Bile acid and phospholipid fatty acid composition in bile of patients with cholesterol and pigment gallstones.

It has been previously reported that patients with cholesterol gallstones have increased biliary deoxycholate and arachidonate content as compared with normal subjects without gallstones. Increased biliary deoxycholate and arachidonate content might be a primary factor in the pathogenesis of cholesterol gallstones or merely an epiphenomenon due to the presence of gallstones. We therefore compared biliary bile acid composition in 46 patients with cholesterol gallstones and 22 patients with pigment stones. In addition, biliary phospholipid fatty acid composition was determined in 44 of these patients (30 cholesterol and 14 pigment stone patients). No significant differences were detected. In particular, the percentage deoxycholic acid (mean +/- SD: 20.3 +/- 8.8% and 21.5 +/- 10.9% respectively) and the percentage arachidonic acid (4.4 +/- 2.0% and 4.5 +/- 2.2%, respectively) were very similar. A significant correlation between age and biliary cholesterol saturation index was found only for the group of patients with pigment stones (R = 0.52, p less than 0.02). In conclusion, the present study does not support a primary role for increased biliary deoxycholic acid or arachidonic acid in the pathogenesis of cholesterol gallstones.     

Renal function in rats with essential fatty acid deficiency

1. Renal function was studied in 50-, 70-, 90- and 200-day-old rats with essential fatty acid deficiency. The pharmacokinetics of tobramycin was investigated in 90-day-old essential fatty acid-deficient rats. 2. A higher glomerular filtration rate and a higher serum concentration of urea were seen in 50-day-old essential fatty acid-deficient rats compared with age-matched controls. Later, the glomerular filtration rate progressively deteriorated in parallel with a decline in effective renal plasma flow and with a concomittant rise in serum levels of urea and creatinine. The serum concentration of protein was lower in the rats with essential fatty acid deficiency and that of sodium was higher than in the control rats. The non-renal clearance of tobramycin was increased in the rats with essential fatty acid deficiency. 3. The early hyperfiltration in essential fatty acid-deficient rats with the subsequent fall in glomerular filtration rate, which was paralleled by a rise in serum levels of urea and creatinine, as well as the increased non-renal clearance of tobramycin, are in accordance with the clinical manifestations of cystic fibrosis. Rats with essential fatty acid deficiency might be a useful model with which to study the pathophysiological renal changes in cystic fibrosis related to the progressive essential fatty acid deficiency in this disease.     

Trace metal and essential fatty acid deficiency during total parenteral nutrition.

While deficiences of trace metals and essential fatty acids are rare in humans fed orally, the widespread use of total parenteral nutrition (TPN) has increased the likelihood of encountering these deficiences. A 14-year-old boy, with total villous atrophy of the small intestine, suffered from severe malnutrition and was placed on a conventional TPN regimen. Although not immediately recognized, he rapidly developed deficiencies of zinc, copper and essential fatty acids. Careful monitoring of the course of the illness and the responses to sequential treatments with zinc, lipid, and copper given intravenously allowed examination of the effects of the deficiencies on skin, intestine, liver, and hemopoietic systems and helped to establish normal requirements for the metals. The progress of the illness suggested that patients with intestinal malabsorption may be especially at risk of developing some of these lesser known complications of TPN.     

Effect of chenodeoxycholic acid and phenobarbital on the rate-limiting enzymes of hepatic cholesterol and bile acid synthesis in patients with gallstones.

The effects of chenodeoxycholic (CDC), 750 mg. per day, phenobarbital (PB), 90 or 180 mg., combined (CDC + PB), and placebo on biliary lipid composition and on the rate-limiting enzymes of hepatic cholesterol synthesis (HMG-CoA reductase) and bile acid synthesis (cholesterol 7alpha-hydroxylase) were studied. Percutaneous liver biopsies were performed after 6 months of therapy in 4 patients from each group participating in a double-blind study of gallstone dissolution. The enzyme activities were also assayed in liver obtained at laparotomy in 7 untreated gallstone patients and 4 without gallstones. 7alpha,12alpha-Dihydroxycholest-4-en-3-one-12alpha-hydroxylase, an enzyme leading to cholic acid synthesis, was determined in 4 untreated gallstone patients and 4 without gallstones. Untreated gallstone patients had 35 per cent greater HMG-CoA reductase (p less than 0.01), 37 per cent less 7alpha-hydroxylase (p less than 0.01), and 40 per cent less 12alpha-hydroxylase (p less than 0.01) than patients without gallstones. CDC, PB, and both increased biliary CDC and decreased the lithogenic index significantly (p less than 0.01) but saturated bile persisted with PB. CDC decreased HMG-CoA reductase 40 per cent (p less than 0.01) and 7alpha-hydroxylase 47 per cent (p less than 0.01). PB increased HMG-CoA reductase 112 per cent (p less than 0.01) and 7alpha-hydroxylase 20 per cent (p less than 0.01). The combination of CDC and PB increased HMGCoA reductase 40 per cent (p less than 0.01) and had no effect on 7alpha-hydroxylase. In conclusion, CDC induced desaturation of bile while decreasing HMG-CoA reductase and increasing CDC in bile. PB reduced the saturation less effectively than CDC; it increased 7alpha-hydroxylase but also increased HMG-CoA reductase.     

Effects of dietary cholesterol on cholesterol and bile acid homeostasis in patients with cholesterol gallstones.

We examined changes in cholesterol and bile acid metabolism produced by dietary cholesterol in gallstone subjects and matched controls. Healthy women were recruited and, after confirming the presence or absence of radiolucent gallstones, they were studied on regular diets and again on the same diet supplemented with five eggs daily for 15-18 d. Studies included plasma lipids, lipoproteins and apolipoproteins, dietary records, cholesterol absorption, cholesterol synthesis, plasma clearance of chylomicron remnants, biliary lipid composition, and secretion and bile acid kinetics. On low cholesterol, gallstone subjects absorbed a slightly lower fraction of dietary cholesterol, synthesized more cholesterol, and had smaller bile acid pools and faster fractional turnover rate (FTR) of bile acids. On high cholesterol, the fraction of cholesterol absorbed decreased in both groups and cholesterol synthesis decreased, especially in the gallstone group. Biliary cholesterol secretion increased in the gallstone group only. FTR of bile acids did not change in either group. Bile acid synthesis and pool tended to increase (P = NS) in the controls, but in gallstone subjects, synthesis and pool size decreased. We concluded that in gallstone subjects cholesterol and bile acid homeostasis is significantly altered, and that increasing dietary cholesterol increases biliary cholesterol secretion and decreases bile acid synthesis and pool, changes associated with cholesterol gallstone formation.     

A possible role for malonyl-CoA in the regulation of hepatic fatty acid oxidation and ketogenesis.

Studied on the oxidation of oleic and octanoic acids to ketone bodies were carried out in homogenates and in mitochondrial fractions of livers taken from fed and fasted rats. Malonyl-CoA inhibited ketogenesis from the former but not from the latter substrate. The site of inhibition appeared to be the carnitine acyltransferase I reaction. The effect was specific and easily reversible. Inhibitory concentrations were in the range of values obtained in livers from fed rats by others. It is proposed that malonyl-CoA functions as both precursor for fatty acid synthesis and suppressor of fatty acid oxidation. As such, it might be an important element in the carbohydrate-induced sparing of fatty acid oxidation.     

Long-chain polyunsaturated fatty acids in rat maternal milk, offspring brain and peripheral tissues in essential fatty acid deficiency.

Fatty acid status in humans is usually related to plasma or red blood cell fatty acid profiles. The aim of the study was to explore whether a maternal deficiency in dietary essential fatty acids would differentially affect lipid fractions in several tissues of the offspring, including brain. Female Wistar rats were fed an essential fatty acid-deficient diet during 3 months before mating. The fatty acid composition of different lipid fractions was examined in maternal milk, and in plasma, red blood cells, liver, adipose tissue, cerebral cortex and hippocampus of the offspring using thin layer and capillary column gas chromatography. Lipid fractions from most tissues of deprived offspring showed a common fatty acid profile characterized by elevated 20:3 omega9/20:4 omega6 ratio, and decreased docosahexaenoic acid and arachidonic acid. However, arachidonic acid was not affected in brain, even though 22:5 omega6 was increased in phospholipids of cerebral cortex and hippocampus. The present results demonstrate different degrees of resistance to essential fatty acid deficiency in lipid fractions and tissues. This suggests a priority distribution of arachidonic acid to preferential areas and shows that blood phospholipid fatty acids do not exactly reflect brain phospholipid status.     

2,4-Dienoyl-coenzyme A reductase deficiency: a possible new disorder of fatty acid oxidation.

Several inherited disorders of fatty acid beta-oxidation have been described that relate mainly to saturated precursors. This study is the first report of an enzyme defect related only to unsaturated fatty acid oxidation and provides the first in vivo evidence that fat oxidation in humans proceeds by the reductase-dependent pathway. The patient was a black female, presenting in the neonatal period with persistent hypotonia. Biochemical studies revealed hyperlysinemia, hypocarnitinemia, normal organic acid profile, and an unusual acylcarnitine species in both urine and blood. The new metabolite was positively identified by mass spectrometry as 2-trans,4-cis-decadienoylcarnitine, derived from incomplete oxidation of linoleic acid. In spite of dietary therapy, the patient died of respiratory acidosis at four months of age. Samples of liver and muscle from the autopsy were assayed for 2,4-dienoyl-coenzyme A reductase activity. Using the substrate 2-trans,4-cis-decadienoylcoenzyme A, the reductase activity was 40% of the control value in liver and only 17% of that found in normal muscle. It is suggested that unsaturated substrates should be used for in vitro testing to cover the full range of potential beta-oxidation defects and that acylcarnitine species identification be used for in vivo detection of this disorder.     

Diet-induced essential fatty acid deficiency in ambulatory patient with type I diabetes mellitus

We report a case of symptomatic essential fatty acid deficiency (EFAD) occurring in a free-living individual with type I diabetes mellitus who was voluntarily following a high-carbohydrate, fat-restricted diet. The patient was 43 yr old with type I diabetes for 18 yr and no chronic complications. His self-imposed diet excluded all red meats, fats, and oils. After several months of this diet, the patient developed lethargy and a pruritic, diffuse, scaly, and erythematous rash. Biochemical studies revealed a mildly elevated SGOT and abnormally low levels of linoleic, linolenic, and arachidonic fatty acids. Treatment with linoleic acid supplementation in his diet improved the rash, normalized SGOT, and corrected the fatty acid profile. We conclude that EFAD may occur in a free-living individual after consuming a very-low-fat diet.     

Biliary lipid composition in patients with cholesterol and pigment gallstones and gallstone-free subjects: deoxycholic acid does not contribute to formation of cholesterol gallstones

BACKGROUND: Four main disturbances have been attributed to cholesterol gallstone disease: hypersecretion of cholesterol from the liver with cholesterol supersaturation in bile; disturbed motility with defective absorption and secretion by the gallbladder; increased crystallisation of cholesterol in the gallbladder bile; and slow intestinal transit with increased amount of deoxycholic acid in the bile acid pool. We aimed to evaluate the biliary lipid composition in a large series of gallstone patients, with emphasis on the amount of deoxycholic acid and with respect to number of stones, compared to gallstone free subjects. MATERIALS AND METHODS: Bile was sampled during operations through puncture of the gallbladder from 145 cholesterol gallstone patients, 23 patients with pigment stones and 87 gallstone free patients undergoing cholecystectomy. Biliary lipid composition, cholesterol saturation, bile acid composition, nucleation time and cholesterol crystals were analysed. RESULTS: The patients with cholesterol gallstones showed higher molar percentage of cholesterol, lower total biliary lipid concentration, higher cholesterol saturation, shorter nucleation time and higher proportion of crystals in bile than the other groups. The nucleation time was significantly shorter in multiple cholesterol gallstone patients, but this was not due to higher cholesterol saturation. Male cholesterol gallstone patients showed higher cholesterol levels, lower total biliary lipid concentration, and higher cholesterol saturation in bile than female patients. There was no difference in biliary content of deoxycholic acid, but significantly lower content of cholic acid in gallstone patients compared to gallstone free patients. CONCLUSIONS: We conclude that deoxycholic acid does not contribute to gallstone formation in cholesterol gallstone patients. The short nucleation time in patients with multiple cholesterol stones is not due to higher cholesterol saturation.     

The development of essential fatty acid deficiency in healthy men fed fat-free diets intravenously and orally.

The hypothesis that clinical and biochemical essential fatty acid deficiency (EFA) might occur from the feeding of eucaloric, fat-free diets was tested in two experiments in healthy men. In Study I, eight men were given fat-free, eucaloric diets containing 80% of calories as glucose and 20% as amino acid hydrolysates by a constant drip over a 24-h period. The diets were fed in succession for periods of 2 wk each, either through a superior vena cava catheter or via a nasogastric tube. EFA deficiency was detected by decreases in linoleic acid and by the appearance of 5, 8, 11-eicosatrienoic acid in lipid fractions of plasma. Linoleic acid decreased significantly during 2 wk of the fat-free diet given intravenously from 48.8 to 9.8% (percent of total fatty acids) in cholesterol esters, from 21.2 to 3.2% in phospholipids, from 9.6 to 2.0% in free fatty acids, and from 14.1 to 2.6% in triglycerides. Eicosatrienoic acid, normally undetectable, appeared 0.6% in cholesterol esters, 2.5% in phospholipids, 0.2% in free fatty acids, and 2.3% in triglycerides. EFA deficiency occurred similarly during the nasogastric feeding. In Study II a subject received the same diet continuously by the nasogastric route for 10 days followed by a 24-h fast. He was then given the fat-free diet intermittently in three meals per day for 3 days. Finally, he was repleted with a diet containing 2.6% linoleic acid. By the 3rd day of the continuous nasogastric feeding, linoleic acid had fallen significantly and eicosatrienoic acid had appeared in plasma lipid fractions as in Study I. These findings were accentuated by day 10. Adipose tissue fatty acid composition did not change. Free fatty acid outflow from adipose tissue was presumably suppressed during the 10 days of continuous feeding. With increased free fatty acid outflow during fasting and intermittent feeding, linoleic acid rose and eicosatrienoic acid decreased. After 13 days of repletion with dietary linoleic acid, the EFA deficiency readily develops when fat-free diets containing glucose are given intravenously or orally as constant 24-h infusions. These diets are similar to the hyperalimentation formulas now being used clinically.     

Prevention of diabetes in the BB rat by essential fatty acid deficiency. Relationship between physiological and biochemical changes

Essential fatty acid (EFA) deficiency exerts a striking protective effect in several animal models of autoimmune disease. We now report that EFA deprivation prevents diabetes in the BB rat, an animal model of human insulin-dependent diabetes mellitus. In diabetes-prone (DP)-BB rats, the incidences of spontaneous diabetes and insulitis (the pathological substrate of autoimmune diabetes) were greatly reduced by EFA deficiency. This beneficial effect of the deficiency state was also seen in diabetes-resistant (DR)-BB rats that, after treatment with antibody to eliminate RT6+ T cells, would otherwise have become diabetic. The susceptibility of EFA-deprived DP-BB rats to spontaneous diabetes was restored when they were given dietary supplements of linoleate at 70 d of age (during the usual period of susceptibility), but not when they were repleted beginning at 120 d (after the peak incidence of diabetes). EFA deficiency did lead to growth retardation, but calorically restricted control rats demonstrated that the protective effect of the deficiency state was not a function of decreased weight. To examine the relationship between the biochemical changes of EFA deficiency and its physiological effects in this system, we compared the fatty acid changes that occurred in EFA-deficient animals that did and did not develop diabetes. Nondiabetic animals had significantly lower levels of (n-6) fatty acids (i.e., linoleate and arachidonate) and higher levels of oleate, an (n-9) fatty acid, than did diabetic animals. Levels of 20:3(n-9), the fatty acid that uniquely characterizes EFA deficiency, were similar in both groups, however. Among diabetic EFA-deficient rats, the age at onset of diabetes was found to correlate inversely with the level of (n-6) fatty acids, the least depleted animals becoming diabetic earliest, whereas there was no correlation with levels of 20:3(n-9). Among animals repleted with linoleate beginning at 70 d, restoration of susceptibility to diabetes correlated with normalization of the level of arachidonate. In summary, EFA deprivation reduced the frequency of diabetes in both DP and RT6-depleted DR-BB rats. This protective effect was strongly associated with depletion of (n-6) fatty acids, particularly arachidonate, but not with accumulation of the abnormal 20:3(n-9). Conjecturally, arachidonate and/or a metabolite may play a key role in mediating inflammatory injury in this animal model of autoimmune diabetes.     

Criteria for essential fatty acid deficiency in plasma as assessed by capillary column gas-liquid chromatography

To develop criteria for deficiency of essential fatty acids (EFA), we used capillary-column gas-liquid chromatography to determine fatty acids (percentage of total fatty acids) in plasma obtained in the fasting state from 56 reference subjects and from 10 patients with intestinal fat malabsorption and suspected EFA deficiency. Fatty acid evaluations (percentage of total fatty acids) that allowed for a clear distinction (P less than 0.01) between reference subjects and patients, based on values two standard deviations below or above the reference mean, included values for linoleic acid (18:2w6) below 27%, and values for palmitic acid (16:0), palmitoleic acid (16:1w7), oleic acid (18:1w9), vaccenic acid (18:1w7), and Mead acid (20:3w9) exceeding 21%, 2.6%, 23.3%, 2.1%, and 0.21%, respectively. Ratios of total EFA to total non-EFA of less than 0.60 and of Mead acid to arachidonic acid of greater than 0.025 also served to identify patients, and were not found in reference subjects. Significant inverse correlations between percentages of plasma EFA and plasma mono-unsaturated fatty acids were noted. Our reference-interval data can be used to assess normality of plasma EFA status.