|Characterization of type I and type III collagens in human tissues:|
|Prev||Chapter 2. Review of the literature||Next|
Atherosclerosis is a response to injury to the endothelium, characterized by a fibroproliferative inflammatory reaction, degenerative changes and accumulation of extracellular matrix and lipids. One of the parameters associated with the endothelial cell dysfunction that results from exposure to oxidized LDL is increased adherence of macrophages and other inflammatory cells. Macrophages are known to localize subendothelially, where they scavenge lipids and turn into large foam cells. Proliferation and migration of vascular smooth muscle cells (SMCs) together with their transition from the “contractile” type into the “synthetic” phenotype are key features in the development of atherosclerosis. SMCs, T-cells and foam cells together form a fatty streak, which in turn may develop into an intermediate, fibrofatty lesion and, finally, into a fibrous plaque with calcification. The whole process is extremely complex, involving growth factors, cytokines, hormones, hemostatic agents and changes in collagen synthesis and smooth muscle cells (see Ross 1993, Rekhter 1999 for reviews).
As in most connective tissues, the quantitatively most abundant group of arterial collagens are the fibril-forming type I and III collagens (see Barnes 1985 for a review). These collagen types together are responsible for tensile strength, but also contribute, together with elastin, somewhat to the extensile properties of the aorta (Dobrin et al. 1984, Dobrin & Mrkwicka 1994). The quantitative ratios of type I and III collagens in normal and atherosclerotic aortas have usually been determined after extraction of the collagens or collagen fragments from tissue specimens and quantification, after separation, into different types (see Barnes 1985, Rauterberg et al. 1993 for reviews).
The increased proportion of type I collagen is thought to be characteristic of the atherosclerotic intimal plaque. In spite of extensive research, our knowledge of the quantitative ratios of type I and III collagens in normal and atherosclerotic aortic wall is incomplete and controversial (see Barnes 1985, Rauterberg et al. 1993). The currently existing data on type I and type III collagen ratios in atherosclerosis are shown summarized in Table 2.
Data from immunohistological studies indicate only small differences in the normal aorta in the distribution of type I and III collagens (see Rauterberg et al. 1993). Both these collagen types are present in the normal fibrillar structures around smooth muscle cells and elastic membranes (McCullagh et al. 1980, Shekhonin et al. 1985, Katsuda et al. 1992). Furthermore, they co-localize with fibronectin (Shekhonin et al. 1987) and stain more intensively in the adventitia than in the media (Katsuda et al. 1992). The type I and III collagens appear together in the thickened intimas of atherosclerotic lesions, where their staining intensity is also greater than in healthy aorta (Katsuda et al. 1992, see Rauterberg et al. 1993).
Table 2. Summary of data on type I and III collagen contents in normal and atherosclerotic human aorta.
|Pepsin digestion,||65 % type I in atherosclerosis||McCullagh & Balian 1975|
|SDS-PAGE||30 % type I in normal aorta|
|Pepsin digestion, SDS-PAGE||66 % type I in atherosclerosis||Ooshima 1981|
|Pepsin /||76 % type I in atherosclerosis||Morton & Barnes 1982|
|CNBr cleavage, SDS-PAGE||No major shift when compared to healthy aorta|
|CNBr cleavage,||58 % type I in normal aorta||Hanson & Bentley 1983|
|CMC chromatography, SDS-PAGE||90 % type I in atherosclerosis|
|Pepsin, DEAE-cellulose chromatography, CNBr, SDS-PAGE||38 % type I in normal aorta||Hill & Harper 1984|
|CNBr, SDS-PAGE||70 % type I in normal aorta||Halme et al. 1986|
|Pepsin,||70 % type I in atherosclerosis||Leuschner &Haust 1986|
|CMC chromatography HPLC, CNBr, SDS-PAGE||No major shift when compared to normal aorta|
|Pepsin, CNBr,||70 % type I collagen, type III ↓||Murata et al.1986|
|SDS-PAGE||with advancing atherosclerosis|
|Pepsin, CNBr,||62 % type I in normal thoracic aorta||Miller et al. 1991|
|HPLC, UV absorbance||47 % type I in normal abdominal aorta|
|CNBr, HPLC,||59 % type I in normal thoracic aorta||Deyl et al. 1997|
|SDS-PAGE, capillary electrophoresis||52 % type I in normal abdominal aorta|
|CMC = carboxymethyl-cellulose; CNBr = cyanogen bromide; DEAE = diethylaminoethyl; HPLC = high-performance liquid chromatography; SDS-PAGE = sodium dodecyl sulfate polyacrylamide gel electrophoresis|
The increased amount of type I collagen-producing cells has been established in all types of human atherosclerotic lesions (Andreeva et al. 1997). In animal models, the collagen content in atherosclerotic lesions has been shown to increase consistently over time (Kratky et al. 1999). The plaque collagen is mainly produced by smooth muscle cells (SMCs), but endothelial cells are also able to synthesize collagen (Canfield et al. 1992). Furthermore, cells different from typical SMCs produce type I collagen in human atherosclerotic lesions (Rekhter et al. 1996, Andreeva et al. 1997, Tintut et al. 1998). SMC phenotype, proliferation, migration and collagen production play an important role in the pathophysiology of atherosclerosis (see Rekhter 1999). There is evidence to indicate that ageing is associated with dysregulation of the proliferative responses of SMCs from old aortas, possibly predisposing to atherosclerotic changes (McCaffrey et al. 1988). In addition, DNA alterations have been found in SMCs of human atherosclerotic lesions, suggesting that mutational events may be part of the atherosclerotic process (De Flora et al. 1997).
At least type I collagen synthesis has been clearly shown to be upregulated in atherosclerotic plaques compared with the media and normal arteries (Rekhter et al. 1993). Recently published data suggest that genes consistent with increased proliferation and collagen production are upregulated in SMCs during atherogenesis (Laury-Kleintop et al. 1999). A number of cytokines and growth factors produced and secreted by macrophages and other inflammatory cells have been described as being involved in the process of atherosclerotic plaque formation, in part by altering the rate of collagen synthesis (see Rekhter 1999 for a review). Transforming growth factor beta (TGF-β ) is the most potent and consistent stimulator of collagen synthesis in vascular SMCs (Amento et al. 1991). There is evidence of genomic instability of the type II TGF-β 1 receptor gene in human atherosclerotic and restenotic vascular cells, probably causing resistance to the antiproliferative effects mediated by this particular receptor type (McGaffrey et al. 1997). Platelet-derived growth factor (PDGF) (Amento et al. 1991), interleukin-1 (IL-1) (Amento et al. 1991), angiotensin-II (Kato et al. 1991), homocysteine (Majors et al. 1997), endothelin-1 (Rizvi et al. 1996) and oxidized LDL (Bachem et al. 1999) have also been reported to stimulate collagen synthesis in SMC culture. Human connective tissue growth factor (CTGF) is undetectable in normal blood vessels, but overexpressed in atherosclerotic lesions, suggesting that CTGF plays a role in atherogenesis (see Oemar & Lüscher 1997). Furthermore, mechanical stress has been shown to have a stimulatory effect on collagen synthesis (Kolpakov et al. 1995a). In contrast, fibroblast growth factor (Majors & Ehrhart 1993, Pickering et al. 1997), nitric oxide (Kolpakov et al. 1995b, Myers & Tanner 1998), interferon gamma (IFN-γ ) (Amento et al. 1991) and prostaglandins (Fitzsimmons et al. 1999) all inhibit collagen production (see Rekhter 1999). In addition, SMC may exhibit different biosynthetic activity depending on the matrix environment, and the extracellular matrix itself is thus able to control collagen synthesis in SMC culture (Thie et al. 1991).
Plaque rupture is responsible for the acute manifestations of atherosclerosis. Rupture prone plaques are characterized by a large lipid pool, a thin fibrous cap, an increased amount of inflammatory cells and a reduced collagen and SMC content (see Lee & Libby 1997, van der Wal & Becker 1999 for thorough reviews). Previous studies (Henney et al. 1991, Galis et al. 1994, Brown et al. 1995, Nikkari et al. 1995, Shah et al. 1995, Li et al. 1996) have implicated matrix metalloproteinases, mostly MMP-1, gelatinases and stromelysins, together with certain tissue inhibitors of metalloproteinases, in the pathogenesis, destabilization and rupture of atherosclerotic lesions. Recently, Sukhova et al. (1999) demonstrated increased levels of MMP-1 and MMP-13 in atheromatous versus fibrous carotid atherosclerotic plaques. A large portion of matrix-degrading enzymes is produced by inflammatory cells, especially macrophages. The numbers of mast cells, T lymphocytes and MMP-9-containing macrophages increase as the atherosclerotic lesions become more severe (Kaartinen et al. 1998). In addition to the MMPs, macrophages in human atheroma have been reported to contain abundant amounts of cathepsins K and S (Sukhova et al. 1998). Certain infections, such as Chlamydia pneumoniae (see Movahed 1999 for a review), have been suggested to play a role in the pathogenesis of atherosclerotic lesions, possibly by inducing the inflammatory process and thus the production of MMPs.
It has been suggested that oxidized low-density lipoprotein (LDL) is a key component in the endothelial injury in atherosclerotic lesions. The type I collagen content in the aortas of cholesterol-fed birds has been shown to be increased compared to controls (Jarrold et al. 1999). LDL may directly injure the endothelium or play a role in the adherence and migration of inflammatory cells (see Ross 1993). In addition, several collagen types are known to bind oxidized lipoproteins, and the accumulation of collagen in the intima may thus promote lipoprotein accumulation (Greilberger et al. 1997). Furthermore, oxidized LDL-containing immunocomplexes have been shown to play an important role in macrophage activation, which, in turn, may result in the secretion of cytokines capable of inducing collagen synthesis (Virella et al. 1995).
The aminoterminal propeptide of type III procollagen, PIIINP, has been found to be elevated in the blood of patients with coronary artery disease (Bonnet et al. 1988). In addition, thrombolytic treatment of patients with acute myocardial infarction results in collagen breakdown (Peuhkurinen et al. 1991, Peuhkurinen et al. 1996). Furthermore, thrombin, a very important peptide in blood coagulation and platelet activation, stimulates procollagen production in cultured SMCs and thus in the arterial wall (Dabbagh et al. 1998).
One of the most common features of advanced atherosclerosis is vascular calcification. Type I collagen has been shown to promote calcification of vascular cells in in vitro conditions (Watson et al. 1998). Type I collagen may also be important in plaque neoangiogenesis (Jackson & Jenkins 1991) and the organization of thrombus (Rekhter et al. 1996). Furthermore, human coronary restenosis involves rapid accumulation of collagen fibers (Pickering et al. 1996). Collagen accumulation has also been found to correlate with the severity of restenosis after angioplasty in an animal model (Lafont et al. 1999). Not only synthesis but also degradation of collagen has an important role in collagen turnover after balloon angioplasty (see Rekhter 1999).
Atherosclerotic plaques are present in aneurysmal walls, and atherosclerosis is thought to be, at minimum, a permissive factor in aneurysm development (see Grange et al. 1997 for a review). There is evidence to suggest that the ratio of type III to type I collagen is not altered in the aneurysmal aorta (Rizzo et al. 1989). However, occasional lower amounts of type III collagen at the tissue level have been described in patients with a family history of aneurysms (Menashi et al. 1987, Powell & Greenhalg 1989). Kuga et al. (1998) suggested that the amount of type III collagen is significantly decreased in atherosclerotic AAAs compared with atherosclerotic aortas without aneurysmal changes. In addition, he found abnormal fragments of type III collagen in AAA tissue. Deak et al. (1992) and van Keulen et al. (1999) reported decreased type III procollagen secretion in fibroblasts in a small group of patients with familial AAA. The expression of type I and III collagens is increased in human abdominal aortic aneurysms (McGee et al. 1991, Mesh et al. 1992, Minion et al. 1993). The increased expression of collagen is located in adventitial fibroblasts, medial smooth muscle cells near the inflammatory infiltrates, and myofibroblasts inside the plaque (Hunter et al. 1996, see Grange et al. 1997). The existing data on the amount of collagen are controversial: there is evidence of increased (Menashi et al. 1987, Rizzo et al. 1989, Baxter et al. 1994, He & Roach 1994), unchanged (McGee et al. 1991) or even decreased (Sumner et al. 1970) proportion of total collagen in AAAs.
Aneurysm formation is a complex remodeling process that involves both synthesis and degradation of matrix proteins (see Ghorpade & Baxter 1996 for a review). The identification of at least the MMPs 1, 2, 3, 9 and, recently, 13 in AAA tissue has been reported by several investigators (Irizarry et al. 1993, Newman et al. 1994, McMillan et al. 1995, Thompson et al. 1995, Sakalihasan et al. 1996, Tamarina et al. 1997, Mao et al. 1999). Freestone et al. (1995) reported more MMP-2 activity in small aneurysms, whereas MMP-9 was significantly increased in large aneurysms. On the other hand, MMP-9 expression has also been reported to be significantly higher in medium-sized AAAs than in either small or large aneurysms (McMillan et al. 1997). In addition, the activity of several cathepsins has been found to be increased in aortic aneurysms (Gacko & Chyczewski 1997). Clinical methods of MMP inhibition are under development. For example, preoperative treatment with doxycycline has been associated with reduction in the aortic wall expression of MMP-2 and MMP-9 (Thompson & Baxter 1999), and marimastat (an MMP inhibitor) has been shown to significantly inhibit matrix degradation and active MMP-2 production in a porcine model of aneurysm disease (Treharne et al. 1999). MMP-9, which is increased in the plasma of AAA patients, could be a potential diagnostic marker for clinical purposes (McMillan & Pearce 1999). Other diagnostic approaches have been studied as well: the aminoterminal propeptide of type III collagen, PIIINP, has previously been found to be clinically useful in the follow-up of patients with abdominal aortic aneurysms (Satta et al. 1997).
Ehlers-Danlos syndrome IV (EDS IV), which is characterized by multiple aneurysms and other connective tissue manifestations, is caused by mutations in the gene encoding the α1-chain of type III collagen (see Kuivaniemi et al. 1997). In addition, there are many studies reporting a strong familial tendency to aneurysms not associated with EDS IV (see Verloes et al. 1996 for a review). In these studies, the proportion of AAA probands with a positive family history has ranged from 12 to 20 %, and the relative risk for siblings has been 10 - 28. Van Keulen et al. (1999) recently reported that 29 % of 56 consecutive AAA patients had a positive family history of aneurysms. The characteristic features of familial aneurysms are an early age of diagnosis and a higher risk of rupture. Furthermore, even though both sporadic and familial AAAs are more common in men, the patients with familial AAA are more likely to be women (see Verloes et al. 1996). Type III collagen gene mutations have been suggested to contribute to the development of AAAs (Kontusaari et al. 1990, Anderson et al. 1996), and other genes, such as those for collagenases and elastin, have also been mapped for abdominal aneurysms (see Verloes et al. 1996). Some of the patients with familial aneurysms show decreased type III collagen production in cultured skin fibroblasts (Deak et al. 1992, van Keulen et al. 1999), or even lack type III collagen in skin and aortic wall (Hamano et al. 1998).