Vitronectin is a substrate for transglutaminases

Vitronectin (VN) was found to be a substrate for both plasma transglutaminase (Factor XIIIa) and guinea pig liver transglutaminase (TG). Incorporation of [3H]-putrescine indicated the presence of reactive glutaminyl residues in VN. When VN was incubated with TG or Factor XIIIa, in the absence of putrescine, multimeric covalent complexes were identified, indicating that VN can also contribute lysyl residues to the bond catalyzed by transglutaminases. Cross-linking of VN by TG and Factor XIIIa may modulate the effects of VN on the complement and coagulation systems in hemostatic plugs and extracellular matrix.     

Gly-Pro-Arg-Pro modifies the glutamine residues in the alpha- and gamma-chains of fibrinogen: inhibition of transglutaminase cross-linking

During blood clotting Factor XIIIa, a transglutaminase, catalyzes the formation of covalent bonds between the epsilon-amino group of lysine and the gamma-carboxamide group of peptide-bound glutamine residues between fibrin molecules. We report that glycyl-L-prolyl-L-arginyl-L-proline (GPRP), a tetrapeptide that binds to the fibrin polymerization sites (D-domain) in fibrin(ogen), inhibits transglutaminase cross-linking by modifying the glutamine residues in the alpha- and gamma-chains of fibrinogen. Purified platelet Factor XIIIa, and tissue transglutaminase from adult bovine aortic endothelial cells were used for the cross-linking studies. Gly-Pro (GP) and Gly-Pro-Gly-Gly (GPGG), peptides which do not bind to fibrinogen, had no effect on transglutaminase cross-linking. GPRP inhibited platelet Factor XIIIa-catalyzed cross-linking between the gamma-chains of the following fibrin(ogen) derivatives: fibrin monomers, fibrinogen and polymerized fibrin fibers. GPRP functioned as a reversible, noncompetitive inhibitor of Factor XIIIa-catalyzed incorporation of [3H]putrescine and [14C]methylamine into fibrinogen and Fragment D1. GPRP did not inhibit 125I-Factor XIIIa binding to polymerized fibrin, demonstrating that the Factor XIIIa binding sites on fibrin were not modified. GPRP also had no effect on Factor XIIIa cross-linking of [3H]putrescine to casein. This demonstrates that GPRP specifically modified the glutamine cross-linking sites in fibrinogen, and had no effect on either Factor XIIIa or the lysine residues in fibrinogen. GPRP also inhibited [14C]putrescine incorporation into the alpha- and gamma-chains of fibrinogen without inhibiting beta-chain incorporation, suggesting that the intermolecular cross-linking sites were selectively affected. Furthermore, GPRP inhibited tissue transglutaminase-catalyzed incorporation of [3H]putrescine into both fibrinogen and Fragment D1, without modifying [3H]putrescine incorporation into casein. GPRP also inhibited intermolecular alpha-alpha-chain cross-linking catalyzed by tissue transglutaminase. This demonstrates that the glutamine residues in the alpha-chains involved in intermolecular cross-linking are modified by GPRP. This is the first demonstration that a molecule binding to the fibrin polymerization sites on the D-domain of fibrinogen modifies the glutamine cross-linking sites on the alpha- and gamma-chains of fibrinogen.     

Isolation and characterization of cDNA clones to mouse macrophage and human endothelial cell tissue transglutaminases.

The deduced amino acid sequences for tissue transglutaminases from human endothelial cells and mouse macrophages have been derived from cloned cDNAs. Northern blot analysis of both tissue transglutaminases shows a message size of approximately 3.6-3.7 kilobases. The molecular weights calculated from the deduced amino acid sequences were 77,253 for human endothelial tissue transglutaminase and 76,699 for mouse macrophage tissue transglutaminase. The deduced amino acid sequence for the human endothelial transglutaminase was confirmed by comparison with the amino acid sequence obtained by cyanogen bromide digestion of the human erythrocyte transglutaminase. The amino acid sequences of both human endothelial and mouse macrophage tissue transglutaminases were compared to other transglutaminases. A very high degree of homology was found between human endothelial, mouse macrophage, and guinea pig liver tissue transglutaminase (greater than 80%). Moreover, human endothelial tissue transglutaminase was compared with human Factor XIIIa and a very high degree of homology (75% identity) was found in the active site region.     

Regulation of the activities of the mammalian transglutaminase family of enzymes

Mammalian transglutaminases catalyze post‐translational modifications of glutamine residues on proteins and peptides through transamidation or deamidation reactions. Their catalytic mechanism resembles that of cysteine proteases. In virtually every case, their enzymatic activity is modulated by elaborate strategies including controlled gene expression, allostery, covalent modification, and proteolysis. In this review, we focus on our current knowledge of post‐translational regulation of transglutaminase activity by physiological as well as synthetic allosteric agents. Our discussion will primarily focus on transglutaminase 2, but will also compare and contrast its regulation with Factor XIIIa as well as transglutaminases 1 and 3. Potential structure–function relationships of known mutations in human transglutaminases are analyzed.     

Post-translational modification of apolipoprotein B by transglutaminases.

The major form of cross-link found in apolipoprotein B was identified as N1N12-bis-(gamma-glutamyl)spermine, a product known to be formed through the catalytic action of transglutaminases (EC 2.3.2.13). N1-(gamma-Glutamyl)spermine was present in a trace amount but epsilon-(gamma-glutamyl)lysine cross-links, which are formed during fibrin formation in plasma, were not detected. In the presence of catalytic amounts of plasma Factor XIIIa (a thrombin-dependent extracellular transglutaminase) or cellular transglutaminase (a cytosolic enzyme), apolipoprotein B and other plasma apolipoproteins (A-I, A-II and C) underwent covalently bridged polymerization and served as amine acceptor substrates. These results suggests that transglutaminases may participate in the covalent modification of apolipoproteins, either in the physiological state or during pathogenesis.     

Screening for the preferred substrate sequence of transglutaminase using a phage-displayed peptide library: identification of peptide substrates for TGASE 2 and Factor XIIIA

Mammalian transglutaminase (TGase) catalyzes covalent cross-linking of peptide-bound lysine residues or incorporation of primary amines to limited glutamine residues in substrate proteins. Using an unbiased M13 phage display random peptide library, we developed a screening system to elucidate primary structures surrounding reactive glutamine residue(s) that are preferred by TGase. Screening was performed by selecting phage clones expressing peptides that incorporated biotin-labeled primary amine by the catalytic reactions of TGase 2 and activated Factor XIII (Factor XIIIa). We identified several amino acid sequences that were preferred as glutamine donor substrates, most of which have a marked tendency for individual TGases: TGase 2, QxPphiD(P), QxPphi, and QxxphiDP; Factor XIIIa, QxxphixWP (where x and phi represent a non-conserved and a hydrophobic amino acid, respectively). We further confirmed that the sequences were favored for transamidation using modified glutathione S-transferase (GST) for recombinant peptide-GST fusion proteins. Most of the fusion proteins exhibited a considerable increase in incorporation of primary amines over that of modified GST alone. Furthermore, we identified the amino acid sequences that demonstrated higher specificity and inhibitory activity in the cross-linking reactions by TGase 2 and Factor XIIIa.     

Human inter-α-inhibitor is a substrate for factor XIIIa and tissue transglutaminase

In this study, we show that inter-α-inhibitor is a substrate for both factor XIIIa and tissue transglutaminase. These enzymes catalyze the incorporation of dansylcadaverine and biotin-pentylamine, revealing that inter-α-inhibitor contains reactive Gln residues within all three subunits. These findings suggest that transglutaminases catalyze the covalent conjugation of inter-α-inhibitor to other proteins. This was demonstrated by the cross-linking between inter-α-inhibitor and fibrinogen by either factor XIIIa or tissue transglutaminase. Finally, using quantitative mass spectrometry, we show that inter-α-inhibitor is cross-linked to the fibrin clot in a 1:20 ratio relative to the known factor XIIIa substrate α2-antiplasmin. This interaction may protect fibrin or other Lys-donating proteins from adventitious proteolysis by increasing the local concentration of bikunin. In addition, the reaction may influence the TSG-6/heavy Chain 2-mediated transfer of heavy chains observed during inflammation.     

Development of selective inhibitors of transglutaminase. Phenylthiourea derivatives

For the purpose of developing a transglutaminase inhibitor which could be effective in physiological and pharmacological studies, a series of phenylthiourea derivatives of alpha, omega-diaminoalkanes were designed, synthesized, and evaluated kinetically as inhibitors of transglutaminases. A homologous series of compounds of the structure phenylthiourea-(CH2)n-NH2, where n = 2, 3, 4, 5, and 6, were tested for the inhibition of both guinea pig liver transglutaminase-catalyzed amine incorporation into various glutamine-containing substrates and plasma transglutaminase (factor XIIIa)-catalyzed amine incorporation into fibrin and fibrin cross-linking. It was found that the inhibitory activity of the compounds increases with increasing number of methylene groups in the side chain up to a maximum of n = 5. A further increase in the length of the methylene side chain to n = 6 results in decreased activity. The Ki value (4.9 X 10(-5) M) of 1-(5-aminopentyl)-3-phenylthiourea (PPTU) (n = 5) for the inhibition of guinea pig transglutaminase-catalyzed amine incorporation into the B chain of oxidized insulin is in close agreement to its Km(app) value (7.1 X 10(-5) M) obtained using 14C-labeled PPTU. PPTU was also found to be a potent inhibitor of plasma transglutaminase-catalyzed fibrin cross-linking. The finding that the specificity of the alkylamines for inhibition is correlated with the length of their methyl side chains is compatible with those reported for aliphatic amines and monodansylcadaverine analogues (where dansyl is 5-dimethylaminonaphthalene-1-sulfonyl). The phenylthiourea derivatives, however, are far less toxic in mice than monodansylcadaverine as indicated by their LD50 values: PPTU, 400 +/- 25 mg/kg; and monodansylcadaverine, 160 +/- 20 mg/kg.     

Quantification of apo[a] and apoB in human atherosclerotic lesions.

Lipoprotein[a] or Lp[a] is a cholesterol-rich plasma lipoprotein that is associated with increased risk for cardiovascular disease. To better understand this association we determined the amount of apo[a] and apoB as possible estimates for Lp[a] and low density lipoprotein (LDL) accumulation in atherosclerotic lesions and in plasma, from patients undergoing vascular surgery, using specific radioimmunoassays for apolipoprotein[a] and apolipoprotein B. Apo[a] and apoB were operationally divided into a loosely bound fraction obtained by extracting minced samples of plaque with phosphate-buffered saline (PBS), and a tightly bound fraction obtained by extracting the residual tissue with 6 M guanidine-HCl (GuHCl). We found that 83% of all apo[a] but only 32% of all apoB in lesions was in the tightly bound fraction. When normalized for corresponding plasma levels, apo[a] accumulation in plaques was more than twice that of apoB. All fractions of tissue apo[a], loosely bound, tightly bound, and total, correlated significantly with plasma apo[a]. However, no significant correlations were found between any of the tissue fractions and plasma apoB. If all apo[a] and apoB had been associated with intact Lp[a] or LDL particles, the calculated mass of tightly bound Lp[a] would actually have exceeded that of tightly bound LDL in five cases with plasma Lp[a] levels above 5 mg apo[a] protein/dl. When PBS and GuHCl extracts of lesions were subjected to one-dimensional electrophoresis, the major band stained for lipid and immunoblotted positively for apo[a] and apoB, suggesting the presence of some intact Lp[a] in these extracts. These results suggest that Lp[a] accumulates preferentially to LDL in plaques, and that plaque apo[a] is directly associated with plasma apo[a] levels and is in a form that is less easily removable than most of the apoB. This preferential accumulation of apo[a] as a tightly bound fraction in lesions, could be responsible for the independent association of Lp[a] with cardiovascular disease in humans.     

Catalytic properties of a calmodulin-regulated transglutaminase from human platelet and chicken gizzard

The kinetic parameters and some enzymatic characteristics of human platelet and chicken gizzard transglutaminases were determined. Activity of the transglutaminases was regulated by calmodulin. These enzymes co-isolated with alpha-actinin and were dissociated from alpha-actinin by gel filtration and absorption onto a calmodulin affinity column. Silver-stained polyacrylamide gels showed that the protein peak eluted by EGTA from this column contained polypeptides of Mr approximately 58,000 and 63,000. The transglutaminases required Ca2+ for incorporation of monodansylcadaverine into casein and actin substrates. Activity was enhanced 3-fold by calmodulin with a biphasic effect, showing stimulation at 10-200 nM and inhibition at concentrations higher than 300 nM. In the presence of 200 nM calmodulin, half-maximal transglutaminase stimulation was obtained with 2.5 microM free [Ca2+]. Chlorpromazine inhibited calmodulin enhancement of the transglutaminases. Activity of the transglutaminases was independent of proteolytic activation, since inhibitors for Ca2+-dependent proteases failed to inhibit filamin cross-linking. For comparison, factor XIIa, a plasma and platelet transglutaminase, required both Ca2+ and thrombin for activation and was insensitive to calmodulin. The cross-linking pattern of fibrin, fibrin monomers, and fibrinogen by the calmodulin-regulated transglutaminases showed, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, disappearance of fibrinogen alpha-chains with no decrease of beta- and gamma-chains or formation of gamma-gamma dimers. By autoradiography, cross-linked products of 125I-fibrinogen revealed heavily labeled high molecular weight polymers and polypeptides of Mr 98,000, 116,000, and 148,000; the latter appeared to be a transient species. However, when fibrin, fibrin monomers, and fibrinogen were used as factor XIIIa substrates, gamma-gamma dimers and alpha-polymers were formed. Formation of gamma-gamma dimers was slower with fibrinogen than with fibrin. Iodoacetamide blocked activity of factor XIIIa but not of the calmodulin-regulated transglutaminases.     

Isolation and partial characterization of apolipoprotein (a) from human lipoprotein (a)

A procedure was developed for the dissociation of apolipoprotein (a) (apo (a)) from pure human lipoprotein (a) (Lp(a)) prepared by density gradient ultracentrifugation and gel filtration. Lp(a) was ultracentrifuged through a layer of saline which was adjusted to a density of 1.182 g/mL and contained 30 mM dithiothreitol (50 mM) and phenylmethylsulfonyl fluoride (1.25 mM). Following centrifugation, the lipid and apolipoprotein B (apo B) were recovered as a lipoprotein (Lp(a) B) in the supernatant fraction, while the apo (a) was recovered as a lipid-poor protein pellet. An investigation of the supernatant lipoprotein by electron microscopy and compositional analysis revealed that it was similar in size and composition to low density lipoprotein (LDL) isolated from the same density range and contained apo B100 with an amino acid and carbohydrate composition which was similar to apo B from LDL. Estimates of the apparent molecular weight of the apo (a) varied amongst individuals but was always greater than apo B100 (congruent to 450,000). The amino acid composition of apo (a), which was very distinct from apo B, was characterized by a higher content of serine, threonine, proline, and tyrosine, but lower amounts of isoleucine, phenylalanine, and lysine when compared with apo B of Lp(a) or LDL. The apo (a) contained a much higher proportion of carbohydrate, in particular N-acetylgalactosamine, galactose, and N-acetylneuraminic acid (which were three- to six-fold higher) than the apo B of Lp(a). It is concluded that apo (a) is distinct from other apolipoproteins owing to its low avidity for lipid and the nature of the interaction with apo B. Lp(a) consists of an LDL-like particle with a carbohydrate-rich apo (a) attached to the surface of apo B.     

Genetic linkage between lipoprotein(a) phenotype and a DNA polymorphism in the plasminogen gene

Coronary heart disease risk correlates directly with plasma concentrations of lipoprotein(a) (Lp(a)), a low-density lipoprotein-like particle distinguished by the presence of the glycoprotein apolipoprotein(a) (apo(a)), which is bound to apolipoprotein B-100 (apoB-100) by disulfide bridges. Size isoforms of apo(a) are inherited as Mendelian codominant traits and are associated with variations in the plasma concentration of lipoprotein(a). Plasminogen and apo(a) show striking protein sequence homology, and their genes both map to chromosome 6q26-27. In a large family with early coronary heart disease and high plasma concentrations of Lp(a), we found tight linkage between apo(a) size isoforms and a DNA polymorphism in the plasminogen gene; plasma concentrations of Lp(a) also appeared to be related to genetic variation at the apo(a) locus. We found free recombination between the same phenotype and alleles of the apoB DNA polymorphism. This suggests that apo(a) size isoforms and plasma lipoprotein(a) concentrations are each determined by genetic variation at the apo(a) locus.     

Lipoprotein [a] is cleared from the plasma primarily by the liver in a process mediated by apolipoprotein [a

The cellular and molecular mechanisms responsible for lipoprotein [a] (Lp[a]) catabolism are unknown. We examined the plasma clearance of Lp[a] and LDL in mice using lipoproteins isolated from human plasma coupled to radiolabeled tyramine cellobiose. Lipoproteins were injected into wild-type, LDL receptor-deficient (Ldlr-/-), and apolipoprotein E-deficient (Apoe-/-) mice. The fractional catabolic rate of LDL was greatly slowed in Ldlr-/- mice and greatly accelerated in Apoe-/- mice compared with wild-type mice. In contrast, the plasma clearance of Lp[a] in Ldlr-/- mice was similar to that in wild-type mice and was only slightly accelerated in Apoe-/- mice. Hepatic uptake of Lp[a] in wild-type mice was 34.6% of the injected dose over a 24 h period. The kidney accounted for only a small fraction of tissue uptake (1.3%). To test whether apolipoprotein [a] (apo[a]) mediates the clearance of Lp[a] from plasma, we coinjected excess apo[a] with labeled Lp[a]. Apo[a] acted as a potent inhibitor of Lp[a] plasma clearance. Asialofetuin, a ligand of the asialoglycoprotein receptor, did not inhibit Lp[a] clearance. In summary, the liver is the major organ accounting for the clearance of Lp[a] in mice, with the LDL receptor and apolipoprotein E having no major roles. Our studies indicate that apo[a] is the primary ligand that mediates Lp[a] uptake and plasma clearance.     

Transglutaminase reactions associated with the rat semen clotting system: modulation by macromolecular polyanions.

The coagulation of rodent semen after ejaculation involves the establishment of ?-(γ-glutamyl)lysyl cross linkages between seminal vesicle secretion proteins as catalyzed by Ca⁺⁺-dependent transglutaminases secreted by the coagulating (anterior prostate) gland. During enzymic clotting of rat vesicular secretion proteins, low molecular weight amines such as putrescine are incorporated into covalent linkage with proteins of both the coagulum and the clot liquor. Bulbourethral gland secretions and certain macromolecular polyanions (notably poly-L-glutamate) enhance the enzymic coagulation of rat vesicular secretion proteins and putrescine incorporation therein. The stimulatory macromolecular polyanions appear to exert their effects by facilitating the ability of vesicular secretion proteins to serve as transglutaminase amine acceptor substrates.     

The linkage with apolipoprotein (a) in lipoprotein (a) modifies the immunochemical and functional properties of apolipoprotein B

Lipoprotein (a) [Lp(a)] was isolated from several donors and its apolipoprotein (a) [apo(a)] dissociated by a reductive treatment, generating the apo(a)-free form of Lp(a) [Lp(a--)] that contains apolipoprotein B (apo B) as its sole protein. Using anti-apo B monoclonal antibodies, the properties of apo B in Lp(a), Lp(a--), and autologous low-density lipoprotein (LDL) were compared. Marked differences in apo B immunoreactivity were found between these lipoproteins, due to the presence of apo(a) in Lp(a). Apo(a) enhanced the expression of two epitopes in the amino-terminal part of apo B while it diminished the immunoreactivity of three other epitopes in the LDL receptor binding domain. Accordingly, the binding of the lipoproteins to the LDL receptor was also decreased in the presence of apo(a). In a different experimental system, the incubation of antibodies that react with 27 distinct epitopes distributed along the whole length of apo B sequence with plastic-bound Lp(a) and Lp(a--) failed to reveal any epitope of apo B that is sterically hindered by the presence of apo(a). Our results demonstrate that the presence of apo(a) modified the organization and function of apo B in Lp(a) particles. The data presented indicate that most likely the modification is not due to a steric hindrance but that some more profound conformational changes are involved. We suggest that the formation of the disulfide bridge between apo B and apo(a) in Lp(a) alters the system of disulfide bonds present in apo B and thereby modifies apo B structure.     

Tissue-type plasminogen activator binds to and is inhibited by surface-bound lipoprotein(a) and low-density lipoprotein

Elevated levels of lipoprotein(a) [Lp(a)] are associated with an increased risk of atherothrombotic disease, but the mechanism(s) by which Lp(a) potentiates atherogenesis is unknown. The extensive homology of apolipoprotein(a) [apo(a)] to plasminogen has led us and others to postulate that Lp(a) may impair fibrinolysis. We have previously shown that Lp(a) inhibits fibrin stimulation of plasminogen activation by tissue-type plasminogen activator (t-PA); however, we and other investigators have been unable to demonstrate direct inhibition of t-PA by Lp(a) in solution. We now report that t-PA binds reversibly and saturably to surface-bound Lp(a) and to low-density lipoprotein (LDL) and that as a result of this binding activation of plasminogen by t-PA is inhibited. The catalytic efficiency (kcat/Km) of t-PA when bound to polystyrene surface-bound fibrinogen increased 2.9-fold compared to t-PA bound to control wells. When bound to surface-bound Lp(a), however, the catalytic efficiency of t-PA was reduced 9.5-fold compared to t-PA bound to control wells; likewise, by binding to surface-bound LDL, the catalytic efficiency of t-PA was reduced 16-fold compared to the control. Studies with defined monoclonal antibodies suggest that major determinants of t-PA binding are its active site, the LDL receptor binding domain of apolipoprotein B-100 (apoB-100), and apo(a). These data suggest a unique mechanism by which Lp(a) and LDL incorporated in an atheroma can inhibit endogenous fibrinolysis and thereby contribute to the genesis of atherothrombotic disease.     

Assembly of lipoprotein (a) in transgenic rabbits expressing human apolipoprotein (a)

The study of human lipoprotein (a) [Lp(a)] has been hampered due to the lack of appropriate animal models since apolipoprotein (a) [apo(a)] is found only in primates and humans. In addition, human apo(a) in transgenic mice can not bind to murine apoB to form Lp(a) particles. In this study, we generated three independent transgenic rabbits expressing human apo(a) in their plasma at 1.8-4.5 mg/dl. In the plasma of transgenic rabbits, unlike the plasma of transgenic mice, about 80% of the apo(a) was covalently associated with rabbit apo-B and was contained in the fractions with density 1.02-1.10 g/ml, indicating the formation of Lp(a). These results suggest that transgenic rabbits expressing human apo(a) exhibit efficient assembly of Lp(a) and can be used as an animal model for the study of human Lp(a).     

脂蛋白(a)合成的调节

脂蛋白(a) ［ LP(a)］是一种与低密度脂蛋白(LDL)结构极其相似的脂蛋白,它由LDL脂质核心、载脂蛋白B100(apoB100)及特异性的成分载脂蛋白(a)［ apo(a)］组成. 大量的研究表明,高LP(a)是动脉粥样硬化独立的危险因素.而LP(a)在血浆中的水平及致病能力取决于其合成的速率及其颗粒的大小. 因此, 如何抑制LP(a)合成,进而从源头减少LP(a) 的血浆水平,对动脉粥样硬化的防治具有重要的意义.本文就当前关于影响LP(a)合成的环节及相关机制进行综述, 从而为降LP(a)药物的研究提供新的视角.     

A microtiter plate transglutaminase assay utilizing 5-(biotinamido)pentylamine as substrate.

Transglutaminases belong to an important family of enzymes involved in hemostasis, skin formation, and wound healing. We describe a technique for the measurement of transglutaminase activity using polystyrene microtiter plates coated with N,N'-dimethylcasein. The substrate 5-(biotinamido)pentylamine is covalently incorporated into N,N'-dimethylcasein by transglutaminase in a calcium-dependent reaction. The biotinylated product is detected by streptavidin-alkaline phosphatase and quantitated by measuring the absorbance at 405 nm following the addition of p-nitrophenyl phosphate. The assay is sensitive, specific, and linear at plasma factor XIIIa concentrations between 0.08 and 1.25 micrograms/ml and at purified guinea pig liver transglutaminase concentrations between 0.05 and 0.8 microgram/ml. The intra-assay coefficient of variation is less than 8%. The solid-phase assay was used to quantitate the transglutaminase activity in Escherichia coli extracts expressing recombinant factor XIII A-chains and to analyze factor XIIIa inhibitors. This method will facilitate the analysis of structure-function relationships of the transglutaminases using recombinant DNA methods. Furthermore, screening of natural and synthetic factor XIIIa inhibitors will be expedited by this solid-phase microtiter plate assay.     

Lipoprotein(a) accelerates atherosclerosis in uremic mice

Uremic patients have increased plasma lipoprotein(a) [Lp(a)] levels and elevated risk of cardiovascular disease. Lp(a) is a subfraction of LDL, where apolipoprotein(a) [apo(a)] is disulfide bound to apolipoprotein B-100 (apoB). Lp(a) binds oxidized phospholipids (OxPL), and uremia increases lipoprotein-associated OxPL. Thus, Lp(a) may be particularly atherogenic in a uremic setting. We therefore investigated whether transgenic (Tg) expression of human Lp(a) increases atherosclerosis in uremic mice. Moderate uremia was induced by 5/6 nephrectomy (NX) in Tg mice with expression of human apo(a) (n = 19), human apoB-100 (n = 20), or human apo(a) + human apoB [Lp(a)] (n = 15), and in wild-type (WT) controls (n = 21). The uremic mice received a high-fat diet, and aortic atherosclerosis was examined 35 weeks later. LDL-cholesterol was increased in apoB-Tg and Lp(a)-Tg mice, but it was normal in apo(a)-Tg and WT mice. Uremia did not result in increased plasma apo(a) or Lp(a). Mean atherosclerotic plaque area in the aortic root was increased 1.8-fold in apo(a)-Tg (P = 0.025) and 3.3-fold (P = 0.0001) in Lp(a)-Tg mice compared with WT mice. Plasma OxPL, as detected with the E06 antibody, was associated with both apo(a) and Lp(a). In conclusion, expression of apo(a) or Lp(a) increased uremia-induced atherosclerosis. Binding of OxPL on apo(a) and Lp(a) may contribute to the atherogenicity of Lp(a) in uremia.