. J. Halayko Mechanical Strain Inhibits Airway Smooth Muscle Gene Transcription via Protein Kinase C Signaling Am. J. Respir. Cell Mol. Biol., July 1, 2004; 31(1): 54 - 61. [Abstract] [Full Text] [PDF]
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M. KJAeR Role of Extracellular Matrix in Adaptation of Tendon and Skeletal Muscle to Mechanical Loading Physiol Rev, April 1, 2004; 84(2): 649 - 698. [Abstract] [Full Text] [PDF]
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 S. Lehoux, C. A. Lemarie, B. Esposito, H. R. Lijnen, and A. Tedgui Pressure-Induced Matrix Metalloproteinase-9 Contributes to Early Hypertensive Remodeling Circulation, March 2, 2004; 109(8): 1041 - 1047. [Abstract] [Full Text] [PDF]
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 N. S. Haque, J. T. Fallon, J. J. Pan, M. B. Taubman, and P. C. Harpel Chemokine receptor-8 (CCR8) mediates human vascular smooth muscle cell chemotaxis and metalloproteinase-2 secretion Blood, February 15, 2004; 103(4): 1296 - 1304. [Abstract] [Full Text] [PDF]
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 K. Heinemeier, H. Langberg, J. L. Olesen, and M. Kjaer Role of TGF-{beta}1 in relation to exercise-induced type I collagen synthesis in human tendinous tissue J Appl Physiol, December 1, 2003; 95(6): 2390 - 2397. [Abstract] [Full Text]
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B.-W. Wang, H. Chang, S. Lin, P. Kuan, and K.-G. Shyu Induction of matrix metalloproteinases-14 and -2 by cyclical mechanical stretch is mediated by tumor necrosis factor-{alpha} in cultured human umbilical vein endothelial cells Cardiovasc Res, August 1, 2003; 59(2): 460 - 469. [Abstract] [Full Text] [PDF]
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E. Mata-Greenwood, B. Meyrick, R. H. Steinhorn, J. R. Fineman, and S. M. Black Alterations in TGF-{beta}1 expression in lambs with increased pulmonary blood flow and pulmonary hypertension Am J Physiol Lung Cell Mol Physiol, July 1, 2003; 285(1): L209 - L221. [Abstract] [Full Text] [PDF]
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K. Grote, I. Flach, M. Luchtefeld, E. Akin, S. M. Holland, H. Drexler, and B. Schieffer Mechanical Stretch Enhances mRNA Expression and Proenzyme Release of Matrix Metalloproteinase-2 (MMP-2) via NAD(P)H Oxidase-Derived Reactive Oxygen Species Circ. Res., June 13, 2003; 92 (11): e80 - e86. [Abstract] [Full Text] [PDF]
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 K. A. Nath, S. K. R. Kanakiriya, J. P. Grande, A. J. Croatt, and Z. S. Katusic Increased Venous Proinflammatory Gene Expression and Intimal Hyperplasia in an Aorto-Caval Fistula Model in the Rat Am. J. Pathol., June 1, 2003; 162(6): 2079 - 2090. [Abstract] [Full Text] [PDF]
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J. Goldman, L. Zhong, and S. Q. Liu Degradation of alpha -actin filaments in venous smooth muscle cells in response to mechanical stretch Am J Physiol Heart Circ Physiol, May 1, 2003; 284(5): H1839 - H1847. [Abstract] [Full Text] [PDF]
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E N T P Bakker, J P Versluis, P Sipkema, J W G E VanTeeffelen, T M Rolf, J A E Spaan, and E VanBavel Differential structural adaptation to haemodynamics along single rat cremaster arterioles J. Physiol., April 15, 2003; 548(2): 549 - 555. [Abstract] [Full Text] [PDF]
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F. Wernig, M. Mayr, and Q. Xu Mechanical Stretch-Induced Apoptosis in Smooth Muscle Cells Is Mediated by {beta}1-Integrin Signaling Pathways Hypertension, April 1, 2003; 41(4): 903 - 911. [Abstract] [Full Text] [PDF]
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N. A. Haseneen, G. G. Vaday, S. Zucker, and H. D. Foda Mechanical stretch induces MMP-2 release and activation in lung endothelium: role of EMMPRIN Am J Physiol Lung Cell Mol Physiol, March 1, 2003; 284(3): L541 - L547. [Abstract] [Full Text] [PDF]
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D. J. Tschumperlin, J. D. Shively, T. Kikuchi, and J. M. Drazen Mechanical Stress Triggers Selective Release of Fibrotic Mediators from Bronchial Epithelium Am. J. Respir. Cell Mol. Biol., February 1, 2003; 28(2): 142 - 149. [Abstract] [Full Text] [PDF]
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 C. A. Houlihan, A. Akdeniz, C. Tsalamandris, M. E. Cooper, G. Jerums, and R. E. Gilbert Urinary Transforming Growth Factor-{beta} Excretion in Patients With Hypertension, Type 2 Diabetes, and Elevated Albumin Excretion Rate: Effects of angiotensin receptor blockade and sodium restriction Diabetes Care, June 1, 2002; 25(6): 1072 - 1077. [Abstract] [Full Text] [PDF]
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R. Al-Jamal and M. S. Ludwig Changes in proteoglycans and lung tissue mechanics during excessive mechanical ventilation in rats Am J Physiol Lung Cell Mol Physiol, November 1, 2001; 281(5): L1078 - L1087. [Abstract] [Full Text] [PDF]
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M. A. Rivera, M. Echegaray, T. Rankinen, L. Perusse, T. Rice, J. Gagnon, A. S. Leon, J. S. Skinner, J. H. Wilmore, D. C. Rao, et al. TGF-{beta}1 gene-race interactions for resting and exercise blood pressure in the HERITAGE Family Study J Appl Physiol, October 1, 2001; 91(4): 1808 - 1813. [Abstract] [Full Text] [PDF]
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H. M. LANGEVIN, D. L. CHURCHILL, and M. J. CIPOLLA Mechanical signaling through connective tissue: a mechanism for the therapeutic effect of acupuncture FASEB J, October 1, 2001; 15(12): 2275 - 2282. [Abstract] [Full Text] [PDF]
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H. D. Intengan and E. L. Schiffrin Vascular Remodeling in Hypertension: Roles of Apoptosis, Inflammation, and Fibrosis Hypertension, September 1, 2001; 38(3): 581 - 587. [Abstract] [Full Text] [PDF]
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 H. Ju, R. Gros, X. You, S. Tsang, M. Husain, and M. Rabinovitch Conditional and targeted overexpression of vascular chymase causes hypertension in transgenic mice PNAS, June 19, 2001; 98(13): 7469 - 7474. [Abstract] [Full Text] [PDF]
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M. J. Davis, X. Wu, T. R. Nurkiewicz, J. Kawasaki, G. E. Davis, M. A. Hill, and G. A. Meininger Integrins and mechanotransduction of the vascular myogenic response Am J Physiol Heart Circ Physiol, April 1, 2001; 280(4): H1427 - H1433. [Abstract] [Full Text] [PDF]
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J.-J. Boffa, P.-L. Tharaux, J.-C. Dussaule, and C. Chatziantoniou Regression of Renal Vascular Fibrosis by Endothelin Receptor Antagonism Hypertension, February 1, 2001; 37(2): 490 - 496. [Abstract] [Full Text] [PDF]
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