Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Ketocholesterols”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed2005

7-Ketocholesterol favors lipid accumulation and colocalizes with Nile Red positive cytoplasmic structures formed during 7-ketocholesterol-induced apoptosis: analysis by flow cytometry, FRET biphoton spectral imaging microscopy, and subcellular fractionation.

BACKGROUND: Oxidized low-density lipoproteins play key roles in atherosclerosis. Their toxicity is at least in part due to 7-ketocholesterol (7KC), which is a potent inducer of apoptosis. In this study on human promonocytic U937 cells, we determined the effects and the interactions of 7KC with cellular lipids during 7KC-induced apoptosis. METHODS: Morphologic and functional changes were investigated by microscopic and flow cytometric methods after staining with propidium iodide, 3,3'-dihexyloxacarbocyanine iodide, and Hoechst 33342. Cellular lipid content was identified by using filipin to quantify free cholesterol and Nile Red (NR), which emit a yellow or orange-red fluorescence in the presence of neutral and polar lipids, respectively. After staining with NR, interactions of 7KC with cellular lipids were identified by fluorescence resonance energy transfer biphoton spectral imaging confocal microscopy and by subcellular fractionation, gas chromatography, and mass spectrometry. RESULTS: During 7KC-induced apoptosis the fluorescence from filipin and the ratio of measured (orange-red vs. yellow) fluorescence of NR were enhanced. Spectral analysis of images obtained in biphoton mode and resulting factor images demonstrated the occurrence of fluorescence resonance energy transfer between 7KC and NR and the subsequent colocalization of 7KC and NR. These data were in agreement with biochemical characterization and demonstrated that 7KC and neutral and polar lipids accumulate in NR-stained cytoplasmic structures. CONCLUSIONS: During 7KC-induced apoptosis, 7KC modifies the cellular content of neutral and polar lipids, favors free cholesterol accumulation, and colocalizes with neutral and polar lipids that are inside NR-stained cytoplasmic structures.

Apoptosis↗
PubMed2004

FRET multiphoton spectral imaging microscopy of 7-ketocholesterol and Nile Red in U937 monocytic cells loaded with 7-ketocholesterol.

OBJECTIVE: To show the effect of 7-ketocholesterol (7KC) on cellular lipid content by means of flow cytometry and the interaction of 7KC with Nile Red (NR) via ultraviolet fluorescence resonance energy transfer (FRET) excitation of NR on U937 monocytic cells by means of 2-photon excitation confocal laser scanning microscopy (CLSM). STUDY DESIGN: Untreated and 7KC-treated U937 cells were stained with NR and analyzed by flow cytometry and CLSM. 3D sequences of images were obtained by spectral analysis in a 2-photon excitation CLSM and analyzed by the factor analysis of medical image sequences (FAMIS) algorithm, which provides factor curves and images. Factor images are the result of the FAMIS image processing method, which handles emission spectra. In FRET analysis, preparations are screened at selected UV wavelengths to avoid emission of NR in the absence of 7KC. RESULTS: During 7KC-induced cell death,flow cytometry and CLSM revealed a modification of the cellular lipid content. Factor images show FRET occurrence and subsequent colocalization of 7KC and NR. CONCLUSION: This investigation established the utility of 2-photon excitation CLSM to assess colocalization of 7KC with NR by FRET and to identify and distinguish polar and neutral lipids stained by NR that accumulate from the effect of 7KC.

Cell Death↗
PubMed2005

The polar nature of 7-ketocholesterol determines its location within membrane domains and the kinetics of membrane microsolubilization by apolipoprotein A-I.

7-Ketocholesterol is an oxidized derivative of cholesterol with numerous physiological effects. In model membranes, 7-ketocholesterol and cholesterol were compared by physical measures of bilayer order and polarity, formation of detergent resistant domains (DRM), phase separation, and membrane microsolubilization by apolipoprotein A-I. In binary mixtures of a saturated phosphatidylcholine (PC), dipalmitoyl-PC (DPPC), and cholesterol or 7-ketocholesterol, the sterols modulate bilayer order and polarity and induce DRM formation to a similar extent. Cholesterol induces formation of ordered lipid domains (rafts) in tertiary mixtures with dioleoyl-PC (DOPC) and DPPC, or DOPC and sphingomyelin (SM). In tertiary mixtures, cholesterol increased lipid order and reduces bilayer polarity more than 7-ketocholesterol. This effect was more pronounced when the mixtures were in a miscible liquid-disordered (L(d)) phase. Substitution of 7-ketocholesterol for cholesterol dramatically reduced the extent of DRM formation in DOPC/DPPC and DOPC/SM bilayers and ordered lipid phase separation in mixtures of a spin-labeled PC with DPPC and with SM. Compared to cholesterol, 7-ketocholesterol decreased the rate for the microsolubilization of dimyristoyl-PC multilamellar vesicles by apolipoprotein A-I. The membrane effects of 7-ketocholesterol were dependent on the phospholipid matrix. In L(d) phase phospholipids, a model for 7-ketocholesterol indicates that the proximity of the 7-keto and 3beta-OH groups puts both polar moieties at the lipid-water interface to tilt the sterol nucleus to the plane of the bilayer. 7-Ketocholesterol was less effective in forming ordered lipid domains, in decreasing the level of bilayer hydration, and in forming phase boundary bilayer defects. Compared to cholesterol, 7-ketocholesterol can differentially modulate membrane properties involved in protein-membrane association and function.

1,2-Dipalmitoylphosphatidylcholine↗
PubMedDate not supplied

Impact of baseline level of 7-ketocholesterol on the rate of coronary reintervention in coronary artery disease patients--results of an observational study with questionnaire-based follow up.

INTRODUCTION: The aim of study was to assess the relation between plasma level of one of the oxysterols--7-ketocholesterol and angiographically evidenced coronary artery disease. MATERIAL AND METHODS: We measured plasma level of 7-ketocholesterol, malondialdehyde, total cholesterol, HDL cholesterol, triglycerides, erythrocyte activity of superoxide dismutase in 233 patients who previously underwent coronary angiography for diagnosis of coronary artery disease. The follow-up of patients (28 months) was done on basis of questionnaires sent out to them. RESULTS: We found that the subgroup of patients with 7-ketocholesterol above the median value (62 ng/mL) has higher rate of coronary reintervention (8/54 pts vs. 5/43 pts in 7-ketocholesterol below 62 ng/mL respectively, p = 0.284 Log-rank test). The finding was more pronounced, yet insignificant, in the subgroup of patients who underwent primary PTCA (6/19 pts vs. 2/20 pts, p = 0.1 Log-rank test). There were 3 patients in high 7-ketocholesterol subgroup who entered the study with previously undergone primary coronary intervention. There were not such patients in low 7-ketocholesterol subgroup. 4 deaths were observed in high 7-ketocholesterol subgroup, and one in low 7-ketocholesterol subgroup (they were predominantly not of cardiovascular origin). CONCLUSIONS: We failed to find a significant relation between 7-ketocholesterol and coronary artery disease, though there is a trend of such relation in patients who undergo PTCA, evidenced by a slightly higher number of repeated revascularizations among patients with high 7-ketocholesterol. The study indicates the areas of further studies with agents capable of lowering oxysterols and assessing also other end points.

Aged↗
PubMed2002

The impairment of endothelium-dependent arterial relaxation by 7-ketocholesterol is associated with an early activation of protein kinase C.

Among components of oxidized low density lipoproteins, cholesterol derivatives oxidized in position 7 inhibit endothelium-dependent arterial relaxation by decreasing the release of the main endothelium-derived relaxing factor, nitric oxide (NO). The aim of the present study was to bring new insights into the molecular mechanism by which 7-ketocholesterol can block the endothelium-dependent arterial relaxation. Superoxide dismutase did not prevent the inhibitory effect of 7-ketocholesterol on endothelium-dependent relaxation, and consistent observations were made whether superoxide dismutase was conjugated or not to polyethylene glycol. In addition, neither glutathione supplementation, nor oxypurinol, i.e. a xanthine oxidase inhibitor could reverse the effect of 7-ketocholesterol, indicating that NO was not inactivated by superoxide anion. A direct alteration of the activity of the calcium-dependent NO synthase could also be ruled out, since identical relaxing effects of the calcium ionophore A23187 were observed whether arterial rings were treated or not with 7-ketocholesterol. 4 Whereas the above observations come in support of an early, inhibitory action of 7-ketocholesterol, the specific blockade of one given subtype of membrane receptors could be discarded, and similar inhibitions were observed when either muscarinic or purinergic receptors were stimulated. Finally, the blockade of protein kinase C activity by chelerythrine arose as the sole relevant tool in preventing the effect of 7-ketocholesterol on the endothelium-dependent relaxation of rabbit aortic rings. In addition, complementary studies on cultured bovine aortic endothelial cells came in direct support of the ability of 7-ketocholesterol to activate PKC. In conclusion, 7-ketocholesterol that is present in human hypercholesterolaemic plasma, in atherosclerotic arteries, and in many processed foods can block the release of NO by vascular endothelial cells through its ability to activate PKC.

Animals↗
PubMed2006

Prevention of 7-ketocholesterol-induced mitochondrial damage and cell death by calmodulin inhibition.

Oxysterols such as 7-ketocholesterol and 25-hydroxycholesterol formed under enhanced oxidative stress in the brain are suggested to induce neuronal cell death. The present study investigated the effect of calmodulin antagonists (trifluoperazine, W-7 and calmidazolium) against the cytotoxicity of 7-ketocholesterol in relation to the mitochondria-mediated cell death process and oxidative stress. PC12 cells exposed to 7-ketocholesterol revealed nuclear damage, decrease in the mitochondrial transmembrane potential, cytosolic accumulation of cytochrome c, activation of caspase-3, increase in the formation of reactive oxygen species and depletion of GSH. N-Acetylcysteine, trolox, carboxy-PTIO and Mn-TBAP reduced the cytotoxic effect of 7-ketocholesterol. Calmodulin antagonists attenuated the 7-ketocholesterol-induced nuclear damage, formation of the mitochondrial permeability transition and cell viability loss in PC12 cells. The results suggest that calmodulin antagonists may prevent the 7-ketocholesterol-induced viability loss in PC12 cells by suppressing formation of the mitochondrial permeability transition, leading to the release of cytochrome c and subsequent activation of caspase-3. The effects seem to be ascribed to their depressant action on the formation of reactive oxygen species and depletion of GSH. The findings suggest that calmodulin inhibition may exhibit a protective effect against the neurotoxicity of 7-ketocholesterol.

Animals↗
PubMed2002

Analysis of oxidative processes and of myelin figures formation before and after the loss of mitochondrial transmembrane potential during 7beta-hydroxycholesterol and 7-ketocholesterol-induced apoptosis: comparison with various pro-apoptotic chemicals.

Among oxysterols oxidized at C7 (7alpha-, 7beta-hydroxycholesterol, and 7-ketocholesterol) 7beta-hydroxycholesterol and 7-ketocholesterol are potent inducers of cell death and probably play central roles in atherosclerosis. As suggested by our previous investigations, 7-ketocholesterol might be a causative agent of vascular damage by inducing apoptosis and enhancing superoxide anion (O2*-) production. To determine the precise relationships between cytotoxicity and oxidative stress, the ability of oxysterols oxidized at C7 to induce apoptosis, to stimulate O2*- production and to promote lipid peroxidation was compared with different pro-apoptotic chemicals: antitumoral drugs (VB, Ara-C, CHX, and VP-16) and STS. All compounds, except 7alpha-hydroxycholesterol, induced apoptosis characterized by the occurrence of cells with fragmented and/or condensed nuclei, loss of mitochondrial potential, caspase-3 activation, PARP degradation, and internucleosomal DNA fragmentation. The highest proportion of apoptotic cells was found with antitumoral drugs and STS, whereas the highest overproduction of O2*- detected before and after the loss of mitochondrial potential was obtained with 7beta-hydroxycholesterol and 7-ketocholesterol. Overproduction of O2*- was always correlated with enhanced lipid peroxidation. Vit E was only capable to significantly counteract apoptosis and oxidative stress induced by 7beta-hydroxycholesterol, 7-ketocholesterol, VB and STS. By electron and fluorescence microscopy, myelin figures evocating autophagic vacuoles were barely observed under treatment with 7beta-hydroxycholesterol and 7-ketocholesterol, and their formation occurring before the loss of mitochondrial potential was reduced by Vit E. In the presence of 7alpha-hydroxycholesterol, no enhancement of O2*- production, no lipid peroxidation, and no formation of myelin figures were observed. Collectively, our data demonstrate, that there can be a more or less important stimulation of oxidative stress during apoptosis. They also suggest that enhancement of O2*- production associated with lipid peroxidation during 7beta-hydroxycholesterol and 7-ketocholesterol-induced apoptosis could contribute to in vivo vascular injury, and that myelin figures could constitute suitable markers of oxysterol-induced cell death.

Analysis of Variance↗
PubMedDate not supplied

7-Ketocholesterol.

7-Ketocholesterol is a major oxidation product of cholesterol found in human atherosclerotic plaque and is more atherogenic than cholesterol in some animal studies. 7-Ketocholesterol can inhibit cholesterol 7 alpha-hydroxylase, the rate-limiting step in bile acid biosynthesis, as well as strongly inhibiting HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. It has even been suggested that 7-ketocholesterol is formed enzymically as an endogenous regulator of cholesterol biosynthesis. However, when tested as a pharmacological cholesterol-lowering agent, inhibition of HMG-CoA reductase was rapidly overcome and the 7-ketocholesterol metabolised. In vitro, 7-ketocholesterol has wide-ranging and potent effects, most of which have the potential to contribute to atherosclerosis. For example, 7-ketocholesterol can be cytotoxic and can induce apoptosis in vascular cells. These effects, either individually or more likely, in combination, all implicate 7-ketocholesterol in the initiation and development of atherosclerosis, but further work is needed to establish whether or not its role is a direct causal one.

Humans↗
PubMed1996

Hydroxypropyl-beta-cyclodextrin-mediated efflux of 7-ketocholesterol from macrophage foam cells.

Atherosclerosis involves the arterial accumulation of lipid-laden "foam cells" containing oxidized and unoxidized sterols and their esters (Mattsson-Hulten, L., Lindmark, H., Diczfalusy, U., Bjorkhem, I., Ottosson, M., Liu, Y., Bondjers, G., and Wiklund, O. (1996) J. Clin. Invest. 97, 461-8). Oxidized sterols are probably critical to atherogenesis because they inhibit cholesterol removal from cells and are cytotoxic. We recently reported that there is deficient induction of cellular cholesterol efflux by apolipoprotein A-I, the main initial acceptor of cellular cholesterol from macrophages loaded in vitro with oxidized low density lipoprotein (Kritharides, L., Jessup, W., Mander, E., and Dean, R. T. (1995) Arterioscler. Thromb. 15, 276-289). There was an even more marked impairment of the release of 7-ketocholesterol which is a major oxysterol in these cells and in human atherosclerotic lesions. Here we show that hydroxypropyl-beta-cyclodextrin can induce selective efflux of 7-ketocholesterol. Efflux of 7-ketocholesterol was time and concentration dependent, and the rate of its removal was 50-fold greater for hydroxypropyl-beta-cyclodextrin than for apolipoprotein A-I. Over a defined range of concentrations (0-5 mg/ml), efflux of 7-ketocholesterol was preferred over that of cholesterol and occurred without cell toxicity. Efflux of free 7-ketocholesterol was associated with decreased intracellular free and esterified 7-ketocholesterol. Hydroxypropyl-beta-cyclodextrin also enhanced efflux of other oxysterols. The physical solubilization of 7-ketocholesterol by the cyclodextrin was much greater than that of cholesterol, in accordance with its differential effects on efflux. These data highlight the importance of extracellular sterol solubilization in the efflux of cellular oxysterols and the mobilization of intracellular free and esterified oxysterol pools in macrophages loaded with oxidized low density lipoprotein. Synthetic sterol-solubilizing agents such as hydroxypropyl-beta-cyclodextrin are thus potential prototypes for the further development of oxysterol-removing agents.

2-Hydroxypropyl-beta-cyclodextrin↗
PubMed1981

Angiotoxic effects of dietary 7-ketocholesterol in chick aorta.

Although dietary 1-ketocholesterol treatment had little effect on plasma and liver lipid metabolism, aortas from chickens treated with 7-ketocholesterol contained numerous dead and dying smooth muscle cells. Chicks receiving dietary cholesterol, in amounts equal to or greater than the amount of 7-ketocholesterol, did not demonstrate significant smooth muscle cell degeneration. After eight weeks, a combined diet of 1% cholesterol plus 10-20 mg/day 1-ketocholesterol produced no larger amounts of aortic lipid deposition, and no greater increases in smooth muscle cell degeneration than a comparable group receiving 7-ketocholesterol alone. The abdominal aorta was always more severely damaged than the ascending arch or thoracic aorta in chicks fed 7-ketocholesterol. The results demonstrate a potent angiotoxic effect of 7-ketocholesterol.

Animals↗
PubMed1997

Induction of similar features of apoptosis in human and bovine vascular endothelial cells treated by 7-ketocholesterol.

Cholesterol oxides have numerous cytotoxic effects and those oxidized in the C7 position have been shown to induce apoptosis in bovine aortic endothelial cells (BAEC). The aim of the present study was to determine whether apoptosis also occurs in human vascular endothelial cells (HUVEC) treated with 7-ketocholesterol. To this end, cultured BAEC and HUVEC were incubated for 48 h with 7-ketocholesterol (concentration range 5-80 micrograms/ml) and the characteristics of cell death were assessed by various methods: counting of adherent and non-adherent cells; analysis of DNA fragmentation pattern; and morphological study by light, fluorescence, and electron microscopy. The 7-ketocholesterol treatment was accompanied by a decrease in the number of adherent cells and an increase in the number of non-adherent cells. Apoptotic cells, recognized by fragmented and/or condensed nuclei after staining with Hoechst 33342 or Giemsa, were mainly detected among non-adherent cells, and agarose gel electrophoresis revealed a typical internucleosomal DNA fragmentation among 7-ketocholesterol-treated cells. The DNA fragmentation was no longer detected when HUVEC and BAEC were simultaneously incubated with 0.5 mmol/l zinc chloride, which is known to inhibit Ca2+/Mg(2+)-dependent endonucleases. Finally, the ultrastructural abnormalities observed by electron microscopy in both 7-ketocholesterol-treated HUVEC and BAEC were remarkably similar and were mainly characterized by condensed chromatin, altered mitochondria, disturbed organization of the cytoskeleton, and vacuoles containing myelin figures and/or cell debris; apoptotic bodies were also frequently detected. It is concluded that 7-ketocholesterol constitutes a potent inducer of apoptosis in endothelial vascular cells of both bovine and human origin, suggesting that cholesterol oxides may be involved in the early steps of the atherosclerotic process in humans.

Animals↗
PubMed2006

7-Ketocholesterol enhances 1-methyl-4-phenylpyridinium-induced mitochondrial dysfunction and cell death in PC12 cells.

The present study investigated the promoting effect of oxysterol 7-ketocholesterol against the cytotoxicity of 1-methyl-4-phenylpyridinium (MPP(+)) in differentiated PC12 cells. 7-Ketocholesterol significantly enhanced the MPP(+)-induced nuclear damage, decrease in the mitochondrial transmembrane potential, cytosolic accumulation of cytochrome c, activation of caspase-3, increase in the formation of reactive oxygen species and depletion of GSH. N-Acetylcysteine, ascorbate, trolox, carboxy-PTIO and Mn-TBAP reduced the cytotoxic effect of MPP(+) in the presence of 7-ketocholesterol. The results indicate that 7-ketocholesterol shows a synergistic effect against the cytotoxic effect of MPP(+). 7-Ketocholesterol may enhance the MPP(+)-induced viability loss in PC12 cells by promoting the mitochondrial membrane permeability change, release of cytochrome c and subsequent activation of caspase-3, which is associated with the increased formation of reactive oxygen species and depletion of GSH. The findings suggest that 7-ketocholesterol as a promoting agent for the formation of mitochondrial permeability transition may enhance the toxic neuronal cell injury.

Animals↗
PubMed2006

Differential modulation of 7-ketocholesterol toxicity against PC12 cells by calmodulin antagonists and Ca2+ channel blockers.

The present study assessed the influence of intracellular Ca2+ and calmodulin against the neurotoxicity of oxysterol 7-ketocholesterol in relation to the mitochondria-mediated cell death process and oxidative stress in PC12 cells. Calmodulin antagonists calmidazolium and W-7 prevented the 7-ketocholesterol-induced mitochondrial damage, leading to caspase-3 activation and cell death, whereas Ca2+ channel blocker nicardipine, mitochondrial Ca2+ uptake inhibitor ruthenium red, and cell permeable Ca2+ chelator BAPTA-AM did not reduce it. Exposure of PC12 cells to 7-ketocholesterol caused elevation of intracellular Ca2+ levels. Unlike cell injury, calmodulin antagonists, nicardipine, and BAPTA-AM prevented the 7-ketocholesterol-induced elevations of intracellular Ca2+ levels. The results show that the cytotoxicity of 7-ketocholesterol seems to be modulated by calmodulin rather than changes in intracellular Ca2+ levels. Calmodulin antagonists may prevent the cytotoxicity of 7-ketocholesterol by suppressing the mitochondrial permeability transition formation, which is associated with the increased formation of reactive oxygen species and the depletion of GSH.

Animals↗
PubMed1994

The effect of dietary 7-ketocholesterol, inhibitor of sterol synthesis, on hepatic microsomal cholesterol 7 alpha-hydroxylase activity in rat.

A group of oxygenated sterols has been identified as physiological regulators of hepatic cholesterol biosynthesis. However, the regulatory effects of these oxysterols on cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme in bile acid biosynthesis, is not clearly elucidated. We administered 0.1% 7-ketocholesterol (15 mg/day), a strong inhibitor of sterol synthesis, to rats orally for 6 days. Then, the levels of accumulated oxysterols in liver microsomes and microsomal 7 alpha-hydroxylase activity were determined. The results were compared to those in the groups of rats treated with either control diet or diets containing 0.1 or 1% cholesterol, 0.1% butylated hydroxytoluene, 3% cholestyramine or 1% taurocholate. 7-Ketocholesterol feeding resulted in significant increase of both 7-ketocholesterol and 7 beta-hydroxycholesterol in microsomal fraction (449.4 +/- 36.8 and 438.2 +/- 46.8 ng/mg protein, respectively; mean +/- S.E.). Hepatic microsomal 7 alpha-hydroxylase activity in the rats fed 7-ketocholesterol was significantly elevated as compared with those of control rats; 44.70 +/- 5.97 vs. 16.57 +/- 2.46 pmol/min per mg protein. Addition of BHT to 7-ketocholesterol reduced the accumulation of 7 beta-hydroxycholesterol, and the stimulatory effect of 7-ketocholesterol on 7 alpha-hydroxylase activity was suppressed. Our results demonstrate that oxysterols do not inhibit but rather stimulate hepatic microsomal 7 alpha-hydroxylase.

Animals↗
PubMed2001

Ceramide generation occurring during 7beta-hydroxycholesterol- and 7-ketocholesterol-induced apoptosis is caspase independent and is not required to trigger cell death.

Biological activities of oxysterols seem tightly regulated. Therefore, the ability to induce cell death of structurally related oxysterols, such as those oxidized at C7(7alpha-, 7beta-hydroxycholesterol, and 7-ketocholesterol), was investigated on U937 cells at different times of treatment in a concentration range of 5-80 microg/ml. Whereas all oxysterols accumulate inside the cells, strong inhibition of cell growth and increased permeability to propidium iodide were observed only with 7beta-hydroxycholesterol and 7-ketocholesterol, which trigger an apoptotic process characterized by the occurrence of cells with fragmented and/or condensed nuclei, and by various cellular dysfunctions: loss of mitochondrial transmembrane potential, cytosolic release of cytochrome c, activation of caspase-9 and -3 with subsequent enhanced activity of caspase-3, degradation of poly(ADP-ribose) polymerase, and increased accumulation of cellular C16 : 0 and C24 : 1 ceramide species. This ceramide generation is not attributed to caspase activation since inhibition of 7beta-hydroxycholesterol- and 7-ketocholesterol-induced apoptosis by Z-VAD-fmk (100 microM), a broad spectrum caspase inhibitor, did not reduce C16 : 0 and C24 : 1 ceramide species accumulation. Conversely, when U937 cells were treated with 7beta-hydroxycholesterol and 7-ketocholesterol in the presence of fumonisin B1 (100 microM), a specific inhibitor of ceramide synthase, C16 : 0 and C24 : 1 ceramide species production was completely abrogated whereas apoptosis was not prevented. Noteworthy, 7alpha-hydroxycholesterol induced only a slight inhibition of cell growth. Collectively, these results are consistent with the notion that the alpha or beta hydroxyl radical position of oxysterols oxidized at C7 plays a key role in the induction of the apoptotic process. In addition, our findings demonstrate that 7beta-hydroxycholesterol- and 7-ketocholesterol-induced apoptosis involve the mitochondrial signal transduction pathway and they suggest that C16 : 0 and C24 : 1 ceramide species generated through ceramide synthase play a minor role in the commitment of 7beta-hydroxycholesterol- and 7-ketocholesterol-induced cell death.

Amino Acid Chloromethyl Ketones↗
PubMed2004

Oxysterol mixtures prevent proapoptotic effects of 7-ketocholesterol in macrophages: implications for proatherogenic gene modulation.

Oxysterols are common components of oxidized low-density lipoprotein and accumulate in the core of fibrotic plaques as a mixture of cholesterol and cholesteryl ester oxidation products. The proapoptotic effects of a biologically representative mixture of oxysterols was compared with equimolar amounts of 7-ketocholesterol and unoxidized cholesterol. The oxysterol mixture in a concentration range actually detectable in hypercholesterolemic patients did not stimulate programmed cell death in cultivated murine macrophages. Unoxidized cholesterol also produced no effect. By contrast, when given alone, 7-ketocholesterol strongly stimulated the mitochondrial pathway of apoptosis with cytochrome c release, caspase-9 activation, and eventually caspase-3 activation. Subsequent experiments showed that when 7-ketocholesterol was administered to cells together with another oxysterol, namely 7betaOH-cholesterol, the strong proapoptotic effect of 7-ketocholesterol was markedly attenuated. As regards the mechanism underlying this quenching, we found that the combined oxysterol treatment counteracted the ability of 7-ketocholesterol, when administered alone, to strongly up-regulate the steady-state levels of reactive oxygen species (ROS) without interfering with sterol uptake. Furthermore, this increase in intracellular ROS appeared to be responsible for the up-regulation of proapoptotic factor, p21, after treatment with 7-ketocholesterol but not in cells challenged with the oxysterol mixture. Competition among oxysterols, apparently at the level of NADPH oxidase, diminishes the ROS induction and direct toxicity that is evoked by specific oxysterols. As a consequence, a more subtle gene modulation by oxysterols becomes facilitated in vascular cells.

Animals↗
PubMed2005

7-Ketocholesterol-induced apoptosis. Involvement of several pro-apoptotic but also anti-apoptotic calcium-dependent transduction pathways.

Oxysterols, and particularly 7-ketocholesterol, appear to be strongly involved in the physiopathology of atherosclerosis. These molecules are suspected to be cytotoxic to the cells of the vascular wall and monocytes/macrophages, particularly by inducing apoptosis. Previous studies have demonstrated that 7-ketocholesterol-induced apoptosis is triggered by a sustained increase of cytosolic-free Ca2+, which elicits the mitochondrial pathway of apoptosis by activation of the calcium-dependent phosphatase calcineurin, leading to dephosphorylation of the 'BH3 only' protein BAD. However, thorough study of the results suggests that other pathways are implicated in 7-ketocholesterol-induced cytotoxicity. In this study, we demonstrate the involvement of two other calcium-dependent pathways during 7-ketocholesterol-induced apoptosis. The activation of the MEK-->ERK pathway by the calcium-dependent tyrosine kinase PYK 2, a survival pathway which delays apoptosis as shown by the use of the MEK inhibitor U0126, and a pathway involving another pro-apoptotic BH3 only protein, Bim. Indeed, 7-ketocholesterol treatment of human monocytic THP-1 cells induces the release of Bim-LC8 from the microtubule-associated dynein motor complex, and its association with Bcl-2. Therefore, it appears that 7-ketocholesterol-induced apoptosis is a complex phenomenon resulting from calcium-dependent activation of several pro-apoptotic pathways and also one survival pathway.

Apoptosis↗
PubMed2004

7-Ketocholesterol enhances the expression of adhesion molecules on human aortic endothelial cells by increasing the production of reactive oxygen species.

The aim of the present study was to assess the expression of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), monocytic adhesion of human aortic endothelial cells (HAECs), and the production of intracellular reactive oxygen species (ROS), when HAECs were stimulated by 7-ketocholesterol. 7-ketocholesterol enhances surface expression of ICAM-1 and VCAM-1 as determined by EIA, induces their mRNA expression by RT-PCR, and stimulates adhesiveness of HAECs to U937 monocytic cells. We confirmed up-regulation of ROS production of HAECs treated with 7-ketocholesterol. Although the surface expression of ICAM-1 and VCAM-1 on HAECs treated with 7-ketocholesterol increased in a time-dependent manner, alpha-tocopherol inhibited this increase of the surface expression of ICAM-1 and VCAM-1. In the monocytic adhesion assay, adhesion of U937 to HAECs treated with 7-ketocholesterol was enhanced, but monoclonal anti-ICAM-1 and VCAM-1 antibodies reduced the endothelial adhesiveness. In conclusion, this study suggests that the endothelial adhesiveness to monocytic cells that was increased by 7-ketocholesterol was associated with enhanced expression of ICAM-1 and VCAM-1 mediated by ROS production.

Aorta↗