The safety and efficacy of oral docosahexaenoic acid supplementation for the treatment of primary sclerosing cholangitis - a pilot study.

BACKGROUND: Primary sclerosing cholangitis (PSC) is characterised by progressive inflammatory and fibrotic destruction of the biliary ducts. There are no effective medical therapies and presently high dose ursodeoxycholic acid is no longer recommended due to significant adverse events in a recent clinical trial. Cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction is associated with PSC in both children and adults. Since CFTR dysfunction leads to altered fatty acid metabolism, specifically reduced docosahexaenoic acid (DHA), we hypothesised that DHA supplementation might be an effective therapy for patients with PSC. AIM: To determine the safety and efficacy of oral DHA supplementation for the treatment of PSC. METHODS: We conducted a 12 month open-label pilot study to evaluate safety of oral DHA and its effects on serum alkaline phosphatase as a primary outcome measure in 23 patients with PSC. DHA was administered orally at 800 mg twice per day. Secondary outcomes included changes in other liver function tests and fibrosis biomarkers. RESULTS: A 1.7-fold increase in serum DHA levels was observed with supplementation. The mean alkaline phosphatase level (±S.E.) at baseline was 357.8 ± 37.1 IU compared to 297.1 ± 23.7 IU (P < 0.05) after 12 months of treatment. There were no changes in other liver function tests and fibrosis biomarkers. No adverse events were reported. CONCLUSIONS: Oral DHA supplementation is associated with an increase in serum DHA levels and a significant decline in alkaline phosphatase levels in patients with PSC. These data support the need for a rigorous trial of DHA therapy in PSC.

Ethyl arachidonate is the predominant fatty acid ethyl ester in the brains of alcohol-intoxicated subjects at autopsy.

The role of fatty acid ethyl esters (FAEE), the nonoxidative ethanol metabolites, as mediators of alcohol-induced organ damage is increasingly being recognized. FAEE are detectable in the blood and in liver and adipose tissue after ethanol ingestion, and on that basis, FAEE can be used as markers of ethanol intake. In this study, 10 samples of human brain were collected at autopsy at the Massachusetts Medical Examiner's Office and analyzed for FAEE. FAEE were isolated and quantified as mass per gram of wet weight. The blood ethanol level was also obtained in each case along with the other drugs detected in routine postmortem toxicology screening tests. Ethyl arachidonate was the predominant FAEE species in the brain, representing up to 77.4% of total FAEE in the brain. The percent age of ethyl arachidonate of the total FAEE in the brain was significantly higher than what has been found in all other organs and tissues previously analyzed. Linoleate, the precursor of arachidonate, was a poor substrate for FAEE synthesis, as the percentage of ethyl linoleate of the total FAEE content was extremely low. Thus, this reflects preferred incorporation of arachidonate into newly synthesized FAEE in the brain. Since arachidonate is derived from linoleate, which is depleted in FAEE while arachidonate is enriched, the synthesis of FAEE may be linked to the desaturation and elongation of linoleate to arachidonate.

Fatty acid ethyl esters in liver and adipose tissues as postmortem markers for ethanol intake.

BACKGROUND: Fatty acid ethyl esters (FAEEs) are nonoxidative metabolites of ethanol. FAEEs are found in liver, pancreas, and adipose tissues up to 24 h after consumption of ethanol, and on that basis, they are potentially useful markers for ethanol intake. In this study with rats, we investigated the efficacy of using FAEEs in liver and in adipose tissue as postmortem markers for premortem ethanol ingestion. METHODS: An animal study was conducted in which test rats received injections of ethanol and control rats received injections of normal saline. The rats were killed 2 h after the injections. The bodies of the animals were stored at 4 degrees C up to 12 h, and samples of liver and adipose tissues were collected at different time intervals and processed for FAEE quantification. In another set of experiments, the rats received injections and were killed as described above, but bodies of animals from both groups were stored at 4, 25, or 37 degrees C for up to 72 h, and liver samples were collected and processed for FAEE quantification. RESULTS: FAEEs were detected up to 12 h after death in liver and adipose tissue samples from the bodies of ethanol-treated animals stored at 4 degrees C; negligible amounts were detected in the bodies of animals that received normal saline. Adipose tissues contained higher amounts of FAEEs than liver, as well as more species: eight FAEE species in adipose tissue and five in liver tissue. Higher concentrations of FAEEs were detected in livers of treated animals stored at 25 degrees C for up to 48 h than in livers of controls stored under the same conditions. CONCLUSIONS: For at least 12 h after death, FAEEs in liver and adipose tissues are useful postmortem markers of premortem ethanol ingestion.

Differences in the fatty acid composition of fatty acid ethyl esters in organs and their secretions.

BACKGROUND: Fatty acid ethyl esters (FAEE) are nonoxidative ethanol metabolites that have been shown to be long term markers of ethanol intake and have been implicated as mediators of ethanol-induced cell injury. Previous studies have indicated that the fatty acid composition of the FAEE found in the plasma of human subjects after ethanol ingestion is predominantly ethyl palmitate and ethyl oleate. This raised the possibility that there is some selectivity toward the fatty acid used for FAEE to be exported from the liver into the blood. METHODS: To address the hypothesis that the fatty acid composition of FAEE secreted from organs, such as the liver and pancreas, differs from the fatty acid composition of FAEE in the organs, this study was performed using rats that received ethanol by intra-arterial infusion. RESULTS: It was found that the fatty acids in FAEE differed significantly in plasma versus liver, bile versus liver, and pancreatic secretions versus pancreas. CONCLUSIONS: These results indicate that organs selectively export certain FAEE species.

Preanalytical variables affecting the quantification of fatty acid ethyl esters in plasma and serum samples.

BACKGROUND: Fatty acid ethyl esters (FAEEs) are cytotoxic nonoxidative ethanol metabolites produced by esterification of fatty acids and ethanol. FAEEs are detectable in blood up to 24 h after ethanol consumption. The objective of this study was to assess the impact of gender, serum or plasma triglyceride concentration, time and temperature of specimen storage, type of alcoholic beverage ingested, and the rate of ethanol consumption on FAEE concentrations in plasma or serum. METHODS: For some studies, subject were recruited volunteers; in others, residual blood samples after ethanol quantification were used. FAEEs were isolated by solid-phase extraction and quantified by gas chromatography-mass spectrometry. RESULTS: For weight-adjusted amounts of ethanol intake, FAEE concentrations were twofold greater for men than women (P /=24 h. The type of alcoholic beverage and rate of consumption did not affect FAEE concentrations. CONCLUSION: These studies advance plasma and serum FAEE measurements closer to implementation as a clinical test for ethanol intake.

The pool of fatty acids covalently bound to platelet proteins by thioester linkages can be altered by exogenously supplied fatty acids.

The goals of this investigation were, first, to develop a chemical strategy to identify and quantitate the mass of fatty acid which is covalently bound to proteins by thioester linkage in unactivated platelets, and, second, to determine whether exogeneously added fatty acids can alter the fatty acid composition of thioester bound fatty acids. Studies with radiolabeled fatty acids cannot identify and quantitate the actual fatty acids bound to proteins because they permit analysis of only the radiolabeled fatty acids added and their metabolites. Therefore, in the absence of metabolic labeling by radiolabeled fatty acids, we isolated the thioester-linked fatty acids from platelet proteins using hydroxylamine at neutral pH to form fatty acid hydroxamates. The hydroxamates were subsequently converted to fatty acid methyl esters by acid methanolysis for quantitation by gas chromatography-mass spectrometry. Using platelet specimens from 14 subjects, 74% of the fatty acid recovered from the unactivated platelet proteins as thioester linked was palmitate. Importantly, however, 22% was stearic acid, and oleate was 4% of the total thioester bound fatty acid. There was minimal variability (2.6-fold at maximum) between the subjects in the amount of the thioester-linked palmitate and thioester-linked stearate. However, there was substantial variability (>100-fold at maximum) between subjects in the amount of thioester-linked oleate. We also demonstrated that incubation of platelets with exogenous fatty acids can alter the profile of fatty acids bound to platelet proteins by thioester linkages. Incubation of platelets with 100 microM palmitate for 3 h increased the amount of thioester-linked palmitate by up to 26%, and incubation of platelets with 100 microM stearate increased the amount of thioester-linked stearate up to 30%. In support of the observation that radiolabeled fatty acids other than palmitate were shown to be capable of binding to platelet proteins by thioester linkage, our results indicate that the fatty acids actually bound to unactivated platelet proteins include a significant amount of stearate, and variable amounts of oleate, as well as palmitate. In addition, the data show that palmitate and stearate can be increased, as a percentage of total protein-bound fatty acid, by incubation with exogenous palmitate and stearate, respectively.

Quantitation of the mass of fatty acid ethyl esters synthesized by Hep G2 cells incubated with ethanol.

Fatty acid ethyl esters (FAEE), esterification products of fatty acids and ethanol, have been increasingly implicated as mediators of ethanol-induced organ damage. The first goal of this study was to determine the mass of FAEE synthesized by Hep G2 cells exposed to a given dose of ethanol. The second goal was to determine whether all fatty acids in cells are equally available for FAEE synthesis. Hep G2 cells and essential fatty acid deficient Hep G2 cells (Hep G2-EFD) were used to study the synthesis of FAEE upon exposure to ethanol. A two-pool fatty acid model was created: (1) a "previously incorporated pool" formed by incubating the cells with 14C-labeled fatty acids for 24 hr; and (2) a "newly incorporated pool" formed by incubating cells with 3H-labeled fatty acids for 0.5 hr. The FAEE production from each pool was then determined. The total production of FAEE within 3 hr by Hep G2 cells in culture was 150 to 250 pmol/mg cell protein. The fatty acids most recently incorporated into the cells were preferred as substrates for FAEE synthesis because a higher percentage of fatty acids from the newly incorporated pool was used for FAEE synthesis than from the previously incorporated pool. Furthermore, a dose-response relationship was observed between the amount of fatty acid in the newly incorporated pool and FAEE production, but not between the amount of fatty acid in the previously incorporated pool and FAEE synthesis. Taken together, the results indicate that a relatively small amount of endogenously synthesized FAEE is generated from specific intracellular pools of fatty acid since not all fatty acids are equally available for FAEE synthesis. This indicates that if endogenous FAEE are toxic, they exert their toxic effect at very low intracellular FAEE concentrations.

Fatty acid ethyl esters in the blood as markers for ethanol intake.

OBJECTIVE: To determine the clinical utility of fatty acid ethyl esters (FAEEs) in the blood as a short-term confirmatory marker for ethanol intake and a longer-term marker for ethanol intake after ethanol is no longer detectable. DESIGN: Single-center controlled clinical trial and a blinded comparison involving 48 blood samples that were positive, negative, or equivocal for blood ethanol. PARTICIPANTS: Seven healthy subjects (4 men and 3 women, aged 21 to 23 years) participated in the clinical trial. Blood samples from participants for the blinded comparison portion of the study were numbered from 1 to 48 and not identified by name. INTERVENTION: The 7 healthy subjects ingested a known amount of ethanol at a fixed rate. The concentration of FAEEs in the blood after ethanol intake was determined for a period of up to 24 hours. There was no intervention in the blinded comparison study. MAIN OUTCOME MEASURES: In the clinical trial, a pharmacokinetic analysis of FAEE concentration in the blood after ethanol intake was completed for 7 individuals whose blood ethanol level was elevated from 25 to 35 mmol/L. In the blinded comparison, the 48 blood samples that were positive, negative, or equivocal for blood ethanol were analyzed for FAEE concentration. RESULTS: In the clinical trial, the disappearance of FAEEs from the blood followed a decay curve that initially resembled the decay curve for blood ethanol. However, because of a very slow secondary elimination phase, the FAEEs were found to persist in the blood for at least 24 hours after ethanol intake was completed. In the blinded comparison, all 20 samples that were positive for ethanol were positive for FAEEs, 7 of 7 samples equivocal for ethanol were positive for FAEEs, and 21 of 21 negative samples for ethanol were negative for FAEEs. CONCLUSIONS: Serum concentration of FAEEs can serve as an excellent short-term confirmatory test for ethanol intake as well as a longer-term marker of ethanol ingestion. Measurement of FAEEs in the blood may be a more sensitive indicator of ethanol ingestion than the measurement of blood ethanol .

Fatty acid ethyl esters are present in human serum after ethanol ingestion.

The aim of the study was to determine whether fatty acid ethyl esters, nonoxidative products of ethanol metabolism selectively present in organs damaged by ethanol abuse, are detectable in the serum after ethanol ingestion. Serum samples of hospital emergency room patients with positive (n = 32) and negative (n = 5) blood ethanol levels were assayed for fatty acid ethyl esters. In a separate study, five healthy subjects received an ethanol dose based on body weight mixed with fruit juice in a 1:2 ratio and administered by measured ingestion. Fatty acid ethyl esters were found in the serum of hospital emergency room patients with positive blood ethanol levels. The concentration of fatty acid ethyl esters in these patients correlated with the concentration of blood ethanol (r = 0.57; 95% confidence interval 0.28 to 0.77; P = 0.0002). In the controlled ethanol ingestion study with five healthy subjects, it was also determined that the serum fatty acid ethyl ester concentration began to decrease within 2 h of the time ethanol ingestion had been stopped. The fatty acid ethyl esters in the serum were bound to lipoprotein and albumin, and there was a higher percentage of saturated fatty acids in the FAEE pool than in the serum free fatty acid and triglyceride pools. These studies indicate that fatty acid ethyl esters, which have been implicated as mediators of ethanol-induced organ toxicity, are present in serum after ethanol ingestion.

Alcohol delays clearance of lipoproteins from the circulation.

Long-term (18-month) consumption of high-dose ethanol ([EtOH] 24% of total calories) by squirrel monkeys results in marked elevations in plasma antiatherogenic high-density lipoprotein (HDL) cholesterol and apolipoprotein (apo) A-1, and atherogenic low-density lipoprotein (LDL) cholesterol and apo B. In an effort to determine whether alterations in lipoprotein turnover could explain the above findings, 131I-HDL apo A-1 and 125I-LDL apo B were injected into EtOH and control animals, following which in-vivo catabolic and production rates were determined. For both lipoproteins, synthetic rates were unaltered, while fractional catabolic rates (FCR) were significantly reduced in EtOH monkeys. Results from this study implicate EtOH-induced changes in hepatic metabolism as the basis for delayed lipoprotein clearance and hence elevated plasma apolipoprotein levels.

Alcohol dose and low density lipoprotein heterogeneity in squirrel monkeys (Saimiri sciureus).

The present study was designed to determine whether normolipidemic male squirrel monkeys (Saimiri sciureus) exhibit low density lipoprotein (LDL) heterogeneity similar to that observed in humans and if present, whether LDL subfractions are altered by consumption of low vs. high dose ethanol (EtOH). Primates were divided into three groups designated control, low, and high EtOH and fed isocaloric liquid diets containing 0%, 12% and 24% of calories as EtOH, respectively, for 6 months. The 12% EtOH caloric level resulted in a modest, non-significant increase in high density lipoprotein (HDL) cholesterol and no change in LDL cholesterol or plasma apolipoprotein B (apo B), while the 24% dose produced significant elevations in plasma, LDL and HDL cholesterol and apo B. Using a single-spin density gradient ultracentrifugation procedure developed for humans, three distinct LDL subclasses designated LDL1a (d = 1.031 g/ml), LDL1b (d = 1.038 g/ml) and LDL 2 (d = 1.046 g/ml) were isolated from all three treatment groups. Monkey LDL subfractions were nearly identical to very light, light and heavy LDL subspecies isolated from human plasma in terms of their: (1) isopycnic densities following ultracentrifugation; (2) co-migration as single bands with beta-electrophoretic mobility in cellulose acetate and agarose electrophoretic gels; (3) size-dependent migration pattern in polyacrylamide gradient electrophoretic gels; (4) co-migration as a single band corresponding to apo B-100, following SDS polyacrylamide gel electrophoresis; and (5) decrease in total cholesterol/protein ratios with increasing LDL subclass density. Although there were no treatment differences in LDL particle size, within each treatment group, mean particle size for each LDL subfraction was significantly different from every other subfraction. Low (12%) dose alcohol had no effect on LDL subfraction mass relative to controls while high alcohol consumption resulted in marked increases in all lipid (except triglyceride) and protein of the larger, buoyant LDL subspecies (LDL1a and LDL1b). Moreover, the best correlation between plasma apo B and LDL subfraction total mass was demonstrated with LDL1b (r = 0.735). Since neither the lipid nor the protein concentration of the small, dense, purportedly more atherogenic, LDL2 changed with the 24% EtOH dose, we propose that the LDL subfraction alterations associated with high alcohol intake in squirrel monkeys (increased LDL1a, increased LDL1b, LDL2 no effect) may represent a compensatory response to modulate the overall atherogenic lipoprotein profile associated with elevations in total LDL cholesterol and plasma apolipoprotein B.

Alcohol produces dose-dependent antiatherogenic and atherogenic plasma lipoprotein responses.

A comprehensive assessment of lipoprotein compositional/metabolic response to incremental caloric ethanol (EtOH) doses ranging from low to moderate to high was undertaken using male squirrel monkeys. Control monkeys were maintained on a chemically defined, isocaloric liquid diet, while experimental primates wee fed increasing doses of alcohol (6, 12, 18, 24, 30, and 36% of energy) substituted isocalorically for carbohydrate at 3-month intervals. Liver function tests and plasma triglyceride were normal for all animals. Plasma cholesterol showed a transient increase at the 12% caloric dose that was attributed solely to an increase in high density lipoprotein (HDL). A more pronounced increase in plasma sterol, beginning at 24% and continuing to 36% EtOH, was the result of increments in both HDL and low density lipoprotein (LDL) cholesterol, although the contribution by the latter was substantial primarily at the 36% dose. Plasma apolipoprotein elevations (HDL apolipoprotein A-I, LDL apolipoprotein B) generally accompanied the lipoprotein lipid increases, although the first atherogenic response for LDL became manifest as a significant increase in apolipoprotein B at 18% EtOH calories. Postheparin plasma lipoprotein lipase was not affected by dietary alcohol, whereas hepatic triglyceride lipase activity showed significant increases at higher (24 and 36%) EtOH doses. Plasma lecithin-cholesterol acyltransferase activity was normal at the 6 and 12% EtOH doses, but exhibited a significant reduction beginning at 18% and continuing to 36% EtOH. Alterations in these key lipoprotein regulatory enzymes may represent the underlying metabolic basis for the observed changes in lipoprotein levels and our earlier findings of HDL2/HDL3 subfraction modifications. Results from our study indicate that in squirrel monkeys, moderate (12%) EtOH caloric intake favors an antiatherogenic lipoprotein profile (increases HDL, normal LDL levels, and lecithin-cholesterol acyltransferase activity), whereas higher doses (24-36%) produce both coronary-protective (increases HDL) and atherogenic (increases LDL) responses. Moreover, the 18% EtOH level represents an important transition dose which signals early adverse alterations in lipoprotein composition (increases apolipoprotein B) and metabolism (decreases lecithin-cholesterol acyltransferase).

Effect of ethanol on low density lipoprotein and platelet composition.

The present study was designed to investigate the effect of ethanol (EtOH) dose on low density lipoprotein (LDL) and platelet composition. Male squirrel monkeys were divided into three groups designated Control, Low, and High EtOH, and fed isocaloric liquid diets containing 0%, 12%, and 24% of calories as EtOH, respectively. After four months of treatment, monkeys fed the 12% alcohol dose had LDL and platelet cholesterol concentrations similar to Controls. By contrast, platelet membranes from High EtOH animals contained significantly more cholesterol which was associated with higher levels of plasma LDL cholesterol and apolipoprotein B. Blood platelet count, size, and mass were similar for all groups and circulating platelet aggregates were absent in the two alcohol cohorts. Despite elevations in platelet cholesterol mass and thromboxane A2 (TXA2) precursor, phospholipid arachidonate, platelet responsiveness, measured as thromboxane formed in response to a collagen challenge in vitro, and the cholesterol/phospholipid molar ratio, were not significantly altered by high dose alcohol. Normal platelet activity in High EtOH monkeys may have resulted from a significant increase in the platelet phospholipid polyunsaturated/saturated fatty acid ratio and a non-significant increase in platelet phospholipid mass, both of which would have a fluidizing effect on platelet membranes. Our data indicate that low EtOH intake has no effect on platelet composition and function while unfavorable platelet cholesterol enrichment following consumption of high dose ethanol may arise from elevations in plasma LDL.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of drinking pattern on plasma lipoproteins and body weight.

The effect of drinking pattern on plasma lipoproteins and body weight was examined in three groups of squirrel monkeys: (1) controls fed isocaloric liquid diet; (2) regular drinkers given liquid diet containing ethanol (EtOH) substituted isocalorically for carbohydrate at 12% of calories daily; and (3) binge drinkers fed 6% EtOH calories daily for a four-day period followed by three days of 20% EtOH to mimic a weekend bout drinking cycle. The number of calories offered per day was the same for all groups, and the average weekly EtOH consumption (12% calories) was identical for the two alcohol treatments. The entire study lasted six months. There were no significant differences in plasma cholesterol, triglyceride or liver function tests. Regular drinkers had the highest high density lipoprotein2/high density lipoprotein3 (HDL2/HDL3) protein and apolipoprotein A-I/B ratios of any group and exhibited a significant elevation in the molar plasma lecithin:cholesterol acyltransferase (LCAT) rate (nmol/min/ml). Binge drinking produced a selective increase in low density lipoprotein (LDL) cholesterol and apolipoprotein B, and a depression in the fractional LCAT rate (% esterified/min). During the course of the study, controls ate 92% of their diet while the alcohol groups each consumed 95% of the liquid diet. Despite this difference, body weight and Quetelet index (weight/height2) decreased progressively in the order controls greater than regular drinkers greater than binge drinkers. Results from our study indicate that moderate, regular daily consumption of EtOH at 12% of calories causes a modest reduction in body weight and produces a coronary protective lipoprotein profile (increases HDL2/HDL3, increases apolipoprotein A-I/B, low LDL cholesterol). By contrast, when this same average weekly dose is concentrated in a binge cycle, unfavorable alterations in lipoprotein composition (increases LDL cholesterol, increases apolipoprotein B) and metabolism (decreases LCAT activity) occur along with weight loss and depletion of body fat. These studies point to the value of the squirrel monkey model in evaluating both favorable and pathophysiological effects of chronic EtOH intake.

Plasma lipoprotein alterations in squirrel monkeys (Saimiri sciureus) during ethanol administration and abstinence.

The time course of lipoprotein changes during ethanol (EtOH) consumption followed by abstinence was examined in 3 groups of male squirrel monkeys: 1) controls fed isocaloric liquid diet; 2) low EtOH monkeys given liquid diet with vodka substituted isocalorically for carbohydrate at 12% of calories; and 3) high EtOH animals fed diet plus vodka at 24% of calories. After 2 weeks, high EtOH monkeys showed significant elevations in total plasma cholesterol which continued to increase at 4 weeks and then declined at 8 weeks. These elevations were the result of increases in both low density (LDL)- and high density lipoprotein (HDL)-cholesterol. Low EtOH monkeys had a modest increase in total cholesterol throughout 8 weeks which was attributed to increments in HDL-cholesterol alone. During abstinence, total, HDL- and LDL-cholesterol concentrations decreased rapidly in the high EtOH group and were similar to control values after 4 days. HDL-cholesterol showed a more gradual decline in animals fed 12% EtOH while LDL-cholesterol remained low and not significantly different from controls. Liver function tests were normal for all animals. Our results indicate that low-dose EtOH favors a coronary protective lipoprotein profile (increases HDL, decreases LDL) in squirrel monkeys while the higher alcohol regimen causes both favorable and unfavorable alterations in plasma lipids which quickly revert to control levels during abstinence.

Effect of ethanol dose on low density lipoproteins and high density lipoprotein subfractions.

Male squirrel monkeys were fed increasing caloric percentages (0, 12, 24, and 36%) of ethanol (ETOH) substituted isocalorically for carbohydrate as part of a chemically defined liquid diet to assess how alcohol dose modifies plasma lipoproteins and liver function. A separate group of primates was used to define the dose at which elevations in plasma apolipoprotein B first occurred and to measure plasma alcohol levels. ETOH caused a dose-related, linear increase in high density lipoprotein (HDL) cholesterol which was primarily the result of increments in coronary protective HDL2 cholesterol. HDL2 total mass (lipid + protein) followed the pattern of HDL2 cholesterol. Animals fed the 12% regimen had plasma ETOH levels of approximately 49 mg/dl, the lowest low density lipoprotein (LDL) cholesterol, and the highest HDL2/HDL3 cholesterol ratio. Significant elevations in apolipoprotein B first appeared at 18% ETOH while higher doses (24 and 36%) caused increases in LDL cholesterol and HDL3, reduced HDL2/HDL3 ratios, and plasma alcohol levels of 142 and 202 mg/dl, respectively. Liver function tests were normal for all animals. Our results indicate that while a moderate ETOH caloric intake (12%) produces an antiatherogenic lipoprotein profile (decreases LDL/HDL, increases HDL2/HDL3), any coronary protection afforded by continued increases in HDL2 at higher doses may be attenuated by concurrent atherogenic alterations (increases LDL cholesterol, increases apolipoprotein B).

Ethanol-induced alterations in lecithin: cholesterol acyltransferase (LCAT) activity in vitro.

Our recent experiments demonstrated that squirrel monkeys fed ethanol (ETOH) at 12% of calories (Low ETOH) had significantly higher plasma lecithin: cholesterol acyltransferase (LCAT) activity than monkeys fed ETOH at 24% of calories (High Ethanol). Control animals had LCAT activity intermediate between that of Low and High ETOH primates. To test whether alcohol directly altered cholesterol esterification in vitro, LCAT activity was measured in pooled primate plasma incubated with ETOH at final concentrations of 60, 80, 160, and 240 mg/dl. A similar experiment was performed using incremental doses of ETOH's major metabolite, acetaldehyde. Peak cholesterol esterification occurred at 60 mg/dl which was comparable to plasma alcohol levels detected in Low ETOH monkeys (63 mg/dl) while LCAT activity was significantly depressed at 160 mg/dl which was similar to blood ETOH monitored in High ETOH primates (159 mg/dl). Maximum cholesterol esterification occurred at an acetaldehyde concentration of 0.45 mumoles/l. Our data indicate that ETOH can either stimulate or inhibit LCAT activity in vitro depending upon concentration and suggest that circulating blood alcohol may induce similar alterations in cholesterol esterification in vivo.