Biological modification of lipoproteins and its role in atherogenesis.
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Animals↗
Biomedical subjects
S Parthasarathy
Publications and source records attributed to S Parthasarathy.
Feature recognition in skull X-rays using digital image processing techniques.
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Algorithms↗
Oxidation of low-density lipoprotein by thiol compounds leads to its recognition by the acetyl LDL receptor.
Reduced glutathione and other compounds with free -SH groups promoted the oxidation of low-density lipoprotein (LDL) in the absence of cells in Ham's F-10 medium. In contrast, compounds in which the thiol groups were oxidized or blocked were ineffective in oxidizing LDL. Thiol-induced modification of LDL did not occur in media lacking in redox metals. It is suggested that thiols react with redox metal, generating thiol- and oxygen-derived free radicals that promote modification of LDL.
Animals↗
Recognition of solubilized apoproteins from delipidated, oxidized low density lipoprotein (LDL) by the acetyl-LDL receptor.
Macrophages express a specific receptor that recognizes acetylated low density lipoprotein (LDL) and certain other chemically modified forms of LDL but not native LDL. LDL oxidatively modified either by incubation with endothelial cells in Ham's F-10 medium or by incubation with 5 microM copper(II) ion in the absence of cells is recognized by this same receptor. This oxidative modification, whether cell-induced or copper-catalyzed, is accompanied by many changes in the physical and chemical properties of LDL, including an increase in density, conversion of phosphatidylcholine to lysophosphatidylcholine, generation of lipid peroxides, and degradation of apolipoprotein B-100. Which changes are essential for eliciting the recognition by the receptor is not known. In the present paper it is shown that fragments of the degraded apolipoprotein from delipidated, oxidized LDL can be almost quantitatively resolubilized using n-octyl beta-D-glucopyranoside. These 125I-labeled, solubilized apoproteins were degraded rapidly by mouse peritoneal macrophages, and that degradation was competitively inhibited by unlabeled acetyl-LDL and endothelial cell-modified LDL but not by native LDL. These results show that the acetyl-LDL receptor recognizes an epitope on the apoprotein moiety, either newly generated or exposed as a result of oxidative modification, rather than some oxidized lipid moiety. Further, the results suggest that the lipids of oxidatively modified LDL do not play an obligatory role in determining the conformation of that epitope.
Animals↗
Oxidatively modified low density lipoproteins: a potential role in recruitment and retention of monocyte/macrophages during atherogenesis.
Previous studies in this laboratory established that low density lipoprotein (LDL) incubated with cultured endothelial cells, smooth muscle cells, or macrophages undergoes free radical-catalyzed oxidative modification that generates lipid peroxides and extensive structural changes in the LDL molecule. The oxidatively modified LDL strongly inhibited chemotactic responses of the mouse resident peritoneal macrophage. The present studies show that this oxidized LDL does not inhibit the motility of mouse monocytes and actually exhibits a chemotactic activity for human monocytes; the chemotactic activity of the oxidized LDL resides in the lipid fraction. These findings allow us to propose a pathogenetic sequence by which elevated plasma LDL levels, followed by oxidative modification in the arterial wall, could sufficiently account for the generation of the lipid-laden foam cells and the initiation of the fatty streak, the earliest well-defined lesion in atherogenesis.
Animals↗
Nonenzymatic oxidative cleavage of peptide bonds in apoprotein B-100.
Incubation of low density lipoprotein (LDL) with endothelial cells converts it to a form that is avidly degraded by macrophages via the acetyl LDL receptor. This modification has previously been shown to be accompanied by extensive breakdown of the major LDL protein (apoB-100) to smaller peptides. ApoB-100 is known to undergo partial degradation during isolation and purification which is commonly attributed to proteolytic enzymes derived from plasma or to contaminant bacteria. In the present studies addition of any of ten different inhibitors of proteolytic enzymes failed to inhibit the endothelial cell-induced degradation of LDL apoB-100 or its subsequent enhanced rate of degradation by macrophages (termed biological modification). Conversely, deliberate digestion of LDL with any of five well-characterized proteolytic enzymes degraded apoB-100 extensively but did not cause biological modification. The disappearance of intact apoB-100 during incubation with endothelial cells paralleled the formation of thiobarbituric acid (TBA)-reactive substances and the breakdown could be completely prevented by the addition of antioxidants or metal chelators. Finally, the incubation of LDL with a free radical-generating system (dihydroxyfumaric acid and Fe3+-ADP) in the absence of cells resulted in the breakdown of apoB-100. These results suggest that the breakdown of apoB-100 during oxidative modification of LDL, whether cell-induced or catalyzed by transition metals, is not mediated by proteolytic enzymes but rather is linked to oxidative attack on the polypeptide chain, either directly or secondary to peroxidation of closely associated LDL lipids.
Amino Acids↗
Probucol inhibits oxidative modification of low density lipoprotein.
Previous studies have established that low density lipoprotein (LDL) incubated with endothelial cells (EC) undergoes extensive oxidative modification in structure and that the modified LDL is specifically recognized by the acetyl LDL receptor of the macrophage. Thus, in principle, EC-modified LDL could contribute to foam cell formation during atherogenesis. Oxidatively modified LDL is also potentially toxic to EC. The present studies show that addition of probucol during the incubation of LDL with EC prevents the increase in the electrophoretic mobility, the increase in peroxides, and the increase in subsequent susceptibility to macrophage degradation. It has also been shown that oxidation of LDL catalyzed by cupric ion induces many of the same changes occurring during EC modification. Addition of probucol (5 microM) also prevented this copper-catalyzed modification of LDL. Most importantly, samples of LDL isolated from plasma of hypercholesterolemic patients under treatment with conventional dosages of probucol were shown to be highly resistant to oxidative modification either by incubation with endothelial cells or by cupric ion in the absence of cells. The findings suggest the hypothetical but intriguing possibility that probucol, in addition to its recognized effects on plasma LDL levels, may inhibit atherogenesis by limiting oxidative LDL modification and thus foam cell formation and/or EC injury. Other compounds with antioxidant properties might behave similarly.
Animals↗
Endothelial cell-derived chemotactic activity for mouse peritoneal macrophages and the effects of modified forms of low density lipoprotein.
Cultured rabbit and bovine aortic endothelial cells generated chemotactic activity for mouse resident peritoneal macrophages, demonstrable in the conditioned medium. This chemotactic activity was heat stable and was not extracted into chloroform/methanol. It was inhibited by addition of endothelial cell-modified low density lipoprotein (EC-modified LDL), a form of LDL shown previously to contain peroxidized lipids, increased lysophosphatidylcholine, and partially degraded apoprotein B. The chemotactic activity was also inhibited by LDL previously oxidized in the absence of cells with 5 microM Cu2+. Inhibitory activity was present in the lipid extract of EC-modified LDL but not in that of native LDL, presumably representing peroxidized lipid components. EC-modified LDL also inhibited the chemotactic activity of zymosan-activated serum. Because EC-modified LDL is taken up in part by way of the acetyl-LDL receptor, the effects of acetyl-LDL were tested. Rather than inhibiting chemotaxis, acetyl LDL showed intrinsic positive chemotactic activity as did also fucoidin and polyinosinic acid, both of which also interact with the acetyl-LDL receptor. These studies suggest mechanisms by which macrophages may be recruited into the subendothelial space by endothelial cell-derived chemotactic factors or by natural polyanions structurally related to fucoidin or polyinosinic acid and then become "trapped" there because of the inhibitory effects of peroxidized lipid components in modified forms of LDL.
Animals↗
Essential role of phospholipase A2 activity in endothelial cell-induced modification of low density lipoprotein.
Previous studies have established that incubation of low density lipoprotein (LDL) with cultured endothelial cells (EC) converts it to a new form (EC-modified LDL) that is now recognized by a specific receptor on macrophages (the acetyl LDL receptor) and is taken up and degraded 3-10 times more rapidly than native LDL (biological modification). The formation of EC-modified LDL depended on generation of free radicals with consequent peroxidation of LDL lipids and was accompanied by extensive hydrolysis of LDL phosphatidylcholine at the 2-position. The present studies show that p-bromophenacyl bromide, a site-specific irreversible inhibitor of phospholipase A2 activity, blocks this hydrolysis and, at the same time, the enhanced macrophage degradation. We show further that during EC modification the apoprotein B of LDL undergoes considerable modification and that this also is prevented by the phospholipase inhibitor. Finally, as reported previously, changes similar to those observed on incubation of LDL with EC can be induced by incubation in the absence of cells but in the presence of a sufficiently high concentration of Cu2+. This also is accompanied by hydrolysis of phosphatidylcholine at the 2-position and breakdown of apoprotein B. These changes are also inhibited by p-bromophenacyl bromide, suggesting the presence of a phospholipase A2 activity associated with LDL as it is isolated. A hypothesis is presented linking lipid peroxidation, phosphatidylcholine hydrolysis, and changes in the LDL apoprotein during EC modification.
Acetophenones↗
Modification of low density lipoprotein by endothelial cells involves lipid peroxidation and degradation of low density lipoprotein phospholipids.
Low density lipoprotein (LDL) incubated with cultured endothelial cells from rabbit aorta or human umbilical vein is altered in several ways (EC-modified): (i) It is degraded by macrophages much faster than LDL similarly incubated in the absence of cells or incubated with fibroblasts. (ii) Its electrophoretic mobility is increased. (iii) Its density is increased. We report here that antioxidants completely prevent these changes. We also report that these changes do not take place if transition metals in the medium are chelated with EDTA. During EC-modification as much as 40% of the LDL phosphatidylcholine is degraded to lysophosphatidylcholine by a phospholipase A2-like activity. When incubation conditions in the absence of cells were selected to favor oxidation--for example, by extending the time of incubation of LDL at low concentrations, or by increasing the Cu2+ concentration--LDL underwent changes very similar to those occurring in the presence of cells, including degradation of phosphatidylcholine. Hence, some phospholipase activity appears to be associated with the isolated LDL used in these studies. The results suggest a complex process in which endothelial cells modify LDL by mechanisms involving generation of free radicals and action of phospholipase (s).
Animals↗
Structural specificity in demyelination induced by lysophospholipids.
The demyelinating activity of lysophosphatidylcholine (lysoPC) and various structural analogs in rat sciatic nerve was evaluated by following electrophysiologic changes within the first hour and 1 week after intraneural injection. The lysophospholipids tested included 1-O-hexadecanoyl-sn-glycero-3-phosphocholine (1-acyl-GPC), 3-O-hexadecanoyl-sn-glycero-1-phosphocholine (3-acyl-GPC), 1-O-hexadecanoylpropanediol-3-phosphocholine (acyl-PPC), 1-O-hexadecylpropanediol-3-phosphocholine (alkyl-PPC) and 1-acyl-sn-glycero-3-phosphoethanolamine (1-acyl-GPE). Changes in conduction velocity, width, amplitude and time integral percentage were measured. Within 1 hour, the highest demyelinating activity was observed for alkyl-PPC, followed by 3-acyl-GPC, 1-acyl-GPC and acyl-PPC. Hydrolysis products of lysoPC (glycerophosphocholine, fatty acid), lysophosphatidylethanolamine (1-acyl-GPE), biradyl choline phospholipids (1,2-di-O-alkyl-rac-glycero-3-phosphocholine, dialkyl-GPC) or sodium deoxycholate proved ineffective in these short-term experiments. One week after intraneural injection, all lysophospholipids tested caused severe electrophysiologic changes, although dialkyl-GPC and sodium deoxycholate did not. Our data suggest (i) that differences in early demyelinating activity by the choline lysophospholipids are related to their rate of turnover, as highest activity was associated with the agents that are not metabolized by lysophospholipase (e.g., alkyl-PPC) or lysolecithin acyltransferase (e.g., 3-acyl-GPC), (ii) that the lysoPC molecule as such and not its products of catabolism causes demyelination, and (iii) that demyelinating activity is not due to the general detergent action of lysoPC, but rather that specific interactions appear to trigger the processes of demyelination induced by lysophospholipids.
Action Potentials↗
Ergonomic considerations in the design of interactive languages for process control.
The paper introduces the concept of software ergonomics and stresses the importance of respecting the working habits of the user while designing the software aids. Language design criteria are presented under two broad categories: those involving the syntax of the language and those involving its semantics. These criteria are then exemplified using an actual experience of designing an interactive language (TOOL) for a large public utility.
Journal Article↗
Technique assessment of demyelination from endoneurial injection.
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Animals↗
Hypolipidemic drugs are inhibitors of phosphatidylcholine synthesis.
Clofibric acid (CPIB) and several other systemic hypolipidemic drugs are shown to block phosphatidylcholine synthesis by inhibiting cholinephosphotransferase (ChoPTase; CDPcholine:1,2-diacylglycerol cholinephosphotransferase, EC 2.7.8.2) and particularly lysolecithin acyltransferase (LLAcylTase; acyl-CoA:1-acylglycero-3-phosphocholine O-acyltransferase, EC 2.3.1.23) of rat liver microsomes. Whereas millimolar drug concentrations are required to affect de novo lecithin synthesis catalyzed by ChoPTase, reacylation of lysolecithin by LLAcylTase is inhibited at micromolar levels. Increasing effectiveness in ChoPTase inhibition is observed in the series CPIB, SaH-42-348, tibric acid, S-321328, WY-14643, S-8527, and DH-990, with IC50 ranging from 22 mM (CPIB) to 0.3 mM (DH-990). LLAcylTase inhibition by the hypolipidemic drugs follows the same general pattern, but IC50 concentrations range from 9 mM (CPIB) to 40 microM (DH-990). The agents inhibit ChoPTase (Ki, 25-0.25 mM) and LLAcylTase (Ki, 10-0.025 mM) noncompetitively. The data suggest that inhibition of phosphatidylcholine synthesis, particularly by the LLAcylTase pathway, may be related to a drug's effectiveness in decreasing serum triglyceride and cholesterol levels by blocking lipoprotein synthesis.
1-Acylglycerophosphocholine O-Acyltransferase↗
A re-examination of the fate of glyceride-glycerol in neutral lipid absorption and transport.
Conventional ideas concerning the unidirectional movement of triacylglycerol from intestinal lumen to lymph with sn-2-monoacylglycerol being the major glyceride-glycerol precursor were challenged by our finding that steady state specific activities of radiolabeled triacylglycerol (glyceryl moiety) in the intestinal mucosa and lumen were greatly reduced as compared to the specific activity of intraduodenally infused triacylglycerol. Investigation of the point at which the radiolabel was diluted was performed in mesenteric lymph duct-cannulated rats with a duodenal cannula through which trioleoyl[3H]glycerol was constantly infused. Both within the bowel lumen and in the intestinal mucosa, monoacylglycerol, diacylglycerol, and triacylglycerol specific activities were 31% or less of the specific activity of the infusate; chylomicron triacylglycerol specific activity was 75%. Efflux of neutral lipid from the mucosa into the bowel lumen was directly demonstrated by finding that when 3H glucose was injected intraperitoneally during triolein infusion, luminal triacylglycerol had a higher specific activity than was present in the mucosa. We conclude that there are two pools of mucosal triacylglycerol. One is rapidly transported and derives most of its glyceride-glycerol from luminal monoacylglycerol. The second is slowly transported; it derives its glyceride-glycerol mainly from endogenous sources and may efflux back into the bowel lumen.
Animals↗
Modulation of phosphatidylcholine synthesis in vitro. Inhibition of diacylglycerol cholinephosphotransferase and lysophosphatidylcholine acyltransferase by centrophenoxine and neophenoxine.
1,2-Diacyl-sn-glycerol : CDPcholine cholinephosphotransferase (EC 2.7.8.2) and acyl-CoA : 1-acyl-sn-glycero-3-phosphocholine acyltransferase (EC 2.3.1.23) activities of rat liver microsomes can be inhibited by centrophenoxine (N,N-dimethylaminoethyl p-chlorophenoxyacetate). This inhibition is brought about by the intact centrophenoxine molecule rather than by the products of hydrolysis. A nonhydrolyzable ether analog of centrophenoxine was synthesized (neophenoxine; N,N-dimethylaminoethyl p-chlorophenoxyethyl ether) and proved most effective in inhibiting the two routes of phosphatidylcholine biosynthesis. While 50% inhibition of the cholinephosphotransferase was attained at 5 mM neophenoxine, 50% inhibition of the acyltransferase required 0.6 mM neophenoxine levels only. Inhibition of the cholinephosphotransferase (Ki approximately 1.5 mM) and the acyltransferase (Ki approximately 1 mM) by neophenoxine was shown to be noncompetitive. Other membrane-bound enzymes, such as glucose-6-phosphatase, monoacylglycerol lipase, alkaline phosphatase or phospholipase A2 were not affected by the inhibitors. Because of this specificity, and because of the high affinity of the microsomal membrane for such agents, centrophenoxine and neophenoxine should prove useful for controlling phosphatidylcholine synthesis and for modulating the phosphatidylcholine deacylation-reacylation cycle.
1-Acylglycerophosphocholine O-Acyltransferase↗
Rapid phospholipase A2 stimulation and diacylglycerol cholinephosphotransferase inhibition in baby hamster kidney cells during initiation of dengue virus infection.
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1-Acylglycerophosphocholine O-Acyltransferase↗
Modulation of diacylglycerol acyltransferase by lysophosphatidylcholine and related monochain phospholipids.
Acyl-CoA : 1,2-diacylglycerol O-acyltransferase (EC 2.3.1.20) activity of rat liver microsomes was found to be stimulated by 1-acyl-sn-glycero-3-phosphocholine at low concentrations, but was inhibited above 0.2 mM. Diacylglycerol acylation was optimal at 75 microM lysophosphatidylcholine, resulting in a more than 2-fold activation of the enzyme. Acyltransferase activity disappeared above 0.5 mM lysophosphatidylcholine levels. 0.05% sodium taurocholate supplementation reduced diacylglycerol acyltransferase activity by approx. 2/3 over the entire range of lysophosphatidylcholine concentrations. 1-O-Hexadecylpropanediol-3-phosphocholine was shown to mimic lysophosphatidylcholine at stimulatory and at inhibitory concentrations in the absence and in the presence of sodium taurocholate, thus ruling out acyl-CoA depletion due to lysophosphatidylcholine acylation as a cause of depressed triacylglycerol synthesis at higher lysophosphatidylcholine levels. 1-Acyl-sn-glycero-3-phosphoethanolamine stimulated diacylglycerol acyltransferase to a lesser extent, without showing inhibition at higher concentrations. The data point towards a direct effect of the lysophospholipids on the acyltransferase system.
Acyltransferases↗