The effect of antioxidant deficiency on tissue lipid composition in the rat. II. Liver

The hepatic phospholipids of the antioxidant-deficient rat fed a source of both linoleate and linolenate showed a progressive net decrease in eicosapentaenoate, a progressive net increase in arachidonate, and there was a concomitant accumulation of fluorescent pigment of the lipofuscin or ceroid type in the tissue. An increased incorporation of isotopically labeled acetate into both the tetraenoic and pentaplus hexaenoic acid fractions was also noted, indicating that the disappearance of polyunsaturated fatty acids was partially countered by increased synthesis. Comparable results were obtained on diets containing either suboptimum or adequate levels of biologically available selenium. Vesicular dilation of the endoplasmic reticulum was noted in animals fed the tocopherol-deficient diet. In separate experiments using a necrogenic diet containing torula yeast, these subcellular alterations were found to be prevented by tocopherol but not by selenium, although selenium supplementation did prevent macroscopically observable damage.     

The effect of antioxidant deficiency on tissue lipid composition in the rat. III. Testes

Production of testicular degeneration in the antioxidant-deficient rat resembles encephalomalacia in the chick in its dependence on essential (ω6) fatty acids and is distinct from the generalized response to all polyunsaturated fatty acids seen in nutritional muscular dystrophy in the rat. The nonessential (ω3) polyunsaturated fatty acids, however, lower the essential fatty acid content of the testicular lipids only slightly, are not themselves incorporated into this tissue to any appreciable degree and thus do not show the inhibitory effect on production of the antioxidant-deficiency sign noted in the studies on encephalomalacia. A direct relationship between the essential fatty acid content of the testes and the rate of testicular degeneration was found, but no effects of biologically available selenium and sulfur amino acids were evident. As the liver and muscle, onset of antioxidant-deficiency is characterized by a decrease in the most highly unsaturated fatty acid in the tissue (22∶5–ω6 in this case) and a net increase in arachidonate.     

The effect of antioxidant deficiency on tissue lipid composition in the rat. IV. Peroxidation and interconversion of polyunsaturated fatty acids in muscle phospholipids

It has been suggested that the net changes which take place in the composition of the muscle phospholipid fatty acids of the antioxidant-deficient rat represent the balance of two opposing processes. To compensate for (A) the preferential peroxidative destruction of the most highly polyunsaturated fatty acids in the tissue there occurs (B) an increase in the conversion of available precursors to the higher polyunsaturated fatty acids. Analysis of the data in terms of peroxidation kinetics indicated that the onset of creatinuria in one group after 3 weeks and in a second group after 7 weeks on an antioxidant-deficient diet occurred in both cases concomitant with the peroxidative “disappearance” of approximately 125 μg of phospholipid polyunsaturated fatty acid per gram wet weight of tissue or 2% of the total muscle phospholipid fatty acids.     

The effect of dietary zinc deficiency on the lipid composition of the rat erythrocyte membrane

The effect of dietary zinc deficiency in the rat on the lipid composition of the erythrocyte membrane was determined. Weanling male Wistar rats were fed an egg whitebased diet containing <1.0 mg Zn/kg dietad libitum. Control rats were either pair-fed orad libitum-fed the basal diet suppelemented with 100 mg Zn/kg diet. A zinc refed group was fed the −Zn diet until day 18 and then pairfed the +Zn diet until day 21. The voluntary feed restriction associated with dietary zinc deficiency resulted in erythrocyte membranes that had depressed phospholipid/protein and elevated cholesterol/phospholipid ratios. Similarly, all feed restricted groups had elevated 22-carbon n−3 polyunsaturated fatty acid (PUFA) and depressed 22-carbon n−6 PUFA concentrations in alkenylacyl and diacyl glycerophosphoethanolamine, phosphatidylserine and phosphatidylcholine; they also had depressed 24∶2n−6 levels in sphingomyelin. The relative concentrations of phospholipids in the membrane was similar between −Zn and +Zn (ad libitum) groups; however, the −Zn group had significantly less phosphatidylserine relative to +Zn (pair-fed) controls.     

Effect of dietary carnosine on plasma and tissue antioxidant concentrations and on lipid oxidation in rat skeletal muscle

The effect of dietary carnosine supplementation on plasma and tissue carnosine and α-tocopherol concentrations and on the formation of thiobarbituric acid reactive substances (TBARS) in rat skeletal muscle homo-genates was evaluated. Plasma, heart, liver and hind leg muscle was obtained from rats fed basal semipurified diets or basal diets containing carnosine (0.0875%), α-tocopheryl acetate (50 ppm), or carnosine (0.0875%) plusα-tocopheryl acetate (50 ppm). Dietary carnosine supplementation did not increase carnosine concentrations in heart, liver and skeletal muscle. Dietary supplementation with both carnosine and α-tocopherol increased carnosine concentrations in liver 1.56-, 1.51- and 1.51-fold as compared with diets lacking carnosine, α-tocopherol or both carnosine and α-tocopherol, respectively. Dietary supplementation with both carnosine and α-tocopherol also increased α-tocopherol concentrations in heart and liver 1.38-fold and 1.68-fold, respectively, as compared to supplementation with α-tocopherol alone. Dietary supplementation with carnosine, α-tocopherol or both car-nosine and α-tocopherol was effective in decreasing the formation of TBARS in rat skeletal muscle homogenate, with dietary α-tocopherol and α-tocopherol plus carnosine being more effective than dietary carnosine alone. The data suggest that dietary supplementation with carnosine and α-tocopherol modulates some tissue carnosine and α-tocopherol concentrations and the formation of TBARS in rat skeletal muscle homogenates.     

The effect of protein deficiency on some rat liver lipid metabolic enzymes and CoA

Two groups of male Wistar rats were fed normal (i.e., 18%) and protein-free diets, respectively, for 7 weeks. In vivo incorporation of [1-¹⁴C] acetate into palmitic, stearic, oleic, and arachidonic acids by the liver was reduced in the protein-deficient rats. In vitro incubation of liver microsomes with labeled palmitate or linoleate revealed no change in the specific activities of chain elongating or desaturating enzymes. Protein deficiency resulted in a decrease in specific activity of short chain acyl-CoA synthetase and in total CoA, accompanied by the virtual disappearance of acyl-CoA and an increase in free CoA. Furthermore, there was less microsomal fatty acid synthetase and mitochondrial β-hydroxybutyrate dehydrogenase activity. These results are discussed in relation to fatty acid synthesis and the changes in liver fatty acid composition.     

Zinc deficiency in the rat alters the lipid composition of the erythrocyte membrane triton shell

The effect of dietary zinc deficiency on the lipid composition of the erythrocyte membrane Triton shell was determined. Weanling male Wistar rats were fed an egg white-based diet containing <1.0 mg Zn/kg dietad libitum. Control rats were either pair-fed orad libitum-fed the basal diet supplemented with 100 mg Zn/kg diet. A Zn refed group was fed the −Zn diet until day 18 and then pairfed the +Zn diet until day 21. Dietary, Zn deficiency caused an increased cholesterol/phospholipid ratio in Triton shells compared to those from pair-fed controls. Zn deficiency caused a decreased double bond index of fatty acids in phosphatidylinositol (PI) and phosphatidylcholine (PC); there was a decreased proportion of 18∶2n−6 and 22∶4n−6 in PC and 20∶4n−6 in PI as compared to that found in pair-fed controls. All glycerophospholipids that were retained in the shell had a lower double bond index and increased content of 16∶0 and/or 18∶0 relative to the phospholipid in the intact membrane.     

Phospholipid fatty acid composition in type I and type II rat muscle

The fatty acid composition of the membrane phospholipids phosphatidylcholine (PC) and phosphatidylethanolamine in insulin-sensitive Type I (soleus) and insulin-resistant Type II (EDL) muscle is not known. In the present studies, soleus and EDL muscles were removed from 250–300 g Sprague-Dawley rats, and the fatty acid composition of total and individual phospholipid (PL) species was quantitated. As expected, triglyceride content was increased twofold in soleus muscle. No quantitative differences in the individual PL species or cholesterol content were found between the two muscles. However, a striking difference in PL fatty acid composition was observed in the PC fraction. An increase in 16∶0 with decreases in 18∶0, 18∶1, 22∶5n-3, and 22∶6n-3 (P<0.001 for each) was observed in the PC fraction of EDL compared to that from soleus, consistent with reduced elongation of PC fatty acids. Inhibition of fatty acid oxidation with the carnitine palmitoyl transferase-1 inhibitor, etomoxir, did not alter the fatty acid pattern in either muscle. We conclude that an alteration in PL fatty acid composition consistent with reduced elongation of both saturated and unsaturated fatty acids is observed in Type II muscle. The restriction of these alterations to the PC fraction has important implications. Deceased (June 28, 1996).     

Effects of dietary saturated andTrans fatty acids on tissue lipid composition and serum LCAT activity in the rat

Groups of rats were fed from weaning with diets containing 5% by wt of hydrogenated coconut oil (HCO), safflower oil, or a concentrate of ethyl elaidate and linolelaidate (TRANS) as the sole source of dietary fat. Fatty acid composition of the lipid classes from serum, liver, heart, and kidney was determined, and the serum lecithin: cholesterol acyl transferase (LCAT) activities were assayed for each animal. Serum LCAT activity was increased by both the HCO and TRANS diets in the early stages of the development of an essential fatty acid (EFA) deficiency but was suppressed in the animals of the TRANS group as they became older. The HCO and TRANS groups exhibited changes in tissue lipid fatty acid composition, as well as reduced growth, characteristic of an EFA deficiency. Conversion of oleic acid to eicosatrienoic acid was impaired in the animals fed the TRANS diet, greatly increasing the octadecenoic acid content of the tissue lipids at the expense of eicosatrienoic acid. The TRANS diet also suppressed incorporation of eicosatrienoic acid into cholesteryl esters of tissue and serum, indicating that, when fed as the sole source of unsaturated fat,trans fatty acids influenced the metabolism of unsaturated fatty acids and cholesterol.     

Effect of vitamin E deficiency on the lipid class and fatty acid composition of rat brain gray and white matter

Three-week old male Sprague-Dawley rats were placed on a control or vitamin E-deficient diet for 9 months. The total lipid and cholesterol contents of brain gray and white matter areas in the vitamin E-deficient group did not differ from controls. The concentration of cerebrosides was lower in white matter but higher in gray matter of deficient animals. However, sulfatide was significantly (P<0.001) higher in white and gray matter of deficient animals compared with controls. Lysolecithin was not found in vitamin E-deficient gray matter but was present in control gray matter lipids. No marked differences were found in the concentrations or relative amounts of sphinogomyelin, phosphatidyl choline, phospholipids of gray or white matter of vitamin E-deficient rats as compared to controls. In addition no remarkable differences were found in the fatty acid composition of total lipid extracts of gray or white matter from vitamin E-deficient rats when compared with controls. Presented in part at the Biochemistry/Biophysics Meeting, Minneapolis, 1974, and the Sixth Meeting of the American Society for Neurochemistry, Mexico City, 1975.     

The effect of dietary fats on the plasma lipid composition of sheep

This study reports on the plasma lipid compositions of sheep fed either a control diet (C), a control diet supplemented with tallow (A) or polyunsaturated fatty acid (B) that had been protected against hydrolysis and hydrogenation in the rumen, or a control diet supplemented with maize oil (D). Diet B considerably increased the 18∶2 content of all the major plasma lipid fractions. Although the feeding of diet D also resulted in an increase in the 18∶2 contents within the cholesteryl ester, unesterified fatty acid, and phospholipid fractions the increases were considerably less than those observed with diet B; the levels of 18∶2 within the triglyceride fraction remained similar to that for the sheep which received the control diet. The effect of feeding diet A was confined solely to the triglyceride fraction where the concentrations of 16∶0 and 18∶1 were increased. The lipoproteins of the plasma were separated into very low density lipoproteins (d<1.006), low density lipoproteins (1.006<d<1.063), and high density lipoproteins (1.063<d<1.21), and the distribution of the major lipids between these lipoprotein fractions was investigated. Diet B increased considerably the proportion of triglyceride found in association with the very low density fraction and the concentrations of 18∶2 within all the lipoprotein fractions; these increases in the concentrations of 18∶2 were not confined to any particular lipid fraction of the lipoproteins. In contrast, the increases in the concentrations of 18∶2 produced as a result of feeding diet D were confined to the low and high density lipoproteins. The effect of feeding diet A was confined to fatty acid changes within the triglycerides of the low and very low density lipoproteins.     

The effect of culture medium composition on ether lipid cytotoxic activity

The effect of a serum-free medium (TNB-100), compared to RPMI 1640 containing 10% fetal bovine serum (FBS), on the lipid composition of HL60 and K562 leukemic cells was investigated. The 10% FBS RPMI medium contained approximately three times more phospholipids (PL), about three times more protein and eight times more cholesterol (CHOL) than did the TNB-100 medium. Cells cultured in TNB-100 medium, referred to as HL60-TNB and K562-TNB cells, were significantly lower in PL and CHOL than 10% FBS RPMI cells, with about a threefold higher PL-to-CHOL ratio; however, these cells were significantly higher in protein content. Cells grown in TNB-100 were also significantly more fluid than 10% FBS RPMI cells and were more sensitive to the fluidizing action of the ether lipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine. The 50% inhibitory dose of the drug was about 50% lower in TNB-grown cells than in 10% FBS RPMI cells.     

Studies on lipid and fatty acid composition of human hepatoma tissue

Studies are reported on the composition of the lipids of human liver and hepatoma tissues from male adults. Liver tissues were obtained from individuals who died from causes other than liver disease or cancer. The hepatoma tissues were obtained from individuals shortly after they succumbed to cancer. The total lipid of each tissue was fractionated quantitatively by silicic acid column chromatography into neutral lipid, glycolipid, and phospholipid fractions. These fractions were analyzed by thin layer chromatography and converted to methyl esters for analysis of their constituent fatty acids by gas liquid chromatography. In comparison to liver tissue, the total amount of lipid in the hepatoma tissues was generally higher and more variable; the lipid of one hepatoma was ca. 92% of the dry wt of the tissue. The greater lipid content of the hepatoma tissues was due to the high percentage of neutral lipid. Except for one specimen, there was ca. the same amount of glycolipid in the hepatoma as in the liver tissues, but the composition of the glycolipid fraction of the hepatoma lipid differed considerably, particularly in the ganglioside fraction. The phospholipid fraction of hepatoma lipid was much lower than that of liver but exhibited only quantitative differences in composition. No glyceryl ether diesters and only traces of plasmalogens of phosphatidyl choline or phosphatidyl ethanolamine were detected in the liver and hepatoma lipids. The levels of monoenoic acids were higher and those of linoleic and polyunsaturated fatty acids lower in the hepatoma lipids. Positional isomers of trienoic acids not normally present in liver tissue were detected in hepatoma lipids. The abnormalities observed in lipid composition indicated interferences in the regulatory processes of lipid metabolism in human hepatoma similar to those observed in animals.     

The lipid composition of microsomal preparations from lactating bovine mammary tissue

The microsomes isolated from lactating bovine mammary tissue contained 4.3 mg lipid per milligram nitrogen. Phospholipids comprised 83% of the lipids. The neutral lipids were composed of triglycerides (20–30%), diglycerides (5–10%), free fatty acids (15–30%, cholesterol (35–40% and cholesterol esters (10–12%, respectively. Phosphatidylcholine was the predominant phospholipid component (>50%), and the remainder consisted of phosphatidylethanolamine (21–13%), phosphatidylserine (4–6%), phosphatidylinositol (8%), sphingomyelin (9%) and lysophosphatidylcholine (2%) respectively. The composition of the microsomal phospholipids was similar to that of isolated mammary cells and tissue homogenates but quite different from milk and fat globule membrane phospholipids. The triglycerides contained short chain fatty acids but their relative concentrations were lower than in milk triglycerides. The various lipid fractions had a variable proportion of saturated fatty acids, i.e., triglycerides (47.7%), diglycerides (86.7%), free fatty acids (70.6%), phosphatidylcholine (50.6%), phosphatidylethanolamine (50.8%), phosphatidylserine (35.3%), phosphatidylinositol (40.5%) and sphingomyelin (82.3%), respectively. The molecular distribution of fatty acids in the microsomal triglycerides and phosphatidylcholine was similar to that occurring in milk, i.e., the short chain and unsaturated fatty acids were concentrated in the primary positions (sn1 andsn3) of the triglycerides, and the unsaturated acids were preferentially located in positionsn2 of the phosphatidylcholine. The compositional data indicate that mammary microsomes are not the direct source of the phospholipids of the milk fat globule.     

Effect of hypothyroidism on the lipid composition of rat plasma and erythrocyte membranes

The effect of hydrothyroidism on plasma and erythrocyte membrane lipid components has been investigated. This pathological state is accompanied by a) a cholesterol increase of about 60% in plasma, and at the same time a 22% reduction in erythroycte membranes; b) 44% and 30% phospholipid level decreases in both plasma and red cell membranes, respectively; and c) almost unaffected phospholipid and fatty acid compositions of both plasma and erythrocyte membranes. All changes were corrected by treatment of the hypothyroid rats with triiodothyronine for two days. These findings suggest that in hypothyroid rats a reduced transfer of cholesterol from plasma to erythrocyte membrane probably takes place. This could explain, at least in part, the increased hematic cholesterol level observed in hypothyroid animals. In red cell membranes, the simultaneous decrease in cholesterol and phospholipid levels does not alter the cholesterol/phospholipid molar ratio, thus avoiding their abnormal function.     

Abnormal lipid composition of fat tissue in human mesenteric panniculitis

Mesenteric fat tissue obtained at autopsy from 6 patients with mesenteric panniculitis (MP) were found to contain significant amounts of cholesteryl esters (CE). In addition, samples from 3 of these cases were found to contain 0.5–1.3% free cholesterol, 0.9–1.9% free fatty acids (FFA), 0.6–2.5% 1-alkyl glyceryl ether diesters and small amounts of squalene. Two of these tissues also contained alk-1-enyl glyceryl ether diesters. The fatty acid compositions of the CE, FFA, triacylglycerides and glyceryl ether diesters (GEDE) were determined and oleic acid (18∶1) was found to be the major fatty acid. The alkyl group composition of the GEDE consisted essentially of 16∶0 and 18∶0 and 18∶1 carbon atoms in both types of ethers.     

The effect of conjugated linoleic acid on the antioxidant enzyme defense system in rat hepatocytes

Short-term effects of physiological concentrations of conjugated linoleic acid (CLA) on membrane integrity, metabolic function, cellular lipid composition, lipid peroxidation, and antioxidant enzymes were examined using rat hepatocyte suspension cultures. Incubation with CLA (5–20 ppm) for 3 h decreased the ability of hepatocyte plasma membranes to exclude trypan blue by approximately 25%, and caused leakage of cytosolic lactate dehydrogenase (LDH) into the medium. The significant decrease (P<0.02) in hepatocyte viability as measured by LDH leakage during cell incubation with 10 and 20 ppm CLA was not associated with significant changes in cellular ATP content. Protein synthesis in hepatocytes was elevated (P<0.05) in the presence of 5 and 10 ppm CLA, but at a higher concentration (20 ppm), protein synthesis was similar to that of control cells. Gluconeogenesis was maintained in cells incubated with lower concentrations of CLA (5 and 10 ppm) but was decreased (P<0.02) at the higher concentration. Incubation with 20 ppm CLA for 3 h did not affect the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme of cholesterol synthesis. Both cis-9,trans-11/trans-9,cis-11, and cis-10,trans-12/trans-10,cis-12 isomers of CLA were incorporated to a similar level into hepatocytes. Levels ranged from 3.9 to 4.1%, respectively, of total fatty acids in neutral lipids, and from 0.7 to 0.8%, respectively, of total fatty acids in phospholipids. Cellular lipid peroxidation remained unchanged in the presence of CLA (5–20 ppm), despite significant inhibition (P<0.05) of superoxide dismutase. Catalase activity was maintained near control levels in the presence of 5 and 10 ppm CLA but was significantly decreased in the presence of 20 ppm CLA. Glutathione peroxidase activity was significantly decreased in the presence of 10 ppm CLA. The apparent sensitivity of the antioxidant enzyme defense system of liver cells to CLA, coupled with the lack of effect of CLA on lipid peroxidation in cells, suggests that cytotoxic effects of CLA as described by LDH leakage and decreased gluconeogenesis were not mediated by a prooxidant action in hepatocytes.     

The effect of soybean moisture during storage on the lipid composition of extracted crude oil

The quality of soybean oil extracted from seed stored under constant temperature and relative humidity for 42 days was evaluated over a wide range of moisture levels. Storage of soybeans at 9, 13 and 18% moisture had little affect on the major lipid components (neutral lipids), even though seed stored at 18% moisture became infected with mold. The level of phospholipid in the extracted crude oil decreased during the last 3 weeks of storage in seeds stored at 13 and 18% moisture from 4 to 2.5% of the total oil. During the same period, the level of free fatty acids, (FFA) (primarily 16∶0 and 18∶2) in these samples increased. This study indicated that the increase in FFA during seed storage at high moisture levels was the result of soybean lipase and possibly phospholipase activity. These findings suggested that soybeans should be kept at less than 13% moisture for long-term on-farm storage to preserve oil quality.     

The effect of dietary arachidonic acid on plasma lipoprotein distributions, apoproteins, blood lipid levels, and tissue fatty acid composition in humans

Normal healthy male volunteers (n=10) were fed diets (high-AA) containing 1.7 g/d of arachidonic acid (AA) for 50 d. The control (low-AA) diet contained 210 mg/d of AA. Dietary AA had no statistically significant effect on the blood cholesterol levels, lipoprotein distribution, or apoprotein levels. Adipose tissue fatty acid composition was not influenced by AA feeding. The plasma total fatty acid composition was markedly enriched in AA after 50 d (P<0.005). The fatty acid composition of plasma lipid fractions, cholesterol esters, triglycerides, free fatty acids, and phospholipid (PL) showed marked differences in the degree of enrichment in AA. The PL plasma fraction from the subjects consuming the low-AA diet contained 10.3% AA while the subjects who consumed the high-AA diet had plasma PL fractions containing 19.0% AA. The level of 22:4n-6 also was different (0.67 to 1.06%) in the plasma PL fraction after 50 d of AA feeding. After consuming the high-AA diet, the total red blood cell fatty acid composition was significantly enriched in AA which mainly replaced linoleic acid. These results indicate that dietary AA is incorporated into tissue lipids, but selectively into different tissues and lipid classes. Perhaps more importantly, the results demonstrate that dietary AA does not alter blood lipids or lipoprotein levels or have obvious adverse health effects at this level and duration of feeding.