Fatty acid specificity for the synthesis of triacylglycerol and phosphatidylcholine and for the secretion of very-low-density lipoproteins and lysophosphatidylcholine by cultures of rat hepatocytes (original) (raw)
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Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism, 1987
The objective of these studies with rat hepatocytes in primary culture was to establish that: (a) membrane phospholipids would become enriched with the specific fatty acid supplemented to the media and (b) hepatocyte monolayer triacylglycerol synthetic rates were dependent on the type of fatty acid enrichment of the membrane phospholipids. Hepatocytes cultured in the absence of media lipid developed a phospholipid fatty acid composition which is indicative of an essential fatty acid deficiency. The extensive rise in 18: l(n-9) content indicated that A'-desaturase was active. The fatty acid composition of phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol in the microsomal-and mitochondrial-enriched fractions was highly dependent upon the type of fatty acid supplemented to the medium. Incorporation of fatty acids into phospholipids was rapid, and a new steady-state in fatty acid composition was achieved within approx. 36 h. Changes in the fatty acid composition of these hepatocyte phospholipid subclasses resulting from media supplementation with 18: 2/20 : 4(n-6) or 20: 5(n-3) were similar, but not identical, to changes which occurred in vivo as a result of consuming diets rich in 18: 2(n-6) or 20: 5(n-3). Hepatocyte lipogenesis was highly dependent upon the type of fatty acid supplemented to the medium. Prior conditioning with 16 : 0 increased triacylglycerol synthesis and secretion. Secretion of triacylglycerol was reduced by polyenoic fatty acid enrichment with 20 : 5(n-3) > 20 : 4/ 18 : 2(n-6). The suppression of triacylglycerol synthesis by 20: 5(n-3) was due to an increased (P < 0.05) diacylglycerol specific activity, which indicates that 20: 5(n-3) suppression of hepatic triacylglycerol production may be caused in part by the inhibition of diacylglycerol acyltransferase.
Essential fatty acid uptake and esterification in primary culture of rat hepatocytes
Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism, 1986
Primary cultures of adult rat hepatocytes were used to compare the uptake and esterification of essential polyunsaturated fatty acids (18 : 2, 20 : 3 and 20 : 4 of the n -6 series) with those of palmitic and oleic acids. The uptake of unesterified fatty acids was linearly related to the free fatty acid/albumin molar ratio for 14 h and did not depend on the unbound free fatty acid level. Whatever the initial free fatty acid/albumin molar ratio, it dropped to 0.5 f 0.1 mM after 14 h, thus showing that hepatocytes have a high capacity for clearing free fatty acids from the medium at high free fatty acid/albumin molar ratios. The free fatty acid uptake become saturable when the free fatty acid and albumin concentrations were raised and the free fatty acid/albumin ratio remained constant. This strongly suggests that albumin-hepatocyte interaction mediates free fatty acid uptake. This uptake was identical whatever the fatty acid tested and did not depend on the relative amounts of fatty acids when they were added simultaneously. Triacylglycerol accumulation and synthesis, monitored by labelled fatty acids, were related to the free fatty acid/albumin molar ratio and exhibited no specificity for the series of fatty acids tested. Triacylglycerols were enriched in all the fatty acids tested by up to 60%, and fatty acid incorporation into diacylglycerols and triacylglycerols reflected the free fatty acid composition of the medium. By contrast, neither the level nor the synthesis of phospholipids varied with free fatty acid/albumin, but the rate of phospholipid turnover depended on the fatty acids tested. Accumulation of these acids was smaller in phospholipids than in triacylglycerols. When linoleic and arachidonic acids were added together, phospholipids (especially phosphatidylethanolamine and phosphatidylinositol) were more enriched in arachidonic acid than triacylglycerols. This might be due to the specificity for fatty acid of the enzymes involved in phospholipid metabolism.
Biochemical Journal, 1999
Hypolipidaemic fatty acid derivatives and polyunsaturated fatty acids decrease concentrations of plasma triacylglycerol by mechanisms that are not fully understood. Because poor susceptibility to β-and\or ω-oxidation is apparently a determinant of the peroxisome proliferating and hypolipidaemic capacity of fatty acids and derivatives, the relative importance of activation of the peroxisome-proliferator-activated receptor α (PPARα), fatty acid oxidation and triacylglycerol synthesis were examined. We have compared the effects of differentially β-oxidizable fatty acids on these parameters in primary cultures of rat hepatocytes. Tetradecylthioacetic acid (TTA), 2-methyleicosapentaenoic acid and 3-thia-octadecatetraenoic acid, which are non-β-oxidizable fatty acid derivatives, were potent activators of a glucocorticoid receptor (GR)-PPARα chimaera. This activation was paradoxically reflected in an substantially increased oxidation of [1-"%C]palmitic acid and\or oleic acid. The incorporation of [1-"%C]palmitic acid and\or oleic acid into cell-associated and secreted triacylglycerol was decreased by 15-20 % and 30 % respectively with these non-β-oxidizable fatty acid derivatives. The CoA ester of TTA inhibited the esterification of 1,2-
The Journal of Lipid Research
The mechanism for the reduced hepatic production of triacylglycerol in the presence of eicosapentaenoic acid was explored in short-term experiments using cultured parenchymal cells and microsomes from rat liver. Oleic, palmitic, stearic, and linoleic acids were the most potent stimulators of tria~yl[~H]glycerol synthesis and secretion by hepatocytes, whereas erucic, a-linolenic, y-linolenic, arachidonic, docosahexaenoic, and eicosapentaenoic acids (in decreasing order) were less stimulatory. There was a linear correlation (r= 0.85, PcO.01) between synthesis and secretion of tria~yl[~H]glycerol for the fatty acids examined. The extreme and opposite effects of eicosapentaenoic and oleic acids on triacylglycerol metabolism were studied in more detail. With increasing number of free fatty acid molecules bound per molecule of albumin, the rate of synthesis and secretion of tria~yl[~H]glycerol increased, most markedly for oleic acid. Cellular uptake of the two fatty acids was similar, but more free eicosapentaenoic acid accumulated intracellularly. Eicosapentaenoic acid caused higher incorporation of ['Hlwater into phospholipid and lower incorporation into triacylglycerol and cholesteryl ester as compared to oleic acid. No difference was observed between the fatty acids on incorporation into cellular free fatty acids, monoacylglycerol and diacylglycerol. The amount of some 16-and 18-carbon fatty acids in triacylglycerol was significantly higher in the presence of oleic acid compared with eicosapentaenoic acid. Rat liver microsomes in the presence of added 1,2-dioleoylglycerol incorporated eicosapentaenoic acid and eicosapentaenoyl-CoA into triacylglycerol to a lesser extent than oleic acid and its CoA derivative. Decreased formatiop of triacylglycerol was also observed when eicosapentaenoyl-CoA was given together with oleoyl-CoA, whereas palmitoyl-CoA, stearoyl-CoA, linoleoyl-CoA, linolenoyl-CoA, and arachidonoyl-CoA had no inhibitory effect.
Biochimica et biophysica acta, 1988
Lipid emulsions were prepared with a similar size and lipid composition to natural lymph chylomicrons, but in which the surface phospholipid was either egg phosphatidylcholine, dioleoyl-, dimyristoyl-, dipalmitoyl- or 1-palmitoyl-2-oleoylphosphatidylcholine (EYPC, DOPC, DMPC, DPPC or POPC). When injected into the bloodstream of conscious rats, the emulsions containing EYPC or POPC were metabolized similarly to natural chylomicrons, consistent with rapid lipoprotein lipase-mediated hydrolysis of triacylglycerols, followed by hepatic uptake of the remnants derived from the emulsions. Phospholipids from the injected emulsions were removed more slowly and became associated with the high-density lipoprotein fractions of the plasma. Emulsions containing DPPC were metabolized differently. Triacylglycerols disappeared very slowly from plasma, indicating lack of hydrolysis by lipoprotein lipase, and phospholipid radioactivity did not transfer to high-density lipoprotein. With emulsions conta...
The Journal of nutrition, 1998
The regulation of plasma lecithin:cholesterol acyltransferase (LCAT) expression is not well understood. Although oleic acid increases both the secretion of triglycerides and LCAT by primary rat hepatocytes, the effect of other fatty acids (FA) on LCAT secretion is not known. This study was designed to examine the effect of FA on the hepatic secretion of LCAT, triglyceride and apolipoprotein A-1 (apoA-1). Primary rat hepatocytes were incubated with serum-free medium, supplemented with individual FA (0-1 mmol/L) for 22-24 h. Preliminary studies indicated a linear secretion of LCAT up to 24 h in both control and FA-treated cells. When hepatocytes were incubated with 1 mmol/L FA, the LCAT secretion increased 50-100% (P < 0.01) in the presence of the 18-carbon FA (stearic, oleic, elaidic and linoleic acids), whereas the presence of butyric, lauric and palmitic acids had no significant effect. LCAT secretion decreased (P < 0.01) in the presence of docosahexaenoic acid (DHA). All FA ...
Biochimica et biophysica acta, 1986
Primary cultures of rat hepatocytes were used to study the effects of eicosapentaenoic and oleic acid on synthesis and secretion of triacylglycerols associated with very low density lipoproteins. From the experiments the following was observed. Oleic acid markedly stimulates secretion as well as synthesis of triacylglycerols, whereas eicosapentaenoic acid causes very little or no increase in secretion or synthesis as compared to a fatty-acid-free medium. The effects could already be observed after 15 min incubation. The inhibitory effect of eicosapentaenoic acid is reversible within 1-2 h. Eicosapentaenoic acid inhibits much of the stimulatory effect of oleic acid on synthesis and secretion of triacylglycerols. The cellular uptake of eicosapentaenoic acid is somewhat higher than that of oleic acid and the metabolism of these fatty acids to acid-soluble materials is similar. Eicosapentaenoic acid does not affect the secretory pathway of triacylglycerols per se. From these results it ...
Factors influencing triacylglycerol synthesis in permeabilized rat hepatocytes
Biochemical Journal, 1992
Rat hepatocytes were treated with Staphylococcus aureus ax-toxin to permeabilize their plasma membrane for lowmolecular-mass compounds. During incubation with I mm labelled fatty acid, phosphatidate and, less clearly, lysophosphatidate rapidly reached a steady state, whereas labelled diacylglycerol accumulated to some extent, at least in the absence of exogenous CDP-choline. Esterification and oxidation were linearly related to the fatty acid concentration, and there was no indication for saturation with acyl-CoA. However, when permeabilized cells were incubated with labelled sn-glycerol 3-phosphate and 1 mm unlabelled fatty acid, glycerolipid synthesis and the level of esterification intermediates reached a plateau between 0.25 and 0.50,umol of the triose phosphate/ml. The synthesis of phosphatidylcholine was dependent on addition of CDP-choline. In presence of the latter, diacylglycerol no longer accumulated and triacylglycerol synthesis was suppressed, although the sum of synthesized diacylglycerol, triacylglycerol and phosphatidylcholine remained constant. This indicates that the same pool of diacylglycerol is shared by cholinephosphotransferase and diacylglycerol acyltransferase and that the relative activity of these enzymes depends on the CDPcholine supply. Comparison of the levels of the esterification intermediates with the activity of the respective steps of the pathway reveals that, at a fixed fatty acid concentration, glycerophosphate acyltransferase determines the esterification rate, whereas lysophosphatidate acyltransferase and, at low CDP-choline levels, diacylglycerol acyltransferase approach saturation at elevated sn-glycerol 3-phosphate concentration. There is, however, no indication for a regulatory role of phosphatidate phosphohydrolase in this system. The significance of these findings for the regulation of triacylglycerol synthesis under conditions in vivo is discussed.