| For medical practitioners and the general public - Cholesterol Journal Article Catalog. |
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| Role of beef and beef tallow, an enriched source of stearic acid, in a cholesterol-lowering diet Denke, M. A. (1994), Am J Clin Nutr 60(6 Suppl): 1044S-1049S. Abstract: The effects of stearic acid on metabolism must be evaluated for stearic acid as an isolated dietary constituent and for stearic acid as a component of a natural fat. Beef products are the most common source of dietary stearic acid in the United States. Two components of beef products, beef protein and beef fat, can potentially impart cholesterol-raising properties. Protein has minimal effects on cholesterol concentrations in humans, but studies suggest that beef fat raises serum cholesterol concentrations. Because beef fat is 19% stearic acid, the cholesterol-raising potential of beef is not as great as predicted by its total saturated fatty acid content. However, beef tallow is hypercholesterolemic compared with fats containing less cholesterol-raising saturated fatty acid. Therefore, curtailment of beef tallow in a cholesterol-lowering diet seems appropriate. Data suggest that lean beef is no more hypercholesterolemic than chicken or fish and, therefore, lean beef need not be eliminated from cholesterol-lowering diets. |
| Role of bile salt-dependent cholesteryl ester hydrolase in the uptake of micellar cholesterol by intestinal cells Shamir, R., W. J. Johnson, et al. (1995), Biochemistry 34(19): 6351-8. Abstract: The bile salt-dependent cholesteryl ester hydrolase (CEH; EC 3.1.1.13) has been proposed to promote the intestinal absorption of both the free and esterified (FC, CE) forms of dietary cholesterol. For example, it was recently reported that in the human intestinal cell line CaCo2, addition of bovine CEH to the medium increased the uptake and intracellular esterification of micellar FC supplied at subphysiological concentrations Lopez-Candales et al. (1993) Biochemistry 32, 12085-12089. To test the ability of CEH to promote micellar cholesterol uptake in a CaCo2 system under more physiological conditions, an in vitro model was developed. Cells stably expressing rat CEH were created by DNA transfection (Tr cells), and the uptake of micellar FC and its intracellular esterification were measured using isotopic methods in Tr and control cells. Experimental parameters that were varied included micellar composition (monoolein or egg PC; FC, CE, or both), the final concentration of micellar cholesterol (1 nM to 50 microM), the origin of CEH (endogenously synthesized vs exogenously added), and the species source of enzyme (rat, pig, man). The uptake of cholesterol that was derived from micellar CE was significantly increased 5-10-fold (p < 0.001) in Tr vs control cells as a result of the hydrolysis of the CE by the CEH and subsequent uptake of the liberated free cholesterol. In contrast, the uptake of micellar FC was not increased by the presence of CEH, whether it was endogenous or exogenous. In addition, based on TLC analysis of extracted cellular lipids, there was no evidence that CEH promoted the esterification of the FC that was taken up.(ABSTRACT TRUNCATED AT 250 WORDS) |
| Role of biliary proteins and non-protein factors in kinetics of cholesterol crystallisation and gallstone growth Jirsa, M. and A. K. Groen (2001), Front Biosci 6: E154-67. Abstract: Numerous biliary proteins as well as non-protein factors influence kinetics of cholesterol crystallisation from supersaturated bile. Cholesterol crystallisation pathways, methods used for quantitative evaluation of liquid-to-solid transition of cholesterol in bile and the classical concept of nucleation defect in pathogenesis of gallstones are reviewed in the first part of this article. The relevance of individual biliary proteins and non-protein factors is discussed in the second part. Finally, a new concept according to which accelerated crystallisation of cholesterol inhibits gallstone growth and protects the gallbladder wall from toxic injury resulting from the increased absorption and accumulation of cholesterol from supersaturated bile is suggested. |
| Role of biliary proteins in pathogenesis of cholesterol gallstones Secknus, R. and R. T. Holzbach (1997), Z Gastroenterol 35(1): 41-6. Abstract: Cholelithiasis is a frequent disease in developed countries with significant economical implications. While the multifactorial pathogenesis of cholesterol gallstones has been widely accepted, the relative importance of the various contributing factors is not yet clear. One focus of research in recent years has been the identification and functional analysis of biliary proteins. Few proteins are synthesized in the biliary tract itself, the majority reflect the composition of serum proteins. Many biliary proteins modify cholesterol crystallization, the initial step of cholesterol gallstone formation. In cholelithiasis, biliary concentration of many pronucleating proteins are increased. However, this may be a consequence rather than the cause of the disease, because a majority of these proteins are acute-phase reactants. They may be secreted into bile at increased concentrations because of--often asymptomatic--cholecystitis due to gallstone disease. A number of protein-lipid interactions have been observed in search of a mechanism of action of biliary effector proteins. Only recently the potential modification of lipid removal from gallbladder bile by Apo A-I was described in addition to its direct antinucleating effect. In conclusion, biliary proteins directly modify cholesterol crystallization by protein-lipid interactions and may influence lipid absorption from the gallbladder. |
| Role of caveolae in cholesterol transport in arterial smooth muscle cells exposed to lipoproteins in vitro and in vivo Thyberg, J., F. Calara, et al. (1998), Lab Invest 78(7): 825-37. Abstract: Arterial smooth muscle cells are able to shift between two major differentiated states with distinct morphologic and functional properties, a contractile phenotype and a synthetic phenotype. Recently, it was demonstrated that contractile smooth muscle cells have numerous caveolae and that these specialized regions of the plasma membrane, to a large extent, are lost when the cells are modified into a synthetic phenotype. At the same time, the levels of the cholesterol-binding membrane protein caveolin remained unchanged and caveolin was redistributed from the cell surface to the perinuclear cytoplasm. In the present investigation, electron microscopy was used to study how smooth muscle cells of different phenotypes react to exposure to low-density lipoprotein and other lipoproteins both in vitro and in vivo. Our findings indicate that contractile cells (present early in primary culture and in the media of normal arterial walls) do not accumulate lipids in the cytoplasm and release excess cholesterol by means of plasma membrane caveolae. Extracellularly, the expelled lipids were built into membranous configurations and piled up as myelin-like deposits. In synthetic cells (formed after a few days in primary culture and as a response to arterial injury), lipids gathered in cytoplasmic droplets and increased amounts of membranous inclusions appeared in endosomes and lysosomes. On the other hand, no signs of extracellular discharge of lipids were detected. The results suggest that contractile smooth muscle cells use caveolin and caveolae to free themselves of excess lipoprotein-derived cholesterol and so manage to maintain a balance in the influx and efflux of cholesterol. Synthetic smooth muscle cells show a Golgi-like immunostaining for caveolin but have an insufficient capacity to use this protein to transport cholesterol to the plasma membrane and out of the cell. Cholesterol will then rather be esterified and collect in lipid droplets, eventually leading to foam cell formation if the uptake of lipoprotein continues. |
| Role of cell cholesterol in modulating antineoplastic ether lipid uptake, membrane effects and cytotoxicity Diomede, L., A. Bizzi, et al. (1990), Int J Cancer 46(2): 341-6. Abstract: Membrane-interactive ether lipids (EL) exert toxic and antiproliferative effects on cancer cells in vitro. They appear to be selectively more toxic to cancer cells than to normal cells and thus they are ideal candidates for bone-marrow purging procedures. However, no conclusive explanation has yet been provided for this property. We now present some data indicating that the cholesterol concentration in the incubation medium modulates EL toxicity against the HL60 leukemic cell line in vitro. Furthermore, model membranes richer in cholesterol take up EL more slowly, and cell cholesterol enrichment of HL60 cells counteracts EL biophysical membrane interaction, but not toxicity, in our experimental model. However, the K562 cell line, a leukemia line less sensitive to EL toxic action, has higher levels of cell cholesterol. Our data provide evidence to explain differences in sensitivity to EL among different cell types and contribute to the understanding of the mechanism of action of EL. |
| Role of cell cholesterol in modulating vincristine uptake and resistance Pallares-Trujillo, J., C. Domenech, et al. (1993), Int J Cancer 55(4): 667-71. Abstract: The relationship between cell-membrane permeability to vincristine and cholesterol/phospholipid levels was studied in L5178Y murine leukemic lymphoblasts and in 2 multidrug-resistant cell sublines, VCR/P60 and VCR/P200, which expressed increasing levels of vincristine resistance. The uptake of 3H-vincristine was measured in all cell lines and in cholesterol-depleted and -reloaded L5178Y and VCR/P200 cells. The initial rate of drug entry in resistant cells was lower than that measured in the parental cell line and it decreased as the relative resistance increased. An increment of cholesterol content, characterized in resistant cells, was directly proportional to the relative resistance to vincristine. Cholesterol depletion in both sensitive and resistant cells resulted in an increase in the rate of vincristine uptake, which reverted to the respective basal levels when each cell line was cholesterol-reloaded. The rate of drug uptake was inversely correlated with the molar ratio of cholesterol to phospholipids. Although both VCR/P cell sublines, but not the sensitive parental cells, expressed the P-glycoprotein in their plasma membrane, there were no differences in drug efflux and retention between resistant and parental cells. These results indicate that cholesterol modulates the permeation of vincristine through the plasma membrane and strongly suggest that increased levels of cholesterol/phospholipid account for the lower drug accumulation and greater resistance in these multidrug-resistant cells. |
| Role of cholestanol in the pathogenesis of cholesterol cholelithiasis Chupin, S. P., L. Tiuriumin Ia, et al. (1992), Ross Med Zh(1): 11-3. Abstract: The proportion of cholestanol/cholesterol in hepatic and cystic bile was evaluated using capillary gas-chromatography in subjects with disturbances in the biliary system (biliary dyskinesia, chronic acalculous cholecystitis, chronic calculous cholecystitis). The proportion was also measured in bile "paste" and cholesterol choleliths. With progression of cholesterol cholelithiasis, the proportion cholestanol/cholesterol increases, reaching its maximum in the stone the center of which accumulates crystals of cholestanol-cholesterol dihydrate. An original conception is proposed on pathogenesis of cholesterol cholelithiasis in man. It considers cholestanol as factor-reason underlying various pathological processes running in cholesterol cholelithiasis. Cholestanol is suggested to mark cholesterol cholelithiasis. |
| Role of cholesterol and polyunsaturated chains in lipid-protein interactions: molecular dynamics simulation of rhodopsin in a realistic membrane environment Pitman, M. C., A. Grossfield, et al. (2005), J Am Chem Soc 127(13): 4576-7. |
| Role of cholesterol and the ganglioside GM(1) in entry and short-term survival of Brucella suis in murine macrophages Naroeni, A. and F. Porte (2002), Infect Immun 70(3): 1640-4. Abstract: Brucella species are gram-negative, facultative intracellular bacteria that infect humans and animals. These organisms can survive and replicate within a membrane-bound compartment inside professional and nonprofessional phagocytic cells. Inhibition of phagosome-lysosome fusion has been proposed as a mechanism for intracellular survival in both types of cells. We have previously shown that the maturation inhibition of the Brucella-containing phagosome appears to be restricted at the phagosomal membrane, but the precise molecular mechanisms and factors involved in this inhibition have yet to be identified. Interestingly, recent studies have revealed that caveolae or lipid rafts are implicated in the entry of some microorganisms into host cells and mediate an endocytic pathway avoiding fusion with lysosomes. In this study, we investigated the role of cholesterol and the ganglioside GM(1), two components of lipid rafts, in entry and short-term survival of Brucella suis in murine macrophages, by using cholesterol-sequestering (filipin and beta-methyl cyclodextrin) and GM(1)-binding (cholera toxin B) molecules. Our results suggest that lipid rafts may provide a portal for entry of Brucella into murine macrophages under nonopsonic conditions, thus allowing phagosome-lysosome fusion inhibition, and provide further evidence to support the idea that the phagosome maturation inhibition is restricted at the phagosomal membrane. |
| Role of cholesterol as a structural and functional effector of the nicotinic acetylcholine receptor Fernandez-Ballester, G., J. Castresana, et al. (1994), Biochem Soc Trans 22(3): 776-80. |
| Role of cholesterol ester mass in regulation of secretion of ApoB100 lipoprotein particles by hamster hepatocytes and effects of statins on that relationship Zhang, Z., K. Cianflone, et al. (1999), Arterioscler Thromb Vasc Biol 19(3): 743-52. Abstract: Our understanding of the factors that regulate the secretion of apoB100 lipoproteins remains incomplete with considerable debate as to the role, if any, for cholesterol ester in this process. This study examines this issue in primary cultures of hamster hepatocytes, a species in which both cholesterol and apoB100 metabolism are very similar to man. Addition of oleate to medium increased the mass of triglyceride and cholesterol ester within the hepatocyte and also increased the secretion of triglycerides, cholesterol ester, and apoB100 into the medium. Next, the responses of hamster hepatocytes to addition of either an HMG-CoA reductase inhibitor (lovastatin) or an acyl-CoA cholesterol acyltransferase inhibitor (58-035) to the medium, with or without added oleate, were determined. Effects of either agent were only evident in the oleate-supplemented medium in which cholesterol ester mass had been increased above basal. If oleate was not added to the medium, neither agent reduced apoB100 secretion; equally important, over the 24-hour incubation, neither agent, at the concentration used, produced any detectable change in intracellular cholesterol ester mass. However, in contrast to the estimates of mass, which were unchanged, under the same conditions radioisotopic estimates of cholesterol ester synthesis were markedly reduced. Any conclusion as to the relation of cholesterol ester mass to apoB100 secretion would therefore depend on which of the 2 methods was used. Overall, the data indicate a close correlation between the mass of cholesterol ester within the hepatocyte and apoB100 secretion from it and they go far to explain previous apparently contradictory data as to this relation. More importantly, though, taken with other available data, they indicate that the primary response of the liver to increased delivery of lipid is increased secretion rather than decreased uptake. These results point, therefore, to a hierarchy of hepatic responses to increased flux of fatty acids and increased synthesis of cholesterol that in turn suggests a more dynamic model of cholesterol homeostasis in the liver than has been appreciated in the past. |
| Role of cholesterol ester pathway in the control of cell cycle in human aortic smooth muscle cells Batetta, B., M. F. Mulas, et al. (2003), Faseb J 17(6): 746-8. Abstract: Cholesterol esterification by acyl-CoA:cholesterol acyltransferase (ACAT) and proliferation of vascular smooth muscle cells (VSMC) are key events in vascular proliferative diseases. Here we performed experiments to ascertain the role of cholesterol ester pathway in the control of human aortic VSMC cycle progression. Results showed that serum-induced VSMC proliferation was preceded by an increased ability of the cells to esterify cholesterol as well as by an increased expression of ACAT and multidrug resistance (MDR1) mRNAs and extracellular related kinases 1/2 (ERK1/2), whereas caveolin-1 levels were markedly decreased. Cell cycle analyses performed in the presence of two inhibitors of cholesterol esterification, directly inhibiting ACAT (Sandoz 58-035) or the transport of cholesterol substrate from plasma membrane to endoplasmic reticulum (progesterone), indicate that each inhibitor suppressed the serum-induced DNA synthesis by accumulation of VSMCs in the G1 phase. The effect was associated with a rapid inhibition of ERK1/2 mitogenic signaling pathway; a down-regulation of cyclin D1, ACAT, and MDR1 mRNA; and an up-regulation of caveolin-1. These data provide a plausible link between cholesterol esterification and control of cell cycle G1/S transition, supporting the hypothesis that cholesterol esterification may accelerate the progression of human vascular proliferative diseases by modulating the rate of the VSMC proliferation. |
| Role of cholesterol esters in triglyceride transport Titov, V. N. (1995), Biokhimiia 60(9): 1371-81. Abstract: Biosynthesis of cholesterol esters in the blood flow and their function are determined by the fact that these compounds are the sole endogenously synthesized molecules whose hydrophobicity is higher than that of triglycerides transferred by the blood. Within the structure of intermediate density lipoproteins, cholesterol esters substitute for triglycerides during association of apoprotein B-100 with the C-terminal lipid-binding domain. Substitution of the liquid-crystalline phase of triglycerides for the corresponding phase of cholesterol esters results in a significant alteration of the apoprotein B-100 conformation and the formation of low density lipoproteins. Within the structure of low density lipoproteins, the N-terminal domain (apoprotein B-100 ligand) acquires an active conformation and an ability to interact with cell membrane receptors, thereby facilitating their absorption of low density lipoproteins. Cholesterol synthesis in the blood is an independent step in triglyceride transport. Inhibition of cholesterol ester synthesis into high density lipoproteins and their conversion into intermediate density lipoproteins is unaccompanied by the acquisition by apoprotein B-100 of a final conformation or by the formation of low density lipoproteins. Remnants of intermediate density lipoproteins having an intermediate conformation of apoprotein B-100 and the inactive position of the domain ligand are accumulated in the blood. This results in the development of IIb type hyperlipoproteinemia. Resumption of conversion of cholesterol esters from high density lipoproteins into intermediate density lipoprotein remnants normalizes the triglyceride transport. The final conformation of apoprotein B-100 can be modelled by substituting triglycerides in intermediate density lipoprotein remnants for another substance whose hydrophobicity is similar to that of cholesterol esters (dolichol, probucol).(ABSTRACT TRUNCATED AT 250 WORDS) |
| Role of cholesterol in cardiovascular dysfunction Saini, H. K., A. S. Arneja, et al. (2004), Can J Cardiol 20(3): 333-46. Abstract: BACKGROUND: Hypercholesterolemia, which is characterized by high levels of lipoprotein-containing cholesterol in plasma, is generally accepted as a major risk factor for the development of atherosclerosis and subsequent myocardial ischemia. The cardiovascular effects of elevated serum cholesterol are predominantly attributed to atherosclerotic lesions in coronary arteries; however, the role of cholesterol in causing heart dysfunction without the occurrence of atherosclerosis is not fully appreciated. OBSERVATIONS: Each type of biological membrane has a specific amount of cholesterol, which is required for proper functioning of the membrane-bound enzymes and cation transporters. High levels of cholesterol have been demonstrated to cause changes in membrane structure and function independent of atherosclerosis. Particularly, alterations in membrane cholesterol content have been shown to affect myocardial contractility, excitability and conduction properties. The activities of cardiac sarcolemmal enzymes such as Na+-K+ ATPase, Mg2+ ATPase and Ca2+ pump ATPase as well as Ca2+-dependent K+ channels and Na+-Ca2+ exchanger are altered as a consequence of changes in the membrane cholesterol content. Furthermore, high levels of cholesterol are known to cause endothelial dysfunction and smooth muscle abnormalities, which occur without visible atherosclerosis lesion development. On the other hand, indirect effects of cholesterol on the cardiovascular system are evident on its oxidative modification and subsequent development of visible atherosclerotic lesions and myocardial ischemia. CONCLUSIONS: Hypercholesterolemia has been shown to cause cardiovascular dysfunction due to direct action on membrane fluidity, enzyme activities and cation transporters in the endothelial cells, vascular smooth muscle cells and cardiomyocytes. On the other hand, development of atherosclerosis by high levels of cholesterol is associated with the oxidation products of cholesterol and is thus considered to affect the cardiovascular system indirectly as a consequence of ischemic heart disease. |
| Role of cholesterol in developing T-tubules: analogous mechanisms for T-tubule and caveolae biogenesis Carozzi, A. J., E. Ikonen, et al. (2000), Traffic 1(4): 326-41. Abstract: Recent work has suggested that caveolae biogenesis and transverse-tubule (T-tubule) formation in muscle cells share similar underlying features. We compared the properties of caveolin-1 (cav-1)-positive caveolae, in epithelial cells, with caveolin-3 (cav-3)-positive precursor T-tubules, in differentiating C2C12 muscle cells, using the cholesterol-binding drug, Amphotericin B (AmphB). Treatment of MDCK epithelial cells with acute high doses or chronic low doses of AmphB caused a loss of surface caveolae and the rapid redistribution of cav-1, and exogenously expressed cav-3, from the cell surface into modified endosomes. This effect was reversible and specific, as the GPI-anchored protein, alkaline phosphatase, was largely unaffected by the treatment unless it had been previously partitioned into caveolar domains. In differentiating C2C12 mouse myotubes, AmphB also caused a complete redistribution of cav-3 from precursor T-tubule elements into enlarged endosomes, morphologically very similar to those seen in MDCK cells. This was accompanied by redistribution of a T-tubule marker and a dramatic reduction in the extent of surface-connected tubular elements. We propose that cholesterol-enriched glycolipid 'raft' domains are involved in the formation and maintenance of diverse membrane systems including caveolae and the T-tubule system of muscle. |
| Role of cholesterol in embryonic development Roux, C., C. Wolf, et al. (2000), Am J Clin Nutr 71(5 Suppl): 1270S-9S. Abstract: We showed previously that 3 distal inhibitors of cholesterol synthesis are highly teratogenic in rats. AY 9944 and BM 15766 inhibit 7-dehydrocholesterol reductase, which catalyzes the last step of cholesterol synthesis, and triparanol inhibits Delta(24)-dehydrocholesterol reductase, which catalyzes the last step in another pathway. These molecules cause holoprosencephalic brain anomalies. Under certain experimental conditions, other anomalies (of the limbs and male genitalia) are also observed. Assays performed by gas chromatography-mass spectrometry (GC-MS) show hypocholesterolemia and an accumulation of precursors. These data indicate that this animal model can be considered a model of Smith-Lemli-Opitz syndrome. Smith-Lemli-Opitz syndrome is a recessive autosomal genetic disease characterized by malformations (microcephaly, corpus callosum agenesis, holoprosencephaly, and mental retardation), male pseudohermaphroditism, finger anomalies, and failure to thrive. The syndrome has been attributed to a deficit in 7-dehydrocholesterol reductase. As assayed by GC-MS, the sterol status of these patients indicates severe hypocholesterolemia and an accumulation of precursors: 7-dehydrocholesterol, 8-dehydrocholesterol, and oxidized derivatives. The presence of 7-dehydrocholesterol in the serum of patients is pathognomonic of the disease. The developmental gene Shh (sonic hedgehog) plays a key role in brain, limb, and genital development; it was shown recently that the Shh protein has to be covalently linked to cholesterol to be active. This is the first time that a posttranslational function has been attributed to cholesterol. There is an obvious relation between Shh dysfunction and the malformations observed in our experiments and in patients with Smith-Lemli-Opitz syndrome. However, the exact relation remains to be clarified. It is clear, however, that the role of cholesterol in embryonic development must be taken into account. |
| Role of cholesterol in formation and function of a signaling complex involving alphavbeta3, integrin-associated protein (CD47), and heterotrimeric G proteins Green, J. M., A. Zhelesnyak, et al. (1999), J Cell Biol 146(3): 673-82. Abstract: Integrin-associated protein (CD47) is a multiply membrane spanning member of the immunoglobulin superfamily that regulates some adhesion-dependent cell functions through formation of a complex with alphavbeta3 integrin and trimeric G proteins. Cholesterol is critical for the association of the three protein components of the supramolecular complex and for its signaling. The multiply membrane spanning domain of IAP is required for complex formation because it binds cholesterol. The supramolecular complex forms preferentially in glycosphingolipid-enriched membrane domains. Binding of mAb 10G2 to the IAP Ig domain, previously shown to be required for association with alphavbeta3, is affected by both the multiply membrane spanning domain and cholesterol. These data demonstrate that cholesterol is an essential component of the alphavbeta3/IAP/G protein signaling complex, presumably acting through an effect on IAP conformation. |
| Role of cholesterol in germ-line development of Caenorhabditis elegans Shim, Y. H., J. H. Chun, et al. (2002), Mol Reprod Dev 61(3): 358-66. Abstract: We investigated the effects of cholesterol starvation on Caenorhabditis elegans development at both embryonic and post-embryonic stages by examining brood size, embryonic lethality, growth rate, and worm size. The brood sizes of worms grown without cholesterol were substantially reduced in subsequent generations as compared to the control group with cholesterol: 13, 33, and 39% at the first, the second, and the third generation, respectively. The growth rate was also reduced by 20%-26%. Worms became adults after 120-130 hr incubation at 20 degrees C. Embryonic lethality was detected in the range of 1.6%-2.9% as compared to 0.8% of the control group. The percent development from an embryo to an adult was lowered by an average of 10%. Further analyses of germ line development to understand the reduction of brood size revealed that both germ line proliferation and differentiation were affected, and the most striking effect was seen in oogenesis. Defective oogenesis resulted in endomitotic oocytes (Emo, 22% at F1, 26% at F2, and 30% at F3). Thus, cholesterol appears to be required for all developmental stages of C. elegans. |
| Role of cholesterol in human immunodeficiency virus type 1 envelope protein-mediated fusion with host cells Viard, M., I. Parolini, et al. (2002), J Virol 76(22): 11584-95. Abstract: In this study we examined the effects of target membrane cholesterol depletion and cytoskeletal changes on human immunodeficiency virus type 1 (HIV-1) Env-mediated membrane fusion by dye redistribution assays. We found that treatment of peripheral blood lymphocytes (PBL) with methyl-beta-cyclodextrin (MbetaCD) or cytochalasin reduced their susceptibility to membrane fusion with cells expressing HIV-1 Env that utilize CXCR4 or CCR5. However, treatment of human osteosarcoma (HOS) cells expressing high levels of CD4 and coreceptors with these agents did not affect their susceptibility to HIV-1 Env-mediated membrane fusion. Removal of cholesterol inhibited stromal cell-derived factor-1alpha- and macrophage inflammatory protein 1beta-induced chemotaxis of both PBL and HOS cells expressing CD4 and coreceptors. The fusion activity as well as the chemotactic activity of PBL was recovered by adding back cholesterol to these cells. Confocal laser scanning microscopy analysis indicated that treatment of lymphocytes with MbetaCD reduced the colocalization of CD4 or of CXCR4 with actin presumably in microvilli. These findings indicate that, although cholesterol is not required for HIV-1 Env-mediated membrane fusion per se, its depletion from cells with relatively low coreceptor densities reduces the capacity of HIV-1 Env to engage coreceptor clusters required to trigger fusion. Furthermore, our results suggest that coreceptor clustering may occur in microvilli that are supported by actin polymerization. |