Developmental changes of the lipidic part of the neutral glycosphingolipids of the rat stomach

Neutral glycolipids were purified from the glandular part of the stomach of rats of different ages from 20 days of gestation to 60 days after birth. The two major glycolipids were identified as glucosylceramide and isogloboside. Free ceramide was also detected. The concentrations of these sphingolipids remained almost stable with development. Monohexosylceramide contained 55 and 68% of 2-hydroxylated fatty acids at 20 and 22 days of gestation, respectively, and 82% in the adult. Its three major bases, C18-sphingenine, C18- and C20-4D-hydroxysphinganine were characterized by gas-liquid chromatography and mass spectrometry of their N-acetyl-O-trimethylsilyl derivatives. The occurrence of the bases changed with development. C18-sphingenine contributed for 26% of the bases at birth and 65% in the adult. Conversely, C18-4D-hydroxysphinganine contributed for 35% of the bases at birth and 9% in the adult. The ceramide part of isogloboside consisted of nonhydroxylated fatty acids and mainly C18-sphingenine throughout development. The percentage of long-chain fatty acids was higher in older animals. These results stressed the specificity of the lipidic part of the rat gastric glycolipids and their specific evolution during the development.     

gAd-globular head domain of adiponectin increases fatty acid oxidation in newborn rabbit hearts

Adiponectin is an adipocyte-derived hormone that has a number of metabolic effects in the body, including the control of both glucose and fatty acid metabolism. The globular head domain of adiponectin, gAd, has also been shown to increase fatty acid oxidation in skeletal muscle. Within days after birth, a rapid increase in fatty acid oxidation occurs in the heart. We examined whether adiponectin or gAd plays a role in this maturation of cardiac fatty acid oxidation. Plasma adiponectin increased in newborn rabbits following birth: 1.2 +/- 0.3 microg/ml in 1-day-old, 6.8 +/- 1.8 microg/ml in 7-day-old, and 45 +/- 5 microg/ml in 6-week-old rabbits. Because plasma insulin levels decrease and remain low throughout the suckling period, and because this decrease may contribute to the maturation of fatty acid oxidation, we examined the effects of adiponectin and gAd on fatty acid oxidation in isolated perfused 1-day-old rabbit hearts in the presence or absence of 100 microunits/ml insulin. Adiponectin (10 microg/ml) did not alter fatty acid oxidation in the presence of insulin. In the absence of insulin, the addition of recombinant gAd (1.5 microg/ml) increased fatty acid oxidation compared with control (129 +/- 18 versus 66 +/- 11 nmol.g dry weight(-1).min(-1), respectively (p < 0.05). In 7-day-old hearts, where fatty acid oxidation rates were 5-fold higher than 1-day-old hearts, gAd did not alter fatty acid oxidation rates. The increase in fatty acid oxidation in 1-day-old hearts occurred independently of changes in 5'-AMP-activated protein kinase, acetyl-CoA carboxylase, or malonyl-CoA. The effect of gAd on fatty acid oxidation was reversed in the presence of 100 microunits/ml insulin. These results suggest that a decrease in plasma insulin and increase in gAd are involved in the increase of cardiac fatty acid oxidation in the immediate newborn period.     

Age and sex characteristics of phospholipid composition of Wistar rat erythrocytes during postnatal ontogenesis

The composition of phospholipids and fatty acids of erythrocyte lipids of intact mature Wistar rats and their progeny in the age of 10, 15, 20, 30, 40, 60, 90 and 110 days was studied with the methods of micro-thin-layer and gas-liquid chromatography. Sex features and age mechanism of erythrocyte lipids composition forming in physiologically standard conditions were established. The fact that the composition of erythrocyte lipids of the untreated grown-up male rats in comparison with the female species is characterised with a lower concentration of phosphatidylinositol, a higher level of phosphatidylcholine and sphingomyelin, a higher saturation level of molecular types of phospholipids, was described. Three age periods of homeostasis of the erythrocyte lipids of the intact rats were distinguished. The postembryonic period (30-40 days after birth) is characterised with forming of the basic pool of erythrocyte lipids: getting to the stability in levels of the basic components of phospholipids and their fatty acids, coming out of phosphatidylinositol by the 40th day after birth. The period of sex-growing (60 days) is accompanied by spasmodic changes in the levels of fatty acids which is most common for female species: rapid increase in saturated and omega 3 polyunsaturated fatty acids, decrease in monoene and omega 6 polyene fatty acids. The period of maturity (110 days) is characterised by a slow metabolism of structural lipids and a stabilisation of their composition.     

De novo lipogenesis in the differentiating human adipocyte can provide all fatty acids necessary for maturation

The primary products of de novo lipogenesis (DNL) are saturated fatty acids, which confer adverse cellular effects. Human adipocytes differentiated with no exogenous fat accumulated triacylglycerol (TG) in lipid droplets and differentiated normally. TG composition showed the products of DNL (saturated fatty acids from 12:0 to 18:0) together with unsaturated fatty acids (particularly 16:1n-7 and 18:1n-9) produced by elongation/desaturation. There was parallel upregulation of expression of genes involved in DNL and in fatty acid elongation and desaturation, suggesting coordinated control of expression. Enzyme products (desaturation ratios, elongation ratios, and total pathway flux) were also correlated with mRNA levels. We used (13)C-labeled substrates to study the pathway of DNL. Glucose (5 mM or 17.5 mM in the medium) provided less than half the carbon used for DNL (42% and 47%, respectively). Glutamine (2 mM) provided 9-10%, depending upon glucose concentration. In contrast, glucose provided most (72%) of the carbon of TG-glycerol. Pathway analysis using mass isotopomer distribution analysis (MIDA) revealed that the pathway for conversion of glucose to palmitate is complex. DNL in human fat cells is tightly coupled with further modification of fatty acids to produce a range of saturated and unsaturated fatty acids consistent with normal maturation.     

Adenovirus-mediated rescue of lipoprotein lipase-deficient mice. Lipolysis of triglyceride-rich lipoproteins is essential for high density lipoprotein maturation in mice.

Lipoprotein lipase (LPL) is the rate-limiting enzyme for the hydrolysis of triglycerides and the subsequent uptake of free fatty acids in extrahepatic tissues. Deficiency of LPL in humans (Type I hyperlipoproteinemia) is associated with massive chylomicronemia, low high density lipoprotein (HDL) cholesterol levels, and recurrent attacks of pancreatitis when not controlled by a strict diet. In contrast to humans, homozygous LPL knock-out mice (L0) do not survive suckling and die between 18 and 24 h after birth. In this study, an adenovirus-based protocol was utilized for the transient expression of LPL during the suckling period in an effort to rescue L0 mice. After a single intraperitoneal injection of 5x10(9) plaque-forming units of LPL-expressing virus immediately after birth, more than 90% of L0 mice survived the first days of life. 3% of L0 mice survived the entire suckling period and lived for up to 20 months, although LPL activity in mouse tissues and postheparin plasma was undetectable in all animals after 6 weeks of age. Adult LPL-deficient mice were smaller than their littermates until 2-3 months of age and exhibited very high triglyceride levels in the fed (4997 +/- 1102 versus 113.4 +/- 18.7 mg/dl) and fasted state (2007 +/- 375 versus 65.5 +/- 7.4 mg/dl). Plasma total cholesterol levels, free fatty acids, and ketone bodies were elevated in L0 mice, whereas plasma glucose was normal. Most strikingly, L0 mice lacked apoA-I-containing prebeta-HDL particles as well as mature HDL resulting in undetectable HDL cholesterol and HDL-apoA-I levels. HDL deficiency in plasma was evident despite normal apoA-I mRNA levels in the liver and normal apoA-I protein levels in plasma, which were predominantly found in the chylomicron fraction. The absence of prebeta-HDL and mature HDL particles supports the concept that the lipolysis of triglyceride-rich lipoproteins is an essential step for HDL maturation.     

Adrenalectomy of Rats Results in Hypomyelination of the Central Nervous System

The effect of adrenalectomy on CNS myelin accumulation was investigated to determine whether glucocorticoids play a role in regulating myelination. When 14-day-old rats were adrenalectomized and sacrificed 7-8 days later, the amount of bulk-isolated myelin in whole brain, as expressed per gram wet weight of brain or per milligram DNA-phosphate, was reduced to about 75% that of sham-operated controls. Both brain weight and DNA content were unchanged by adrenalectomy. Examination of individual brain regions also revealed decreased amounts of myelin in adrenalectomized animals. Brain glycerol 3-phosphate dehydrogenase specific activity was reduced in adrenalectomized animals to 40-60% that of controls, and serum corticosterone levels were less than 0.6% of control levels. The amount of cerebral myelin in animals adrenalectomized on day 21 and sacrificed 9 days later was not significantly reduced. This suggests a possible role of glucocorticoids during the early period of rapid myelination.     

Life-long effect of a single neonatal treatment with estradiol or progesterone on rat uterine estrogen receptor binding capacity.

Rats treated with a single dose of 17 beta-estradiol or progesterone within 24 h of birth were subjected to ovariectomy at 8 weeks of age and were nine days later examined for the binding capacity of the uterine estradiol receptors by saturation and competition tests (with diethylstilbestrol used as competitor). The Bmax value of the neonatally estradiol-treated rats (6.78 x 10(-10) M) was significantly decreased relative to the control (1.99 x 10(-9) M). The competition analysis affirmed these results. Neonatal progesterone treatment also accounted for a significant decrease (1.25 x 10(-9) M) in receptor concentration relative to the control (1.66 x 10(-9) M). Considering the competition analysis the decrease was less than in the case of estradiol and not even significant by saturation analysis. The uterine mass did not differ between the experimental and control rats, but part of those treated with estradiol developed ovarian cysts. It follows that not only synthetic steroids (DES, allylestrenol), but also an excessive presence of the physiological steroid hormone during the critical period of receptor maturation can account for a decrease in uterine receptor concentration in adulthood.     

Influences of gonadal and adrenal androgens on the side glands of Suncus murinus.

In order to know the contribution of adrenal and gonadal androgens to the development of the side gland of Suncus murinus, we studied the effects of gonadectomy and adrenalectomy on gland thickness and the plasma levels of testosterone, androstenedione (delta 4-dione) and dehydroepiandrosterone (DHA). In males, castration decreased gland thickness to 71% of the control. The plasma levels of delta 4-dione and testosterone were also decreased from 4.16 +/- 0.50 and 0.65 +/- 0.10 ng/ml to 1.44 +/- 0.17 and 0.12 +/- 0.02 ng/ml, respectively. Adrenalectomy following castration caused no notable additional decrease in gland thickness, although the plasma levels of delta 4-dione and DHA were further decreased by this treatment. In females, ovariectomy affected neither gland thickness nor plasma androgen levels, except for a peculiar rise in the plasma concentration of delta 4-dione. In contrast, adrenalectomy in addition to ovariectomy decreased gland thickness to 63% of the control and the plasma concentrations of delta 4-dione and DHA from 1.43 +/- 0.26 and 0.43 +/- 0.05 ng/ml to 0.37 +/- 0.11 and 0.10 +/- 0.04 ng/ml, respectively. Therefore, testicular androgens are required for the male side gland to fully develop, whereas in the female adrenal androgens are important for the maintenance of sebaceous gland activity and delta 4-dione is quantitatively more important than DHA. One hour after the intraperitoneal administration of [3H]delta 4-dione, dihydrotestosterone was found to be the major androgen bound to nuclei of the side gland. Thus, the side gland can utilize delta 4-dione as a precursor of a more active androgen.(ABSTRACT TRUNCATED AT 250 WORDS)     

Oxidative stress in newborn infants with and without asphyxia as measured by plasma antioxidants and free fatty acids

A rapid perfusion of oxygen in infants at birth may cause an increase of oxidative stress. To assess this possibility, we measured levels of blood plasma antioxidants and free fatty acids in 20 normal infants at 0, 1, 3, and 5 days after birth. Plasma levels of the most reactive antioxidant, ascorbic acid, decreased daily to equilibrium values at days 3 and 5. Percentages of oxidized form of coenzyme Q-10 (%CoQ-10) in total coenzyme Q, another good marker of oxidative stress, in infants (25-31%) were significantly higher than those in healthy young adults (4.5%). Plasma levels of total free fatty acids (FFA) in normal infants were highest at day 1 and decreased rapidly thereafter. The content of polyunsaturated fatty acids (PUFA) in total FFA was lowest at day 1 and then increased. Since PUFA are susceptible to oxidation, these changes in FFA composition suggest that oxidative stress is most evident at the initial day of neonatal life. Furthermore, it appears that mono-unsaturated fatty acids such as oleic and palmitoleic acids increase in response to the oxidative loss of PUFA. Similar changes in plasma antioxidants, FFA levels, and FFA compositions were observed in 9 infants with asphyxia. Values of %CoQ-10 in infants with asphyxia were significantly greater than those in normal infants, suggesting that infants with asphyxia have elevated oxidative stress.     

Changes in free fatty acids and diglycerides in mouse brain at birth and during anoxia

Abstract: Within the first few hours of life in the mouse, marked changes were seen in brain endogenous free fatty acids (FFA). A 21% decrease in the total FFA pool occurred during the 1st h of life, and a constant value was maintained thereafter to 10 h. Polyunsaturated fatty acids displayed a different pattern of change. There was 27% less free ararhidonic acid at birth (0 h) than 1 h later. Similar values were obtained for docosahexaenoic acid at birth and at 10 h, although palmitoleic and oleic acids decreased markedly after 1 h. The polyunsaturated fatty acyl chains of diglycerides (DG) showed a statistically significant increase as a function of time after birth, despite an unchanged total DG pool size. The brains of pups subjected to 40 min of N₂-anoxia immediately after delivery exhibited a decrease in FFA, especially the monoenoic components, but 60 min of anoxia yielded higher FFA levels. Anoxia induced at 10 h increased FFA and arachidonic acid was higher than when anoxia was induced at 0 h. FFA accumulation was further stimulated by raising the environmental temperature during anoxia. When anoxia was induced, DG exhibited a net increase in palmitate, oleate, and palmitoleate at 0 and 10 h. No arachidonoyl-DG accumulated at 0 h, even after 60 min of anoxia, and stearate was unchanged at 0 and 10 h. The lipid changes observed in the brain during the first hours of life suggest that the enzymatic reactions that promote accumulation of free arachidonic and docosahexaenoic acids and arachidonoyl-DG in the mature brain are present at low levels at the time of delivery. The sluggish modifications found in our study may be related to the longer resistance of newborns to oxygen deficiency.     

Conformational changes in human serum albumin studied by fluorescence and absorption spectroscopy. Distance measurements as a function of pH and fatty acids.

pH- and fatty acid-induced conformational changes in human serum albumin were investigated by fluorescence-energy transfer, determining the distance between Trp-214 and bound bilirubin at 25 degrees C. This distance changes significantly with the pH, being 2.52 +/- 0.01 nm at pH 6, 2.31 +/- 0.04 nm at pH 9, 2.13 +/- 0.07 nm at pH 11.0 and 2.77 nm at pH 11.9. The influence of different fatty acids on the distance was also determined. At pH 7.4 medium-chain fatty acids seem to increase this distance, whereas long-chain fatty acids, at low concentrations, decrease the distance between the two chromophores. The contraction of the protein carrying long-chain saturated fatty acids is even more pronounced at pH 9.     

Human cecal bile acids: concentration and spectrum

To obtain information on the concentration and spectrum of bile acids in human cecal content, samples were obtained from 19 persons who had died an unnatural death from causes such as trauma, homicide, suicide, or drug overdose. Bile acid concentration was measured via an enzymatic assay for 3alpha-hydroxy bile acids; bile acid classes were determined by electrospray ionization mass spectrometry and individual bile acids by gas chromatography mass spectrometry and liquid chromatography mass spectrometry. The 3alpha-hydroxy bile acid concentration (mumol bile acid/ml cecal content) was 0.4 +/- 0.2 mM (mean +/- SD); the total 3-hydroxy bile acid concentration was 0.6 +/- 0.3 mM. The aqueous concentration of bile acids (supernatant after centrifugation) was identical, indicating that most bile acids were in solution. By liquid chromatography mass spectrometry, bile acids were mostly in unconjugated form (90 +/- 9%, mean +/- SD); sulfated, nonamidated bile acids were 7 +/- 5%, and nonsulfated amidated bile acids (glycine or taurine conjugates) were 3 +/- 7%. By gas chromatography mass spectrometry, 10 bile acids were identified: deoxycholic (34 +/- 16%), lithocholic (26 +/- 10%), and ursodeoxycholic (6 +/- 9), as well as their primary bile acid precursors cholic (6 +/- 9%) and chenodeoxycholic acid (7 +/- 8%). In addition, 3beta-hydroxy derivatives of some or all of these bile acids were present and averaged 27 +/- 18% of total bile acids, indicating that 3beta-hydroxy bile acids are normal constituents of cecal content. In the human cecum, deconjugation and dehydroxylation of bile acids are nearly complete, resulting in most bile acids being in unconjugated form at submicellar and subsecretory concentrations.     

Octanoate metabolism in isolated hepatocytes and mitochondria from fetal, newborn and adult rabbit. Evidence for a high capacity for octanoate esterification in term fetal liver

Ketogenesis from endogenous fatty acids or from exogenous octanoate has been studied in isolated hepatocytes from fetal. 24-h-old newborn and adult rabbit. In fed adult rabbits, endogenous ketogenesis is low and increases sixfold in the presence of 2 mM octanoate. At birth, endogenous ketogenesis is low and markedly increases 24 h after birth but, in both cases, the addition of 2 mM octanoate does not increase the rates of ketone body production. Hepatocytes isolated from 24-h-old newborn or fed adult rabbits and incubated with [1-14C]octanoate show a preferential channeling of fatty acid into oxidation (84-92% of octanoate metabolized). In contrast, esterification represents 43% of the amount of octanoate metabolized at birth. Chromatographic analysis of labelled triacylglycerols shows that 76 +/- 2% of labelled fatty acids are identified as octanoate and all of the radioactivity in the octanoate peak is due to the carboxyl carbon. In hepatocytes from term fetus, the low capacity for octanoate oxidation is associated with a high capacity for esterification, whatever the octanoate concentration in the medium. Octanoate activated to octanoyl-CoA in the cytosol of fetal hepatocyte is not oxidized in the mitochondria since carnitine acyltransferase I has a low activity at birth in the rabbit. This suggests that only a part of the octanoate pool is activated outside the mitochondria. Factors involved in the direct esterification of octanoate into triacylglycerols in term fetal hepatocytes are discussed.     

Increased microsomal oxidation of hydroxyl radical scavenging agents and ethanol after chronic consumption of ethanol

The ability of the neonatal rat to oxidize the branched-chain amino acids leucine and valine and their corresponding keto acids was evaluated. In vivo, about 20% of orally administered labeled amino or keto acids were oxidized in 6 h, after which time little further oxidation occurred. In perfused neonatal liver the amino acids were oxidized at only 5-10% the rate of the keto acids. The oxidation of the keto acids showed a saturable dependence on concentration. The decarboxylation of ketoisocaproate (KIC) had a maximal rate of 40.1 +/- 1.6 mumol/h/g liver with an apparent Km of 0.27 +/- 0.03 mM, and decarboxylation of ketoisovalerate (KIV) had a maximal rate of 37.9 +/- 1.9 mumol/h/g liver and an apparent Km of 0.28 +/- 0.04 mM. KIC was ketogenic, producing mainly acetoacetate at a maximal rate of 44.5 +/- 1.6 mumol/h/g liver with an apparent Km of 0.27 +/- 0.03 mM. On the other hand, KIV was not gluconeogenic, although the perfused neonatal liver was able to produce glucose from lactate. During liver perfusion, KIV did not produce measurable quantities of either propionic or beta-aminoisobutyric acids, which are possible end products of KIV metabolism. Decanoic acid inhibited the decarboxylation of both keto acids to the same extent with a maximal effect at 0.4 mM fatty acid. At saturating levels, KIC was less ketogenic than decanoate. Inhibition of endogenous fatty acid oxidation by 2-tetradecylglycidic acid had no effect on keto acid oxidation. These data suggest that branched-chain amino acids derived from milk proteins are probably not quantitatively significant sources of either ketone bodies or glucose in the neonatal rat.     

Lipid composition of alveolar macrophage plasma membrane during postnatal development

This study was undertaken to define the age-related alterations in lipid composition that resident rabbit alveolar macrophages (AM) undergo during postnatal development. The eventual goal is to correlate these changes with the functional maturation of these cells. The number of AM recorded from total lung lavages rose markedly during the first 14 days of life, from 4.9 X 10(5) to 1.1 X 10(7). Adult lungs yielded 1.1 X 10(8) AM. A gradual but significant increase in fluorescence polarization (P) was observed during development when purified AM plasma membranes were tagged with the probe 1,6-diphenyl-1,3,5 hexatriene trimethyl ammonium. The rise ranged from a mean P value of less than or equal to 0.22 to 0.24 (p less than 0.001) for AM plasma membranes from rabbits 1- or 7-day-old to 30- or 150-day-old rabbits, respectively. This finding suggests that the fluidity of the AM plasma membrane decreased during postnatal development. Palmitic, stearic, oleic, and linoleic acids were the most prevalent fatty acids found in the neutral lipid fraction of the AM plasma membrane throughout development. The content of stearic acid rose from 10 to 16%, arachidonic acid rose from 2.8 to 9%, myristic acid decreased from 3.2 to 1.3%, palmitic acid decreased from 42 to 36%, and oleic and linoleic acids changed relatively little during the first 30 days of life. The levels of docosatetraenoic and docosapentaenoic increased gradually during the first 14 days of life, and by 30 days of life the levels declined to that observed at birth. The sum of these changes resulted in an increase in the ratio of unsaturated to saturated fatty acids (1 to 1.15) in the neutral lipid fraction. During the first month of life, the neutral lipid fatty acid pool in the total lipid fraction of AM plasma membrane increased from 12 to 18 mole %, cholesterol increased from 7 to 14 mole %, and total phospholipids decreased from 81 to 67 mole %. These changes resulted in increasing the cholesterol to phospholipid ratio from 0.09 at birth to 0.23 by 150 days of life. The levels of all three major lipid fractions were comparable at 30 days and 150 days of life. Adult levels of choline phosphoglycerides, the predominant phospholipid, were observed by 7 days of life to have decreased from 47 to 34.5 mole %, and the levels of ethanolamine phosphoglycerides and sphingomyelin increased from 17.5 to 25 mole % and from 9 to 13 mole %, respectively. Adult levels of lyso-bis-phosphatidic acid were reached by 30 days of life increasing from 8.2 to 17.8 mole %.(ABSTRACT TRUNCATED AT 400 WORDS)     

Hypoxia and ion activities within the brain stem of newborn rabbits

Eleven rabbits between the 1st and 28th days of life were anesthetized (ketamine 40 mg/kg and acepromazine 3 mg/kg im) thoracotomized, paralyzed, and artificially ventilated with 50% O2 and 10% O2 in N2 or 100% N2. Three-barreled ion-sensitive microelectrodes were used to measure direct-current potentials, potassium (aK+o) and calcium (aCa2+o) activities, and tissue PO2. During control, mean levels of aK+o and aCa2+o were 4.4 +/- 1.1 and 1.3 +/- 0.3 mM, respectively. During hypoxia, changes in aCa2+o were inconsistent, and aK+o revealed three phases: slow (phase I) and fast (phase II) rate of rise and a saturation level (phase III) at the group mean of 6.8 +/- 2.3 mM. Durations of phases I and II decreased, and their slopes increased with maturation. Hypoxia-related excitation of phrenic nerve activity (PHR) occurred during phase I, and gasplike PHR and/or apnea occurred during phases II and III. During recovery after hypoxia, PHR was independent of aK+o levels. Vagal nerve stimulation caused a rapid increase in aK+o followed by a continuous decay even though stimulation continued. Hypoxia had no significant effect on maximal aK+o increase. We concluded that ion homeostasis is less sensitive to the reduced availability of O2 shortly after birth than it is later in life. This age dependence may have an important role in the high resistance to lack of O2 during the early postnatal period in mammals.     

Marked acceleration of exogenous fatty acid incorporation into cellular triglycerides by fetuin.

Fetuin belongs to a group of fetal glycoproteins whose specific function is not known. In this study we investigated the effect of bovine fetuin on exogenous fatty acid incorporation into lipid classes by fetal rabbit aortic smooth muscle cells (SMC) and human fetal skin fibroblasts. When compared with albumin, the addition of fetuin to the culture medium caused a dramatic increase in labeled fatty acid incorporation (nanomoles/mg of protein) by SMC into triglycerides (albumin (control) 2.8 +/- 0.3 + fetuin 178.3 +/- 13.7). This effect was noted at a wide range of fetuin concentrations (0.2-5%) at oleate:fetuin molar ratios of 3.3-0.13, respectively. Similar effects were noted using human fetal skin fibroblasts with both labeled oleic and arachidonic acids (0.1 mM) as substrates (arachidonic acid incorporation into triglycerides, albumin (control) 76.9 +/- 16.2 + fetuin 684.6 +/- 64.1). Stimulation of fatty acid incorporation into di- and monoglycerides was also noted. Although the amount of unbound fatty acid in the presence of fetuin was greater than with albumin, experiments done under conditions that create identical unbound oleate levels (by varying fatty acid concentration) still showed increased fatty acid incorporation into triglycerides by SMC when exposed to fetuin. This marked effect of fetuin on triglyceride accumulation in cells was confirmed by lipid analysis, strong positive staining with oil red O, and transmission of electron microscopy. Furthermore, the potential physiological role of fetuin in terms of fatty acid and transport was attested by (a) the presence of significant amounts of free fatty acids associated with fetuin; and (b) by the stimulatory effect of fetuin, even when added to culture media containing other fatty acid carriers. These results show that (a) fetuin is far more efficient than albumin in incorporating fatty acids into cells; and (b) this might represent a novel function for fetuin during development.     

Carnitine stimulation of glucose oxidation in the fatty acid perfused isolated working rat heart.

The effects of L-carnitine on myocardial glycolysis, glucose oxidation, and palmitate oxidation were determined in isolated working rat hearts. Hearts were perfused under aerobic conditions with perfusate containing either 11 mM [2-3H/U-14C]glucose in the presence or absence of 1.2 mM palmitate or 11 mM glucose and 1.2 mM [1-14C]palmitate. Myocardial carnitine levels were elevated by perfusing hearts with 10 mM L-carnitine. A 60-min perfusion period resulted in significant increases in total myocardial carnitine from 4376 +/- 211 to 9496 +/- 473 nmol/g dry weight. Glycolysis (measured as 3H2O production) was unchanged in carnitine-treated hearts perfused in the absence of fatty acids (4418 +/- 300 versus 4547 +/- 600 nmol glucose/g dry weight.min). If 1.2 mM palmitate was present in the perfusate, glycolysis decreased almost 2-fold compared with hearts perfused in the absence of fatty acids. In carnitine-treated hearts this drop in glycolysis did not occur (glycolytic rates were 2911 +/- 231 to 4629 +/- 460 nmol glucose/g dry weight.min, in control and carnitine-treated hearts, respectively. Compared with control hearts, glucose oxidation rates (measured as 14CO2 production from [U-14C]glucose) were unaltered in carnitine-treated hearts perfused in the absence of fatty acids (1819 +/- 169 versus 2026 +/- 171 nmol glucose/g dry weight.min, respectively). In the presence of 1.2 mM palmitate, glucose oxidation decreased dramatically in control hearts (11-fold). In carnitine-treated hearts, however, glucose oxidation was significantly greater than control hearts under these conditions (158 +/- 21 to 454 +/- 85 nmol glucose/g dry weight.min, in control and carnitine-treated hearts, respectively). Palmitate oxidation rates (measured as 14CO2 production from [1-14C]palmitate) decreased in the carnitine-treated hearts from 728 +/- 61 to 572 +/- 111 nmol palmitate/g dry weight.min. This probably occurred secondary to an increase in overall ATP production from glucose oxidation (from 5.4 to 14.5% of steady state myocardial ATP production). The results reported in this study provide direct evidence that carnitine can stimulate glucose oxidation in the intact fatty acid perfused heart. This probably occurs secondary to facilitating the intramitochondrial transfer of acetyl groups from acetyl-CoA to acetylcarnitine, thereby relieving inhibition of the pyruvate dehydrogenase complex.