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(Circulation Research. 2001;88:458.)
© 2001 American Heart Association, Inc.

Editorial

Regulation of Vascular Cell Behavior by Collagen

Form Is Function

J. Geoffrey Pickering

From the John P. Robarts Research Institute (Vascular Biology Group), London Health Science Centre and Departments of Medicine (Cardiology), Biochemistry, and Medical Biophysics, University of Western Ontario, London, Ontario, Canada.

Correspondence to J. Geoffrey Pickering, MD, PhD, FRCP(C), London Health Science Centre, 339 Windermere Rd, London, Ontario N6A 5A5, Canada. E-mail gpickering{at}rri.on.ca

Key Words: collagen • smooth muscle cell • gene expression • migration

	Introduction

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The vascular extracellular matrix (ECM) is a complex mixture of collagens, elastin, glycoproteins, and proteoglycans. These constituents not only provide mechanical integrity to the vessel wall but comprise a repertoire of insoluble ligands that can signal the cell to control proliferation, migration, differentiation, and survival. In the normal adult artery wall, the basement membrane is the primary ECM compartment that interacts with the vascular smooth muscle cell (SMC), and its components are believed to be important in maintaining a stable and well-differentiated SMC.¹ However, during conditions of arterial restructuring, the ECM can be quickly remodeled through a combination of synthesis of new ECM molecules, regulated assembly of these molecules, and proteolytic degradation and editing of existing structures. Together, these actions provide a new and dynamic set of ECM stimuli that can have a profound effect on SMC behavior.

One of the most abundant ECM molecules of both healthy and diseased arteries is type I collagen, a fibril-forming, heterotrimeric molecule comprised of two 1(I) collagen chains and one 2(I) collagen chain. These chains are synthesized as soluble propeptides that wind around each other in the endoplasmic reticulum to form type I procollagen, with its long triple helical domain. As type I procollagen molecules are transported to the cell surface, they associate with each other.² These aggregates nonetheless remain in solution, by virtue of the nonhelical domains of procollagen at the amino and carboxy termini. Outside the cell, the propeptide termini are proteolytically removed, on which collagen molecules fall out of solution and polymerize into a highly ordered collagen fibril.³ The size of the fibrils and their organization within the ECM will vary depending on mechanical influences, the stage of tissue development, and interactions with other ECM molecules, such as decorin, type III collagen, and lumican.^{4 5 6} Nevertheless, as discussed below, the fibrils are a key determinant of SMC function.

The interplay between vascular cells and type I collagen is often examined using tail or skin collagen that has been solubilized in acid and then coated onto the culture dish as a planar surface of monomers or oligomers. This is a convenient approach to eliciting cell-collagen interactions and has yielded important insights. For example, it has been determined that vascular SMCs use ₂ß₁ integrin to crawl on type I collagen⁷ and that this motility can be enhanced through localized proteolysis of the collagen substrate.⁸ However, the planar substrate does not provide a close representation of the collagen fibril network that assembles in vivo. Moreover, there is considerable evidence that polymerized collagen fibrils can influence cell behavior differently than a monolayer of collagen. Fibroblasts in a collagen fibril gel, in contrast to those on planar collagen, were found to stop replicating, reduce their expression of type I and III collagens, and induce the expression of collagenase-1 (matrix metalloproteinase-1 [MMP-1]).^{9 10} It has also been shown in various cell types that polymerized collagen fibrils can induce the expression of collagenase-3 (MMP-13)¹¹ and the ₂-integrin subunit,¹² suppress the expression of heat shock protein (Hsp) 47,¹³ activate mitogen-activated protein kinases and protein kinase C-,^{11 12} and attenuate responses to growth factors.¹⁴ Furthermore, type I collagen in its polymerized, but not monomeric, form is a potent inhibitor of SMC proliferation, a process mediated by upregulation of cyclin-dependent kinase-2 (cdk-2) inhibitors.¹⁵ Thus, in regulating SMC behavior, collagen form dictates collagen function.

In this issue of Circulation Research, Ichii et al¹⁶ broaden our understanding of the interactions between polymerized collagen fibrils and vascular SMCs. These investigators cultured human SMCs for 24 hours on bovine skin collagen in either a monomeric or polymerized form and assessed the differential consequences for gene expression using suppressive subtraction hybridization. Twenty-two differentially expressed transcripts were found, and the authors provide good confirmation of the differences by Northern blot analyses. They also showed that 9 of the transcripts that were downregulated by fibrillar collagen, relative to monomeric collagen, were upregulated in the rat carotid artery after balloon injury. This does not prove that the shifts in gene expression in the artery wall were attributable to altered cellular interaction with fibrillar collagen. It does, however, provide a meaningful in vivo context for the specific genes identified by the in vitro screen.

As with any expression screen, definitive statements regarding the function, or even importance, of the individual genes identified cannot be made. However, the patterns of gene expression themselves can be informative and hypothesis-generating, and the findings of Ichii et al¹⁶ illustrate this. For example, whereas polymerized collagen suppressed the expression of 18 genes relative to monomeric collagen, it stimulated the expression of only 4. This net suppression may reflect the more physiological environment afforded by the collagen gel and implicates collagen fibrils as structures that can terminate some of the cellular processes that were activated during vascular remodeling. At the same time, it may be particularly informative to pursue the significance of the minority, upregulated genes, including tissue-type plasminogen activator, angiopoietin-related protein-2, and microfibril-associated glycoprotein 4.

It is also noteworthy that more than half of the differentially expressed transcripts could be categorized as encoding either ECM proteins or cytoskeleton-associated proteins. The authors established that fibronectin, thrombospondin-1, and tenascin-C were downregulated by polymerized collagen, highlighting a phenomenon whereby the ECM environment can regulate its own constituents. Furthermore, the associated change in cytoskeletal gene expression is consistent with a linkage between the ECM, SMC shape, and gene expression. Indeed, the differences in gene expression between SMCs on monomeric or polymerized collagen may be due to differences in cell geometry and in the different mechanical forces exerted on the cell surface by the 2 substrates.

Ichii et al¹⁶ also show that SMCs that have been cultured on fibrillar collagen subsequently manifest a repressed ability to directionally migrate to thrombospondin-1 on a substrate of vitronectin or osteopontin. The authors suggest that because SMC migration on vitronectin or osteopontin is mediated, at least in part, by _vß₃ integrin and because polymerized collagen had no effect on _vß₃ integrin expression, the repressed motility may be due to inhibition of _vß₃ integrin function. The authors observed that SMCs on polymerized collagen had smaller focal adhesions with less -actinin, and they postulate that this may repress _vß₃ integrin–mediated migration. This remains speculative, because, depending on the initial force with which the cell is attached to the substrate, a reduction in focal adhesion assembly can actually favor motility over stationary adhesion.¹⁷ Nevertheless, the findings are consistent with an interesting paradigm whereby one ECM constituent suppresses the cellular responses to another. This filtering-out of specific signals may be an important mode of cell regulation in vascular disease. Mechanistically, a form of crosstalk between integrins or other ECM-initiated signaling components seems possible and warrants additional investigation.

Clearly, there is a wealth of information in the insoluble ECM of the artery wall. However, as highlighted by the data of Ichii et al,¹⁶ the mere presence of a given ECM molecule at the surface of the cell may not define the consequences for that cell. Higher levels of control must be considered, including the extent of ECM assembly and a dynamic reciprocity between the cell and the substrate that regulates both the constituents of the ECM and the responsiveness of the cell to those constituents. Elucidating the complex networks that underlie this control remains an exciting challenge.

	Footnotes

 
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.

	References

Top Introduction References

 

1. Thyberg J, Hultgardh-Nilsson A. Fibronectin and the basement membrane components laminin and collagen type IV influence the phenotype properties of subcultured rat aortic smooth muscle cells differently. Cell Tissue Res. 1994;276:263–271.[Medline] [Order article via Infotrieve]
2. Bonfanti L, Mironov AA, Martinez-Menarguez JA, Martella O, Fusella A, Baldassarre M, Buccione R, Geuze HJ, Luini A. Procollagen traverses the Golgi stack without leaving the lumen of cisternae: evidence for cisternal maturation. Cell. 1998;95:993–1003.[Medline] [Order article via Infotrieve]
3. Kadler KE, Holmes DF, Trotter JA, Chapman JA. Collagen fibril formation. Biochem J. 1996;316:1–11.
4. Danielson KG, Baribault H, Holmes DF, Graham H, Kadler KE, Iozzo RV. Targeted disruption of decorin leads to abnormal collagen fibril morphology and skin fragility. J Cell Biol. 1997;136:729–743.[Abstract/Free Full Text]
5. Romanic AM, Adachi E, Kadler KE, Hojima Y, Prockop DJ. Copolymerization of pNcollagen III and collagen I: pNcollagen III decreases the rate of incorporation of collagen I into fibrils, the amount of collagen I incorporated, and the diameter of the fibrils formed. J Biol Chem. 1991;266:12703–12709.[Abstract/Free Full Text]
6. Chakravarti S, Magnuson T, Lass JH, Jepsen KJ, LaMantia C, Carroll H. Lumican regulates collagen fibril assembly: skin fragility and corneal opacity in the absence of lumican. J Cell Biol. 1998;141:1277–1286.[Abstract/Free Full Text]
7. Skinner MP, Raines EW, Ross R. Dynamic expression of ₁ß₁ and ₂ß₂ integrin receptors by human vascular smooth muscle cells: ₂ß₁ integrin is required for chemotaxis across type I collagen coated membranes. Am J Pathol. 1994;145:1070–1081.[Abstract]
8. Li S, Chow LH, Pickering JG. Cell surface-bound collagenase-1 and focal substrate degradation stimulate the rear release of motile vascular smooth muscle cells. J Biol Chem. 2000;275:35384–35392.[Abstract/Free Full Text]
9. Mauch C, Hatamochi A, Scharffetter K, Krieg T. Regulation of collagen synthesis in fibroblasts within a three-dimensional collagen gel. Exp Cell Res. 1988;178:493–503.[Medline] [Order article via Infotrieve]
10. Mauch C, Adelmann-Grill B, Hatamochi A, Krieg T. Collagenase gene expression in fibroblasts is regulated by a three-dimensional contact with collagen. FEBS Lett. 1989;250:301–305.[Medline] [Order article via Infotrieve]
11. Ravanti L, Heino J, Lopez-Otin C, Kahari VM. Induction of collagenase-3 (MMP-13) expression in human skin fibroblasts by three-dimensional collagen is mediated by p38 mitogen-activated protein kinase. J Biol Chem. 1999;274:2446–2455.[Abstract/Free Full Text]
12. Xu J, Clark RA. A three-dimensional collagen lattice induces protein kinase C- activity: role in ₂ integrin and collagenase mRNA expression. J Cell Biol. 1997;136:473–483.[Abstract/Free Full Text]
13. Rocnik E, Saward L, Pickering JG. HSP47 expression by smooth muscle cells is increased during arterial development and lesion formation and is inhibited by fibrillar collagen. Arterioscler Thromb Vasc Biol. 2001;21:40–46.[Abstract/Free Full Text]
14. Clark RA, Nielsen LD, Welch MP, McPherson JM. Collagen matrices attenuate the collagen-synthetic response of cultured fibroblasts to TGF-ß. J Cell Sci. 1995;108:1251–1261.[Abstract]
15. Koyama H, Raines EW, Bornfeldt KE, Roberts JM, Ross R. Fibrillar collagen inhibits arterial smooth muscle proliferation through regulation of Cdk2 inhibitors. Cell. 1996;87:1069–1078.[Medline] [Order article via Infotrieve]
16. Ichii T, Koyama H, Tanaka S, Kim S, Shioi A, Okuno Y, Raines EW, Iwao H, Otani S, Nishizawa Y. Fibrillar collagen specifically regulates human vascular smooth muscle cell genes involved in cellular responses and the pericellular matrix environment. Circ Res. 2001;88:460–467.[Abstract/Free Full Text]
17. Palecek SP, Loftus JC, Ginsberg MH, Lauffenburger DA, Horwitz AF. Integrin-ligand binding properties govern cell migration speed through cell-substratum adhesiveness. Nature. 1997;385:537–540.[Medline] [Order article via Infotrieve]

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