Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.2.2.7 (heparinase)
1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study examines the role of L-selectin in monocyte adhesion to arterial endothelium, a key pathogenic event of atherosclerosis. Using a nonstatic (rotation) adhesion assay, we observed that monocyte binding to bovine aortic endothelium at 4 degrees C increased four to nine times upon endothelium activation with tumor necrosis factor (TNF)-alpha. mAb-blocking experiments demonstrated that L-selectin mediates a major part (64 +/- 18%) of monocyte attachment. Videomicroscopy experiments performed under flow indicated that monocytes abruptly halted on 8-h TNF-alpha-activated aortic endothelium, approximately 80% of monocyte attachment being mediated by L-selectin. Flow cytometric studies with a L-selectin/IgM heavy chain chimeric protein showed calcium-dependent L-selectin binding to cytokine-activated and, unexpectedly, unactivated aortic cells. Soluble L-selectin binding was completely inhibited by anti-L-selectin mAb or by aortic cell exposure to trypsin. Experiments with cycloheximide, chlorate, or neuraminidase showed that protein synthesis and sulfate groups, but not sialic acid residues, were essential for L-selectin counterreceptor function. Moreover, heparin lyases partially inhibited soluble L-selectin binding to cytokine-activated aortic cells, whereas a stronger inhibition was seen with unstimulated endothelial cells, suggesting that cytokine activation could induce the expression of additional ligand(s) for L-selectin, distinct from heparan sulfate proteoglycans. Under flow, endothelial cell treatment with heparinase inhibited by approximately 80% monocyte attachment to TNF-alpha-activated aortic endothelium, indicating a major role for heparan sulfate proteoglycans in monocyte-endothelial interactions. Thus, L-selectin mediates monocyte attachment to activated aortic endothelium, and heparan sulfate proteoglycans serve as arterial ligands for monocyte L-selectin.
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PMID:Monocyte adhesion to activated aortic endothelium: role of L-selectin and heparan sulfate proteoglycans. 904 58

Vessel wall subendothelial extracellular matrix, a dense mesh formed of collagens, fibronectin, laminin, and proteoglycans, has important roles in lipid and lipoprotein retention and cell adhesion. In atherosclerosis, vessel wall heparan sulfate proteoglycans (HSPG) are decreased and we therefore tested whether selective loss of HSPG affects lipoprotein retention. A matrix synthesized by aortic endothelial cells and a commercially available matrix (Matrigel; , Rutherford, NJ) were used. Treatment of matrix with heparinase/heparitinase (1 U/ml each) increased LDL binding by approximately 1.5-fold. Binding of lipoprotein (a) [Lp(a)] to both subendothelial matrix and Matrigel(R) increased 2-10-fold when the HSPG were removed by heparinase treatment. Incubation of endothelial cells with oxidized LDL (OxLDL) or lysolecithin resulted in decreased matrix proteoglycans and increased Lp(a) retention by matrix. The effect of OxLDL or lysolecithin on endothelial PG was abolished in the presence of HDL. The decrease in matrix HSPG was associated with production of a heparanase-like activity by OxLDL-stimulated endothelial cells. To test whether removal of HSPG exposes fibronectin, a candidate Lp(a) binding protein in the matrix, antifibronectin antibodies were used. The increased Lp(a) binding after HSPG removal was inhibited 60% by antifibronectin antibodies. Similarly, the increased Lp(a) binding to matrix from OxLDL-treated endothelial cells was inhibited by antifibronectin antibodies. We hypothesize that atherogenic lipoproteins stimulate endothelial cell production of heparanase. This enzyme reduces HSPG which in turn promotes Lp(a) retention.
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PMID:Subendothelial retention of lipoprotein (a). Evidence that reduced heparan sulfate promotes lipoprotein binding to subendothelial matrix. 925 86

Lipoprotein lipase (LPL)-mediated lipolysis of very low density lipoprotein (VLDL) has been demonstrated to increase U937 monocyte adhesion to endothelial cells. In the present study, we evaluated the ability of LPL to enhance human monocyte adhesion to bovine aortic endothelial cells (BAEC) in the absence of exogenous lipoproteins. Exposure of BAEC to 1 microgram/ml LPL at 37 degrees C resulted in a significant increase in monocyte adhesion over control values. Addition of VLDL in the culture media further enhanced the LPL effect. A significant increase in monocyte adhesion was also observed when BAEC were incubated with LPL at 4 degrees C. Heparin or heparinase treatment of BAEC totally abolished the LPL stimulatory effect on monocyte adhesion. In addition, incubation of monocytes with heparinase suppressed the ability of LPL to stimulate monocyte adhesion to endothelial cells. These treatments also markedly decreased LPL binding to the monocyte and endothelial cell surfaces. In contrast to native LPL, heat inactivated or phenylmethylsulfonyl fluoride (PMSF)-treated LPL did not increase monocyte adhesion to BAEC. Finally, incubation of LPL in the presence of the 5D2 antibody resulted in a total suppression of the LPL-induced monocyte adhesion to BAEC. Taken together, these data demonstrate that LPL activity plays an important role in LPL-induced monocyte adhesion and that LPL binding to heparan sulfate proteoglycans expressed on both monocytes and endothelial cells surfaces is required for the enhanced monocyte adhesion. These results suggest a new mechanism by which LPL may promote the development of atherosclerosis, that of facilitating monocyte adhesion to the endothelium.
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PMID:Lipoprotein lipase enhances human monocyte adhesion to aortic endothelial cells. 932 82

This study evaluated whether human monocyte-derived macrophages synthesize specific types of proteoglycans with lipoprotein-binding capability that could contribute to lipid retention in the arterial wall. After labeling with either [35S]SO4 or [35S]methionine, macrophages secreted a high molecular mass proteoglycan, with glycosaminoglycan chains of approximately 18 kDa and core protein bands of approximately 100 and 55 kDa. Both core protein bands were recognized by an antibody to PG-100, an antibody that recognizes the proteoglycan form of macrophage colony-stimulating factor (PG-100/PG-MCSF). The interaction between PG-100/PG-MCSF and low density lipoproteins (LDL) was examined by gel mobility shift. In this system, PG-100/PG-MCSF was resolved further into two forms. The two forms had the same core proteins but differed in their overall size and glycosaminoglycan content. The larger form contained glycosaminoglycan chains that were entirely chondroitin ABC lyase-sensitive, whereas the smaller form contained chains that were sensitive to both chondroitin ABC lyase and heparinase. Both forms bound native LDL with high affinity, but the larger form bound LDL with higher affinity than the smaller form. The glycosaminoglycan chains of PG-100/PG-MCSF, but not the core proteins, were responsible for binding to native LDL. Mildly oxidized LDL and methyl-LDL, which have an electrophoretic charge similar to that of native LDL, also bound PG-100/PG-MCSF. In contrast, extensively oxidized LDL and acetyl-LDL, which are more electronegative than native LDL, did not bind to either form of PG-100/PG-MCSF. The demonstration of two forms of human monocyte-derived macrophage PG-100/PG-MCSF which bind LDL may represent an additional role for macrophages in the extracellular trapping of lipoproteins in atherosclerosis.
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PMID:Human monocyte-derived macrophages secrete two forms of proteoglycan-macrophage colony-stimulating factor that differ in their ability to bind low density lipoproteins. 963 47

Lp(a) is a major inherited risk factor for premature atherosclerosis. The mechanism of Lp(a) atherogenicity has not been elucidated, but likely involves both its ability to interfere with plasminogen activation and its atherogenic potential as a lipoprotein particle after receptor-mediated uptake. We demonstrate that Lp(a) stimulates production of vascular cell adhesion molecule 1 (VCAM-1) and E-selectin in cultured human coronary artery endothelial cells (HCAEC). This effect resulted from a rise in intracellular free calcium induced by Lp(a) and could be inhibited by the intracellular calcium chelator, BAPTA/AM. The involvement of the LDL and VLDL receptors in Lp(a) activation of HCAEC were ruled out since Lp(a) induction of adhesion molecules was not prevented by an antibody (IgGC7) to the LDL receptor or by receptor-activating protein, an antagonist of ligand binding to the VLDL receptor. Addition of alpha2-macroglobulin as well as treatment with heparinase, chondroitinase ABC, and sodium chlorate did not decrease levels of VCAM-1 and E-selectin stimulated by Lp(a), suggesting that neither the low density lipoprotein receptor-related protein nor cell-surface proteoglycans are involved in Lp(a)-induced adhesion molecule production. Neither does the binding site on HCAEC responsible for adhesion molecule production by Lp(a) appear to involve plasminogen receptors, as levels of VCAM-1 and E-selectin were not significantly decreased by the addition of glu-plasminogen, the lysine analog epsilon-aminocaproic acid, or by trans-4-(aminomethyl)-cyclohexanecarboxymethylic acid (tranexamic acid), which acts by binding to the lysine binding sites carried on the kringle structures in plasminogen. In contrast, recombinant apolipoprotein (a) [r-apo(a)] competed with Lp(a) and attenuated the expression of VCAM-1 and E-selectin. In summary, we have identified a calcium-dependent interaction of Lp(a) with HCAEC capable of inducing potent surface expression of VCAM-1 and E-selectin that does not appear to involve any of the known potential Lp(a) binding sites. Because leukocyte recruitment to the vessel wall appears to represent one of the important early events in atherogenesis, this newly described endothelial cell-activating effect of Lp(a) places it at a crucial juncture in the initiation of atherogenic disease and may lead to a better understanding of the role of Lp(a) in the vascular biology of atherosclerosis.
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PMID:Expression of adhesion molecules by lp(a): a potential novel mechanism for its atherogenicity. 983 67

Aggregated low density lipoprotein (LDL) is taken up by macrophages at enhanced rate, leading to macrophage cholesterol accumulation and foam cell formation. Since macrophages were shown to mediate self aggregation of modified forms of LDL, we sought to study the effect of macrophages on the susceptibility of native LDL to aggregation. Incubation of LDL (100 microg of protein/ml) with J-774A.1 macrophage-like cell line for 18 h at 37 degrees C, led to a 114 and 56% enhanced susceptibility of LDL to aggregation by vortexing and by Bacillus cereus SMase respectively. Macrophage conditioned media (MCMs) that were obtained from J-774A.1 cells also enhanced the susceptibility of LDL to aggregation by vortexing and SMase by 134 and 75% respectively, suggesting the involvement of macrophage secretory products in the enhanced aggregation of LDL. As proteoglycans were shown to be involved in lipoprotein aggregation, we analyzed the possible involvement of macrophage-released proteoglycans in LDL aggregation. Incubation of LDL (100 microg protein/ml) with 25 microg of proteoglycans that were isolated from MCM led to a dose-dependent enhanced susceptibility of LDL to aggregation by vortexing or by SMase by up to 62 and 77% respectively. The stimulatory effect of the MCMs on LDL aggregation was markedly reduced upon MCMs treatment with the glycosaminoglycan hydrolyzing enzyme chondroitinase ABC, chondroitinase AC, but not heparinase. On the contrary, incubation of LDL (100 microg of protein/ml) with increasing concentrations (up to 50 microg/ml) of chondroitin sulfate, or heparan sulfate enhanced the susceptibility of LDL to aggregation by up to 98 or by only 18% respectively, in comparison with non-treated LDL. Since macrophages under atherogenic conditions (cholesterol-loading, cellular lipid peroxidation and activation) demonstrate enhanced secretion of proteoglycans, we finally studied the effect of J-774A.1 macrophages on the susceptibility of native LDL to aggregation under the above atherogenic conditions. Incubation of LDL with cholesterol-loaded macrophages led to a 62% enhanced susceptibility of LDL to undergo aggregation by vortexing, in comparison with LDL that was incubated with non-loaded cells. Macrophage activation with phorbol myristate acetate (5 microM of PMA) also significantly increased cell-mediated aggregation of LDL by 50%, in comparison with non-activated cells. Lipid peroxidized macrophages obtained by cell treatment with either FeSO4 (50 microM), or angiotensin II (10(-7) M) enhanced the susceptibility of LDL to aggregation by 22 or by 39% respectively. These results suggest that under atherogenic conditions, macrophages release proteoglycans, and mainly chondroitin sulfate, which can contribute to cell-mediated formation of aggregated LDL, a potent inducer of macrophage foam cells which are the hallmark of early atherogenesis.
Atherosclerosis 1999 Jan
PMID:Macrophage released proteoglycans are involved in cell-mediated aggregation of LDL. 992 May 6

Lipoprotein lipase (LPL) and apolipoprotein E (apo E) independently enhance binding and uptake of lipoproteins to cells. A coordinate effect of LPL and apo E has been previously described in human hepatozytes where simultaneous addition of both proteins resulted in an additive increase of chylomicron binding and uptake. The role of lipoprotein receptors and proteoglycans in this coordinate effect was now analysed using various cell types and heparinase treatment. To investigate a pathophysiological relevance, the effect of LPL and normal apo E-3 was compared to LPL and four apo E variants, associated with type III hyperlipoproteinemia (HLP). Apo E-3 and LPL increased the binding and uptake of chylomicrons and beta-very low density lipoproteins (VLDL) in an additive way in all cell types analysed, except proteoglycan deficient Chinese hamster ovary (CHO)-cells. Heparinase treatment almost completely abolished the effect of apo E and LPL. Addition of LPL to the apo E variants resulted in significant compensation of their defective function in mediating beta-VLDL binding to low density lipoprotein (LDL)-receptor defective fibroblasts. These findings indicate that the coordinate effect of apo E and LPL is mediated by proteoglycans and lipoprotein receptors, independent of the LDL receptor. LPL may compensate for the defective function of apo E variants by enhancing lipoprotein binding to these receptors. Defects in this mechanism may explain how mutations in the LPL molecule contribute to the manifestation of type III HLP in addition to the presence of a defective apo E.
Atherosclerosis 1999 Jul
PMID:Lipoprotein lipase compensates for the defective function of apo E variants in vitro by interacting with proteoglycans and lipoprotein receptors. 1042 96

Lipoprotein interactions with macrophage proteoglycans (PGs) is believed to play an important role in the cellular uptake of lipoproteins and in macrophage cholesterol accumulation. Recently, we have shown the participation of macrophage plasma membrane glycosaminoglycans (GAGs) in the cellular uptake of oxidized LDL (Ox-LDL). The aim of the present study was to identify the specific cell surface proteoglycans involved in this interaction. J-774 A.1 macrophage-like cell line plasma membrane proteoglycans were isolated by anion exchange chromatography from cells that were prelabeled with [35S]sodium sulfate. Using Sepharose 6B chromatography, cell surface major proteoglycans were identified as chondroitin sulfate (CS) proteoglycans (77%) and heparan sulfate (HS) proteoglycans (23%). Binding rates of these 35S-labeled proteoglycans to Ox-LDL and to native LDL were analyzed by their ability to bind lipoproteins coupled to a CnBr-activated Sepharose CL-4B chromatography. Of the total labeled cell surface proteoglycans added to the column, 57% were bound to the Sepharose-coupled Ox-LDL, whereas 73% of the cell surface proteoglycans were bound to the Sepharose-coupled native LDL. Binding of the plasma membrane macrophage 35S-labeled proteoglycans to Ox-LDL was inhibited by adding increasing concentrations of non-labeled chondroitin sulfate, or by pretreatment of the 35S-labeled proteoglycans fraction with chondroitinase ABC. In contrast, neither the addition of non-labeled heparan sulfate, nor pretreatment of the labeled proteoglycans fraction with heparinase III, had any significant effect on proteoglycan binding to Ox-LDL. These findings were further supported by using mutant cells characterized by specific glycosaminoglycan deficiencies. Ox-LDL binding and degradation by mutant 745 CHO cells which are characterized by a deficiency in both heparan sulfate and chondroitin sulfate, was decreased by 28 and 27% respectively, compared to the binding of Ox-LDL to the wild-type CHO cells. Ox-LDL binding and degradation by mutant 677 CHO cells, which lack heparan sulfate but have increased levels of chondroitin sulfate, however, was found to be increased by 29 and 19%, respectively, compared to Ox-LDL binding to the wild-type CHO cells. Finally, analysis of the cell surface proteoglycans in macrophages that were subjected to oxidative stress, by their preincubation with angiotensin II, exhibited a 51-59% increase in their cell surface proteoglycan content, with a major effect on chondroitin sulfate proteoglycans. The present study thus demonstrated that Ox-LDL can specifically bind to macrophage surface chondroitin sulfate proteoglycans, and the macrophage content of this proteoglycan is increased under oxidative stress. The interaction between macrophage chondroitin sulfate proteoglycans and Ox-LDL can contribute to enhanced uptake of Ox-LDL with the formation of cholesterol-loaded foam cells, and accelerated atherosclerosis.
Atherosclerosis 2000 Mar
PMID:Macrophage plasma membrane chondroitin sulfate proteoglycan binds oxidized low-density lipoprotein. 1070 9

Aggregated low-density lipoprotein (LDL) was shown to be present in the atherosclerotic lesion, but the mechanism responsible for its formation in vivo is not known yet. To find out whether LDL aggregation occurs in the arterial wall during atherogenesis, LDLs were extracted from the aortas of apolipoprotein E-deficient (E(0)) mice during their aging (and the development of atherosclerosis), and were analyzed for their aggregation states, in comparison to LDLs isolated from aortas of control mice. LDL isolated from aortas of E(0) mice was already aggregated at 1 month of age and its aggregation state substantially increased with age, with 3-fold elevation at 6 months of age compared to younger, 1-month-old, mice. Only minimal aggregation could be detected in LDL derived from control mice. Electron microscopy examination revealed that LDL particles from aortas of the E(0) mice were heterogeneous in their size, ranging between 20 and 300 nm. The mouse aortic LDL contained proteoglycans (PGs) and their content increased with the age of the mice, with about 2-fold higher levels than those found in LDLs derived from aortas of control mice. Macrophage-released PGs were previously demonstrated to enhance LDL aggregation in vitro. However, their involvement in LDL aggregation in vivo has not been studied yet. Thus, we next studied the effect of arterial macrophage-released PGs on the susceptibility of plasma LDL to aggregation by Bacillus cereus sphingomyelinase (SMase). Foam cell macrophages were isolated from aortas of the atherosclerotic E(0) mice at 6 months of age and were found to be loaded with cholesterol and to contain oxidized lipids. To analyze the effect of macrophage-released PGs on LDL aggregation, PGs were prelabeled by cell incubation with [35S]sulfate, followed by incubation of macrophage-released PGs with E(0) mouse plasma LDL (200 microg protein/ml) for 1 h at 37 degrees C. [35S]Sulfated PGs were found to be LDL-associated and the susceptibility of PG-associated LDL to aggregation by SMase was increased by up to 45% in comparison to control LDL. Similar results demonstrating the involvement of PGs in LDL aggregation were obtained upon incubation of LDL with increasing concentrations of PGs that were isolated from the entire aorta of E(o) mice (rather than the isolated macrophages). The stimulatory effect of macrophage-released PGs on LDL aggregation was markedly reduced when the PGs were pretreated with the glycosaminoglycan-hydrolyzing enzymes, chondroitinase ABC or chondroitinase AC, and to a much lesser extent with heparinase. We thus conclude that macrophage-released chondroitin sulfate PG can contribute to the formation of atherogenic aggregated LDL in the arterial wall.
Atherosclerosis 2000 May
PMID:Macrophage-released proteoglycans enhance LDL aggregation: studies in aorta from apolipoprotein E-deficient mice. 1078 39

Vascular smooth muscle cell (VSMC) proliferation is a key event in the development and progression of atherosclerotic lesions. Accumulating evidence suggests that lipoprotein lipase (LPL) produced in the vascular wall may exert proatherogenic effects. The aim of the present study was to examine the effect of LPL on VSMC proliferation. Incubation of growth-arrested human VSMCs with purified endotoxin-free bovine LPL for 48 and 72 hours, in the absence of any added exogenous lipoproteins, resulted in a dose-dependent increase in VSMC growth. Addition of VLDLs to the culture media did not further enhance the LPL effect. Treatment of growth-arrested VSMCs with purified human or murine LPL (1 microg/mL) led to a similar increase in cell proliferation. Neutralization of bovine LPL by the monoclonal 5D2 antibody, irreversible inhibition, or heat inactivation of the lipase suppressed the LPL stimulatory effect on VSMC growth. Moreover, preincubation of VSMCs with the specific protein kinase C inhibitors calphostin C and chelerythrine totally abolished LPL-induced VSMC proliferation. In LPL-treated VSMCs, a significant increase in protein kinase C activity was observed. Treatment of VSMCs with heparinase III (1 U/mL) totally inhibited LPL-induced human VSMC proliferation. Taken together, these data indicate that LPL stimulates VSMC proliferation. LPL enzymatic activity, protein kinase C activation, and LPL binding to heparan sulfate proteoglycans expressed on VSMC surfaces are required for this effect. The stimulatory effect of LPL on VSMC proliferation may represent an additional mechanism through which the enzyme contributes to the progression of atherosclerosis.
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PMID:Proliferative effect of lipoprotein lipase on human vascular smooth muscle cells. 1103 Dec 6


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