Extracellular matrix-mediated remodeling and mechanotransduction in large vessels during development and disease

The vascular extracellular matrix (ECM) is synthesized and secreted during embryogenesis and facilitates the growth and remodeling of large vessels. Proper interactions between the ECM and vascular cells are pivotal for building the vasculature required for postnatal dynamic circulation. The ECM serves as a structural component by maintaining the integrity of the vessel wall while also regulating intercellular signaling, which involves cytokines and growth factors. The major ECM component in large vessels is elastic fibers, which include elastin and microfibrils. Elastin is predominantly synthesized by vascular smooth muscle cells (SMCs) and uses microfibrils as a scaffold to lay down and assemble cross-linked elastin. The absence of elastin causes developmental defects that result in the subendothelial proliferation of SMCs and inward remodeling of the vessel wall. Notably, elastic fiber formation is attenuated in the ductus arteriosus and umbilical arteries. These two vessels function during embryogenesis and close after birth via cellular proliferation, migration, and matrix accumulation. In dynamic postnatal mechano-environments, the elastic fibers in large vessels also serve an essential role in proper signal transduction as a component of elastin-contractile units. Disrupted mechanotransduction in SMCs leads to pathological conditions such as aortic aneurysms that exhibit outward remodeling. This review discusses the importance of the ECM—mainly the elastic fiber matrix—in large vessels during developmental remodeling and under pathological conditions. By dissecting the role of the ECM in large vessels, we aim to provide insights into the role of ECM-mediated signal transduction that can provide a basis for seeking new targets for intervention in vascular diseases.     

Targeted disruption of fibulin-4 abolishes elastogenesis and causes perinatal lethality in mice

Elastic fibers provide tissues with elasticity which is critical to the function of arteries, lungs, skin, and other dynamic organs. Loss of elasticity is a major contributing factor in aging and diseases. However, the mechanism of elastic fiber development and assembly is poorly understood. Here, we show that lack of fibulin-4, an extracellular matrix molecule, abolishes elastogenesis. fibulin-4-/- mice generated by gene targeting exhibited severe lung and vascular defects including emphysema, artery tortuosity, irregularity, aneurysm, rupture, and resulting hemorrhages. All the homozygous mice died perinatally. The earliest abnormality noted was a uniformly narrowing of the descending aorta in fibulin-4-/- embryos at embryonic day 12.5 (E12.5). Aorta tortuosity and irregularity became noticeable at E15.5. Histological analysis demonstrated that fibulin-4-/- mice do not develop intact elastic fibers but contain irregular elastin aggregates. Electron microscopy revealed that the elastin aggregates are highly unusual in that they contain evenly distributed rod-like filaments, in contrast to the amorphous appearance of normal elastic fibers. Desmosine analysis indicated that elastin cross-links in fibulin-4-/- tissues were largely diminished. However, expression of tropoelastin or lysyl oxidase mRNA was unaffected in fibulin-4-/- mice. In addition, fibulin-4 strongly interacts with tropoelastin and colocalizes with elastic fibers in culture. These results demonstrate that fibulin-4 plays an irreplaceable role in elastogenesis.     

mRNA expression of lysyl oxidase and matrix metalloproteinase-12 in mouse skin

Elastic fibers in the dermis play an important role in skin elasticity. The desmosine crosslinking structure constructed of lysyl oxidase (LOX) in elastic fibers contributes to elasticity, while elastic fibers are primarily degraded by one of the matrix metalloproteinases (MMPs), MMP-12. We investigated the gender differences and diurnal variation of these enzymes. Gender-based differences in LOX mRNA expression were detected, and were significantly lower in females. In contrast, higher MMP-12 mRNA expression was observed in the light period, suggesting that elastic fibers might be degraded in the light rather than the dark period.     

Genetically engineered chondrocytes overexpressing elastin improve cell retention and chondrogenesis in a three-dimensional GelMA culture system

Elastic cartilage possesses many elastic fibers and has a high degree of elasticity. However, insufficient elastic fiber production remains unsolved in elastic cartilage tissue engineering. Exogenous elastin is difficult to degrade and violates cell proliferation and migration during cartilage regeneration. Moreover, exogenous elastic fibers are difficult to assemble with endogenous extracellular matrix components. We produced genetically engineered chondrocytes overexpressing elastin to boost endogenous elastic fiber production. After identifying that genetic manipulation hardly impacted the cell viability and chondrogenesis of chondrocytes, we co-cultured genetically engineered chondrocytes with untreated chondrocytes in a three-dimensional gelatin methacryloyl (GelMA) system. In vitro study showed that the co-culture system produced more elastic fibers and increased cell retention, resulting in strengthened mechanics than the control system with untreated chondrocytes. Moreover, in vivo implantation revealed that the co-culture GelMA system greatly resisted host tissue invasion by promoting elastic fiber production and cartilage tissue regeneration compared with the control system. In summary, our study indicated that genetically engineered chondrocytes overexpressing elastin are efficient and safe for promoting elastic fiber production and cartilage regeneration in elastic cartilage tissue engineering.     

Effects of physical exercise on the elasticity and elastic components of the rat aorta

To evaluate the effects of exercise on aortic wall elasticity and elastic components, young male rats underwent various exercise regimes for 16 weeks. In the exercised rats, the aortic incremental elastic modulus decreased significantly when under physiological strain. The aortic content of elastin increased significantly and the calcium content of elastin decreased significantly in the exercised group. The accumulated data from the exercised and sedentary groups revealed that the elastin calcium content was related positively to the incremental elastic modulus. We concluded that physical exercise from an early age decreases the calcium deposit in aortic wall elastin and that this effect probably produced in the exercised rats a distensible aorta.     

36-kDa microfibril-associated glycoprotein (MAGP-36) is an elastin-binding protein increased in chick aortae during development and growth

MAGP-36 was discovered in porcine aorta in 1989 and is thought to be one of the microfibril-associated proteins. MAGP-36 has been localized on the surface of elastic fibers or laminae in immunohistochemical studies. However, its functional role in the aorta is obscure. Herein, we report on the binding activity of MAGP-36 to components of the aortic wall and its accumulation pattern in the aorta during development and growth. In vitro, MAGP-36 bound to elastin and collagen in a Ca(2+)-dependent manner, and mediated the adhesion of human aortic smooth muscle cells. This cell adhesion mostly depended on the RGD-containing domain of MAGP-36. We examined the accumulation of MAGP-36 with quantitative Western blot analysis and immunoelectron microscopy in chick aortae during development and growth. The amount of MAGP-36 increased on the surface of elastic fibers or laminae between days 14 and 34 after the start of incubation, and reached a plateau at about 53 days. This accumulation of MAGP-36 roughly correlated with an increase in blood pressure for this period. Thus, MAGP-36 might be a bridging protein that connects elastin to other components of the aortic wall and might play a role in maintaining the integrity of the aortic structure under arterial pressure.     

The infrastructure of aortic elastic fibers

Elastic fibers are composed of a central core of elastin that is amorphous and electron-lucent in conventional transmission electron micrographs and peripheral microfibrils. A complex infrastructure within the amorphous elastin of mature rat aorta is made visible by fixation and staining with a glutaraldehyde-ruthenium red mixture in phosphate buffer or osmium-ruthenium red in cacodylate buffer. The infrastructure is composed of at least two interlacing but distinct elastic structural components; a framework of circumferentially orientated microfibrils and a three-dimensional meshwork of filaments that permeate the fiber. The latter resembles a reticulum that has previously been observed in freeze-fractured and negatively stained elastin and attributed to the supramolecular organization of elastin. Microfibrils also extend from the core of the elastic fiber into the surrounding matrix where they appear to function as anchoring fibers. These observations indicate that the elastic properties of the arterial wall are an integrated function of both elastin and microfibrils.     

Electron microscopic study of the prenatal development of the thoracic aorta in the rat

Prenatal development of the thoracic aorta of the rat during the period ranging from gestational days 12 to 21 was examined by transmission electron microscopic and morphometric studies. The process of wall formation occurred in four major phases. At phase I (gestational day 12), the dorsal aorta consists of an endothelium and loosely surrounding mesenchymal cells. Collagen fibrils and fine filamentous materials are sparsely present in the intercellular space. At phase II (days 13 to 16), the mesenchymal cells begin to differentiate to myoblasts, which have small clusters of myofilaments with dense bodies, rough endoplasmic reticulum, and a discontinuous basal lamina. The differentiating cells form a few compact cell layers around the endothelium. Elastic fibers first occur sparsely in juxtacellular spaces at days 13-14. The thickness of the aorta increases rapidly from 1-3 layers of cells at day 13 to 5-8 layers at day 17, leading to a maximum of 5-9 cell layers at day 20. The differentiation of myoblasts and elastogenesis are initiated in the inner layers, and later progress toward the outer layer of the aortic wall. At phase III (days 17 to 19), the myoblasts continue to develop into typical smooth muscle cells, and elastic fibers rapidly increase in both size and number. At phase IV (day 20 and later), smooth muscle cells have well-developed myofilaments in the cell periphery, and rough endoplasmic reticulum and other organelles tend to accumulate in the apical portion of the cytoplasm. Elastic laminae appear in a few inner layers of the aortic wall.     

Unraveling the mechanism of elastic fiber assembly: The roles of short fibulins

Evolution of elastic fibers is associated with establishment of the closed circulation system. Primary roles of elastic fibers are to provide elasticity and recoiling to tissues and organs and to maintain the structural integrity against mechanical strain over a lifetime. Elastic fibers are comprised of an insoluble elastin core and surrounding mantle of microfibrils. Elastic fibers are formed in a regulated, stepwise manner, which includes the formation of a microfibrillar scaffold, deposition and integration of tropoelastin monomers into the scaffold, and cross-linking of the monomers to form an insoluble, functional polymer. In recent years, an increasing number of glycoproteins have been identified and shown to be located on or surrounding elastic fibers. Among them, the short fibulins-3, -4 and -5 particularly drew attention because of their potent elastogenic activity. Fibulins-3, -4 and -5 are characterized by tandem repeats of calcium binding EGF-like motifs and a C-terminal fibulin module, which is conserved throughout fibulin family members. Initial biochemical characterization and gene expression studies predicted that fibulins might be involved in structural support and/or matrix–cell interactions. Recent analyses of short fibulin knockout mice have revealed their critical roles in elastic fiber development in vivo. We review recent findings on the elastogenic functions of short fibulins and discuss the molecular mechanism underlying their activity in vitro and in vivo.     

Traction Force Measurements of Human Aortic Smooth Muscle Cells Reveal a Motor-Clutch Behavior

The contractile behavior of smooth muscle cells (SMCs) in the aorta is an important determinant of growth, remodeling, and homeostasis. However, quantitative values of SMC basal tone have never been characterized precisely on individual SMCs. Therefore, to address this lack, we developed an in vitro technique based on Traction Force Microscopy (TFM). Aortic SMCs from a human lineage at low passages (4-7) were cultured 2 days in conditions promoting the development of their contractile apparatus and seeded on hydrogels of varying elastic modulus (1, 4, 12 and 25 kPa) with embedded fluorescent microspheres. After complete adhesion, SMCs were artificially detached from the gel by trypsin treatment. The microbeads movement was tracked and the deformation fields were processed with a mechanical model, assuming linear elasticity, isotropic material, plane strain, to extract the traction forces formerly applied by individual SMCs on the gel. Two major interesting and original observations about SMC traction forces were deduced from the obtained results: 1. they are variable but driven by cell dynamics and show an exponential distribution, with 40% to 80% of traction forces in the range 0-10 μN. 2. They depend on the substrate stiffness: the fraction of adhesion forces below 10 μN tend to decrease when the substrate stiffness increases, whereas the fraction of higher adhesion forces increases. As these two aspects of cell adhesion (variability and stiffness dependence) and the distribution of their traction forces can be predicted by the probabilistic motor-clutch model, we conclude that this model could be applied to SMCs. Further studies will consider stimulated contractility and primary culture of cells extracted from aneurysmal human aortic tissue.     

Elastic and collagenous networks in vascular diseases

Supravalvular aortic stenosis (SVAS), Marfan syndrome (MFS) and Ehlers-Danlos syndrome type IV (EDS IV) are three clinical entities characterized by vascular abnormalities that result from mutations of structural components of the extracellular matrix (ECM). Analyses of naturally occurring human mutations and of artificially generated deficiencies in the mouse have provided insights into the pathogenesis of these heritable disorders of the connective tissue. SVAS is associated with haploinsufficiency of elastin, one of the two major components of the elastic fibers. SVAS is characterized by narrowing of the arterial lumen due to the failure of regulation of cellular proliferation and matrix deposition. Mutations in fibrillin 1 are the cause of dissecting aneurysm leading to rupture of the ascending aorta. Fibrillin-1 is the building block of the microfibrils that span the entire thickness of the aortic wall and are a major component of the elastic fibers that reside in the medial layer. The vascular hallmark of EDS IV is rupture of large vessels. The phenotype is caused by mutations in type III collagen. The mutations ultimately affect the overall architecture of the collagenous network and the biomechanical properties of the adventitial layer of the vessel wall. Altogether, these genotype-phenotype correlations document the diversified contributions of distinct extracellular macroaggregates to the assembly and function of the vascular matrix.     

Glycosaminoglycans contribute to extracellular matrix fiber recruitment and arterial wall mechanics

Elastic and collagen fibers are well known to be the major load-bearing extracellular matrix (ECM) components of the arterial wall. Studies of the structural components and mechanics of arterial ECM generally focus on elastin and collagen fibers, and glycosaminoglycans (GAGs) are often neglected. Although GAGs represent only a small component of the vessel wall ECM, they are considerably important because of their diverse functionality and their role in pathological processes. The goal of this study was to study the mechanical and structural contributions of GAGs to the arterial wall. Biaxial tensile testing was paired with multiphoton microscopic imaging of elastic and collagen fibers in order to establish the structure–function relationships of porcine thoracic aorta before and after enzymatic GAG removal. Removal of GAGs results in an earlier transition point of the nonlinear stress–strain curves \((p<0.05)\). However, stiffness was not significantly different after GAG removal treatment, indicating earlier but not absolute stiffening. Multiphoton microscopy showed that when GAGs are removed, the adventitial collagen fibers are straighter, and both elastin and collagen fibers are recruited at lower levels of strain, in agreement with the mechanical change. The amount of stress relaxation also decreased in GAG-depleted arteries \((p<0.05)\). These findings suggest that the interaction between GAGs and other ECM constituents plays an important role in the mechanics of the arterial wall, and GAGs should be considered in addition to elastic and collagen fibers when studying arterial function.     

Crosslinked elastic fibers are necessary for low energy loss in the ascending aorta

In the large arteries, it is believed that elastin provides the resistance to stretch at low pressure, while collagen provides the resistance to stretch at high pressure. It is also thought that elastin is responsible for the low energy loss observed with cyclic loading. These tenets are supported through experiments that alter component amounts through protease digestion, vessel remodeling, normal growth, or in different artery types. Genetic engineering provides the opportunity to revisit these tenets through the loss of expression of specific wall components. We used newborn mice lacking elastin (Eln^−/−) or two key proteins (lysyl oxidase, Lox^−/−, or fibulin-4, Fbln4^−/−) that are necessary for the assembly of mechanically-functional elastic fibers to investigate the contributions of elastic fibers to large artery mechanics. We determined component content and organization and quantified the nonlinear and viscoelastic mechanical behavior of Eln^−/−, Lox^−/−, and Fbln4^−/− ascending aorta and their respective controls. We confirmed that the lack of elastin, fibulin-4, or lysyl oxidase leads to absent or highly fragmented elastic fibers in the aortic wall and a 56–97% decrease in crosslinked elastin amounts. We found that the resistance to stretch at low pressure is decreased only in Eln^−/− aorta, confirming the role of elastin in the nonlinear mechanical behavior of the aortic wall. Dissipated energy with cyclic loading and unloading is increased 53–387% in Eln^−/−, Lox^−/−, and Fbln4^−/− aorta, indicating that not only elastin, but properly assembled and crosslinked elastic fibers, are necessary for low energy loss in the aorta.     

Heterogeneous distribution of isoactins in cultured vascular smooth muscle cells does not reflect segregation of contractile and cytoskeletal domains.

We have previously demonstrated that alpha-smooth muscle (alpha-SM) actin is predominantly distributed in the central region and beta-non-muscle (beta-NM) actin in the periphery of cultured rabbit aortic smooth muscle cells (SMCs). To determine whether this reflects a special form of segregation of contractile and cytoskeletal components in SMCs, this study systematically investigated the distribution relationship of structural proteins using high-resolution confocal laser scanning fluorescent microscopy. Not only isoactins but also smooth muscle myosin heavy chain, alpha-actinin, vinculin, and vimentin were heterogeneously distributed in the cultured SMCs. The predominant distribution of beta-NM actin in the cell periphery was associated with densely distributed vinculin plaques and disrupted or striated myosin and alpha-actinin aggregates, which may reflect a process of stress fiber assembly during cell spreading and focal adhesion formation. The high-level labeling of alpha-SM actin in the central portion of stress fibers was related to continuous myosin and punctate alpha-actinin distribution, which may represent the maturation of the fibrillar structures. The findings also suggest that the stress fibers, in which actin and myosin filaments organize into sarcomere-like units with alpha-actinin-rich dense bodies analogous to Z-lines, are the contractile structures of cultured SMCs that link to the network of vimentin-containing intermediate filaments through the dense bodies and dense plaques.     

Structural strain energy function applied to the ageing of the human aorta

Stiffening of the aorta with progressing age leads to decrease of aortic compliance and thus to an increase of pulse pressure amplitude. Using a strain energy function (SEF) which takes into account the composition of the arterial wall, we have studied the evolution of key structural components of the human thoracic aorta using data obtained from the literature. The SEF takes into account the wavy nature of collagen, which upon gradual inflation of the blood vessel is assumed to straighten out and become engaged in bearing load. The engagement of the individual fibers is assumed to be distributed log-logistically. The use of a SEF enables the consideration of axial stretch (lambda(z)) and residual strain (opening angle) in the biomechanical analysis. Both lambda(z) and opening angle are known to change with age. Results obtained from applying the SEF to the measurements of aortic pressure-diameter curves indicate that the changes in aortic biomechanics with progressing age are not to be sought in the elastic constants of elastin and collagen or their volume fractions of the aortic wall but moreover in alterations of the collagen mesh arrangement and the waviness of the collagen fibers. In old subjects, the collagen fiber ensemble engages in load bearing much more abruptly than in young subjects. Reasons for this change in collagen fiber dynamics may include fiber waviness remodeling or cross-linkage by advanced glycation end-products (AGE). The abruptness of collagen fiber engagement is also the model parameter that is most responsible for the decreased compliance at progressed ages.     

Angiogenesis inhibitor endostatin is a distinct component of elastic fibers in vessel walls.

Theendothelial cell inhibitor endostatin (22 kDa) is part of the carboxyl-terminal globular domain of collagen XVIII and shows a widespread tissue distribution. Immunohistology of adult mouse tissues demonstrated a preferred localization in many vessel walls and some other basement membrane zones. A strong immunogold staining was observed across elastic fibers in the multiple elastic membranes of aorta and other large arteries. Staining was less strong along sparse elastic fibers of veins and almost none was observed in the walls of arterioles and capillaries. Strong evidence was also obtained for some intracellular and basement membrane associations. Immunogold double staining of elastic fibers showed a close colocalization of endostatin with fibulin-2, fibulin-1, and nidogen-2, but not with perlecan. Reasonable amounts of endostatin could be extracted from aorta and skin by EDTA, followed by detergents, with aorta being the richest source of the inhibitor identified so far. Solubilizations with collagenase and elastase were approximately fivefold less efficient. Immunoblots of aortic extracts detected major endostatin components of 22-25 kDa whereas skin extracts also contained some larger components. Solid-phase assays demonstrated distinct binding of recombinant mouse endostatin to the fibulins and nidogen-2, consistent with their tissue colocalization. Together, the data indicate several different ways for endostatin to be associated with the extracellular matrix, and its release may determine biological activation. This also defines a novel function for some elastic tissues.     

New insights into elastic fiber assembly

Elastic fibers provide recoil to tissues that undergo repeated stretch, such as the large arteries and lung. These large extracellular matrix (ECM) structures contain numerous components, and our understanding of elastic fiber assembly is changing as we learn more about the various molecules associated with the assembly process. The main components of elastic fibers are elastin and microfibrils. Elastin makes up the bulk of the mature fiber and is encoded by a single gene. Microfibrils consist mainly of fibrillin, but also contain or associate with proteins such as microfibril associated glycoproteins (MAGPs), fibulins, and EMILIN-1. Microfibrils were thought to facilitate alignment of elastin monomers prior to cross-linking by lysyl oxidase (LOX). We now know that their role, as well as the overall assembly process, is more complex. Elastic fiber formation involves elaborate spatial and temporal regulation of all of the involved proteins and is difficult to recapitulate in adult tissues. This report summarizes the known interactions between elastin and the microfibrillar proteins and their role in elastic fiber assembly based on in vitro studies and evidence from knockout mice. We also propose a model of elastic fiber assembly based on the current data that incorporates interactions between elastin, LOXs, fibulins and the microfibril, as well as the pivotal role played by cells in structuring the final functional fiber.     

The three-dimensional micro- and nanostructure of the aortic medial lamellar unit measured using 3D confocal and electron microscopy imaging.

Changes in arterial wall composition and function underlie all forms of vascular disease. The fundamental structural and functional unit of the aortic wall is the medial lamellar unit (MLU). While the basic composition and organization of the MLU is known, three-dimensional (3D) microstructural details are tenuous, due (in part) to lack of three-dimensional data at micro- and nano-scales. We applied novel electron and confocal microscopy techniques to obtain 3D volumetric information of aortic medial microstructure at micro- and nano-scales with all constituents present. For the rat abdominal aorta, we show that medial elastin has three primary forms: with approximately 71% of total elastin as thick, continuous lamellar sheets, 27% as thin, protruding interlamellar elastin fibers (IEFs), and 2% as thick radial struts. Elastin pores are not simply holes in lamellar sheets, but are indented and gusseted openings in lamellae. Smooth muscle cells (SMCs) weave throughout the interlamellar elastin framework, with cytoplasmic extensions abutting IEFs, resulting in approximately 20 degrees radial tilt (relative to the lumen surface) of elliptical SMC nuclei. Collagen fibers are organized as large, parallel bundles tightly enveloping SMC nuclei. Quantification of the orientation of collagen bundles, SMC nuclei, and IEFs reveal that all three primary medial constituents have predominantly circumferential orientation, correlating with reported circumferentially dominant values of physiological stress, collagen fiber recruitment, and tissue stiffness. This high resolution three-dimensional view of the aortic media reveals MLU microstructure details that suggest a highly complex and integrated mural organization that correlates with aortic mechanical properties.     

主动脉夹层血管壁中氧化应激蛋白NOX1 表达及意义

目的:探讨主动脉夹层(aortic dissection,AD)血管组织中氧化应激蛋白NOX1的表达及意义。方法:收集2014年~2015年在武汉大学人民医院AD患者升主动脉血管组织标本(AD组)及多器官捐献患者升主动脉血管组织标本(对照组)各12例。采用维多利亚蓝染色观察主动脉中膜弹性纤维形态结构;应用SP免疫组织化学法及Western blot法对标本组织中的NOX1进行检测分析。结果:AD组主动脉中膜弹性纤维形态和排列不规则、破碎、丢失,结构紊乱。免疫组织化学显示NOX1阳性表达见于主动脉壁平滑肌细胞胞质中,AD组与对照组比较,NOX1表达明显升高。Western blot蛋白印迹示AD组NOX1表达明显增高,差异有统计学意义(P0.05)。结论:NOX1在AD主动脉中膜中表达上调,可能在AD的发生中发挥作用。     

A comparison of oxidized LDL-induced collagen secretion by graft and aortic SMCs: role of PDGF

Smooth muscle cells (SMCs) from prosthetic vascular grafts constitutively secrete higher levels of collagen than aortic SMCs. Lipid oxidation products accumulate in grafts, and we postulated that they stimulate SMC production of collagen. The effect of oxidized low-density lipoprotein (oxLDL) on type I collagen secretion by aortic and graft SMCs was compared. SMCs isolated from the canine thoracic aorta or Dacron thoracoabdominal grafts (n = 10) were incubated with native LDL or oxLDL (0-400 microg cholesterol/ml) for 72 h. Type I collagen in the conditioned medium was measured by ELISA. OxLDL increased collagen production by graft SMCs from 4.1 +/- 0.3 to 11.0 +/- 0.4 ng/microg DNA and by aortic SMCs from 2.3 +/- 0.1 to 3.5 +/- 0.2 ng/microg DNA. Native LDL had little effect. LY-83583, a superoxide generator, stimulated a dramatic increase in collagen secretion by graft SMCs and a smaller but significant elevation by aortic SMCs. OxLDL has been shown to increase PDGF production by graft SMCs, and PDGF can stimulate collagen production. Anti-PDGF antibody inhibited the increase in collagen production by graft SMCs that was stimulated by oxLDL, implicating PDGF as one mechanism of oxLDL-induced collagen production. Lipid oxidation products that accumulate in prosthetic vascular grafts can cause an oxidative stress that stimulates PDGF production by graft SMCs that in turn stimulates collagen production, contributing to the progression of intimal hyperplasia.