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Cellular adhesion molecules on vascular smooth muscle cells

Marina Braun, Peter Pietsch, Karsten Schrör, Gert Baumann, Stephan B. Felix
DOI: http://dx.doi.org/10.1016/S0008-6363(98)00302-2 395-401 First published online: 1 February 1999

Abstract

Several studies during the last years have shown that, in addition to endothelial cells, vascular smooth muscle cells also express the cellular adhesion molecules ICAM-1 and VCAM-1 in atherosclerosis, restenosis and transplant vasculopathy. In vitro studies have characterized stimulatory and inhibitory factors that regulate the expression of ICAM-1 and VCAM-1 on cultured smooth muscle cells. There is evidence for a role of adhesion molecules on smooth muscle cells for leukocyte accumulation and activation of mononuclear cells. Some recent data suggest that the expression of adhesion molecules on smooth muscle cells are cell cycle-dependent and influence smooth muscle cell proliferation and differentiation. Therefore, ICAM-1 and VCAM-1 on smooth muscle cells may contribute to the inflammatory reaction in the vascular wall and may actively be involved in the progression and stability of atherosclerotic plaques.

Keywords
  • Smooth muscle cells
  • Atherosclerosis
  • Cytokines
  • Restenosis
  • Transplant vasculopathy

Time for primary review 26 days.

1 Introduction

It is well established that interactions of endothelial cells and leukocytes via cell adhesion molecules play an important role for leukocyte recruitment in atherogenesis [1, 2]. Besides selectins, the adhesion molecules of the immunoglobulin family intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) contribute to the adhesion of leukocytes to activated endothelium [3]. ICAM-1 binds to LFA-1 (CD18/CD11a) or Mac-1 (CD18/CD11b) and mediates the adhesion of monocytes, lymphocytes and neutrophils to endothelial cells. In addition to its role in cell–cell adhesion, ICAM-1 also serves as a receptor for soluble fibrinogen and hyaluronic acid [4]. VCAM-1 binds to VLA4 (integrin α4/β1) on lymphocytes and monocytes [5]. Elevated levels of soluble adhesion molecules have been measured in patients with atherosclerosis [6]. High levels of soluble ICAM-1 were observed in variant angina [6] and correlated with the risk of future myocardial infarction [7].

Some years ago, the expression of ICAM-1 and VCAM-1 was also detected on intimal smooth muscle cells in the atherosclerotic vascular wall [2]. This was a surprising finding that raised the question of the biological relevance of these molecules.

This review gives an overview of the data available on the regulation and biological functions of ICAM-1 and VCAM-1 on smooth muscle cells and its possible implications in vascular diseases.

2 Expression of cellular adhesion molecules on smooth muscle cells in human vascular diseases

2.1 Atherosclerosis

Evidence for the expression of adhesion molecules on smooth muscle cells in human aortae and coronary arteries came from immunohistochemical studies using antibodies against ICAM-1 or VCAM-1 and cell-specific antigens. Whereas ICAM-1 expression could not be detected on smooth muscle cells in the normal adult aorta, expression of this adhesion molecule was observed on smooth muscle cells in the intima of atherosclerotic lesions [8, 9]. In addition, ICAM-1 expression was occasionally seen in some medial smooth muscle cells adjacent to atherosclerotic plaques [10]. Similarly, VCAM-1 expression was detected on intimal smooth muscle cells in atherosclerotic coronary arteries [11] as well as in aortae and carotid arteries [12]. Several investigations demonstrated a correlation between ICAM-1 [11] and VCAM-1 expression [11–13] on smooth muscle cells and mononuclear cell infiltration suggesting that mediators derived from these mononuclear cells may contribute to the induction of adhesion molecules in smooth muscle cells in vivo. On the other hand, the expression of adhesion molecules on smooth muscle cells may facilitate the accumulation of transmigrated leukocytes within the vascular wall [14]. ICAM-1 and VCAM-1 expression on smooth muscle cells in the intima and media was most prominent in fibrous plaques and advanced atherosclerotic lesions [15]. In atherosclerotic coronary arteries, the expression of VCAM-1 was prevalent in smooth muscle cells and macrophages in the base of the plaques and in intimal neovasculature [11–13]. There was a strong association of the intensity of immunostaining of intimal neovessels for VCAM-1 and the mononuclear cell infiltration, suggesting a role of the neovasculature in leukocyte recruitment to the vascular wall [13].

There are discrepant findings on the onset of adhesion molecule expression in smooth muscle cells in different experimental models of atherosclerosis. In hypercholesterolemia-induced atheromatosis in rabbits, VCAM-1 expression on endothelial cells was a very early observation whereas VCAM-1 expression on intimal smooth muscle cells and occasionally on medial smooth muscle cells adjacent to atheromae occurred later in the development of the disease [16]. In contrast, in ApoE- and LDL-receptor-deficient mice, a striking VCAM-1 expression in the media in areas prone to lesion development was observed, whereas endothelial immunostaining was diffuse [17]. The expression of VCAM-1 on medial smooth muscle cells occurred prior to or coincident with mononuclear cell infiltration. In atherosclerotic vessels, VCAM-1 expression was detected in medial smooth muscle cells at the base of the lesions as well as in the fibrous cap [17].

Recently, a strong focal expression of ICAM-1 in smooth muscle cells and endothelial cells in the outer wall of the internal carotid artery in man was observed preceding mononuclear cell infiltration [18]. This vascular region has a high risk for atherosclerotic lesion development because of hemodynamic characteristics such as low shear stress and oscillations of flow [18]. These findings suggest, that ICAM-1 does not only serve as a leukocyte adhesion molecule but may have additional functions in atherosclerotic-prone regions.

2.2 Transplant vasculopathy

There are only a few investigations of adhesion molecule expression in allograft vasculopathy. Biopsies of rejected kidney allografts demonstrated an upregulation of endothelial cell adhesion molecules and a marked de novo expression of VCAM-1 on intimal as well as medial smooth muscle cells [19, 20]. In a murine model of cardiac allograft vasculopathy endothelial expression of ICAM-1 and VCAM-1 was also observed in control hearts whereas the expression of VCAM-1 on intimal and medial smooth muscle cells was unique for the cardiac allograft [21]. The positive correlation of VCAM-1 expression and mononuclear cell infiltration in kidney allografts suggests direct interactions of smooth muscle cells and leukocytes which might contribute to the progression of the disease [20].

2.3 Restenosis

Data on the expression of adhesion molecules on smooth muscle cells in restenosis came exclusively from animal studies. In the rabbit aorta, induction of ICAM-1 expression on endothelial cells was observed 2 days after balloon injury. In neointimal smooth muscle cells, however, a marked expression of ICAM-1 was detected after 5 to 10 days and remained constant for at least 30 days after the injury [22]. In contrast to the strong endothelial expression of VCAM-1 in this model, neointimal smooth muscle cells demonstrated only a weak immunostaining [23].

Balloon injury of rat carotid arteries resulted in a strong expression of ICAM-1 on medial smooth muscle cells after 1 and 2 days [24]. The expression of ICAM-1 on neointimal smooth muscle cells and regenerating endothelial cells was observed later, 5 to 7 days after the injury. At this time-point, medial ICAM-1 expression was decreased [24]. Treatment of the animals with an antibody against ICAM-1 for 6 days resulted in a significant reduction of neointima formation in this model [24]. Interestingly, no reduction of mononuclear cell infiltration was observed. Consistent with this observation was the lack of effect of an antibody against LFA-1, suggesting LFA-1-independent functions of ICAM-1 in the development of neointima proliferation in this model [24]. The time-course of treatment suggests ICAM-1 expression on medial smooth muscle cells as the primary target of the beneficial effects obtained by the antibody treatment [25].

3 Regulation of adhesion molecule expression on smooth muscle cells

3.1 Role of cytokines

The correlation between the extent of adhesion molecule expression in the atherosclerotic vessel wall and mononuclear cell infiltration suggests a potential role of mononuclear cell-derived mediators in the induction of adhesion molecules [2, 11]. The mononuclear cell infiltration in the atherosclerotic vessel wall mainly consists of macrophages and T lymphocytes [1, 26, 27]. In atherosclerotic plaques, T lymphocytes are often located in the fibrous cap together with smooth muscle cells [28]. A substantial amount of the T cells is activated as indicated by HLA-DR and VLA1 expression [29]. Activated T lymphocytes and macrophages generate and release several cytokines with a number of biological effects on neighbouring cells [1]. Significant amounts of TNFα and IL-1β, mainly secreted by macrophages, as well as interferon (IFN)-γ, an important T cell derived cytokine, have been detected in the atherosclerotic vessel wall [29–32].

Several in vitro studies have investigated the effects of cytokines on the expression of ICAM-1 and VCAM-1 on cultured human smooth muscle cells. The table demonstrates the effects of cytokines on adhesion molecule expression on human smooth muscle cells isolated from different vascular regions [33–42]. The induction of adhesion molecules by cytokines is based on de novo synthesis [38]. IL-6, IL-8, TNFβ, IL-1α had no effects on the expression of both adhesion molecules [38].

View this table:
Table 1

Effects of cytokines on the expression of adhesion molecules on human smooth muscle cells in vitro

+=induction; −=no effect; n.d.=not determined.

ReferenceOriginTNFα ICAM-1/VCAM-1II-1β ICAM-1/VCAM-1II-4 ICAM-1/VCAM-1IFNγ ICAM-1/VCAM-1
Stemme et al., 1992uterine arteriesn.d.n.d.+n.d.n.d.n.d.+n.d.
Couffinhal et al., 1993aorta+n.d.n.d.n.d.n.d.n.d.n.d.n.d.
Li et al., 1993aorta, saph. veinn.d.-n.d.n.d.n.d.+n.d.+
Couffinhal et al., 1994aorta+++-n.d.n.d.+n.d.
Morisaki et al., 1994shunt arteries+-+++-n.d.n.d.
Yang et al., 1994aorta+-+-n.d.n.d.n.d.-
Braun et al., 1995coronary artery++++--+-
Gamble et al., 1995saph. veinn.d.+n.d.+n.d.+n.d.+
Wang et al., 1995pulmonary arteryn.d.n.d.++n.d.n.d.n.d.n.d.
Thorne et al., 1996coronary artery++++n.d.n.d.n.d.n.d.
Braun et al., 1997saphenous vein++n.d.n.d.-++n.d.
  • + = induction; − = no effect; n.d. = not determined.

Table 1 demonstrates some discrepant effects of some of these cytokines on human vascular smooth muscle cells in the different studies. Different cell isolation and cultivation procedures, different number of cell passages, different experimental protocols as well as detection methods may account for these discrepancies. In addition, there is evidence from our laboratory for differences in the response of smooth muscle cells of different vascular origin. IL-4 had no effect on ICAM-1 expression in smooth muscle cells from human coronary arteries as well as saphenous veins [38]. In contrast, Il-4 induced the expression of VCAM-1 on cells from saphenous vein whereas it had no effect on coronary smooth muscle cells [42].

3.2 Role of growth factors

Few studies deal with the effects of growth factors on adhesion molecule expression on smooth muscle cells. In cells isolated from shunt arteries, PDGF-BB and -AB but not -AA induced the expression of ICAM-1. This stimulation, however, was only observed when cells were used in a subconfluent state whereas confluent cells did not respond [36]. TGFβ markedly inhibited basal and TNFα-induced expression of VCAM-1 on human cultured smooth muscle cells isolated from saphenous veins [39]. In contrast, TGFβ had no significant effect on VCAM-1 expression in umbilical vein endothelial cells [43].

3.3 Role of low density lipoproteins

Oxidized and minimally modified low density lipoproteins (LDLs) increased the adhesion of monocytes to human coronary smooth muscle cells, whereas native LDLs had no effect [41]. However, an upregulation of adhesion molecules was not induced on smooth muscle cells by modified LDLs. Therefore, the observed monocyte adhesion involves pathways different from ICAM-1 and VCAM-1 and their respective ligands [41]. In contrast to these findings, both, native and oxidized LDLs, elicited adhesion molecule expression on endothelial cells [44–46].

3.4 Inhibitors of adhesion molecule expression in smooth muscle cells

Recently it has been demonstrated that co-cultivation of smooth muscle cells and endothelial cells inhibits basal and TNFα-induced expression of VCAM-1 on smooth muscle cells [39]. A similar effect was achieved by exogenously added TGFβ. Thus, endothelium-derived TGFβ has been suggested to be the responsible endothelial mediator. However, the effect of endothelial cells could not be prevented by an antibody against TGFβ [39].

Besides TGFβ, the roles of two other endothelium-derived mediators, nitric oxide (NO) and prostacyclin were not investigated in this study. Recently, the NO donors SIN-1 and sodium nitroprusside have been shown to inhibit IL-1-induced expression of ICAM-1 and IFNγ-induced expression of VCAM-1 on smooth muscle cells isolated from human aorta and saphenous vein [47]. This effect of NO was independent of stimulation of the soluble guanylate cyclase [47]. The effect of NO donors occurred at the transcriptional level and involved inhibition of both basal and IL-1-stimulated NFκB activity [47].

We have recently shown that the prostacyclin mimetic cicaprost significantly inhibits TNFα-and Il-1β-induced expression of VCAM-1 and to a lesser extent of ICAM-1 on smooth muscle cells from human coronary arteries and saphenous veins. This inhibition occurred at the transcriptional level and was mediated by an increase in cAMP levels [48]. In contrast, there was no effect of cicaprost and other cAMP elevating substances on IFNγ-induced expression of ICAM-1. Therefore, the observed effects of cAMP involve inhibition of certain signaling pathways that are shared by TNFα and Il-1β but not by IFNγ [48].

Taken together, these experimental data suggest that endothelial cells may control smooth muscle cell expression of adhesion molecules via three different mediators, TGFβ, prostacyclin and NO. This modulatory action of endothelial mediators may be physiologically important. Under pathophysiological conditions, such as atherosclerosis, endothelial cells change their repertoire of functional activities which might contribute to the expression of adhesion molecules on smooth muscle cells. In addition to other well characterized effects, the inhibition of adhesion molecule expression on smooth muscle cells may contribute to the established beneficial effects of the NO precursor l-arginine [49] as well as the prostacyclin mimetic cicaprost in experimental atherosclerosis models [50].

4 Biological functions of adhesion molecules on smooth muscle cells

4.1 Interactions of leukocytes and smooth muscle cells

The adhesion of monocytes and lymphocytes to cytokine-stimulated smooth muscle cells could be antagonized by neutralizing antibodies against ICAM-1 or VCAM-1 in adhesion assays in vitro [33, 34, 39, 41].

The biological consequences of adhesion of leukocytes to smooth muscle cells are not well characterized. In vitro, the production of macrophage inflammatory protein 1 (MIP-1) was activated in monocytes cultured on ICAM-1-coated plates [51]. Recently, an increase in procoagulant activity of monocytes by adhesion to smooth muscle cells has been described. This effect was associated with an increase in tissue factor mRNA and protein synthesis by monocytes and was specifically blocked by an antibody against ICAM-1 [52].

ICAM-1 and VCAM-1 can act as co-stimulatory signals in T cell stimulation during antigen presentation [53]. Several studies have demonstrated that smooth muscle cells in the atherosclerotic vessel wall can express MHC II proteins and, therefore, a role of these cells in antigen presentation to T lymphocytes has been postulated [26, 28, 54]. Detailed experimental studies, however, revealed that MHC II positive smooth muscle cells effectively stimulate pre-activated alloreactive T cells but failed to stimulate resting T cells suggesting a lack of important co-stimulators [55, 56].

Recently, it has been shown that induction of adhesion molecules on endothelial cells by activated leukocytes can be observed independently of cytokines by an interaction between CD40Ligand on leukocytes and CD40 on endothelial cells [57]. Similarly, a recent observation demonstrated that the expression of ICAM-1 and VCAM-1 on smooth muscle cells can be induced by CD40–CD40Ligand interactions [Regitz-Zagrosek, personal communication].

4.2 Differentiation of smooth muscle cells

ICAM-1 [58] and VCAM-1 [59] are expressed on immature fetal smooth muscle cells during embryogenesis. In embryonic mice, a strong expression of VCAM-1 was observed especially in the mesenchyme which is assumed to be the source of developing smooth muscle cells [60]. Knockout mice for VCAM-1 or its ligand VLA4 showed alterations in the development of the heart and circulation such as failure of the formation of epicardium and the coronary vasculature and poorly developed myocardium [61, 62]. ICAM-1 and VCAM-1 expression decreases during ontogeny and both adhesion molecules are absent on differentiated contractile smooth muscle cells in the media of adult arteries [58, 59]. In atherosclerosis, transplant vasculopathy and restenosis smooth muscle cells undergo a phenotypic change from the contractile to a “synthetic” cell type resembling fetal smooth muscle cells [2, 21, 63]. The expression of adhesion molecules in this context could, therefore, be interpreted as a marker of phenotypic change and re-expression of fetal genes [2]. It has recently been shown that in addition to VCAM-1, fetal smooth muscle cells also express the respective ligand VLA4. In accordance, intimal smooth muscle cells in atherosclerotic vessels also expressed both the α4-integrin and VCAM-1 [59]. Human aortic smooth muscle cells cultured in the presence of serum exhibited a synthetic phenotype with a low expression of contractile proteins. Serum deprival induced the expression of contractile proteins in these cells [59]. Prior to the induction of contractile proteins, a strong expression of VCAM-1 and a redistribution of the α4-integrin was observed. Investigations with monoclonal antibodies clearly demonstrated, that interactions between VCAM-1 and VLA4 are essential for the induction of contractile proteins [59].

4.3 Adhesion molecules and smooth muscle proliferation and migration

Recently, it has been shown that the expression of VCAM-1, induced by the cytokine TNFα, is cell cycle-dependent [64]. The induction of VCAM-1 is much weaker in proliferating cells compared to quiescent cells. Cell cycle arrest in the S phase of proliferating cells resulted in a significant inhibition of VCAM-1 expression [64].

In addition to β2 integrins, there are different ligands for ICAM-1, i.e. fibrinogen, which has been suggested to mediate cell proliferation via this adhesion molecule [25]. ICAM-1 also has at least two binding domains for hyaluronic acid [65]. Hyaluronic acid and its binding proteins have been implicated in cell growth, migration and differentiation [65]. The specific role of ICAM-1–hyaluronic acid interactions have not been clearly defined. However, the association of ICAM-1 with the cytoskeleton suggests that this adhesion molecule may mediate effects of hyaluronic acid on cell adhesion and migration [65]. There is evidence for an important role of hyaluronic acid for smooth muscle cell migration in restenosis [66, 67]. Recently, a marked upregulation of ICAM-1 and hyaluronic acid expression in smooth muscle cells after mechanical injury in vitro has been reported. An antibody against ICAM-1 significantly reduced injury-induced smooth muscle cell DNA synthesis [68]. Therefore, the inhibition of ICAM-1–hyaluronic acid interactions may contribute to the beneficial effects of antibodies against ICAM-1 in experimental restenosis models [24, 68].

5 Summary and conclusions

Cellular adhesion molecules are expressed on smooth muscle cells in embryogenesis as well as in vascular diseases. Studies on cultured cells have clearly defined certain cytokines as potent inducers and TGFβ, NO and prostacyclin mimetics as inhibitors of cellular adhesion molecule expression in smooth muscle cells. ICAM-1 expression in medial smooth muscle cells in atherosclerosis-prone regions prior to mononuclear cell infiltration suggests an active role in lesion development. Interactions of ICAM-1 and ligands distinct from LFA-1 or Mac-1 may contribute to smooth muscle cell migration and proliferation not only in neointima formation after mechanical injury but also in atherogenesis. In cultured smooth muscle cells, interactions of VCAM-1 and its ligand VLA4 are essential for phenotypic modulation resulting in a more contractile cell type. As both, VCAM-1 and VLA4 have been demonstrated on smooth muscle cells in atherosclerotic plaques, similar interactions may influence the phenotype and synthetic capacity of smooth muscle cells in vivo. Experimental studies clearly demonstrated the functional relevance of adhesion molecule expression on smooth muscle cells in adhering monocytes and lymphocytes. Therefore, they may contribute to leukocyte accumulation within the vascular wall and thereby influence the cellular composition of atherosclerotic plaques. There are first reports demonstrating a role of ICAM-1 in the activation of mononuclear cells.

Smooth muscle cell migration and proliferation and extracellular matrix production are relevant for the stability of atherosclerotic plaques whereas the accumulation of macrophages and T cells and their production of proteases and certain cytokines are correlated with plaque instability [69]. Therefore, taking into account all data presented here, adhesion molecules on smooth muscle cells may not only be important in plaque progression but also influence plaque stability.

In summary, data on the regulation of adhesion molecules on vascular smooth muscle cells in vitro and in vivo are accumulating. However, further studies are still necessary to complete our knowledge of their role in the pathophysiology of vascular diseases and to encourage considerations on new therapeutic strategies.

Acknowledgements

The authors are grateful to Erika Lohmann for expert secretarial assistance.

References

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