Biomedical subjects
J R Patsch
Publications and source records attributed to J R Patsch.
[Structure and metabolism of high-density lipoproteins].
Epidemiological studies have established an inverse correlation between the levels of high density lipoproteins (HDL) in the blood and coronary heart disease (CHD). However, no cause-effect relationship has been established for this important discovery, nor are the mechanisms known by which HDL may provide protection. A recent study has established a strong association between the plasma HDL2 level and the ability to clear dietary fat from the circulation. This article discusses recent studies that demonstrate the important role of chylomicron catabolism as a determinant of HDL2 levels in the circulation.
Postprandial lipemia. A key for the conversion of high density lipoprotein2 into high density lipoprotein3 by hepatic lipase.
In this study, we have investigated the effects of alimentary lipemia in 15 normotriglyceridemic individuals on high density lipoproteins2 (HDL2) with respect to structure, composition, and substrate efficacy for hepatic lipase in vitro. In the study subjects, HDL2 levels ranged widely from 4.7 to 151.7 mg/dl plasma. HDL2 were isolated in the postabsorptive (pa) state and in the postprandial (pp) state, i.e., 7 h after ingestion of a standard fatty meal. In going from the pa state to the pp state, HDL2 exhibited higher flotation rates and lower densities due to a decreased proportion of protein (38.7----36.2%) and a higher abundance in phospholipid (32.5----34.9%). There was a variable increase in triglyceride at the expense of cholesteryl esters; this increase was correlated positively with the magnitude of pp lipemia (r = 0.69, P less than 0.01) and inversely with HDL2 levels (r = -0.72, P less than 0.01). Hdl2 fractions were incubated with human hepatic lipase in vitro. Product lipoproteins formed from lipolysis of pa-HDL2 and triglyceride-poorer pp-HDL2 were reduced in phospholipid content (by 25 and 50%, respectively) but remained in the size and density range of native HDL2. By contrast, a major fraction of triglyceride-richer pp-HDL2 was converted to particles with density, size, and apoprotein composition of native HDL3. Changes consistent with these findings in vitro were observed in vivo also, where 15 h postprandially, individuals with high-level lipemia showed a decrease in HDL2 and rise in HDL3, while those with lower-level lipemia did not. This study indicates that the magnitude of postprandial lipemia determines the proportion of triglyceride in pp-HDL2, which in turn determines whether or not HDL2 are converted to HDL3 by hepatic lipase action.
Exercise, high density lipoproteins, and fat tolerance.
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Inverse relationship between blood levels of high density lipoprotein subfraction 2 and magnitude of postprandial lipemia.
Triglyceridemic response to a standard oral fat meal was determined in 28 healthy subjects and related to the levels of several lipids, lipoproteins, and apolipoproteins in the post-absorptive plasma. A fatty test meal was administered orally, and postprandial plasma triglyceride levels were determined. In the fasting blood samples, concentrations of apolipoproteins (apo) A-I, A-II, and B were determined by radioimmunoassay, and those of high density lipoprotein (HDL) subfractions HDL2 and HDL3, by zonal ultracentrifugation. The magnitude of triglyceridemic response showed a negative correlation with the plasma levels of HDL2 (r = -0.860, P less than 0.001), HDL-associated cholesterol (r = -0.605, P less than 0.001), and apoA-I (r = -0.459, P less than 0.02). No correlation was found between the triglyceridemic response and the levels of total cholesterol, HDL3, and apoA-II. Triglyceridemic response was correlated positively with fasting triglyceride concentrations (r = 0.450, P less than 0.02) and apoB levels (r = 0.396, P less than 0.03). In two subjects followed for 3 yr, when HDL2 levels rose or fell, the triglyceridemic response decreased or increased, respectively (r = -0.944; r = -0.863). Our data indicate that normolipidemic individuals with high HDL2 levels in the plasma are able to clear alimentary fat at a faster rate than normolipidemic subjects with low HDL2 levels. The pronounced difference in severity and duration of postprandial lipemia among subjects with varying HDL2 levels may help to explain the negative correlation between the risk of atherosclerosis and HDL cholesterol levels.
Effect of cholesterol feeding on the distribution of plasma lipoproteins and on the metabolism of apolipoprotein E in the rabbit.
Changes in lipoprotein distribution and in apolipoprotein E metabolism were studied in rabbits fed a diet containing 1% cholesterol. Lipoprotein distribution was monitored by rate zonal ultracentrifugation, gel filtration chromatography, and electrophoretic techniques. Normolipidemic rabbit plasma contained VLDL, IDL, LDL, and HDL. Within 1 week on the 1% cholesterol diet, the d less than 1.006 g/ml material was greatly elevated, and the lipoproteins of higher density (LDL and HDL) decreased below levels of detection. Cholesteremic d less than 1.006 g/ml material was cholesteryl ester-rich, triglyceride-poor, and contained particles of Sf 20 to greater than 400. Upon diet normalization, the LDL and HDL reappeared within 2-4 weeks accompanied by a decrease in the d less than 1.006 g/ml material. The metabolism of apoE was studied by injecting purified and 125I-labeled apoE into rabbits and following the clearance of the tracer. ApoE in the normolipemic rabbit demonstrated a fractional catabolic rate (FCR) of 0.132 hr-1 and a half-life (t1/2) of 10.3 hr. ApoE in the hypercholesterolemic rabbit demonstrated an FCR of 0.055 hr-1 and a t1/2 of 49.5 hr. ApoE concentrations in the plasma as estimated by electroimmunoassay were 19.5 mg/dl in the control rabbit and 199 mg/dl in the hypercholesterolemic rabbit. From these data, absolute synthetic rates of 20.3 mg/kg per day and 86.1 mg/kg per day were calculated for the control and the hypercholesterolemic rabbit, respectively. We conclude that the cholesterol-supplemented diet caused pronounced elevation of apoE in the plasma due to overproduction of the protein.
The high density lipoproteins.
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Plasma lipoprotein changes resulting from immunologically blocked lipolysis.
The role of lipoprotein lipase (LPL) in the generation of low density lipoprotein (LDL) and high density lipoprotein (HDL) was investigated. Intravenous injections of high titer goat antiserum against highly purified chicken LPL into fasted roosters quantitatively blocks the removal of plasma VLDL triglyceride (1976. J. Lipid Res. 17: 498-505). Analyses of the chemical components of lipoproteins after 8 hr of LPL inhibition showed that the very low density lipoprotein (VLDL) concentration increased over 10-fold, while LDL and HDL concentrations decreased by 5-fold and 48%, respectively. LDL and HDL cholesterol levels decreased logarithmically over the 8-hr period, with half-lives of 2.4 and 6 hr, respectively. The composition of these lipoprotein fractions on a percent weight basis changed significantly. Experimental LDL contained 37% less phospholipid, 64% less cholesterol, and 2.3-fold more triglyceride than control LDL. Experimental HDL contained 3.1-fold more triglyceride and 50% less unesterified cholesterol than control HDL. The Stokes' radii of HDL were determined by gel filtration on Biogel A5M and Ultrogel AcA 22: the radius of experimental HDL (44.9 A) was smaller than that of control HDL (55.4 A). These measurements were confirmed by electron microscopy (43 and 54 A, respectively). After rate zonal ultracentrifugations of plasma samples, control LDL was clearly resolved, while no LDL could be detected in the experimental samples. Rate zonal ultracentrifugations of plasma samples also indicated that control HDL had a higher flotation rate than experimental HDL. Equilibrium zonal ultracentrifugation showed experimental HDL to be more dense than control HDL with hydrated densities of 1.118 and 1.113 g/ml, respectively. These experiments provide in vivo evidence that LDL is a direct metabolic product of VLDL and that LPL plays a role in the transfer of surface constituents from VLDL to HDL.-Behr, S. R., J. R. Patsch, T. Forte, and A. Bensadoun. Plasma lipoprotein changes resulting from immunologically blocked lipolysis.
Oxandrolone and plasma triglyceride reduction: effect on triglyceride-rich and high density lipoproteins.
Oxandrolone, an anabolic androgenic steroid, has been shown repeatedly to lower plasma triglycerides in hypertriglyceridemic patients. This study was performed to determine which of seven subfractions of triglyceride-rich lipoproteins are affected by the action of oxandrolone with respect to both their plasma levels and composition. Concurrently, we have determined the levels and composition of HDL subfractions and the plasma levels of the major HDL apoprotein, apoA-I. Oxandrolone was administered to two hypertriglyceridemic subjects, one with type III and one with type V hyperlipoproteinemia until plasma triglycerides were below the target level of 270 mg/dl. Two months and two weeks were required for the type III and type V patients, respectively. In both subjects, the treatment caused a reduction in the plasma levels of all seven subclasses of triglyceride-rich lipoproteins without altering their overall composition. LDL were at least temporarily increased. The reduction of VLDL subfractions caused by oxandrolone was accompanied by a progressive and consistent effect on HDL subfractions in both hypertriglyceridemic subjects; in the type III patient, oxandrolone reduced HDL2 from low pretreatment levels further until they became undetectable. The type V subject had no detectable HDL2 levels prior to treatment. In both subjects, oxandrolone lowered the levels of HDL3. This lowering effect was caused by a preferential reduction of the less dense, major HDL3 subfraction, i.e. HDL3L, causing the denser, smaller HDL3 subfraction, HDL3D, to become the predominant HDL class. The lowering of HDL levels was reflected by a decrease in the plasma levels of the major HDL apoprotein, apoA-I. This first report on the simultaneous reduction of VLDL and the larger, less dense HDL subclasses suggest that oxandrolone lowers plasma triglycerides by a mechanism other than increased lipolysis.
Lipoprotein-X: carbon-13 nuclear magnetic resonance studies on native, reconstituted, and model systems.
Lipoprotein-X (LP-X), a lipoprotein isolated from human cholestatic plasma by ethanol--acetate precipitation and zonal ultracentrifugation, has been studied by 13C NMR at 67.9 MHz. Spectra of LP-X and its three subfractions are markedly different from those of normal human high-density lipoprotein3 (HDL3) or low-density lipoprotein (LDL). Spectra of LP-X are characterized by the presence of unusually broad resonance lines, especially those attributable to C6 of unesterified cholesterol (160--260 Hz) and to C beta of phospholipid glyceride (240--290 Hz). In contrast, the CH2O, CH2N, and N(CH3)3 choline resonances have line widths comparable to those of normal LDL and HDL3. For the subfraction LP-X1, spin--lattice relaxation times (T1) of the fatty acyl olefin resonances at 129.8 and 128.0 ppm and of the unesterified cholesterol C6 at 120.1 ppm were measured to be 675, 766, and 162 ms, respectively. These times are comparable to those measured for the corresponding resonances in single bilayer vesicles whose lipid composition approximates that of LP-X. The three LP-X subfractions isolated by zonal ultracentrifugation gave spectra which are identical, within experimental error, as judged qualitatively from their appearance and quantitatively from the line widths of selected resonances. In addition, 13C NMR spectra of sonicated total LP-X lipids are similar to spectra of the intact native lipoprotein. This study suggests (a) that motions of lipids in LP-X as probed by 13C NMR are similar to the motions of lipids found in model vesicular systems, (b) that the motions of the cholesterol rings and phospholipid fatty acyl chains are significantly more restricted in LP-X than in HDL3 and LDL, and (c) that the motions of the phosphoryl moieties in all three systems are similar.
Lipoprotein-X: proton and phosphorus-31 nuclear magnetic resonance studies on native, reconstituted, and model systems.
LP-X, a lipoprotein present in the low-density range (d 1.006--1.063 g/mL) of cholestatic human plasma, has been studied with its normal counterpart (LDL) by 1H and 31P nuclear magnetic resonance. The 220-MHz 1H spectrum of LP-X contains four major lines: the choline CH2N and N+(CH3)3 resonances and the cholesteryl--acyl CH2 and CH3 envelopes. The widths of these four lines at 37 degrees C are approximately 24, 10, 124, and 48 Hz, respectively. The latter two line widths are much greater than the corresponding ones of LDL (28 and 20 Hz), suggesting the much more restricted motion of acyl chains and/or cholesteryl rings in LP-X. This difference persists over the temperature range 15--52.5 degrees C. The microscopic fluidity of LP-X and LDL was compared by titration with 2,2,6,6-tetramethylpiperidinyl-1-oxy (Tempo), a paramagnetic amphiphile which distributes between the bulk aqueous phase and the fluid lipid phase of lipoproteins. Tempo is much less effective in broadening the 1H resonances of LP-X than of LDL, indicating the lower permeability/fluidity of the former. The 40.5-MHz 31P spectrum of LP-X consists of a single resonance whose line width is approximately 20 Hz and whose spin--lattice relaxation time is 2.23 +/- 0.15 s. Titration of LP-X with Pr3+ ions splits this resonance into two lines, one remaining at the chemical shift of the original resonance and the other paramagnetically shifting downfield. The ratio of integrated areas for these two lines was 1:1.72. Titration of phosphatidylcholine--cholesterol vesicles alone, vesicles containing apolipoprotein-C and albumin, or vesicles containing apolipoprotein-X gave results similar to those obtained with native LP-X, suggesting the presence of a single bilayer structure in all of these systems.
Stimulation and suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase in normal human fibroblasts by high density lipoprotein subclasses.
Plasma concentrations of high density lipoproteins, (HDL3), a subclass of HDL, are relatively constant, while those of HDL2 are variable. We report that HDL2 suppress, while HDL3 stimulate 3-hydroxy-3-methylglutaryl-CoA reductase (EC 1.1.1.34) activity in normal human fibroblasts. HDL3, which contained no detectable HDL2 or low density lipoproteins, stimulated 3-hydroxy-3-methylglutaryl-CoA reductase activity 2-fold from 60 to 120 pmol/mg per min. The induction, which exhibited saturation kinetics with maximum stimulation at 150 microgram HDL phospholipid/ml medium, occurred only in the late-log and stationary phases of cell growth and was abolished by 0.1 mM actinomycin D or cycloheximide.l Apoliproprotein HDL3 did not stimulate enzyme activity, whereas the total lipid extract of HDL3 was about 1.7 times more potent than were the native HDL3 in stimulating enzyme activity. HDL2 consistently suppressed 3-hydroxy-3-methylglutaryl-CoA reductase in normal fibroblasts by 20-50% at 80 microgram HDL2 protein/ml. Mixtures of HDL2 and HDL3 suppressed when HDL2 were greater than 35% of the total HDL. The suppressive effects of HDL2 were abolished by treatment with 0.1 M cyclohexanedione and restored by regeneration of arginyl residues, suggesting an apolipoprotein-mediated suppressive mechanism. The total lipid extract of HDL2 stimulated 3-hydroxy-3-methylglutaryl-CoA reductase 2-fold at 3 microgram lipid phosphorus/ml. Moreover, HDL2 and HDL3 both stimulated 3-hydroxy-3-methylglutaryl-CoA reductase activity in receptor-negative fibroblasts. HDL2 have both the ability to suppress 3-hydroxy-3-methylglutaryl-CoA reductase activity in cells which possess low density lipoproteins receptors and to activate the enzyme in receptor-negative cells. These results show that variations in culture conditions and differences in the proportions of HDL subclasses must be considered in the interpretation of studies investigating cellular responses to HDL.
Characterization of human high density lipoproteins by zonal ultracentrifugation.
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Dynamics of lipid motions in high-density lipoprotein subfractions HDL2 and HDL3: magnetic resonance studies.
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Effects of HDL subclasses on 3-hydroxy-3-methylglutaryl coenzyme A reductase in human fibroblasts.
Monolayer cultures of normal human fibroblasts were used to study the effects of the main subclasses of high-density lipoproteins, HDL2 and HDL3, on 3-hydroxy-3-methylglutaryl CoA reductase (EC 1.1.1.34) activity. In this system, HDL3 (d = 1.125-1.210 g/cm3) specifically induced HMG-CoA reductase activity. Evaluation of culture dynamics revealed that enzyme induction was restricted to the stationary phase of growth. When the cells were incubated with HDL2 (d = 1.063-1.125 g/cm3), suppression of reductase activity was observed. Mixtures of HDL2 and HDL3 suppressed reductase activity when HDL2 was greater than 35% of the total HDL. The suppressive effects of HDL2 were abolished by treatment with cyclohexanedione and restored by regeneration of the arginyl residues, suggesting an apoprotein-mediated suppressive mechanism. These observations show that the cellular effects of HDL depend upon the stage of cell growth and the ratio of HDL subclasses in HDL as usually isolated.
Effects of saturated and polyunsaturated fat diets on the chemical and physical properties of lipoproteins from type V hyperlipoproteinemic subjects.
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The measurement of apolipoprotein A-I in human plasma by electroimmunoassay.
A variable proportion of the total apolipoprotein A-I (apo A-I) present in plasma or high density lipoproteins (HDL) is normally detectable by immunochemical methods. This has been attributed to masking of some of the immunoreactive sites of apo A-I by lipid in the intact HDL particle. This difficulty has been circumvented by heating or delipidation. We find that exposure of plasma to concentrations of urea greater than about 7.0 M in the barbital buffer used to dilute plasma samples for estimation by electroimmunoassay enables the complete detection of Apo A-I, as judged by comparison with samples delipidated with tetramethylurea. The need for time-consuming heating or delipidation is avoided.
Influence of an atherogenic diet on the structure of swine low density lipoproteins.
Five groups of three swine each were fed a basal diet supplemented with 15% tallow and either 0.0, 1.0, 1.5, 2.0%, or 2.5% cholesterol. The animals were studied over a period of 9 weeks to observe changes in plasma lipids and low density lipoproteins (LDL). At the end of the study period, LDL was analyzed by rate zonal ultracentrifugation, characterized chemically, and examined by differential scanning calorimetry. Within 3 weeks of initiation of the cholesterol-supplemented diets, there was an increase in the plasma levels of cholesterol and total LDL. LDL from swine fed the basal diet and the basal diet plus 1.0% cholesterol appeared in two LDL populations (LDL1 and LDL2) when analyzed by rate zonal ultracentrifugation. After diets containing 1.5, 2.0, and 2.5% dietary cholesterol, there was an increase in the mean flotation rate of total LDL which shifted to a lower density. LDL1 from the high cholesterol diets had a decreased triglyceride content when compared to those of the low cholesterol diets. When examined by differential scanning calorimetry, the LDL1 from the animals fed at least 1.5% cholesterol had phase transitions above body temperature, whereas the LDL from those fed 0 and 1.0% cholesterol had phase transitions below 37 degrees C. In contrast, the thermal behavior and fatty acid compositions of the extracted cholesteryl esters of the LDL obtained after the five different diets were not remarkably different. Since LDL triglyceride and cholesteryl esters are predicted to coexist in a common phase in the LDL core, the different thermal behavior of the LDL obtained after diets with different cholesterol contents is due to differences in triglyceride content which are a secondary effect of cholesterol-feeding. From these data we conclude that dietary cholesterol increases plasma LDL content, decreases LDL triglyceride content, and alters the particle structure. These changes in lipoprotein structure may contribute to the known development of atherosclerosis in cholesterol-fed swine.