| For medical practitioners and the general public - Cholesterol Journal Article Catalog. |
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| Comparison of the uptake and processing of cholesterol from chylomicrons of different fatty acid composition in rats fed high-fat and low-fat diets Bravo, E., L. Flora, et al. (1997), Eur J Biochem 246(1): 92-102. Abstract: The fate of 3Hcholesterol carried in chylomicrons prepared from rats given a meal of palm oil (rich in long-chain saturated fatty acids), olive oil (rich in monounsaturated fatty acids) or corn oil (rich in n-6 polyunsaturated fatty acids) was investigated in vivo in rats fed a low-fat diet or a diet supplemented with the corresponding oil (to provide 40% of the calories) for 21 days. In the low-fat-fed groups, radioactivity was removed from the blood and secreted into bile over 180 min more rapidly when the chylomicrons were derived from corn oil as compared to palm or olive oil. After feeding the corresponding high-fat diets, however, both parameters were decreased in rats fed palm and corn oil, but not olive oil. As a result of these changes, the rates of removal of radioactivity from the blood and secretion into bile were similar in animals given the olive oil and corn oil diets, and higher than those in rats fed the palm oil diet. All the high-fat diets tended to increase the proportion of the radioactivity in the plasma found in the 1.006-1.050-g/ml fraction (low-density lipoprotein) and decrease that in the 1.050-1.25-g/ml (high-density lipoprotein) fraction in comparison to the respective low-fat diet groups, but the transfer of radioactivity to the plasma high-density lipoprotein fraction was particularly slow in palm-oil-fed rats. These findings indicate that diets high in saturated or n-6 polyunsaturated fat retard the metabolism of chylomicron cholesterol in comparison to diets low in fat, while those high in monounsaturated fat do not have this effect. As a consequence of this, the rate of removal of cholesterol of dietary origin from the body is slower in animals fed saturated as compared to monounsaturated or n-6 polyunsaturated fat. Thus, differential metabolism of chylomicron cholesterol clearly plays an important role in the hyper- and hypo-cholesterolaemic effects of these dietary fats. |
| Comparison of three Knowledge Representation formalisms for encoding the NCEP Cholesterol Guidelines Starren, J. and G. Xie (1994), Proc Annu Symp Comput Appl Med Care: 792-6. Abstract: Although many Knowledge Representation (KR) formalisms have been used to encode care guidelines, there are few direct comparisons among different formalisms. In order to compare their suitability for encoding care guidelines, three different KR formalisms were used to encode the National Cholesterol Education Panel (NCEP) guideline. PROLOG, a First Order Logic system, CLASSIC, a frame-based representation system, and CLIPS, a production rule system, were used in the comparison. All three representations allowed accurate encoding of the guideline. PROLOG produced the most compact representation, but proved the most difficult to debug. The lack of arbitrary disjunction in CLASSIC greatly increased the complexity of the encoding. Overall, the CLIPS representation was the most intuitive and easiest to use. |
| Comparison of three species of dietary fish: effects on serum concentrations of low-density-lipoprotein cholesterol and apolipoprotein in normotriglyceridemic subjects Gerhard, G. T., B. D. Patton, et al. (1991), Am J Clin Nutr 54(2): 334-9. Abstract: Limited information is available comparing the effect of different species of fish on serum low-density-lipoprotein-cholesterol (LDL-C) and apolipoprotein B (apo B) concentrations. We fed 21 normotriglyceridemic males diets containing different species of fish (200 g Dover sole, Chinook salmon, or sablefish) for 18 d in a three-period crossover design. Concentrations of apo B and LDL-C rose on the salmon and sablefish diets compared with the sole diet (P = 0.02 for apo B, 0.08 for LDL-C). These increases were parallel to each other: apo B rose 14% and LDL-C rose 16% on the salmon diet and 17% and 14%, respectively, on the sablefish diet compared with the diet consumed before the study. These results suggest that the consumption of fish with a moderate amounts of n-3 fatty acids may cause a deleterious rise in LDL-C and apo B concentrations in normotriglyceridemic males. |
| Comparison of total cholesterol and full lipid panel values in identifying at-risk individuals in a community screening program Hoffman, C. J., C. Lee, et al. (1993), Am J Health Promot 7(4): 260-2. |
| Comparison of total cholesterol measurement using reflotron capillary analysis and laboratory venous blood analysis Heilbraun, J., A. M. Snelling, et al. (1995), Am J Health Promot 10(1): 12-4. |
| Comparison of two assays for measuring LDL cholesterol Maitra, A., S. V. Hirany, et al. (1997), Clin Chem 43(6 Pt 1): 1040-7. Abstract: The purpose of this study was to evaluate the LipiDirect assay (L-LDL) against the Direct LDL immunoseparation method (D-LDL) and beta quantification (BQ-LDL) for measurement of LDL cholesterol (LDL-C) in patients with normo- and hypertriglyceridemia. Samples from 156 patients triglyceride (Tg) range 0.61-9.95 g/L were assayed for LDL-C concentrations with the three methods. An additional seven patients with type III hyperlipidemia and 25 paired sera from fasting and nonfasting individuals also were analyzed by the three methods. Both assays displayed excellent precision. The mean LDL-C value from L-LDL was significantly higher than BQ-LDL and D-LDL in normo- and hypertriglyceridemic samples (P < 0.001). The mean absolute bias of L-LDL vs BQ-LDL was 12.7% for Tg < 4 g/L and 30.6% for Tg > or = 4 g/L, compared with 6.2% and 12.5%, respectively, for D-LDL. L-LDL correctly classified only 68% of patients with LDL-C < 1.30 g/L and 57% of patients with LDL-C between 1.30-1.59 g/L as compared with 98% and 93%, respectively, for D-LDL (P < 0.001). In patients with type III hyperlipidemia, L-LDL had a 130% positive bias with BQ-LDL as compared with a 14% negative bias for D-LDL. With all three methods there were no significant differences between samples from fasting and nonfasting individuals. On the basis of these findings, the D-LDL assay appears to be superior to the L-LDL assay. |
| Comparison of two direct methods for HDL cholesterol measurement with an indirect precipitation method in diabetic patients Saeed, B. O., P. Smart, et al. (2002), Diabetes Nutr Metab 15(3): 169-72. Abstract: The conventional precipitation method for measuring HDL cholesterol involves a centrifugation step which prevents automation of the method. Several methods have been introduced for measuring HDL cholesterol without the need for a centrifugation step. These new methods are therefore automatable and can process a large number of samples in a short period of time. Measuring HDL cholesterol is an important aspect of management of diabetes mellitus. In this study, we compared 2 direct methods for measuring HDL cholesterol with a conventional precipitation technique in 63 patients with either Type 1 or Type 2 diabetes mellitus. Both direct methods showed acceptable precision but they both showed positive bias compared to the conventional precipitation method. The greatest degree of bias occurs at low HDL cholesterol levels, which are more important for Type 2 patients. Such differences may affect cardiovascular risk calculation in patients with diabetes. Further studies are required to investigate if a correction factor needs to be introduced when these direct assays are used to measure HDL cholesterol in patients with Type 2 diabetes mellitus. |
| Comparison of two homogeneous assays with a precipitation method and an ultracentrifugation method for the measurement of HDL-cholesterol Arranz-Pena, M. L., J. Tasende-Mata, et al. (1998), Clin Chem 44(12): 2499-505. Abstract: We report on the performance of four HDL-cholesterol assays: a homogeneous method based on polyethylene glycol-modified enzymes/alpha-cyclodextrin sulfate (PEGME; Kyowa); a homogeneous method based on polyanion-polymer/detergent (PPD; Daiichi); the usual precipitation method with phosphotungstic acid/MgCl2 (PTA); and an ultracentrifugation (UC) procedure. The homogeneous HDL-cholesterol assays (performed with automated analyzers) were precise and correlated well with the PTA and UC assays. The specificity and accuracy of the PEGME method were better than those of the PPD method. |
| Comparison of two homogeneous HDL cholesterol methods in a large population study Chittamma, A., W. L. Roberts, et al. (2004), Clin Biochem 37(9): 745-9. Abstract: OBJECTIVES: High-density lipoprotein cholesterol (HDL-C) is an independent risk factor for coronary heart disease. Data are lacking on the performance of homogeneous methods using a large number of samples. DESIGN AND METHODS: We compared the performance of two HDL-C direct assays, the Dimension RxL (the Dade method) and the COBAS INTEGRA (the Roche method), for population screening. Performance was assessed using 4214 sera obtained from the International Collaborative Study on Atherosclerosis and Stroke In Asia (InterASIA) participants. RESULTS: The method comparison results demonstrated that both methods were highly correlated (r = 0.972). Deming regression analysis showed a slope of 1.009 +/- 0.007, an intercept of 0.048 +/- 0.009 and a S(y/x) of 0.08. The means were 1.29 +/- 0.33 and 1.23 +/- 0.33 mmol/l for the Roche and Dade methods, respectively, and the range of observed values were 0.30-3.05 and 0.19-2.86 mmol/l, respectively. The 95% confidence interval for the mean of the method differences was -0.10 to 0.22 mmol/l. Percentages of low (> or = 1.55 mmol/l), normal (1.03-1.54 mmol/l), and high risk (< 1.03 mmol/l) results were 15.5, 55.4, 29.0 for the Dade and 19.3, 59.3, 21.4 for the Roche method. The percentage of concordantly classified subjects at each cut point was 77.1%, 84.4%, and 95.5%. The percentage of overall consistency subjects was 85.4%. Thirteen percent of subjects were discordantly classified into the higher-risk group while the 1.6% of subjects were discordantly classified into the lower-risk group. CONCLUSIONS: Both homogeneous HDL-C methods were correlated and agree well with one another. The percentage of concordantly classified subjects was high. Thus, either method is suitable for large population studies. |
| Comparisons of freshly isolated strains of Lactobacillus acidophilus of human intestinal origin for ability to assimilate cholesterol during growth Buck, L. M. and S. E. Gilliland (1994), J Dairy Sci 77(10): 2925-33. Abstract: Fecal isolates of Lactobacillus acidophilus were obtained from human volunteers and tested for bile tolerance, the ability to deconjugate bile salts, and the ability to assimilate (take up) cholesterol during growth. One hundred and twenty-three of the 304 isolates of lactobacilli obtained were identified as L. acidophilus. In most cases, isolates of L. acidophilus from the same volunteer varied significantly in the amount of cholesterol assimilated, bile salt deconjugated, and bile tolerance. The two cultures from each of nine volunteers that assimilated the most cholesterol were compared as a group to select the most active cultures. Lactobacillus acidophilus ATCC 43121 (an isolate from the intestines of a pig, which in an earlier study aided significantly in controlling serum cholesterol in pigs) was included in this comparison. Significant variation in the ability to assimilate cholesterol was observed among these isolates from different volunteers. Eight of 17 isolates assimilated numerically but not significantly more cholesterol than L. acidophilus ATCC 43121, and 4 isolates assimilated significantly less. Bile tolerance and bile salt deconjugation also varied significantly among the selected isolates. Six of the selected isolates were quantitatively but not significantly better able to deconjugate bile salts than L. acidophilus ATCC 43121, but none was significantly more bile tolerant. Based on characteristics tested, isolates B7, D3, L1, 016, and 017 have the most potential of those included in this study for use as dietary adjuncts to lower human serum cholesterol. |
| Compartmental isolation of cholesterol participating in the cytoplasmic cholesteryl ester cycle in Chinese hamster ovary 25-RA cells Klansek, J. J., G. J. Warner, et al. (1996), J Biol Chem 271(9): 4923-9. Abstract: Using the Chinese hamster ovary cell line, 25-RA, we have demonstrated that lipoprotein-derived cholesterol and endogenously synthesized cholesterol are selectively differentiated with respect to their cellular locations. These cells lack sterol-mediated regulation, spontaneously storing large amounts of esterified cholesterol, which turns over with a half-time of 7.5 h. When 3Hcholesterol was provided to the cells in serum to trace cellular cholesterol, the specific activities of cellular free and esterified cholesterol (6238 +/- 273 and 5128 +/- 277 cpm/ microg, respectively) failed to equilibrate, indicating that bulk cellular free cholesterol is isolated from that participating in the cholesteryl ester cycle. Using 3Hacetate to trace the fate of endogenously synthesized cholesterol, a failure of equilibration was also observed (specific activities of free and esterified cholesterol = 280 +/- 37 and 458 +/- 8 cpm/ microg, respectively). The lower specific activity of the precursor indicates that endogenously synthesized cholesterol is preferentially esterified. When cells radiolabeled with 3Hacetate were post-incubated in the absence of radiolabel, the specific activity of the esterified cholesterol pool remained significantly higher than that of the free cholesterol, suggesting that cholesterol derived from hydrolysis of esterified cholesterol is preferentially re-esterified. |
| Compartmentalization of cholesterol biosynthesis. Conversion of mevalonate to farnesyl diphosphate occurs in the peroxisomes Biardi, L. and S. K. Krisans (1996), J Biol Chem 271(3): 1784-8. Abstract: We have recently demonstrated that mevalonate kinase and farnesyl diphosphate (FPP) synthase are localized predominantly in peroxisomes. This observation raises the question regarding the subcellular localization of the enzymes that catalyze the individual steps in the pathway between mevalonate kinase and FPP synthase (phosphomevalonate kinase, mevalonate diphosphate decarboxylase, and isopentenyl diphosphate isomerase). These enzyme are found in the 100,000 x g supernatant fraction of cells or tissues and have been considered to be cytoplasmic proteins. In the current studies, we show that the activities of mevalonate kinase, phosphomevalonate kinase, and mevalonate diphosphate decarboxylase are equal in extracts prepared from intact cells and selectively permeabilized cells, which lack cytosolic enzymes. We also demonstrate structure-linked latency of phosphomevalonate kinase and mevalonate diphosphate decarboxylase that is consistent with a peroxisomal localization of these enzymes. Finally, we show that cholesterol biosynthesis from mevalonate can occur in selectively permeabilized cells lacking cytosolic components. These results suggest that the peroxisome is the major site of the synthesis of FPP from mevalonate, since all of the cholestrogenic enzymes involved in this conversion are localized in the peroxisome. |
| Compartmentalization of cholesterol metabolism and cellular growth in cultured intestinal crypt cells Reimann, F. M., G. Herold, et al. (1991), Biochim Biophys Acta 1085(3): 315-21. Abstract: Growth of rat intestinal crypt derived cells IEC-6 ceased when the key enzyme of cholesterol synthesis, hydroxymethylglutaryl-CoA reductase, was blocked by the competitive inhibitor mevinolin. This effect was reversed by the addition of mevalonolactone. LDL suppressed reductase activity as well as cholesterol synthesis from 14Coctanoate and stimulated acyl-CoA cholesterol acyltransferase, but failed to support cell growth despite rapid receptor mediated degradation even in the presence of low mevalonolactone concentrations. Inhibition of cholesterol esterification by Sandoz-Compound 58-035 enhanced cell growth in the presence of mevinolin, but did not promote proliferation in the additional presence of low-density lipoproteins. HDL3 but not HDL2 or tetranitromethane-modified HDL3 totally reversed the mevinolin induced inhibition of cell growth. This rescue by HDL3 was overcome by an increased dose of mevinolin. HDL3 derepressed reductase, stimulated cholesterol synthesis and reduced cholesterol esterification, but did not reverse the cholesterol synthesis inhibition by mevinolin. It is concluded that IEC-6 cells preferentially use endogenously synthesized cholesterol for membrane formation rather than low-density lipoprotein cholesterol. High-density lipoproteins appear to normalize cell growth in the presence of mevinolin by inhibition of cholesterol esterification and probably by inducing the formation of non sterol products of mevalonate. |
| Compartmentation of cholesterol within the cell Liscum, L. and J. R. Faust (1994), Curr Opin Lipidol 5(3): 221-6. Abstract: Mammalian cells tightly regulate their cholesterol content and the intracellular disposition of cholesterol. Most cellular free cholesterol resides in the plasma membrane, where it exists in lateral domains. Mechanisms governing cellular cholesterol levels and compartmentation are still largely unknown. In this review, we highlight recent studies documenting the compartmentation of cholesterol, especially those that have relevance to the regulation of cholesterol content. |
| Compensatory phospholipid digestion is required for cholesterol absorption in pancreatic phospholipase A(2)-deficient mice Richmond, B. L., A. C. Boileau, et al. (2001), Gastroenterology 120(5): 1193-202. Abstract: BACKGROUND AND AIMS: Numerous studies have suggested phospholipid inhibition of dietary cholesterol absorption through the gastrointestinal tract. This study addressed the importance of luminal phospholipid hydrolysis in this process. METHODS: The effect of phospholipase inhibition on cholesterol transport from intestinal lumen to the lymphatics was evaluated in lymph fistula rats. Cholesterol and phospholipid absorption efficiency in intact animals was evaluated in control and phospholipase A(2) (PLA2) gene-targeted mice. RESULTS: The PLA2 inhibitor FPL 67047XX retarded cholesterol absorption in a lymph fistula rat model. Under basal chow-fed dietary conditions, cholesterol absorption efficiency from a single bolus meal, and plasma lipid levels, were similar among PLA2+/+, PLA2+/-, and PLA2-/- mice. Interestingly, the nonhydrolyzable phospholipid dioleoyl ether phosphatidylcholine suppressed cholesterol absorption by 10% to 18% in mice without regard to their PLA2 genotype. When 1-palmitoyl-2-(14)Coleoyl-phosphatidylcholine was used as the substrate, the radiolabeled phospholipid was found to be hydrolyzed and absorbed with equal efficiency in PLA2+/+, PLA2+/-, and PLA2-/- mice. CONCLUSIONS: These results suggested that although phospholipid digestion in the intestinal lumen is a prerequisite for efficient cholesterol absorption, additional enzyme(s) can compensate for pancreatic PLA2 in catalyzing phospholipid digestion and facilitating cholesterol absorption in PLA2 knockout mice. |
| Competitive carotenoid and cholesterol incorporation into liposomes: effects on membrane phase transition, fluidity, polarity and anisotropy Socaciu, C., R. Jessel, et al. (2000), Chem Phys Lipids 106(1): 79-88. Abstract: Pure 1,2-dipalmitoyl-sn-glycero-3-phosphorylcholine (DPPC) or mixed DPPC:1,2-dipalmitoyl phosphatidyletanolamine (DPPE):1,2-dipalmitoyl diphosphatidylserine (DPPS) (17:5:3) liposomes were incorporated with 5 mol% dietary carotenoids (beta-carotene, lutein and zeaxanthin) or with cholesterol (16 and 48 mol%) in the absence or presence of 15 mol% carotenoids, respectively. The carotenoid incorporation yields ranged from 0.42 in pure to 0.72 in mixed phospholipid liposomes. They decreased significantly, from 3 to 14%, in the corresponding cholesterol-doped liposomes, respectively. Highest incorporation yields were achieved by zeaxanthin and lutein in phospholipid liposomes while in cholesterol-containing liposomes, lutein was highest incorporated. The effects on membrane structure and dynamics were determined by differential scanning calorimetry, steady-state fluorescence and anisotropy measurements. Polar carotenoids and cholesterol cause similar, dose-dependent effects: ordering and rigidification revealed by broadening of the transition peak, and increase of anisotropy. Membrane hydrophobicity is determined by cholesterol content and carotenoid polarity. In cholesterol-doped liposomes, beta-carotene is less incorporated than in cholesterol-free liposomes. Our observations suggest effects of carotenoids, even at much lower effective concentrations than cholesterol (8 to 80-fold), on membrane structure and dynamics. Although they are minor constituents of animal membranes, carotenoids may act as modulators of membrane phase transition, fluidity, polarity and permeability, and therefore, can influence the membrane physiology and pathology. |
| Complement dependent and independent liposome uptake by peritoneal macrophages: cholesterol content dependency Huong, T. M., H. Harashima, et al. (1998), Biol Pharm Bull 21(9): 969-73. Abstract: The uptake mechanisms of liposomes by rat peritoneal macrophages (PMs) were investigated. Incubation of liposomes with fresh rat serum enhanced the uptake of liposomes depending on the liposome size and cholesterol (CH) content. The binding of liposomes was also enhanced by serum, and this increase depended on the size and CH content as in the case of liposome uptake, which suggested that the binding of opsonized liposomes with PMs govern the extent in liposome uptake. The rate constant for the internalization (k(int)) was calculated by measuring both uptake and binding. The k(int) cannot explain the variation of liposome uptake for different sizes and CH contents. The kint values for liposomes with high (44%) and medium (33%) CH contents were constant (2.5 h(-1)), while those for liposomes with low (22%) CH content were significantly elevated (5-9 h(-1)). These results indicate the presence of at least two kinds of uptake mechanisms of liposomes. Treatment of serum with anti-C3 antibody completely inhibited the enhanced uptake of CH-high, large liposomes, which suggested that complement receptor-mediated phagocytosis may be an uptake mechanism for CH-high and -medium liposomes. In addition, complement-independent enhanced uptake was suggested for CH-low liposomes, since no inhibition was observed for CH-low liposomes by anti-C3 antibody and these liposomes were disintegrated in serum via complement-independent pathway. These results provided evidence that PMs take up liposomes via complement-dependent and independent mechanisms depending on the CH content of the liposomes. |
| Complete down-regulation of low-density-lipoprotein-receptor activity in the human hepatoma cell line Hep G2 by beta-migrating very-low-density lipoprotein and non-lipoprotein cholesterol. Different cellular regulatory pools of cholesterol Kamps, J. A. and T. J. van Berkel (1992), Eur J Biochem 206(3): 973-8. Abstract: Regulation of low-density-lipoprotein-receptor activity by low-density lipoprotein (LDL), cholesteryl-ester-rich beta-migrating very-low-density lipoprotein (beta-VLDL) and non-lipoprotein cholesterol was investigated in the human hepatoma cell line Hep G2. Competition studies indicate that LDL and beta-VLDL are bound to the same recognition site, tentatively the LDL receptor. The regulatory response of the LDL receptor upon prolonged incubation with LDL or beta-VLDL was, however, markedly different. 22 h preincubation of Hep G2 cells with excess LDL caused a partial down regulation to 31% of the initial level of the high-affinity association of LDL and 26% of the high-affinity degradation of LDL, while with beta-VLDL a complete down regulation of the LDL-receptor activity is observed. Preincubation of Hep G2 cells with beta-VLDL for 22 h led to a fourfold increase in intracellular cholesterol esters and a twofold increase in acyl-coA:cholesterol acyltransferase activity. With LDL, the amount of intracellular cholesterol esters is increased 1.6-fold. The more effective down regulation of LDL receptors by beta-VLDL as compared to LDL can be explained by the more effective intracellular cholesterol delivery with beta-VLDL than with LDL. Preincubation of Hep G2 cells for 22 h with acetylated LDL hardly influenced the LDL-receptor activity. Non-lipoprotein cholesterol, however, caused a complete down regulation of LDL-receptor activity at even lower extracellular cholesterol concentrations than with beta-VLDL. The complete down regulation of LDL receptors by non-lipoprotein cholesterol is not accompanied by a significant increase in acyl-coA:cholesterol acyltransferase activity, while the intracellular cholesterol ester concentration is only increased 1.6-fold. It is suggested that the effectiveness of non-lipoprotein cholesterol to regulate LDL receptors is caused by its efficiency to reach the sterol regulatory site. The inability of LDL to down regulate its receptor completely can thus be explained by the inability of LDL to deliver cholesterol adequately at the intracellular regulatory site of the LDL receptor. The observed complete down regulation of the LDL receptor by beta-VLDL may be responsible for the cholesterol-rich-diet induced, complete down regulation of LDL-receptor-mediated clearance of LDL in vivo. |
| Complete mapping of crystallization pathways during cholesterol precipitation from model bile: influence of physical-chemical variables of pathophysiologic relevance and identification of a stable liquid crystalline state in cold, dilute and hydrophilic bile salt-containing systems Wang, D. Q. and M. C. Carey (1996), J Lipid Res 37(3): 606-30. Abstract: Using complementary physical-chemical techniques we defined five different crystallization pathways as functions of time (30 days) and increasing lecithin (egg yolk) content in pathophysiologically relevant model biles super-saturated (cholesterol saturation indices, 1.2 - 2.7) by dilution of approximately equal to 29 g/dl bile salt-lecithin-cholesterol micellar solutions. As evidenced by quasi-elastic light-scattering spectroscopy, supersaturation was heralded by the appearance of unilamellar vesicles. With the lowest lecithin contents, arc-like crystals with habit and density (d 1.030 g/mL) consistent with anhydrous cholesterol appeared first and evolved via helical and tubular crystals to form plate-like cholesterol monohydrate crystals (d 1.045 g/mL). With higher lecithin fractions, cholesterol monohydrate crystals appeared earlier than arc and other transitional crystals. With typical physiological lecithin contents, early liquid crystals (d 1.020 g/mL) were followed by cholesterol monohydrate crystals and subsequent appearances of arc and other intermediate crystals. With higher lecithin contents, liquid crystals were followed by cholesterol monohydrate crystals only, and at the highest lecithin mole fractions, liquid crystals appeared that did not generate solid crystals. Added calcium increased solid crystal number in proportion to its concentration (5 - 20 mM) but did not influence appearance times, crystallization pathways, or micellar cholesterol solubilities. Decreases in temperature (37 degrees --> 4 degrees C), total lipid concentration (7.3 --> 2.4 g/dL), and bile salt hydrophobicity (3 alpha, 12 alpha --> 3 alpha, 7 alpha, 12 alpha --> 3 alpha, 7 beta hydroxylated taurine conjugates) progressively shifted all crystallization pathways to lower lecithin contents, retarded crystallization, and decreased micellar cholesterol solubilities. The lecithin content of mother biles decreased markedly during crystallization especially where liquid crystals were a coexisting phase at equilibrium. This systematic study provides a framework for understanding cholesterol crystallization in human and animal biles and for examining factors that influence the kinetics of phase separation. |
| Complete replacement of membrane cholesterol with 4,4',14-trimethyl sterols in a human T cell line defective in lanosterol demethylation Buttke, T. M. and T. M. Folks (1992), J Biol Chem 267(13): 8819-26. Abstract: A3.01 is a hypoxanthine/aminopterin/thymidine-sensitive, human immunodeficiency virus-susceptible, human T cell line derived by Folks et al. (Folks, T., Benn, S., Rabson, A., Theodore, T., Hoggan, M. D., Martin, M., Lightfoote, M., and Sell, K. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 4539-4543) following exposure of CEM cells to 8-azaguanine. In the present study, it is shown that A3.01 also contains a heretofore unrecognized mutation in cholesterol biosynthesis. A3.01 cells grown in the presence of 10% fetal bovine serum (FBS) contain primarily cholesterol in their membranes, but based on 14Cacetate labeling, synthesize only lanosterol and 24,25-dihydrolanosterol. Reduction in the amount of FBS provided resulted in decreased cellular levels of cholesterol with corresponding increases in the two 4,4',14-trimethyl sterols. In A3.01 cells cultured in 1% FBS medium, lanosterol and 24,25-dihydrolanosterol accounted for 7 and 45%, respectively, of total cellular sterols. Following dilution of the 1% FBS-grown cells into serum-free media, the level of membrane cholesterol gradually declined, such that after three passages it became virtually undetectable, whereas the proportions of lanosterol and 24,25-dihydrolanosterol rose to 25 and 75%, respectively. Even after eight passages in the serum-free media, A3.01 cells displayed a complete absence of cholesterol with no obvious effect on cell growth. Membranes isolated from A3.01 cells grown in the presence or absence of 10 micrograms/ml of cholesterol displayed similar phospholipid:sterol ratios, but membranes from the unsupplemented cells contained only approximately 5% as much cholesterol as the supplemented cell membranes. Finally, A3.01 cells grown in the absence of cholesterol were extremely resistant to the cytotoxic effects of amphotericin B, whereas cells cultured in the combined presence of 1% FCS and 10 micrograms/ml of cholesterol were sensitive to the drug. Collectively, these results demonstrate that 4,4',14-trimethyl sterols can effectively replace cholesterol in a human T cell lineage, indicating that not all mammalian cells have a requirement for cholesterol, per se. The A3.01 T cell lineage should prove useful in defining the role of cholesterol in membrane fusion and human immunodeficiency virus-mediated syncitia formation and cytopathic effects. |