Possible involvement of increased glycoxidation and lipid peroxidation of elastin in atherogenesis in haemodialysis patients
Yuji Yamamoto1,2,
Noriyuki Sakata1,,
Jing Meng1,
Masaya Sakamoto2,
Akiko Noma2,
Iori Maeda2,
Kouji Okamoto2 and
Shigeo Takebayashi1
1 Second Department of Pathology, School of Medicine, Fukuoka University, Fukuoka, Japan and
2 Department of Biochemical Engineering and Science, Kyushu Institute of Technology, Iizuka, Japan
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Abstract
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Background. Glycoxidation and lipid peroxidation products accumulate in collagen of various tissues in haemodialysis patients with end-stage renal disease (ESRD). The purpose of this study was to test the hypothesis that increased glycoxidation and lipid peroxidation of aortic elastin is implicated in the cardiovascular complications, particularly atherosclerosis, of chronic haemodialysis patients.
Methods. Post-mortem aortic samples were obtained from 16 deceased subjects, including chronic haemodialysis patients (group 1 n=6, age 64.7±11.4 years) and control subjects (group 2 n=10, age 61.1±10.4 years). The samples were divided into three vessel wall sites: atherosclerotic intima, lesion-free intima, and media. They were sequentially treated with 0.01 M phosphate-buffered saline, collagenase, and elastase to obtain three fractions, namely soluble (SF), collagen (CF), and elastin (EF) fractions, respectively. Using spectrophotofluorometry, the pentosidine- and malondialdehyde (MDA)-linked fluorescence of these fractions was measured at wavelengths 335/385 and 390/460 (excitation/emission), respectively.
Results. Samples from haemodialysis patients (group 1) exhibited a significant increase in both pentosidine- and MDA-linked fluorescence of EF in atherosclerotic intima, lesion-free intima, and media samples, compared with samples from control subjects (group 2). In group 1, the levels of pentosidine- and MDA-linked fluorescence of EF were highest in atherosclerotic intima among the three aortic sites. Interestingly, in both groups, the levels of pentosidine- and MDA-linked fluorescence of EF were significantly higher than those of CF in all aortic sites. There was a strong correlation between the levels of pentosidine- and MDA-linked fluorescence in CF and EF for all aortic sites. In group 1, the pentosidine- and MDA-linked fluorescence levels of EF correlated significantly with the duration of haemodialysis in lesion-free intima and media.
Conclusions. Our study provides the first biochemical evidence for a close link between aortic elastin glycoxidation and lipid peroxidation. In addition, we demonstrated high levels of these products in the aortic elastin of haemodialysis patients with ESRD. Our findings support the hypothesis that modification of aortic elastin by glycoxidation and lipid peroxidation may contribute to the development of vascular complications, particularly atherosclerosis, in patients with end-stage renal failure.
Keywords: cardiovascular complications; elastin; end-stage renal disease; glycoxidation; haemodialysis; lipid peroxidation
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Introduction
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Cardiovascular complications are the principal cause of morbidity and mortality in patients with end-stage renal disease (ESRD) requiring haemodialysis. Risk factors for cardiovascular complications include left ventricular hypertrophy, hypertension, dyslipidaemia, and glucose intolerance. These patients have accelerated courses of two different arterial diseases, arteriosclerosis and atherosclerosis, which are the major lesions underlying the cardiovascular complications [1]. Several studies have demonstrated an increased concentration of pentosidine, a major glycoxidation product, in the tissues and plasma of uraemic patients [2]. We have recently demonstrated increased accumulation of N
-(carboxymethyl)lysine, another glycoxidation product, in the extracellular matrix of atherosclerotic lesions in subjects with ESRD [3]. Increased levels of oxidative stress have also been established in patients with ESRD, as evidenced by the accelerated formation of malondialdehyde (MDA) in low-density lipoprotein and red blood cell membranes [4,5]. Furthermore, glycoxidation and lipid peroxidation of low-density lipoprotein have been shown to synergistically promote development of atherosclerotic lesions [6]. Therefore, glycoxidation and lipid peroxidation may be intrinsically linked and may play an important role in the pathogenesis of cardiovascular complications of haemodialysis patients with ESRD. However, the mechanism by which glycoxidation and lipid peroxidation contributes to the development of cardiovascular complications, particularly atherosclerosis of haemodialysis patients with ESRD, remains unclear.
Collagen is the major extracellular matrix component in various tissues, including the dura mater, skin, and kidney, and is modified by pentosidine in association with aging, diabetes, and renal dysfunction [2]. We demonstrated previously the presence of high levels of advanced glycation end-products (AGEs) in arterial collagen in association with aging and diabetes [7]. Elastin is another extracellular matrix protein in various tissues, such as arteries and lung, which seems a less likely target for glycation, as it contains relatively few
-amino groups. However, elastin has the lowest turnover rate of all matrix components with a half-life of approximately 40 years [8]. Recent studies have shown that AGEs, such as N
-(carboxymethyl)lysine and pentosidine, accumulate in the elastin fibres of human skin and yellow ligament [9,10]. Furthermore, in vitro incubation of elastin with ribose induces the formation of pentosidine [11]. However, no information is available on the modification of aortic elastin by glycoxidation and lipid peroxidation in haemodialysis patients with ESRD.
The present study was undertaken to investigate whether glycoxidation and lipid peroxidation are increased in aortic elastin of haemodialysis patients with ESRD, and how these modifications of elastin contribute to the development of atherosclerosis, the lesion which underlies most cardiovascular complications.
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Subjects and methods
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Reagents
HRP-conjugated rabbit IgG antibody was purchased from Dako (Denmark). Albumin-conjugated desmosine and anti-desmosine-KLH antibody were obtained from Elastin Products Co. (Pacific, MO). o-Phenylenediamine dihydrochloride, acetonitrile, triethylamine, and bacterial collagenase (type VII) were purchased from Sigma Chemical Co. (St Louis, MO). The BCA (bicinchoninic acid) protein assay kit was purchased from Pierce (Rockford, IL). Elastase (from porcine pancreas, 326 U/mg) was obtained from Eisai Co. (Tokyo, Japan). All other chemicals were of the best grade available from commercial sources.
Subjects and samples
Samples were collected from the aortas of 16 autopsy cases, which included six patients who died of ESRD, classified as group 1, and 10 age-matched control subjects without ESRD (group 2). In each case, an autopsy was performed within 26 h of death. The aorta was opened longitudinally and extensively rinsed with 0.01 M phosphate-buffered saline (PBS) to wash out any blood. The ascending and thoracic aortas were then examined to the level of the renal arteries by the same pathologist (N.S.). Arterial samples were divided into three sites under a dissecting microscope: media, visibly raised atheromatous intima, and adjacent intima free of atheromatous plaque, as described previously [11]. Atheroma was then stripped away from the visibly raised atheromatous intima under a dissecting microscope. The tissue samples were cut into small pieces and rinsed twice with a buffer containing 4 mmol/l EDTA, 0.1 mol/l phenylmethylsulfonyl fluoride, 1 mmol/l N-ethylmaleimide, and 0.1 µg/ml pepstatin A. After centrifugation at 5000 r.p.m. at 4°C for 15 min, the samples were rapidly frozen in liquid nitrogen and stored at -40°C until use.
Preparation of soluble, collagen, and elastin fractions
Samples were homogenized in the same buffer, and then centrifuged at 5000 r.p.m. for 15 min. The pellets were delipidated with a chloroform:methanol mixture (2:1, vol:vol) by gentle shaking at 4°C for 24 h. After centrifugation at 5000 r.p.m. for 15 min, pellets were washed sequentially once with methanol and three times with deionized water. After drying with a freeze dryer (EYELA FD-80, Tokyo Rikakikai) for 24 h, the pellets were stored at -40°C until use. Dry pellets (5 mg) were immersed in 0.01 M PBS and then incubated at 4°C for 24 h. After centrifugation at 15000 r.p.m. at 4°C for 20 min, the supernatants were collected as the soluble fraction (SF). The precipitate was washed twice with HEPES buffer (pH 7.4) and then incubated with 1 ml of bacterial collagenase (type VII, 150 U/ml) in HEPES buffer (pH 7.4) containing 0.1 mol/l CaCl2 at 37°C for 24 h. After centrifugation at 15000 r.p.m. at 4°C for 20 min, the supernatants were collected as the collagen fraction (CF). The precipitate was washed three times with deionized water and then incubated with 1 ml of elastase (100 µg/ml) in TrisHCl buffer (0.1 M, pH 7.4) at 37°C for 24 h. After centrifugation at 15000 r.p.m. at 4°C for 20 min, the supernatants were collected as the elastin fraction (EF). Toluene:chloroform (1:1) mixture (4 µl) was added to the reaction mixture to prevent bacterial growth during collagenase or elastase digestion. Histopathological examination of delipidated tissues before and after collagenase and elastase digestions was performed on the frozen sections. After fixation with 10% buffered formalin, sections were subjected to Masson trichrome stain for collagen fibres and Wiegert's elastica stain for elastin fibres.
Assay of soluble protein, collagen, and elastin
The protein concentration in the SF was measured with a BCA protein assay kit using human albumin as a standard, according to the instructions provided by the manufacturer. The collagen content was determined according to Stagemann and Stalder [12]. Briefly, 50 µl of sample was acid-hydrolysed in 1 ml of 6 N HCl at 100°C for 24 h. After evaporation of the HCl, 4-hydroxyproline was measured in the dry residue with a spectrophotometer (Shimazu, Tokyo) at a wavelength of 550 nm. The collagen content was calculated on the assumption that hydroxyproline constitutes 14% of collagen. The elastin content was determined by measuring desmosine in acid-hydrolysed samples by competitive enzyme-linked immunosorbent assay (ELISA). Briefly, the wells of microtitre plates were coated with 1 µg/ml of albumin-conjugated desmosine in PBS. After incubation at 4°C for 24 h, the wells were washed three times with washing buffer (PBS containing 0.05% Tween 20). After incubation with 0.5% gelatin for 60 min to block non-specific binding, the wells were washed again with washing buffer. The sample (60 µl) was mixed with an equal volume of diluted rabbit anti-desmosine-KLH antibody. A portion of the mixture (100 µl) was added to each well, followed by incubation for 60 min. The wells were then washed three times with washing buffer and incubated with HRP-conjugated anti-rabbit IgG antibody for 60 min. After washing three times with washing buffer, the reactivity of peroxidase was determined by incubation with o-phenylenediamine dihydrochloride for appropriate intervals, and the absorbance at 492 nm was read on a micro-ELISA plate reader. A standard curve was constructed with known concentrations of desmosine. The elastin content was calculated on the assumption that desmosine constitutes 0.9% of elastin.
Spectrofluorometry of pentosidine and MDA
The pentosidine- and MDA-linked fluorescence of each fraction was measured at wavelengths of 335/385 and 390/460 nm (excitation/emission), respectively, using a spectrophotofluorometer (Shimadzu, Japan) [13]. Data were expressed in arbitrary units (AU) per mg protein (AU=fluorescence intensity of sample/fluorescence intensity of blank solution).
Data analysis
The numerical data were expressed as mean±SD. The MannWhitney U test was used to evaluate differences between two groups. Pearson's correlation coefficient was used to identify the correlation between different variables, including the pentosidine- and MDA-linked fluorescence and the duration of haemodialysis. A P value <0.05 was considered significant.
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Results
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Subjects
Group 1 comprised six haemodialysis patients with ESRD caused by nephrosclerosis (n=2), chronic glomerulosclerosis (n=2), secondary amyloidosis (n=1), and haemolytic uraemic syndrome (n=1). Group 2 included 10 age-matched subjects without ESRD as controls, whose pathological diagnoses were malignant neoplasm (n=7), cerebral haemorrhage (n=1), bronchopneumonia (n=1), and ethylene glycol poisoning (n=1). There was no clinical or pathological evidence for diabetes mellitus in any subject. The mean ages of groups 1 and 2 were 64.7±11.4 and 61.1±10.4 years, respectively. The mean duration of haemodialysis in group 1 was 8.66±6.12 years. Significant differences were observed in the levels of blood urea nitrogen (group 1, 70.3±24.5; group 2, 23.7±10.9 mg/dl), serum creatinine (group 1, 8.7±5.42; group 2, 1.2±0.7 mg/dl) and C-reactive protein (CRP) (group 1, 18.9±16.3; group 2, 5.24±6.51 mg/dl) between the two groups, but not in that of fasting blood sugar (group 1, 123.3±41.7; group 2, 97.2±8.71 mg/dl).
Histopathological examination
Representative sections of delipidated tissues before and after collagenase and elastase digestions are shown in Figure 1
. All delipidated tissues prior to digestion contained blue stained collagen fibres (Masson trichrome stain) and dark blue stained elastin fibres (Wiegert's elastica stain). After digestion, in contrast, no collagen or elastin fibres were detected in the tissues.

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Fig. 1. Representative photomicrograph of delipidated aortic media before and after digestion. Note the preservation of collagen and elastin fibres in delipidated tissues before collagenase and elastase digestions (a and c). In contrast, no collagen or elastin fibres were detected in tissues digested by collagenase and elastase (b and d). (a and b) Masson trichrome stain; (c and d) Wiegert's elastica stain. Scale bar, 20 µm.
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Biochemical analysis
Figure 2
shows representative competitive ELISA of CF and EF, using anti-desmosine-KLH antibody. The immunoreactivity of anti-desmosine-KLH antibody was inhibited by EF, but not by CF. This suggests that the EF contained desmosine, whereas the CF did not. The soluble protein, collagen, and elastin contents of each fraction are shown in Table 1
. The absolute and relative contents of total matrix protein, defined as the sum of collagen and elastin, did not differ between the two groups or between different sites, including atherosclerotic intima, lesion-free intima, and media. In both groups, the elastin content of atheromatous intima was significantly smaller than that of lesion-free intima and media. Conversely, the collagen content was greater in atheromatous intima than in lesion-free intima and media. Consequently, the ratio of collagen to elastin was significantly higher in atheromatous intima compared with that in lesion-free intima and media. The levels of pentosidine-linked fluorescence of various tissue samples are shown in Figure 3
. In all sites of aorta, including atheromatous intima, lesion-free intima, and media, the pentosidine-linked fluorescence level of elastin was significantly higher in haemodialysis patients with ESRD (group 1) than in control subjects (group 2) (a, P<0.05). In contrast, the pentosidine-linked fluorescence level of collagen did not differ between the two groups. In addition, both haemodialysis patients with ESRD (group 1) and control subjects (group 2) exhibited significantly higher levels of pentosidine-linked fluorescence of elastin than of collagen in all aortic sites (b, P<0.05). The mean pentosidine-linked fluorescence levels of elastin in haemodialysis patients with ESRD (group 1) were 340±204, 243±132, and 187±100 (AU/mg elastin) in atheromatous intima, lesion-free intima, and media, respectively, whereas the corresponding levels in control subjects (group 2) were 87±22, 80±21, and 102±32 (AU/mg elastin), respectively. The mean pentosidine-linked fluorescence levels of collagen in group 1 were 62±22, 53±10, and 56±8 (AU/mg collagen) in atheromatous intima, lesion-free intima, and media, respectively, whereas the corresponding levels in group 2 were 46±8, 42±11, and 40±9 (AU/mg collagen), respectively. The pentosidine-linked fluorescence level of soluble protein in group 1 was significantly higher in atheromatous intima and media than that in group 2 (a, P<0.05), but no difference was observed between the levels in lesion-free intima between the two groups. The levels of MDA-linked fluorescence of various aortic tissue samples are shown in Figure 4
. The MDA-linked fluorescence level of collagen and elastin in any site was significantly increased in haemodialysis patients with ESRD (group 1) compared with control subjects (group 2) (a, P<0.05). In addition, in both groups, the level of MDA-linked fluorescence of elastin was significantly higher than that of collagen in any site (b, P<0.05). The mean MDA-linked fluorescence levels of elastin in group 1 were 513±265, 403±242, and 265±118 (AU/mg elastin) in atheromatous intima, lesion-free intima, and media, respectively, whereas the corresponding values in group 2 were 105±38, 98±25, and 128±44 (AU/mg elastin), respectively. The mean MDA-linked fluorescence levels of collagen in group 1 were 30±15, 32±7, and 37±8 (AU/mg collagen), in atheromatous intima, lesion-free intima, and media, respectively, while in group 2 the corresponding levels were 9±4, 15±3, and 16±5 (AU/mg collagen), respectively. Haemodialysis patients (group 1) exhibited significantly higher MDA-linked fluorescence levels of elastin in atheromatous intima, than in media (c, P<0.05), but no difference was observed between atheromatous intima and lesion-free intima. Table 2
shows the correlation between the pentosidine- and MDA-linked fluorescence levels of each fraction in various sites of the aorta of groups 1 and 2. There was a significant correlation between the levels of pentosidine- and MDA-linked fluorescence in collagen and in elastin in all aortic sites, including atheromatous intima, lesion-free intima, and media. The level of pentosidine-linked fluorescence of soluble protein correlated with that of MDA-linked fluorescence in media, but not in atheromatous intima and lesion-free intima. As shown in Table 3
, in haemodialysis patients with ESRD, both the pentosidine- and MDA-linked fluorescence levels of elastin demonstrated a significantly positive correlation with duration of haemodialysis in lesion-free intima and media. In contrast, the pentosidine- and MDA-linked fluorescence levels of elastin and collagen did not correlate with duration of haemodialysis in atheromatous intima.

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Fig. 2. Representative competitive ELISA using anti-desmosine-KLH antibody. Each well was coated with 1 µg/ml of albumin-conjugated desmosine. Test samples were mixed with anti-desmosine antibody and a portion of the mixture was added to each well, followed by incubation for 60 min. After washing, antibodies bound to the well were detected by reaction with HRP-conjugated second antibody, followed by o-phenylenediamine dihydrochloride. Solid triangles: desmosine standard; open squares: elastin fraction; open circles: collagen fraction.
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Table 1. Soluble protein, collagen, and elastin contents extracted from various tissue samples of aorta in haemodialysis patients with ESRD (group 1) and control subjects (group 2)
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Fig. 3. Pentosidine-linked fluorescence levels in atheromatous intima (A), lesion-free intima (I), and media (M) in haemodialysis patients (hatched bars) and control subjects (open bars). Data are expressed as the mean±SD of AU/mg protein, AU/mg collagen, and AU/mg elastin in soluble fraction (SF), collagen fraction (CF), and elastin fraction (EF), respectively. a, P<0.01 vs control subjects (group 2); b, P<0.01 vs CF.
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Fig. 4. MDA-linked fluorescence levels in atheromatous intima (A), lesion-free intima (I), and media (M) in haemodialysis patients (hatched bars) and control subjects (open bars). Data are expressed as mean±SD of AU/mg protein, AU/mg collagen, and AU/mg elastin in soluble fraction (SF), collagen fraction (CF), and elastin fraction (EF), respectively. a, P<0.01 vs control subjects (group 2); b, P<0.01 vs CF; c, P<0.05 vs media.
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Table 2. Correlation between the level of pentosidine- and MDA-linked fluorenscence in soluble protein, collagen and elastin in various tissue samples, including atheromatous intima, lesion-free intima, and media
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Table 3. Correlation between duration of haemodialysis and level of pentosidine- and MDA-linked fluorescence of soluble protein, collagen and elastin in various tissue samples, including atheromatous intima, lesion-free intima, and media in haemodialysis patients with ESRD
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Discussion
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The major findings of the present study were as follows. First, haemodialysis patients with ESRD exhibited increased pentosidine- and MDA-linked fluorescence of elastin in the aorta. Secondly, among various aortic sites in haemodialysis patients, atheromatous intima had the highest level of pentosidine- and MDA-linked fluorescence. Thirdly, in both haemodialysis patients with ESRD and control subjects without ESRD, pentosidine- and MDA-linked fluorescence levels of elastin were significantly higher in any site than those of collagen. Finally, the level of pentosidine-linked fluorescence of elastin and collagen significantly correlated with that of MDA-linked fluorescence in any aortic site.
Extracellular matrices, such as collagen and elastin, alter their biochemical properties when they undergo glycation and oxidation. These changes include loss of flexibility, altered conformation and susceptibility to enzymatic digestion [14]. In this study, we used enzymatic digestion with collagenase and elastase to obtain collagen and elastin from aortic tissues of haemodialysis patients and control subjects. Histopathological examination of delipidated aortic tissues obtained from haemodialysis patients and control subjects revealed that the fibrillar structure of collagen and elastin disappeared after enzymatic digestion. As shown in Table 1
, there was no difference in the content of total matrix protein digested by collagenase and elastase between haemodialysis patients and controls. These results suggest that aortic tissues from haemodialysis patients and control subjects were well digested by collagenase and elastase. Competitive ELISA analysis confirmed the presence of desmosine in the EF but not in the CF. In our preliminary study, in vitro glycoxidation of collagen molecules decreased their content of lysine and arginine, but did not alter that of hydroxyproline. Thus, we established an analysis for determination of collagen and elastin in aortic tissues obtained from haemodialysis patients and control subjects. However, the effects of factors other than glycoxidation on biochemical properties of matrix proteins have yet to be elucidated.
Collagen is the major structural component of the cardiovascular system, and is modified by glycoxidation and lipid peroxidation [13,14]. Haemodialysis patients with ESRD exhibit an increase in glycoxidation and lipid peroxidation products in the collagen matrix of various tissues, including heart, skin, and kidney [4,13]. Elastin has the lowest turnover rate of all matrix components, and has been shown to accumulate glycoxidation products in skin and yellow ligament [9,10]. However, little is known about modification of aortic elastin by glycoxidation and lipid peroxidation. In the present study, aortic elastin exhibited significantly higher levels of pentosidine- and MDA-linked fluorescence than collagen in haemodialysis patients with ESRD and in control subjects. This suggests that aortic elastin can be modified by glycoxidation and lipid peroxidation.
Increased levels of glycoxidation and lipid peroxidation products have been well documented in patients with ESRD [4,13]. Tissue and plasma concentrations of pentosidine are elevated in diabetic and non-diabetic patients with ESRD [4,5]. Oxidative stress is also enhanced in patients with ESRD, as evidenced by increased MDA contents in low-density lipoprotein and red blood cell membranes [15]. Moreover, oxidative stress occurs both at the lipid and protein levels [16]. We recently demonstrated increased modification of cardiac collagen by glycoxidation and lipid peroxidation in haemodialysis patients [13]. However, little is known about these modifications of elastin in haemodialysis patients. The present study demonstrated, for the first time, increased glycoxidation and lipid peroxidation products in elastin of aortic tissues in haemodialysis patients with ESRD. Moreover, among different aortic sites in these patients, atheromatous intima exhibited the highest levels of glycoxidation and lipid peroxidation products in elastin. Accumulation of N
-(carboxymethyl) lysine, a glycoxidative product, is noted in the extracellular matrix of atherosclerotic lesions in haemodialysis patients with ESRD, which correlates with the duration of haemodialysis [3]. Thus, the increased modification of aortic elastin by glycoxidation and lipid peroxidation may relate to development of vascular complications in haemodialysis patients with ESRD, including atherosclerosis.
Glycoxidation and lipid peroxidation reactions may be intrinsically linked and may play a causal role in the cardiovascular complications of patients with ESRD [13]. The formation of two glycoxidation products, N
-(carboxymethyl)lysine and pentosidine, is enhanced by a metal-catalysed reaction, as evidenced by the presence of increased metal-mediated oxidant stress in uraemia [17]. We recently demonstrated that glycoxidation and lipid peroxidation of low-density lipoprotein may synergistically promote the development of atherosclerotic lesions [6]. Glycated matrix proteins, such as elastin and collagen, have recently been shown to bind increased amounts of transition metals, which can participate in the oxidation of ascorbic acid [18]. In the present study, the level of pentosidine-linked fluorescence of elastin and collagen correlated significantly with that of MDA-linked fluorescence in any aortic site. This finding suggests that the modification of matrix proteins, including elastin and collagen, by glycoxidation and lipid peroxidation, may synergistically enhance the development of cardiovascular complications in haemodialysis patients with ESRD.
The biological consequences of increased modification of elastin by glycoxidation and lipid peroxidation in patients with ESRD could not be elucidated in the present study. Elastin is one of the major extracellular matrix components of the aorta and is implicated in various arterial diseases, including atherosclerosis and aneurysm, either through the loss of its mechanical properties or by acting as a substrate for calcification and lipid deposition. In vivo and in vitro glycation of connective tissues, including collagen and elastin, causes an increase in their stiffness [11]. Calcium deposits have been shown to increase proportionately with accumulation of glycated elastin in the aorta of diabetic rats [19]. We have recently demonstrated that haemodialysis patients with ESRD show accumulation of calcium deposits in the media layer of the aorta, and that such deposits were preferentially found around pentosidine-modified elastin fibres (data not shown). The arterial system in patients with ESRD undergoes structural remodelling, characterized by diffuse dilatation, hypertrophy, and stiffening of the large arteries [1]. Thus, we speculate that these modifications of elastin may, at least in part, be involved in the development of vascular complications in haemodialysis patients with ESRD through alterations in viscoelastic properties of arterial walls.
The development of atherosclerosis is associated with inflammatory processes occurring within arterial intima. CRP, the prototype acute-phase protein, has been shown to co-localize with the terminal complement complex within atherosclerotic lesions [20]. Our study demonstrated high levels of plasma CRP in haemodialysis patients, who exhibited increased modification of elastin and collagen by glycoxidation and lipid peroxidation in aorta. Although the biological function of CRP is largely unknown, this finding implicates the inflammatory process in atherogenesis and increased modification of elastin by glycoxidation and lipid peroxidation in haemodialysis patients.
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Acknowledgments
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This work was supported in part by a Grant-in-Aid for Scientific Research (No. 10670183) from the Ministry of Education, Science and Culture of Japan, and in part by funds from the Central Research Institute of Fukuoka University.
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Notes
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Correspondence and offprint requests to: Noriyuki Sakata, MD, Second Department of Pathology, School of Medicine Fukuoka University 45-1, 7-chome Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan. Email: nysakata{at}fukuoka\|[hyphen]\|u.ac.jp 
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Received for publication: 5. 5.01
Revision received 20.10.01.