Cytokine-Induced Accumulation of Very Long-Chain Fatty Acids in Rat C6 Glial Cells: Implication for X-Adrenoleukodystrophy

Abstract: X-Adrenoleukodystrophy (X-ALD) is an inherited metabolic disorder of very long-chain fatty acids (VLCFA) with subsequent manifestation of neuroinflammatory disease. To investigate the possible role of proinflammatory cytokines in the X-ALD disease process, we examined the effect of cytokines on the metabolism of VLCFA in C6 glial cells expressing oligodendrocyte-like properties. C6 glial cells under serum-free conditions were treated with different combinations of cytokines (tumor necrosis factor-α, interleukin-1β, interferon-γ) or cytokine with bacterial lipopolysaccharide (LPS). Cytokine-treated C6 cells had higher concentrations of VLCFA, measured as percent weight and also as C_26:0/C_22:0 ratio, which were 300–400% as compared with the controls. We also found increased levels of C_26:1 in cytokine-treated cells. The accumulation of VLCFA paralleled the decrease (35–55%) in peroxisomal β-oxidation activity and a 12- to 14-fold increase in the production of nitric oxide (NO). Individual cytokines were unable either to produce NO or to increase the levels of VLCFA in C6 cells. Inhibition of cytokine-induced NO production by l -N-methylarginine, an inhibitor of NO synthase (NOS), and N-acetylcysteine, an inhibitor of cytokine-mediated induction of inducible NOS, normalized the peroxisomal β-oxidation activity and the levels of VLCFA, suggesting a role for the proinflammatory cytokines and NO toxicity in the neuropathological changes associated with abnormal VLCFA metabolism (e.g., X-ALD). X-ALD is a peroxisomal disease having deficient oxidation of VLCFA, resulting in the excessive accumulation of VLCFA in all tissues but especially in brain. We observed greater increase in levels of VLCFA in the inflammatory region of ALD brain (in the demyelinating plaque and the area around the plaque) than in the normal-looking area away from the plaque; this also indicates that cytokines in the proinflammatory region may augment the VLCFA defect caused by the inherited abnormality in X-ALD brain. Although C6 glial cultured cells do not reflect the X-ALD model precisely, the observed relationship between the cytokine-induced inhibition of the oxidation of VLCFA, excessive accumulation of VLCFA, and excessive production of NO and their normalization by inhibitors of NOS in C6 glial cells suggests that NO-mediated toxicity may play a role in VLCFA-associated neuroinflammatory diseases (e.g., X-ALD).     

Metabolic Engineering of Plants to Produce Very Long-chain Polyunsaturated Fatty Acids

Very long-chain polyunsaturated fatty acids (VLCPUFAs) are essential for human health and well-being. However, the current sources of these valuable compounds are limited and may not be sustainable in the long term. Recently, considerable progress has been made in identifying genes involved in the biosynthesis of VLCPUFAs. The co-expression of these genes in model systems such as plant embryos or yeast provided many valuable insights into the mechanisms of VLCPUFA synthesis. The recent successful reconstitution of pathways leading to the synthesis of arachidonic acid, eicosapentaenoic acid and finally docosahexaenoic acid in oil-seed plants indicates the feasibility of using transgenic crops as alternative sources of VLCPUFAs. The various approaches used to attain these results and the specific constraints associated with each approach are discussed.     

Effects of Cyclohexanonic Long-Chain Fatty Alcohol, tCFA15 on Amino Acids in Diabetic Rat Brain: A Preliminary Study

The aim of this study was to evaluate the effects of streptozotocin-induced type 1diabetes and a subchronic treatment with cyclohexanonic long-chain fatty alcohol, 3-(15-hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen 1-one (tCFA15) on contents of amino acids including aspartate, glutamate, glutamine, GABA, glycine, taurine, alanine, serine, threonine, and arginine in the prefrontal cortex, hippocampus and striatum. Levels of glutamate, threonine, taurine, alanine, arginine, and the ratio of glutamate/glutamine were altered region-differently in the brain of diabetic rats. However, tCFA15 region-specifically antagonized the changes in taurine and arginine levels and the ratio of glutamate/glutamine. The alteration in glutamate/glutamine ratio may indicate that experimental models of type 1 diabetes have abnormalities of neuron-gria interaction in brain.     

Co-production of lactic acid and chitin using a pelletized filamentous fungus Rhizopus oryzae cultured on cull potatoes and glucose

Aims: This paper developed a novel process for lactic acid and chitin co-production of the pelletized Rhzious oryzae NRRL 395 fermentation using underutilized cull potatoes and glucose as nutrient source. Methods and Results: Whole potato hydrolysate medium was first used to produce the highest pelletized biomass yield accompanying the highest chitin content in biomass. An enhanced lactic acid production then followed up using batch, repeated batch and fed batch culture with glucose as carbon source and mixture of ammonia and sodium hydroxide as neutralizer. The lactic acid productivity peaked at 2·8 and 3 g l⁻¹ h⁻¹ in repeated batch culture and batch culture, respectively. The fed batch culture had the highest lactate concentration of 140 g l⁻¹. Conclusions: Separation of the biomass cultivation and the lactic acid production is able to not only improve lactic acid production, but also enhance the chitin content. Cull potato hydrolysate used as a nutrient source for biomass cultivation can significantly increase both biomass yield and chitin content. Significance and Impact of the Study: The three-step process using pelletized R. oryzae fermentation innovatively integrates utilization of agricultural residues into the process of co-producing lactic acid and chitin, so as to improve the efficiency, revenues and cost of fungal lactic acid production.     

Hydrocarbons, Fatty Acids, and Lipids of Freshwater Grasses of the Potamogetonaceae Family

The composition of hydrocarbons, fatty acids, and of total, polar, and neutral lipids was studied in freshwater Potamogetonaceae grasses collected in two different regions of the Volga river. More than 40 fatty acids and hydrocarbons were separated and identified by chromato-mass spectrometry. The variability of lipid characteristics of plants of the same family is discussed.     

Molecular Species of Phosphatidylcholine Containing Very Long Chain Fatty Acids in Human Brain: Enrichment in X-Linked Adrenoleukodystrophy Brain and Diseases of Peroxisome Biogenesis Brain

Molecular species of phosphatidylcholine containing unsaturated (i.e., monoenoic and polyenoic) 32- to 40-carbon (very long chain) fatty acids (VLCFA-PC) are present in normal human brain, the fatty acid composition changing significantly with development. There is a marked increase in the concentration and a change in the polyenoic VLCFA composition of these molecular species in brains of patients with inherited defects in peroxisomal biogenesis [Zellweger's syndrome, neonatal adrenoleukodystrophy (ALD), and infantile Refsum's disease]. In contrast, there is a marked increase in monoenoic VLCFA-PC in X-linked ALD whereas molecular species containing polyenoic VLCFA are minor components.     

Utilization of Uniformly Labeled 13C-Polyunsaturated Fatty Acids in the Synthesis of Long-Chain Fatty Acids and Cholesterol Accumulating in the Neonatal Rat Brain

Abstract: Polyunsaturated fatty acids are needed for normal neonatal brain development, but the degree of conversion of the 18-carbon polyunsaturated fatty acid precursors consumed in the diet to their respective 20-and 22-carbon polyunsaturates accumulating in the brain is not well known. In the present study, in vivo ¹³C nuclear magnetic resonance spectroscopy was used to monitor noninvasively the brain uptake and metabolism of a mixture of uniformly ¹³C-enriched 16-and 18-carbon polyunsaturated fatty acid methyl esters injected intragastrically into neonatal rats. In vivo NMR spectra of the rat brain at postnatal days 10 and 17 had larger fatty acid signals than in uninjected controls, but changes in levels of individual fatty acids could not be distinguished. One day after injection of the U-¹³C-polyunsaturated fatty acid mixture, ¹³C enrichment (measured by isotope ratio mass spectrometry) was similar in brain phospholipids, free fatty acids, free cholesterol, and brain aqueous extract; ¹³C enrichment remained high in the phospholipids and cholesterol for 15 days. ¹³C enrichment was similar in the main fatty acids of the brain within 1 day of injection but 15 days later had declined in all except arachidonic acid while continuing to increase in docosahexaenoic acid. These changes in ¹³C enrichment in brain fatty acids paralleled the developmental changes in brain fatty acid composition. We conclude that, in the neonatal rat brain, dietary 16-and 18-carbon polyunsaturates are not only elongated and desaturated but are also utilized for de novo synthesis of long-chain saturated and monounsaturated fatty acids and cholesterol.     

Role of the Blood-Brain Barrier in the Formation of Long-Chain ω-3 and ω-6 Fatty Acids from Essential Fatty Acid Precursors

Elongated, more highly polyunsaturated derivatives of linoleic acid (18:2 omega-6) and linolenic acid (18:3 omega-3) accumulate in brain, but their sites of synthesis and mechanism of entry are not well characterized. To investigate the role of the blood-brain barrier in this process, cultured murine cerebromicrovascular endothelia were incubated with [1-14C]18:2 omega-6 or [1-14C]18:3 omega-3 and their elongation/desaturation products determined. The major metabolite of 18:2 omega-6 was 20:4 omega-6, whereas the primary product from 18:3 omega-3 was 20:5 omega-3. Although these products were found primarily in cell lipids, they were also released from the cells and gradually accumulated in the extracellular fluid. Eicosanoid production was observed from the 20:4 omega-6 and 20:5 omega-3 that were formed. No 22:5 omega-6 or 22:6 omega-3 fatty acids were detected, suggesting that these endothelial cells are not the site of the final desaturation step. Although the uptake of 18:3 omega-3 and 18:2 omega-6 was nearly identical, 18:3 omega-3 was more extensively elongated and desaturated. Competition experiments demonstrated a preference for 18:3 omega-3 by the elongation/desaturation pathway. These findings suggest that the blood-brain barrier can play an important role in the elongation and desaturation of omega-3 and omega-6 essential fatty acids during their transfer from the circulation into the brain.     

Age‐Related Changes in Fatty Acids in Obese Offspring of Streptozotocin‐Induced Diabetic Rats

Objective: The long‐term effects of fetal hyperinsulinemia, time course of changes in liver and very‐low‐density lipoprotein (VLDL) lipid levels and fatty acid compositions were investigated in obese offspring of streptozotocin‐induced mildly diabetic rats. Research Methods and Procedures: Mild hyperglycemia in pregnant rats was induced by intraperitoneal injection of streptozotocin on day 5 of gestation. Control pregnant rats were injected with citrate buffer. Liver and VLDL lipids and fatty acids were analyzed in offspring at different ages. Results: At birth, obese pups had higher VLDL triglyceride levels, saturated fatty acids, and C20:4n‐6. They also had lower C18:2n‐6 proportions in VLDL triglycerides, phospholipids, and cholesteryl esters than controls pups. In 1‐month‐old male and female obese rats, VLDL and liver lipid amounts were similar to those in their respective controls; however, high levels of C18:2n‐6 and C20:4n‐6 were noted in liver and VLDL lipids. At the age of 2 months, liver and VLDL triglyceride levels were higher in obese females than in control females. Fatty acid abnormalities seen in obese rats included low C18:3n‐3 and high C22:6n‐3 proportions in liver triglycerides and phospholipids. At the age of 3 months, obese rats, both males and females, compared with control animals, had higher VLDL and hepatic lipids with reduced C20:4n‐6 levels and polyunsaturated/saturated fatty acids ratios in hepatic and VLDL triglycerides and phospholipids. Discussion: Fetal obesity, associated with alterations in VLDL lipid fatty acid composition, represents an important risk factor for adult obesity and diabetes.     

Immobilization and stability of a Rhizopus oryzae lipase expressed in Pichia pastoris: comparison between native and recombinant variants

The stability of a soluble extract containing a recombinant lipase from Rhizopus oryzae (Cursive) lipase (rROL) produced by Pichia pastoris (Cursive), as well as that for the commercial extract containing the lipase produced by the native organism (nROL), was investigated. The results showed higher residual activity values of the commercial protein compared with the recombinant one. Moreover, two different kinds of support, the polypropylene powder EP100 and Eupergit®C, were tested to immobilize the enzymes. The residual activity of the immobilizated derivatives was also tested to determine whether their stability was enhanced. The results showed a slight improvement in rROL using both supports but a decrease in nROL using Eupergit®C. The study of the residual activity of soluble and immobilized enzymes was performed by means of a central composite rotatable experiment design. In addition, EP100 adsorption isotherms were determined. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Relationships of Plasma Leptin Levels to Changes in Plasma Free Fatty Acids in Women Who Are Lean and Women Who Are Abdominally Obese

HENNES, MAGDA MI, ARNAVAZ DUA, DIANA L MAAS, GABRIELE E SONNENBERG, GLENN R KRAKOWER, AHMED H KISSEBAH. Relationships of plasma leptin levels to changes in plasma free fatty acids in women who are lean and women who are abdominally obese. Regulation of leptin production by the hormonal and metabolic milieu is poorly understood. Because abdominal obesity is commonly associated with elevated plasma free fatty acid (FFA) flux, we examined the effects of augmenting FFA on plasma leptin levels in women who were lean and of suppressing FFA in women with abdominal obesity. In study 1, nine subjects who were lean, after a 12-hour overnight fast, received either intravenous saline or Intralipid plus heparin to increase the plasma FFA concentration to approximately 1000 μmol/ L. After 3 hours of additional fasting, subjects underwent 3-hour hyperglycemic clamps. In study 2, seven subjects with abdominal obesity were evaluated by a similar protocol, but lipolysis and plasma FFA flux were instead maximally suppressed by acipimox. In the individuals who were lean, leptin levels were unchanged during clamping. Increasing plasma FFA reduced plasma leptin from 7.66 ± 0.66 to 7.05 ±0 0.66 (p=0.03), but 3 hours of hyperglycemia plus hyperinsulinemia had no additional effect on leptin levels (7.15 ± 0.71). Basal leptin levels, 4-fold higher in the subjects with obesity, were reduced from 34.6 ± 2.4 μg/L to 32.3 ± 1.1 μg/L (p=0.004) during the clamp period. When plasma FFA flux was suppressed, however, plasma leptin levels after clamped hyperglycemia/hyperinsulinemia were increased to 38.9 ± 1.2 μg/L (p=0.014 vs. time 0 and 0.001 vs. saline protocol). Changes in leptin concentrations are not correlated with changes in FFA. These results suggest that plasma FFA concentration does not regulate plasma leptin levels in basal, extended fasting, or hyperglycemic/hyperinsulinemic states.     

Fatty Acid Composition of Mitochondrial Membrane Lipids in Cultivated (Zea mays) and Wild (Elymus sibiricus) Grasses

The fatty acid composition of mitochondrial membrane lipids from seedlings of two grass species differing in their chilling tolerance, elymus (Elymus sibiricus) and maize (Zea mays), was studied by gas-liquid chromatography. An increased chilling tolerance of elymus, a perennial wild species, seems to be caused by a high content of unsaturated fatty acid residues in the total membrane lipids; linoleic (41.9%) and -linolenic (23.4%) acids predominated among these fatty acids. The contents of linoleic and -linolenic acids in the total lipids of maize membranes were 57.5% and 5.8%, respectively. Polar lipids of elymus mitochondrial membranes were characterized by about similar contents of linoleic (33.6%) and -linolenic (31.3%) acids. Linoleic acid (51.7%) predominated in maize mitochondrial polar lipids, whereas the -linolenic acid content was 9.0%. A high level of 3-desaturase activity in the mitochondrial membranes seems to be an important factor of elymus chilling tolerance. This high activity seems to be developed as an adaptation in the course of evolution of this species.     

Concomitant Increases in the Levels of Choline and Free Fatty Acids in Rat Brain: Evidence Supporting the Seizure-Induced Hydrolysis of Phosphatidylcholine

The main objective of this study was to determine whether the excitotoxic cholinesterase inhibitor soman increases the catabolism of phospholipids in rat brain. Injections of soman (70 micrograms/kg, s.c.), at a dose that produced toxic effects, increased the levels of both free fatty acids (175-250% of control) and free choline (250% of control) in rat cerebrum 1 h after administration. All fatty acids contained in brain phosphatidylcholine were elevated significantly including palmitic (16:0), stearic (18:0), oleic (18:1), arachidonic (20:4), and docosahexaenoic (22:6) acids. The changes observed were consistent with those reported to occur following ischemia and the administration of other convulsants. Pretreatment of rats with the anticonvulsant diazepam (4 mg/kg, i.p.) prevented both the signs of soman toxicity and the soman-induced increase of choline and free fatty acids. Diazepam alone did not affect the levels of choline or free fatty acids, cholinesterase activity, or soman-induced cholinesterase inhibition, suggesting that soman toxicity involves a convulsant-mediated increase in phosphatidylcholine catabolism. In addition, administration of the convulsant bicuculline, at a dose that produces seizures and increases the levels of free fatty acids in brain, significantly increased the levels of choline. Results suggest that excitotoxic events enhance the hydrolysis of phosphatidylcholine in brain as evidenced by a concomitant increase in the levels of choline and free fatty acids.     

醋豆与大豆中脂肪酸含量的比较及营养学意义

目的:分析比较醋浸泡后大豆中的脂肪酸(FA)含量的变化,为大豆开发提供实验数据.方法:用石油醚提取总脂肪,甲基化后用气相色谱分析脂肪酸的含量分布.结果:醋浸泡28d后,醋豆总脂肪的相对含量趋于稳定,其中醋豆和大豆相比,软脂酸下降了1.21%,硬脂酸下降了0.58%,油酸下降了3.01%,亚油酸增加了1.32%,亚麻酸增加了6.32%.结论:饱和脂肪酸(SFA)和单不饱和脂肪酸的相对含量下降,多不饱和脂肪酸(PUFA)相对含量显著升高.醋大豆及大豆的多不饱和脂肪酸与饱和脂肪酸相对含量的比值分别为4.11和3.2.上述变化的从营养学或生理学的角度,更符合人体需求.     

Free Fatty Acids and Energy Metabolites in Ischemic Cerebral Cortex with Noradrenaline Depletion

Abstract: We tested whether cerebral noradrenaline (NA) may play a central role in mediating the increased production of free fatty acids (FFAs) during cerebral ischemia. Levels of FFAs, cyclic AMP, and NA, as well as ATP, ADP, and AMP, were measured in cerebral cortex during decapitation ischemia in rats 2 weeks after unilateral locus ceruleus lesion. Comparisons were made between the results obtained from the contralateral cortex with normal NA content and the NA-depleted ipsilateral cortex. Although NA depletion was associated with a diminished transient rise of cyclic AMP in response to ischemia, it failed to influence the magnitude of FFA increase or the decline of energy state within the 15-min period of ischemia. A more than twofold increase of total FFAs (sum of palmitic, stearic, oleic, arachidonic, and docosahexaenoic acids) was observed in both hemispheres at 1 min after decapitation, when energy failure became manifest. The increased production of FFAs continued throughout the 15 min of ischemia, with a preferential rise in the levels of stearic and arachidonic acids. There was an inverse correlation between FFA levels and total adenylate pool. The results do not support a major role for NA and cyclic AMP in increasing cortical FFAs during complete ischemia. Instead, they are consistent with the view that impaired oxidative phosphorylation activates deacylating enzymes. Disturbance of reacylation due to energy depletion is probably another factor contributing to the continuous increase of FFAs during prolonged ischemia.     

Effect of Hydroperoxy Fatty Acids on Acylation and Deacylation of Arachidonoyl Groups in Synaptic Phospholipids

The effect of hydroperoxy fatty acids on reactions involved in the acylation-deacylation cycle of synaptic phospholipids was studied in vitro, using nerve ending fraction isolated from rat forebrain. 15-Hydroperoxyeicosatetraenoic acid (15-HPETE), 13-hydroperoxylinoleic acid (13-HP 18: 2), and hydroperoxydocosahexaenoic acid (22:6 Hpx), at 25 microM final concentration, all inhibited the incorporation of [1-14C]arachidonate into synaptosomal phosphatidylinositol (PI), phosphatidylcholine (PC), and triacylglycerides by 50-80%. The lowest effective concentration of 15-HPETE and 13-HP 18:2 resulting in significant inhibition of the reacylation of PI was 5 microM, whereas the inhibition of [1-14C]arachidonate incorporation into PC required 10 and 5 microM hydroperoxy fatty acids, respectively. Cumene hydroperoxide and tert-butyl hydroperoxide at concentrations of 100 microM did not inhibit reacylation of PI and PC. Synthesis of labeled arachidonoyl-CoA from [1-14C]arachidonate was decreased by about 50% by 25 microM hydroperoxy fatty acids both in synaptosomes and in the microsomal fraction. Use of [1-14C]arachidonoyl-CoA as a substrate, to bypass the fatty acid activation reaction, revealed that activity of acyltransferase was not affected significantly by 25 microM 15-HPETE and 13-HP 18:2. At the same time, however, the hydrolysis of labeled arachidonoyl-CoA was substantially enhanced. Exposure of synaptosomes to 25 microM fatty acid hydroperoxides did not affect significantly the endogenous concentrations of five major free fatty acids. It is concluded that (1) among synaptic phospholipids, reacylation of PI and PC is the most susceptible to the inhibitory action of fatty acid hydroperoxides, and (2) the enzymes affected by these compounds in nerve endings are arachidonoyl-CoA synthetase and hydrolase.     

Alteration in Fatty Acid Composition of Adult Rat Brain Capillaries and Choroid Plexus Induced by a Diet Deficient in n-3 Fatty Acids: Slow Recovery After Substitution with a Nondeficient Diet

Wistar rats were fed for three generations with a semisynthetic diet containing either 1.5% sunflower oil (940 mg% of C18:2n-6, 6 mg% of C18:3n-3) or 1.9% soya oil (940 mg% of C18:2n-6, 130 mg% of C18:3n-3). At 60 days of age, the male offspring of the third generation were killed. The fatty acyl composition of isolated capillaries and choroid plexus was determined. The major changes noted in the fatty acid profile of isolated capillaries were a reduction (threefold) in the level of docosahexaenoic acid and, consequently, a fourfold increase in docosapentaenoic acid in sunflower oil-fed animals. The total percentage of polyunsaturated fatty acids was close to that in the soya oil-fed rats, but the ratio of n-3/n-6 fatty acids was reduced by threefold. In the choroid plexus, the C22:6n-3 content was also reduced, but by 2.6-fold, whereas the C22:5n-6 content was increased by 2.3-fold and the ratio of n-3/n-6 fatty acids was reduced by 2.4-fold. When the diet of sunflower oil-fed rats was replaced with a diet containing soya oil at 60 days of age, the recovery in content of n-6 and n-3 fatty acids started immediately after diet substitution; it progressed slowly to reach normal values after 2 months for C22:6n-5 and 2.5 months for C22:6n-3. The recovery in altered fatty acids of choroid plexus was also immediate and very fast. Recovery in content of C22:5n-6 and C22:6n-3 was complete by 46 days after diet substitution.     

Modulation by Unsaturated Fatty Acids of Norepinephrine- and Adenosine-Induced Formation of Cyclic AMP in Brain Slices

Abstract: The effect of linoleic acid on the formation of cyclic AMP in the slices of guinea pig cerebral cortex was examined. Treatment of the slices with linoleic acid resulted in an increase of basal and of norepinephrine-stimulated formation of cyclic AMP. The stimulatory effect on the basal level of cyclic AMP was not specific for linoleic acid: the potency of the fatty acid was related to the magnitude of unsaturation. In contrast, the enhancement of norepinephrine-stimulated formation of cyclic AMP seemed relatively specific for linoleic acid and arachidonic acid. Linoleic acid markedly enhanced the stimulated formation of cyclic AMP by histamine and adenosine, as well that by norepinephrine, without affecting that by excitatory amino acids and veratridine. Theophylline, adenosine deaminase, and 2'-deoxyadenosine antagonized the effect of linoleic acid. Linoleic acid enhanced the maximum responses to norepinephrine and adenosine without altering the ED₅₀ values for these agonists. When linoleic acid-treated slices were washed with Krebs-Ringer containing defatted bovine serum albumin, both enhancement of the response to norepinephrine and the amount of [¹⁴C]linoleic acid incorporated in a free form significantly diminished.     

Metabolic Turnover of Fatty Acids and Acylglycerols in Rat Sciatic Nerve

To explain the discrepancy between the low level and high metabolic activity of endoneurial free fatty acids (FFAs) and triacylglycerol (TG), levels of de novo synthesized FFA and acylglycerols were measured in rat sciatic endoneurium at various intervals after endoneurial microinjection of [14C]acetate. Soon after injection (less than 10 min), the [14C]acetate was metabolized to FFA and incorporated into diacylglycerol (DG), TG, sterols, ceramides, and various phospholipids. The proportions of 14C-labeled FFA, DG, TG, and ceramides to total 14C-labeled lipids decreased, whereas those of phospholipids and cerebrosides increased with time after injection. These findings suggest that rapid turnover of FFA and TG may contribute to their low level in sciatic endoneurium. The de novo synthesized fatty acids were largely incorporated into phosphatidylcholine (approximately 50% of total 14C-labeled phospholipids), probably via the cytidine nucleotide pathway using 1,2-DG as a metabolic intermediate. Hydrolysis of [14C]phosphatidylcholine revealed that fatty acids were labeled at both the C-1 (approximately 43%) and C-2 (approximately 57%) positions. On the other hand, a temporal association between decreased amounts of 14C-label in ceramides and increased amounts of 14C-label in sphingomyelin and galactocerebrosides supports the hypothesis that peripheral nerve galactocerebroside is derived, in vivo, from ceramide via acylation of sphingosine. This exclusive labeling of endoneurial lipids by endoneurial microinjection of labeled precursor provides a unique model for studying synthesis and metabolic turnover of membrane lipids in experimental neuropathies.