Effect of specific dietary fatty acids on lipogenesis in the livers and mammary glands of lactating mice

The effects of linoleic, linolenic and columbinic acids fed as 4% of a high carbohydrate (50% glucose) diet on the activities and the amounts of several enzymes associated with fatty acid synthesis in livers and mammary glands of lactating mice were compared with those for stearic and oleic acids. Fatty acid synthesis, measured in vivo, was significantly lower in livers of mice ingesting all 3 polyunsaturated fatty acids (PUFA), whereas in mammary glands synthesis was lower only in mice receiving columbinic acid. The activities of fatty acid synthetase (FAS) and acetyl CoA carboxylase were significantly reduced in liver by all 3 PUFA, as were activities of glucose-6-phosphate dehydrogenase, malic enzyme (ME) and citrate cleavage enzyme (CCE), also associated with lipogenesis. In mammary gland, on the other hand, the activities of these enzymes were unaffected by dietary PUFA. The tissue contents of FAS, ME and CCE, measured by rocket immunoelectrophoresis, were found to be significantly reduced in liver by linoleate, linolenate and columbinate but were not significantly altered in mammary gland. The decrease in hepatic lipogenesis observed was principally due to a decrease in the amounts of these enzymes induced by the dietary PUFA but the inhibition in mammary gland caused by columbinate could not be accounted for by a reduction in enzyme contents and therefore may be due to allosteric effects which occur when fatty acid synthesis is measured with³H₂O. The fatty acid composition in liver and mammary gland of dams and in liver and kidney of pups completely reflected dietary fatty acids. Columbinate made up ca. 20% of the total fatty acids in both tissues of the columbinic acid-fed mice and ca. 15% in the pup tissues. This suggests that columbinate is incorporated into milk lipids of dams and is easily absorbed by pups. The elevated ratios of 16/16∶1 and 18/18∶1 in liver and mammary gland of dams and liver and kidney of the pups from dams fed linoleate, linolenate and columbinate suggest that each of these polyunsaturated fatty acids in the diet can inhibit the activity of Δ9 desaturase.     

Free fatty acid inhibition of the insulin induction of glucose-6-phosphate dehydrogenase in rat hepatocyte monolayers

Rat hepatocytes in monolayer culture were utilized to determine if the decrease in glucose-6-phosphate dehydrogenase (G6PD) activity resulting from the ingestion of fat can be mimicked by the addition of fatty acids to a chemically, hormonally defined medium. G6PD activity in cultured hepatocytes was induced several-fold by insulin. Dexamethasone or T3 did not amplify the insulin induction of G6PD. Glucose alone increased G6PD activity in cultured hepatocytes from fasted donors by nearly 500%. Insulin in combination with glucose induced G6PD an additional two-fold. The increase in G6PD activity caused by glucose was greater in hepatocytes isolated from 72 hr-fasted rats as compared to fed donor rats. Such a response was reminiscent of the “overshoot” phenomenon in which G6PD activity is induced well above the normal level by fasting-refeeding rats a high glucose diet. Addition of linoleate to the medium resulted in a significant suppression of insulin’s ability to induce G6PD, but linoleate had no effect on the induction of G6PD activity by glucose alone. A shift to the right in the insulin-response curve for the induction of G6PD also was detected for the induction of malic enzyme and acetyl-CoA carboxylase. Arachidonate (0.25 mM) was a significantly more effective inhibitor of the insulin action than linoleate was. Apparently rat hepatocytes in monolayer culture can be utilized as a model to investigate the molecular mechanism by which fatty acids inhibit the production of lipogenic enzymes. In part, this mechanism of fatty acid inhibition involves desensitization of hepatocytes to the lipogenic action of insulin.     

Selectivity in incorporation,utilization and retention of oleic and linoleic acids by human skin fibroblasts

Fetal human fibroblasts were grown in culture medium containing 10% fetal bovine serum supplemented with [1-¹⁴C] linoleate or [1-¹⁴C] oleate. At all concentrations of exogenous fatty acids, the incorporation of oleate was greater than that of linoleate. With increased medium fatty acid concentrations, linoleate in triacylglycerol (TAG) could be increased from 13 to 75% of the total incorporated; at each concentration, relatively more linoleate than oleate was in TAG. When the cells were exposed to exogenous oleate/linoleate mixtures, the composition of the mixture determined the extent of incorporation of both fatty acids. When the mixture was primarily linoleate, scarce oleate was used preferentially for phospholipids (PL); no such specificity for scarce linoleate was observed. Addition of exogenous fatty acids resulted in a shift of previously incorporated¹⁴C fatty acids from phospholipid into TAG; retention of oleate in PL was greater than that of linoleate. Incorporation of oleate into phospholipids was also higher than that of linoleate from exogenous fatty acid mixtures which were 80% saturated. It is suggested that normal human fibroblasts have adapted to the low levels of exogenous polyunsaturated fatty acids in culture media by increased use of oleate in phospholipid. Even when the cells aresupplemented with linoleate, the preferential use of oleate in phospholipid groups is retained. Presented in part at the ASBC Meeting, Dallas, april 1979.     

Specific inhibition of hepatic fatty acid synthesis exerted by dietary linoleate and linolenate in essential fatty acid adequate rats

Dietary linoleate and linolenate were investigated for their ability to specifically inhibit liver and adipose tissue lipogenesis in meal-fed (access to food 900-1,200 hr), essential fatty acid (EFA) adequate rats. Supplementing a high carbohydrate diet containing 2.5% safflower oil with 3% palmitate 16∶0, oleate 18∶1, or linoleate 18∶2 did not affect in vivo liver or adipose tissue fatty acid synthesis. However, 18∶2 addition to the basal diet did result in a significant (P<0.05) decline of liver fatty acid synthetase (FAS) and glucose-6-phosphate dehydrogenase (G6PD) activities. When the safflower oil content of the basal diet was reduced to 1%, the addition of 3% 18∶2 or linolenate 18∶3 significantly (P<0.05) depressed hepatic FAS, G6PD, and in vivo fatty acid synthesis by 50%. Addition of 18∶1 caused no depression in hepatic FAS activity but did result in a significant (P<0.05) decline in liver G6PD activity and fatty acid synthesis which was intermediate between basal and basal +18∶2-or+18∶3-fed animals. Adipose tissue rates of lipogenesis were completely unaffected by dietary fatty acid supplementation. Similarly, the addition of 3 or 5% 18∶3 to a basal diet for only one meal resulted in no change in lipogenesis relative to that in animals fed the basal diet. The data indicate that, like rats fed EFA-deficient diets, dietary 18∶2 and 18∶3 exert a specific capacity to depress rat liver FAS and G6PD activities and rate of fatty acid synthesis. Michigan Agricultural Experiment station Journal Article No. 7581. D.R. Romsos is the recipient of Career Development Award K04 AM 00112     

Enhancement of liver microsomal acyl CoA-lysophospholipid acyltransferase activity in pyridoxine-deficient rats

In rats deficient in pyridoxine and essential fatty acids, liver phospholipids contained less arachidonic acid and more oleic and eicosatrienoic acids than those from animals only deficient in essential fatty acids. This pattern persisted after the animals were supplemented with linoleate for 6 days. Liver oleyl and arachidonyl CoA-lysophospholipid acyltransferase activities were significantly higher in pyridoxine-deficient animals. Supplementation with linoleate for 6 days resulted in a marked increase in arachidonyl CoA-acyltransferase activity in pyridoxine-deficient rats but a relatively small increase in the supplemented animals. Differences in fatty acid composition between pyridoxine-deficient and supplemented rats can not be ascribed to lower liver transacylase activity in the deficient animals.     

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.     

Effect of polyunsaturated fatty acids in obese mice

Genetically obese (ob/ob) mice display a variety of metabolic differences from lean litter mates. In the obese state, fatty acid desaturation-elongation in brown adipose tissue mitochondria is apparently altered, resulting in differences in membrane fatty acid composition. This change in membrane lipid environment appears to influence GDP binding and there-fore the activity of the proton conductance pathway associated with regulation of energy expenditure in these animals. In liver, binding of insulin to the nuclear membrane is increased by feeding a high polyunsaturated/saturated (P/S) diet fat. Consumption of a high P/S diet decreased mRNA levels for fatty acid synthase, acetyl-CoA carboxylase, malic enzyme, and pyruvate kinase in obese and lean animals. Expression of mRNA for these lipogenic enzymes was higher in obese animals and suggests that obese mice may be resistant to polyunsaturated fatty acid feedback control of gene expression.     

Lipid composition of hepatocyte plasma membranes from geese overfed with corn

Twelve-week-old Landes male geese were overfed with corn for 21 d in order to induce liver steatosis (fatty liver). Lipid composition of hepatocyte plasma membranes from fatty livers was compared to that of lean livers obtained from geese fed a normal diet. The ratio cholesterol/phospholipids was higher in fatty hepatocyte plasma membranes (0.63 vs. 0.47), whereas the phospholipid/protein ratio was less than half. Overfeeding induced changes in fatty acid composition of hepatocyte plasma membranes, including a greater than twofold increase in the percentage of oleic acid (29.7 vs. 13.8%) and a somewhat lesser increase in lauric, palmitic, and palmitoleic acid contents of plasma membrane lipids of fatty livers. A concomitant reduction in the proportion of stearic acid (18.4 vs. 25.1%) was also observed. In fatty livers, the increased ratio of saturated to polyunsaturated fatty acids (PUFA) (1.5 vs. 1.0) was related to a significant decrease in PUFA content. Among all the PUFA, only the eicosatrienoic acid (20∶3n−9) percentage was increased by liver steatosis. Overfeeding with corn appeared to induce competition between de novo synthesized and dietary fatty acids incorporated in hepatocyte plasma membranes. This resulted in an accumulation of de novo synthesized monounsaturated and derived fatty acids in plasma membranes from overfed birds. A defect in the incorporation of linoleic acid and linoleic- and linolenic-derived PUFA was observed despite the high proportion of these essential fatty acids in the diet. It was conclued that in overfed palmipeds, de novo hepatic lipogenesis prevails over dietary lipid intake to modulate lipid composition of the fatty liver plasma membrane.     

The hypotriglyceridemic effect of eicosapentaenoic acid in rats is reflected in increased mitochondrial fatty acid oxidation followed by diminished lipogenesis

The effect of eicosapentaenoic acid (EPA) on fatty acid oxidation and on key enzymes of triglyceride metabolism and lipogenesis was investigated in the liver of rats. Repeated administration of EPA to normolipidemic rats resulted in a time-dependent decrease in plasma triglycerides, phospholipids and cholesterol. The triglyceride-lowering effect was observed after one day of feeding whereas lowering of plasma cholesterol and phospholipids was observed after five days of treatment. The triglyceride content of liver was reduced after two-day treatment. At that time, increased mitochondrial fatty acid oxidation occurred whereas mitochondrial and microsomal glycerophosphate acyltransferase was inhibited. The phosphatidate phosphohydrolase activity was unchanged. Adenosine triphosphate:citrate lyase, acetyl-CoA carboxylase, fatty acid synthetase and glucose-6-phosphate dehydrogenase were inhibited during the 15 d of EPA treatment whereas peroxisomal β-oxidation was increased. At one day of feeding, however, when the hypotriglyceridemic effect was established, the lipogenic enzyme activities were reduced to the same extent in palmitic acid-treated animals as in EPA-treated rats. In cultured rat hepatocytes, the oxidation of [¹⁴C]palmitic acid to carbon dioxide and acid-soluble products was stimulated in the presence of EPA. These results suggest that the instant hypolipidemia in rats given EPA could be explained at least in part by a sudden increase in mitochondrial fatty acid oxidation, thereby reducing the availability of fatty acids for lipid synthesis in the liver for export,e.g., in the form of very low density lipoproteins, even before EPA induced peroxisomal fatty acid oxidation, reduced triglyceride biosynthesis and diminished lipogenesis.     

Liver subcellular fatty acid profiles of chicks fed diets containing hydrogenated fats and varying linoleate levels

Day-old male broiler chickens were fed semipurified diets containing 5% lipid from one of four different lipid sources: corn oil (CO), partially hydrogenated soybean oil (HSBO), a spent restaurant grease (SRG) and a purified mixture of triolein, tripalmitin and tristearin (OPS). Diets CO and HSBO contained adequate amounts of linoleic acid, but diets SRG and OPS were deficient in linoleate. In addition, SRG and HSBO containedtrans isomers of 16∶1 and 18∶1. The diets were fed for 3 wk to determine the effects of low linoleate levels andtrans isomers on fatty acid profiles in liver microsomes, mitochondria and cytosol. Chicks fed HSBO had the highest body weights, while those fed SRG and OPS had the lowest. The incidence and severity of dermatitis were similar for all treatments. The proportions of linoleate and arachidonate in lipids from liver subcecullar fractions were reduced significantly in chicks fed OPS and SRG; however levels of 20∶3ω9 were not increased. Feeding HSBO, which is high in both linoleate and linolenate, resulted in higher levels of 18∶3ω3 and 20∶5ω3 in liver subcellular fractions and lower levels of 20∶4ω6 than those seen in chicks fed CO. The isomeric forms of 18∶1 present in the partially hydrogenated fats (HSBO and SRG) appeared to be incorporated into the lipids of liver fractions. The results of this study show that dietary lipids influence fatty acid, profiles of chick liver microsomes, mitochondria and cytosol. Decreases in linoleate and arachidonate in these organelles occur before overt essential fatty acid (EFA) deficiency signs in chicks fed EFA-deficient diets. Published as Scientific Paper No. 7512, College of Agriculture and Home Economics Research Center, Project No. 4723, Washington State University, Pullman, WA.     

Conjugated linoleic acid modulates hepatic lipid composition in mice

Conjugated linoleic acid (CLA) is a chemoprotective fatty acid that inhibits mammary, colon, forestomach, and skin carcinogenesis in experimental animals. We hypothesize that the ubiquitous chemoprotective actions of dietary CLA in extrahepatic tissues are dependent upon its role in modulating fatty acid composition and metabolism in liver, the major organ for lipid metabolism. This study begins to evaluate the role of CLA in lipid metabolism by determining the modulation of fatty acid composition by CLA. Female SENCAR mice were fed semipurified diets containing 0.0% (Diet A), 0.5% (Diet B), 1.0% (Diet C), or 1.5% (Diet D) CLA (by weight) for six weeks. Mice fed Diets B, C, and D exhibited lower body weights and elevated amounts of extractable total lipid in livers compared with mice fed diets without CLA (Diet A). Analyses of the fatty acid composition of liver by gas chromatography revealed that dietary CLA was incorporated into neutral and phospholipids at the expense of linoleate in Diets B, C, and D; oleate increased and arachidonate decreased in neutral lipids of CLA diet groups. In addition, increasing dietary CLA was associated with reduced linoleate in hepatic phospholipids. In an in vitro assay, CLA was desaturated to an unidentified 18:3 product to a similar extent as linoleate conversion to γ-linolenate (9.88, and 13.63%, respectively). These data suggest that CLA may affect metabolic interconversion of fatty acids in liver that may ultimately result in modified fatty acid composition and arachidonate-derived eicosanoid production in extrahepatic tissues. In addition to determining how dietary CLA modulates eicosanoid synthesis, further work is needed to identify enzymatic products that may result from desaturation of CLA.     

Effects of lovastatin on hepatic fatty acid metabolism

Thein vitro andin vivo effects of lovastatin on fatty acid metabolism were studied in isolated rat hepatocytes. When addedin vitro to cell incubations, lovastatin stimulatedde novo fatty acid synthesis and acetyl-CoA carboxylase activity, whereas fatty acid synthase, activity was unaffected. Lovastatin depressed palmitate, but not octanoate, oxidation. This may be attributed to the lovastatin-induced increase, in intracellular malonyl-CoA levels, as no concomitant changes of carnitine palmitoyltransferase I (CPT-I) specific activity was detected. Lovastatin had no effect on the synthesis and secretion of triacylglycerols and phospholipids in the form of very low density lipoproteins (VLDL). When rats were fed a diet supplemented with 0.1% (w/w) lovastatin for one week, both acetyl-CoA carboxylase activity andde novo fatty acid synthesis were reduced compared to pair-fed controls, whereas fatty acid synthase activity was unaffected. Palmitate oxidation was enhanced in the lovastatin-fed group. There was an increase in CPT-I activity but no change in intracellular concentration of malonyl-CoA. Lovastatin feeding and no significant effect either on the esterification of exogenous palmitic acid into both cellular and VLDL triacylglycerols and phospholipids or on hepatic lipid accumulation. Thein vitro andin vivo effects of lovastatin were not significantly different between periportal and perivenous hepatocytes. The results indicate that: (i) the administration of lovastatin increased the fatty acid-oxidative capacity of the liver at the expense of its lipogenic capacity, (ii) the rate ofde novo cholesterol synthesis did not seem to be a limiting factor in the synthesis and secretion of VLDL and (iii) lovastatin produced opposite effects on hepatic fatty acid metabolismin vitro andin vivo.     

Influence of fasting-refeeding and dietary linoleate on liver glucose-6-phosphate dehydrogenase and phospholipid fatty acid composition in rats adapted to a purified diet

Responses to refeeding after fasting were studied in male rats fed a purified casein-sucrose diet containing 5% safflower oil. After a 48 hr fast, the rats were fed either the same diet or the same diet minus oil (fat-free diet). These experiments were designed to distinguish changes due to fasting and refeeding alone without a change of diet from those changes caused by refeeding a diet of different composition. In the first experiment, rats were refed for 3 or 7 days. In rats refed either diet, liver glucose-6-phosphate dehydrogenase (G6PD) activity was elevated above refasting levels, but after 7 days, activity in rats refed the 5% safflower oil diet was significantly lower than in those refed the fatfree diet. The amount of liver arachidonate in rats refed the safflower oil diet was the same during refeeding as before fasting. In the second experiment, rats were refed the fat-free diet for 1, 2, 3, or 7 days. Liver G6PD and fatty acid synthetase were measured, as well as fatty acids in liver total lipids and phospholipids. G6PD activity increased above prefasting levels after one day refeeding and continued to increase, for 7 days. Fatty acid synthetase activity increased for the first 3 days of refeeding, with no additional increase after 7 days. In all rats refed the fat-free diet, the proportions of arachidonate and linoleate in liver phospholipids diminished with time, and eicosatrienoate appeared. These results show that (a) maintenance of liver phospholipid arachidonate did not prevent increased G6PD activity in early refeeding, but the elevated G6PD activity later declined when phospholipid arachidonate was maintained by feeding a source of linoleate; (b) the metabolic state of fastedrefed rats had not returned to prefasting conditions even after 7 days of refeeding a linoleate-rich diet to which the rats were adapted before fasting.     

Lipogenic enzyme activities in primary cultures of adult mouse hepatocytes

The effects of various unsaturated fatty acids such as oleic (18∶1n−9), linoleic (18∶2n−6) and arachidonic (20∶4n−6) on the activities of fatty acid synthetase (FAS), malic enzyme (ME), glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH) all were determined in primary cultures of mouse hepatocytes. Activities of FAS and ME were found to decrease with time in culture regardless of whether hepatocyte donors were fed diets containing polyunsaturated fatty acid-free hydrogenated cottonseed oil (HCTO) or corn oil (CO). On the other hand, while G6PDH activity also declined in cultured hepatocytes obtained from HCTO-fed mice, the activity of this enzyme increased in cells cultured from CO-fed mice. 6PGDH activity was found to increase in hepatocytes obtained from both diet groups. Neither 18∶2 nor 20∶4 when added to media could alter FAS or ME activities compared with those observed with either 18∶1-containing or fatty acid-free media. Since lactic dehydrogenase activity and the rate of incorporation of [³H] leucine into FAS protein were unaltered with time in hepatocyte cultures, the decreased activities of FAS and ME cannot be attributed to a loss in cell viability during culture but rather appear to be specific for those enzymes which respond to diet hormones in vivo. Examination of the fatty acid contents of the cells after the culture period showed that the values for the ratios of 16∶0/16∶1 and of 18∶0/18∶1 were elevated when either 18∶2 or 20∶4 was added to the medium even though there was no evidence for elongation of the added 18∶2 or for 20∶4 being converted to 22∶4. This result suggest that Δ9-desaturase activity was inhibited by these polyunsaturated fatty acids and that conversion of 18∶2 to 20∶4 was not required for such action. The rate of synthesis determined by the relative rate of incorporation of [³H]leucine into FAS was two to five times higher in hepatocytes prepared from mice fed the HCTO diet than in hepatocytes from mice fed the CO diet. We have concluded that the mechanisms for long-term regulation may not be contained entirely within the liver.     

Relative incorporation of linoleic and arachidonic acid in phospholipids and triglycerides of different rat tissues

Fat-deficient rats were fed different amounts of methyl linoleate for increasing periods of time. The fatty acid composition of triglycerides and phospholipids of epididymal fat pad, epirenal fat depot, intestinal fat depot, liver, and the pool of heart, kidney, lungs and pancreas was determined. The distribution of the total amount of linoleic and arachidonic acid incorporated into phospholipids and triglycerides per rat was calculated. Phospholipids and triglycerides of depot tissues presented different fatty acid compositions. Although the phospholipids of liver and the pool of heart, kidney, lung and pancreas specifically incorporated linoleic acid at the beginning they very rapidly attained a rather steady composition, whereas triglycerides went on incorporating the acid. The amount of linoleic acid incorporated into the phospholipids of depot tissues was rather small. The triglycerides undoubtedly contributed in the highest proportion to the total pool of linoleic acid. However, the highest proportion of arachidonic acid was found in the total pool of phospholipids. The total amount of linoleic acid incorporated into the phospholipids was an approximately lineal function of the amount of phospholipids independent of period of administration and doses of methyl linoleate. Besides presenting two lineal functions of the amount of phospholipids, arachidonic acid showed a vertical increase coincident with a vertical decrease of the amount of eicosa-5,8,11-trienoic acid. At this period no change in the amount of the phospholipid was shown. This phenomenon is explaioned as a possible direct replacement of eicosatrienoic acid by arachidonic acid.     

Effect of nutritional status on the fatty acid composition of rat liver and cultured hepatocytes

The lipid concentration and fatty acid composition of the whole liver and of cultured hepatocytes isolated from the livers of rats fed ad libitum (fed), fasted for 24 hr (fasted), or fasted for 48 hr and then refed a fat-free, high carbohydrate diet for 48 hr (refed) was studied. Hepatocytes were maintained as monolayer cultures in serum-free, lipid-free media and their fatty acid composition was analyzed at 3, 24, 48, 72 and 96 hr. The livers of fed animals, as well as their hepatocytes, contained less total lipid than those from animals on either of the other dietary regimes. Livers of fasted animals had three times the amount of lipid found in the livers of fed animals, and the livers of refed animals contained five times the amount of lipid as the livers of fed animals (all based on mg lipid/g wet weight of liver). The fatty acid composition of hepatocytes after 3 hr of culturing was very similar to that of fresh liver when compared in each of the dietary regimes. However, while the fatty acid compositions of livers and hepatocytes from fed and fasted animals were similar, the pattern in liver of refed animals was quite distinct from that of the fed animals. In the fed and fasted animals palmitic acid (16∶0), stearic acid (18∶0), oleic acid (18∶1[n-9]), linoleic acid (18∶2[n-6]) and arachidonic acid (20∶4[n-6]) were the major fatty acids of the liver; in refed animals 16∶0, palmitoleic acid (16∶1[n-7]), 18∶0, 18∶1(n-9) andcis-vaccenic acid (the n-7 isomer of oleic acid) were the major fatty acids. During maintenance in culture the 18∶1(n-9) content of the hepatocytes increased in cells from livers of animals on all three dietary regimes. The polyunsaturated fatty acid content was similar in fresh livers and isolated hepatocytes in all samples when compared on the basis of μg fatty acid/mg of hepatocyte or liver protein. It was also found that the polyunsaturated fatty acid content of hepatocytes was remarkedly stable with time of culture when the cells were incubated in serum-free, lipid-free medium. Thus, isolated hepatocytes maintained in serum-free medium appear to be a possible system for the evaluation of the effects of prior nutritional status on fatty acid metabolism in the whole animal, not subject to hormonal and other somatic influences which often complicate the interpretation of such nutritional studies.     

ω-3 Fatty acids in smooth muscle cell phospholipids increase membrane cholesterol efflux

The aim of our work was to determine whether fatty acid modifications in smooth muscle cell phospholipids affect cholesterol efflux and desorption. [³H]Cholesterol was used to label cholesterol pools in the whole cell or selectively in the plasma membrane. Cells were incubated for 12 h in order to increase oleate, linoleate, arachidonate, eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) in phospholipids. Cholesterol efflux was monitored using native or tetranitromethane modified high-density lipoprotein₃ (HDL₃). When all cholesterol pools were labeled, the efflux from cells treated with different fatty acids were not different. Plasma membrane cholesterol efflux remained unchanged after oleate, linoleate or arachidonate treatments, but was markedly increased after EPA and DHA enrichment, both with native HDL₃ and with tetranitromethane-high-density lipoprotein. These results suggest that the positive effects of n−3 fatty acid consumption on the atherosclerotic process could be linked in part to an increase in plasma membrane cholesterol efflux from vascular smooth muscle cells.     

Fatty acid metabolism in L1210 murine leukemia cells: Differences in modification of fatty acids incorporated into various lipids

L1210 leukemia cells can utilize all of the main fatty acids that normally are present in the ascites fluid in which they grow. This finding is consistent with the view that L1210 cells derive most of their fatty acids from the ascites fluid. From 80–90% of each fatty acid was incorporated into cell lipids without structural modification, suggesting that the lipid composition of these cells can be altered by changing the type of fatty acids to which they are exposed. Most importantly, the palmitate that was subsequently incorporated into total cell phospholipids was elongated and desaturated somewhat more than that incorporated into triglycerides. This difference was due primarily to more extensive modification of the palmitate incorporated into the ethanolamine phosphoglycerides fraction. Although there was no difference between total phospholipids and triglycerides with linoleate, more of the linoleate incorporated into ethanolamine phosphoglycerides was elongated and further desaturated than that incorporated into choline phosphoglycerides and triglycerides. These findings indicate that fatty acids incorporated into various cell lipid fractions are not structurally modified to the same extent. There appears to be greater modification of fatty acid used for ethanolamine phosphoglyceride synthesis as compared with triglyceride and choline phosphoglyceride synthesis.     

Dietary fat composition and tocopherol requirement: III. Quantitative studies on the relationship between dietary linoleate and vitamin E

The present study has involved biologically titrating linoleate vs. vitamin E using the male rat as the indicator. In the first of the titration studies, the dietary tocopherol level was held constant, while in the second study the linoleate intake was held constant. The investigation was conducted with male rats since these have a much higher linoleate requirement than females. By first depleting such animals of their stores of essential fatty acids by feeding a fat-free diet from weaning, a sensitive test organism was provided. These animals have an immediate need for linoleate during the repletion periods. If an imbalance between linoleate and vitamin E content existed in any of the dietary regimens, such an imbalance would have been more likely noted in test animals actively metabolizing the ingested linoleate. Based upon various nutritional and biochemical indices, the amount of tocopherol ordinarily included in the basic diets fed to our rats, 0.01% as dl-alphatocopheryl acetate, was adequate even when the diet provided up to 5% linoleate; an amount corresponding to ca. 12% of the total calories and providing a ratio of linoleate to the tocopherol of ca. 500:1. In the reverse biological titration with all test diets now providing the constant level of 5% linoleate, ratios of linoleate to vitamin E were satisfactory even in a ratio of as much as 2500:1 (or 0.4 mg gram of vitamin E per polyunsaturated fatty acid). The control animals continued on the fat-free diet indicated that there is a need for added tocopherols even in the absence of linoleate according to a number of biochemical indices. Based upon a number of accepted bioanalytical approaches, the minimum requirement for linoleate by the fat-depleted male rat was found to be between 100–200 mg/day or ca. 1–2% of the caloric intake. Although the fatty acid composition of tissue lipid fractions is markedly affected by the amount of linoleate in the diet, dietary tocopherol supplements have little effect on these values.     

Age-strain interrelations in lipid metabolism of rats

Various aspects of lipid metabolism were compared in Fisher 344 (F) and Sprague-Dawley (SD) rats aged 2, 6, 12, 18 and 24 months. The analyses included free and total cholesterol of serum and liver, LCAT, hepatic HMG-CoA reductase, cholesterol 7α-hydroxylase, fatty acid synthetase, acetyl CoA carboxylase and cholesterol synthesis from acetate or mevalonate. The body weight of SD rats increases with age whereas that of F rats plateaus at 9–12 months. Liver and aorta cholesterol levels were comparable for the 2 strains. Serum cholesterol varied but was usually lower in F rats. HMG-CoA reductase and cholesterol 7α-hydroxylase activities were not significantly different. Cholesterol synthesis from acetate was significantly higher only in 2-month-old F rats; synthesis from mevalonate was similar at each level. Acetyl CoA carboxylase and fatty acid synthetase activity were generally higher in F rats at every age level. The major difference between F and SD rats is in their pattern of weight gain with age. Differences in lipid metabolism are most marked between the young (2-month) rats.