| For medical practitioners and the general public - Cholesterol Journal Article Catalog. |
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| Free cholesterol fiber-optic biosensor for serum samples with simplex optimization Marazuela, M. D., B. Cuesta, et al. (1997), Biosens Bioelectron 12(3): 233-40. Abstract: An optical fiber biosensor for free cholesterol monitoring in serum samples is described. Silicone-entrapped tris(4,7-diphenyl-1,10-phenanthroline) ruthenium(II) complex, the luminescence of which is sensitive to oxygen changes, is used as an optical transducer of the oxidation of cholesterol by cholesterol oxidase. The biocatalyst is entrapped in a graphite powder layer deposited onto the dyed silicone film. Optimization of some interdependent chemical variables which affect the performance of the biosensor has been achieved by application of a super-modified simplex method. The dynamic range of the biosensing membranes is found to be 0.15-3.0 mM of free cholesterol. Studies of the reproducibility, stability and interferences of the device, as well as the application of the sensor to measurements in serum samples, are reported. Simplex optimization has proven to be a very useful tool in the search for the optimal conditions for performing analyses with the optical fiber biosensor. |
| Free cholesterol induces activation but not translocation of protein kinase C in cultured ascites tumour cells Haeffner, E. W. and U. Wittmann (1994), Cell Signal 6(2): 201-7. Abstract: Ascites tumour cells, cultivated in a serum replacement medium with low mitogenic activity supplemented with free cholesterol or cholesterol complexed to digitonine, were used to study protein kinase C (PKC) activation and its translocation from the cytosol to the membrane compartment. Activity of partially purified PKC was assessed by the Ca2+/phospholipid-dependent phosphorylation of histone as substrate. Staurosporine-sensitive PKC activity was increased by a factor of about three in the membrane compartment as the result of cholesterol supplementation with about 90% of total activity in the membrane, and the remaining 10% in the cytosolic fraction. A translocation of enzyme mass from the cytosol to the membrane compartment was not induced under these conditions, as documented by the enzyme-linked immuno adsorbent assay using a monoclonal anti-PKC antibody. In cells cultivated in the presence of the cholesterol-digitonide complex PKC activation was not observed, suggesting an inadequate utilization of cholesterol as membrane constituent. |
| Free cholesterol loading of macrophages induces apoptosis involving the fas pathway Yao, P. M. and I. Tabas (2000), J Biol Chem 275(31): 23807-13. Abstract: Macrophage death is an important feature of atherosclerosis, but the cellular mechanism for this process is largely unknown. There is increasing interest in cellular free cholesterol (FC) excess as an inducer of lesional macrophage death because macrophages accumulate large amounts of FC in vivo, and FC loading of macrophages in culture causes cell death. In this study, a cell culture model was used to explore the cellular mechanisms involved in the initial stages of FC-induced macrophage death. After 9 h of FC loading, some of the macrophages exhibited externalization of phosphatidylserine and DNA fragmentation, indicative of an apoptotic mechanism. Incubation of the cells with Z-DEVD-fluoromethylketone blocked these events, indicating dependence upon effector caspases. Macrophages from mice with mutations in either Fas or Fas ligand (FasL) demonstrated substantial resistance to FC-induced apoptosis, and FC-induced death in wild-type macrophages was blocked by an anti-FasL antibody. FC loading had no effect on the expression of cell-surface Fas but caused a small yet reproducible increase in cell-surface FasL. To determine the physiological significance of this finding, unloaded and FC-loaded Fas-deficient macrophages, which can only present FasL, were compared for their ability to induce apoptosis in secondarily added Fas-bearing macrophages. The FC-loaded macrophages were much more potent inducers of apoptosis than the unloaded macrophages, and this effect was almost completely blocked by an inhibitory anti-FasL antibody. In summary, during the early stages of FC loading of macrophages, a fraction of cells exhibited biochemical changes that are indicative of apoptosis. An important part of this event is FC-induced activation of FasL that leads to Fas-mediated apoptosis. In light of recent in vivo findings that show that apoptotic macrophages in atherosclerotic lesions express both Fas and FasL, we present a cellular model of Fas-mediated death in lesional foam cells. |
| Free cholesterol loading of macrophages is associated with widespread mitochondrial dysfunction and activation of the mitochondrial apoptosis pathway Yao, P. M. and I. Tabas (2001), J Biol Chem 276(45): 42468-76. Abstract: Macrophage death in advanced atherosclerotic lesions leads to lesional necrosis and possibly plaque rupture and acute vascular occlusion. Among the likely causes of lesional macrophage death is intracellular accumulation of excess free cholesterol (FC), which is known to occur in vivo. We recently showed that FC loading of macrophages causes apoptosis, approximately 50% of which is mediated by activation of cell-surface FasL and triggering of the Fas pathway (Yao, P. M., and Tabas, I. (2000) J. Biol. Chem. 275, 23807-23813). To elucidate other pathways of death in FC-loaded macrophages, we investigated mitochondrial transmembrane potential (DeltaPsi(m)) and the mitochondrial apoptosis pathway in FC-loaded mouse peritoneal macrophages. Starting between 3 and 6 h of FC loading, DeltaPsi(m) was markedly decreased in the majority of macrophages and was independent of the Fas pathway. The decrease in DeltaPsi(m) by FC loading was not prevented by GSH, thus distinguishing it from 7-ketocholesterol-induced mitochondrial dysfunction. Cytochrome c release into the cytosol was noted by 4 h of FC loading, and activation of caspase-9 and effector caspases was observed at 6 h. Finally, we found that both cellular and mitochondrial levels of the pro-apoptotic protein Bax were increased severalfold as early as 4 h after FC loading. Thus, FC loading, perhaps via increased levels of Bax and/or cholesterol overloading of mitochondria, triggers cytochrome c release and activation of caspase-9 and the effector caspases, leading to macrophage apoptosis. These findings and our previous data support a model in which FC loading of macrophages promotes a dual program of caspase-mediated death. |
| Free cholesterol loading of macrophages stimulates phosphatidylcholine biosynthesis and up-regulation of CTP: phosphocholine cytidylyltransferase Shiratori, Y., A. K. Okwu, et al. (1994), J Biol Chem 269(15): 11337-48. Abstract: Atheroma macrophages accumulate large amounts of free cholesterol (FC) as well as cholesteryl ester (CE). An important adaptive response to FC loading might be increased cellular phospholipid to accommodate the excess FC. To explore this idea, J774 macrophages were incubated for 48 h without lipid, with acetyl-low density lipoprotein to induce mostly CE loading, or with acetyl-low density lipoprotein plus an acyl-CoA:cholesterol O-acyltransferase inhibitor (58035) to induce marked FC loading. The total phospholipid content approximately doubled in FC-loaded versus control or CE-loaded macrophages, with phosphatidylcholine showing the largest increase (approximately 2.5-fold versus control). Electron micrographs revealed the presence of multiple intracellular membrane whorls in the FC-loaded macrophages but not in the control or CE-loaded macrophages. 3HCholine incorporation into phosphatidylcholine was also greater in FC-loaded macrophages versus control or CE-loaded macrophages, whereas 3Hphosphatidylcholine degradation was similar in all of the macrophages. In these experiments and in others that used non-lipoprotein cholesterol, there was a very close correlation between cellular FC content and phosphatidylcholine biosynthesis. To determine the mechanism of increased phosphatidylcholine synthesis, FC-loaded and CE-loaded macrophages were pulsed with 3Hcholine, then chased and assayed for labeled phosphatidylcholine biosynthetic precursors. The only major differences were a 2-fold greater disappearance of label from 3Hcholine phosphate and a 5-fold greater appearance of label in CDP-3Hcholine in the FC-loaded macrophages. These data suggest a stimulation of CTP:phosphocholine cytidylyltransferase (CT), which was confirmed by microsomal CT assays. Further studies revealed that the increase in phosphatidylcholine biosynthesis in FC-loaded macrophages was: (a) reversible under conditions of high density lipoprotein3-mediated cellular cholesterol efflux; (b) not blocked by cycloheximide-induced protein synthesis inhibition; and (c) not associated with increased CT mRNA levels. Thus, FC loading of macrophages leads to an increase in phosphatidylcholine mass which is caused by increased phosphatidylcholine biosynthesis. The mechanism appears to be FC-mediated post-translational activation of CT. This adaptive response may be important for atheroma macrophage survival, and disruption of the response may lead to macrophage necrosis and lesion progression. |
| Free cholesterol transfer from human lower-density lipoproteins (d less than 1.063) to lipoprotein-deficient serum and high-density lipoproteins Velazquez, E., A. Montes, et al. (1990), Metabolism 39(12): 1263-6. Abstract: The in vitro transfer of free cholesterol (FC) between human serum lipoproteins in the absence of lecithin:cholesterol acyltransferase (LCAT) activity has been examined. The results show that the amount of FC that the high-density lipoprotein (HDL) and lipoprotein-deficient serum (LDS) fractions were able to capture from low-density lipoproteins (LDL) and very-low-density lipoproteins (VLDL) was proportional to the amount of FC present in d less than 1.063 lipoproteins. The presence of HDL increased this transfer markedly. These results indicate that, in the absence of LCAT activity, FC can transfer from lower-density lipoproteins to higher-density serum fractions, and this transfer might increase under hypercholesterolemic conditions. The possible importance of this phenomena in regard to the exchange of FC between serum lipoproteins and tissue cells is discussed. |
| Free cholesterol-loaded macrophages are an abundant source of tumor necrosis factor-alpha and interleukin-6: model of NF-kappaB- and map kinase-dependent inflammation in advanced atherosclerosis Li, Y., R. F. Schwabe, et al. (2005), J Biol Chem 280(23): 21763-72. Abstract: Two key features of atherosclerotic plaques that precipitate acute atherothrombotic vascular occlusion ("vulnerable plaques") are abundant inflammatory mediators and macrophages with excess unesterified, or "free," cholesterol (FC). Herein we show that FC accumulation in macrophages leads to the induction and secretion of two inflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6). The increases in TNF-alpha and IL-6 mRNA and protein were mediated by FC-induced activation of the IkappaB kinase/NF-kappaB pathway as well as activation of MKK3/p38, Erk1/2, and JNK1/2 mitogen-activated protein kinases (MAPK). Activation of IkappaB kinase and JNK1/2 was needed for the induction of both cytokines. However, MKK3/p38 signaling was specifically involved in TNF-alpha induction, and Erk1/2 signaling was required for IL-6. Most interestingly, activation of all of the signaling pathways and induction of both cytokines required cholesterol trafficking to the endoplasmic reticulum (ER). The CHOP branch of the unfolded protein response, an ER stress pathway, was required for Erk1/2 activation and IL-6 induction. In contrast, one or more other ER-related pathways were responsible for activation of p38, JNK1/2, and IkappaB kinase/NF-kappaB and for the induction of TNF-alpha. These data suggest a novel scenario in which cytokines are induced in macrophages by endogenous cellular events triggered by excess ER cholesterol rather than by exogenous immune cell mediators. Moreover, this model may help explain the relationship between FC accumulation and inflammation in vulnerable plaques. |
| Free fatty acids modulate intermembrane trafficking of cholesterol by increasing lipid mobilities: novel 13C NMR analyses of free cholesterol partitioning Johnson, R. A., J. A. Hamilton, et al. (2003), Biochemistry 42(6): 1637-45. Abstract: Cholesterol and free fatty acids in membranes modulate major biological processes, and their cellular metabolism and actions are often coordinately regulated. However, effects of free fatty acid on cholesterol-membrane interactions have proven difficult to monitor in real time in intact systems. We developed a novel (13)C NMR method to assess effects of free fatty acids on molecular interactions of cholesterol within--and transfer between--model membranes. An important advantage of this method is the ability to acquire kinetic data without separation of donor and acceptor membranes. Large unilamellar phospholipid vesicles (LUV) with phosphatidylcholine/cholesterol ratios of 4:1 served as cholesterol donors. Small unilamellar vesicles (SUV) made with phosphatidylcholine were acceptors. The (13)C(4)-cholesterol peak is narrow in SUV, but very broad in LUV, spectra; the increase in intensity of this peak over time monitored transfer. Oleic acid and other long chain free fatty acids saturated (C12-18) and unsaturated (C18) dose-dependently increased mobilities of lipids in LUV (phospholipid and cholesterol) and cholesterol transfer rates, whereas short (C8-10) and very long (C24) chain free fatty acids did not. Decreasing pH from 7.4 to 6.5 (+/-oleic acid) had no effect on cholesterol transfer, and 5 mol % fatty acyl-CoAs increased transfer rates, demonstrating greater importance of the fatty-acyl tail over the headgroup. In LUV containing sphingomyelin, transfer rates decreased, but the presence of oleic acid increased transfer 1.3-fold. These results demonstrate free fatty acid-facilitated cholesterol movement within and between membranes, which may contribute to their multiple biological effects. |
| Free phytosterols effectively reduce plasma and liver cholesterol in gerbils fed cholesterol Hayes, K. C., A. Pronczuk, et al. (2002), J Nutr 132(7): 1983-8. Abstract: The potential of free phytosterols (including 20% stanols) to lower plasma and liver lipids was assessed in three experiments with gerbils fed diets containing cholesterol. The first explored the ability of phytosterols (0.5%) to block absorption of 0.05, 0.10, and 0.5% cholesterol. The second assessed the importance of consuming phytosterols (0.75%) simultaneously with cholesterol (0.15%). The third compared free phytosterols (0.75%) with similar levels of esterified sterols or stanols using diets containing 0.15% cholesterol. A 5:1 ratio of phytosterols:cholesterol effectively blocked cholesterol absorption when the dietary cholesterol load was moderate. Consuming a 5:1 ratio with every meal was more effective than receiving equal phytosterols in a 10:1 ratio every other day. Finally, free phytosterols dissolved in fat were as effective as esterified sterols and stanols in lowering plasma and liver cholesterol, and all were equally effective at blocking cholesterol absorption as shown by increased fecal cholesterol output. Plant sterol accumulation in the liver was minimal for all test groups. |
| Free phytosterols facilitate excretion of endogenous cholesterol in gerbils Hayes, K. C., A. Pronczuk, et al. (2005), J Nutr Biochem 16(5): 305-11. Abstract: To determine whether phytosterols (PST) facilitate excretion of whole body cholesterol and whether dietary fat or enhancing gallbladder contraction with curcumin might influence this process, four experiments were conducted in gerbils. In Experiment 1, naive gerbils received cholesterol-free purified diets with 30% energy from fat and 0% or 0.75% free PST from tall oil for 4 weeks. In Experiment 2, body cholesterol pools were expanded by feeding a diet containing 0.3% cholesterol for 3 weeks. Subsequently, PST was provided in either fat-free or normal-fat diets without cholesterol for only 2 h each morning, followed by a low-fat diet for the rest of the day and food restriction overnight. In Experiment 3, gerbils were preloaded with cholesterol, followed by either PST alone or PST+curcumin to enhance gallbladder contraction. In Experiment 4, curcumin or curcumin+PST were fed with 30% as fat and 0.15% cholesterol throughout the study. Because of the small whole body cholesterol pool in Experiment 1, the impact of PST was limited. When whole body cholesterol was expanded in Experiments 2 and 3, subsequent reductions of liver esterified cholesterol by PST were significant. In the presence of dietary fat, PST caused a greater reduction (23%) than in a fat-free diet (8%) compared to respective controls. Curcumin (Experiments 3 and 4) proved ineffective in reducing liver or plasma cholesterol pools, and the 3:1 ratio between PST/diet cholesterol was less effective at blocking cholesterol absorption than a 5:1 ratio previously employed. Thus, free PST removed whole body cholesterol, which was enhanced by concomitant fat intake, but was unaffected by a gallbladder contracting agent. |
| Free radical modification of lipoproteins and cholesterol accumulation in cells upon atherosclerosis Panasenko, O. M., T. V. Vol'nova, et al. (1991), Free Radic Biol Med 10(2): 137-48. Abstract: An electron spin probe study was made of the effect of lipid peroxidation (LPO) on the structure of surface proteolipid layer of human serum low-density lipoproteins (LDL). The results obtained with a positively charged spin label and stearic acid spin probes with doxyl labels at positions 5, 12, and 16 revealed that LPO caused a decrease in phospholipid molecule mobility both in the region of polar heads and in the region of acyl chains till the depth of at least 1.7 mm from water-lipid interface. Under relatively high levels of oxidation (more than 6 mumol MDA/g LDL phospholipid) the polarity of lipid phase increased. The decrease in efficiency of tryptophan fluorescence quenching by nitroxide fragments incorporated in hydrophobic regions at the depth of approximately 2 nm from water-lipid interface indicated that lipid-protein interaction was disturbed as a result of oxidation of LDL lipids. In addition, the LPO-induced modification of apo-B, the main protein of LDL, was examined with maleimide spin label. LPO led to increase in mobility of strongly immobilized maleimide labels and in the number of weakly immobilized ones. Oxidized LDL revealed decreased ability to incorporate spin-labeled steroid (androstane) as compared to native ones. LPO-induced structural changes of LDL surface are supposed to be a reason of enhanced accumulation of cholesterol in human monocytes during their incubation with oxidized LDL. The cholesterol content in red cells was shown to be directly correlated to MDA content in apo-B containing lipoproteins but not in whole serum. Our findings suggest that free radical modification of serum lipoproteins but not solely an increased level of LPO products in blood is one important cause for cholesterol accumulation in cells and, apparently, for their transformation into foam cells during atherosclerosis. |
| Freeze-fracture cytochemistry of membrane cholesterol in Blastocystis hominis Yoshikawa, H. and A. Hayakawa (1996), Int J Parasitol 26(10): 1111-4. Abstract: Membrane cholesterol in Blastocystis hominis was detected by freeze-fracture methods using a polyene antibiotic, filipin. Since the intramembrane particles (IMP) were distributed heterogeneously on both plasma and vacuole membranes, many IMP-free areas were observed. Even in filipin-treated cells, filipin-cholesterol complexes were not detected in IMP-free areas on the plasma membrane, whereas on the central vacuole membrane the complexes were mainly observed in IMP-free regions. These results indicate the different organization of the membrane cholesterol between the plasma membrane and central vacuole membrane. Most of the granules in the central vacuole were densely labeled with filipin, indicating the accumulation of cholesterol in the vacuole. |
| Frequency distribution of serum cholesterol levels in patients with panic disorder: comparison with normal controls Shioiri, T., K. Fujii, et al. (1998), Psychiatry Clin Neurosci 52(6): 601-4. Abstract: We compared the frequency distribution of total cholesterol (TC) levels in 103 patients with panic disorder (PD) with that in 173 gender- and age-matched normal controls (NC). There was no significant difference in the mean TC level between the PD and the NC groups. The distribution of TC levels in the PD group was similar to that in the NC group. As a whole distribution pattern, there is no association between high serum cholesterol levels and panic disorder. However, four male PD patients had very high TC levels of more than 260 mg/dL, and two of them had obviously deviated values from the frequency distribution of TC levels in the NC group. Our findings are supportive of the view that male PD patients with high TC levels have excess mortality due to cardiovascular diseases. |
| Frequency of citation and outcome of cholesterol lowering trials Anderson, J. (1992), Bmj 305(6850): 421-2. |
| Frequency of citation and outcome of cholesterol lowering trials Burr, M. L., A. M. Fehily, et al. (1992), Bmj 305(6850): 422. |
| Frequency of citation and outcome of cholesterol lowering trials Durrington, P. N., M. F. Laker, et al. (1992), Bmj 305(6850): 420-1; author reply 422. |
| Frequency of citation and outcome of cholesterol lowering trials Game, F. L. and R. H. Neary (1992), Bmj 305(6850): 421; author reply 422. |
| Frequency of citation and outcome of cholesterol lowering trials Goodwin, J. F. (1992), Bmj 305(6850): 421; author reply 422. |
| Frequency of citation and outcome of cholesterol lowering trials Ravnskov, U. (1992), Bmj 305(6855): 717. |
| Frequency of citation and outcome of cholesterol lowering trials Thompson, G. R. (1992), Bmj 305(6850): 422. |