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Biomedical subjects

E Shrago

Publications and source records attributed to E Shrago.

At least 37 records · Page 2Linked to original sources
PubMed1989

Specific labeling of beef heart mitochondrial ADP/ATP carrier with N-(3-iodo-4-azidophenylpropionamido)cysteinyl- 5-(2'-thiopyridyl)cysteine-coenzyme A (ACT-CoA), a newly synthesized 125I-coenzyme A derivative photolabel.

An azido-125I-CoA photolabel was synthesized from N-(3-iodo-[125I]4-azidophenylpropionamido)cysteinyl- 5-(2'thiopyridyl) cysteine ([125I]ACTP) and CoASH, separated by chromatography on a silica gel TLC, and identified by autoradiography. Synthesis of [125I]ACT-CoA from [125I]ACTP and CoA was further confirmed by monitoring the release of one of the products, thiopyridone, at 343 nm and concurrent formation of [125I] ACT-CoA. Beef heart mitochondria were incubated in the presence of the 125I-CoA derivative with or without specific inhibitors and/or substrates of the ADP/ATP carrier, and immediately photolyzed for 5 s. Sodium dodecyl sulfate-gel electrophoresis and autoradiography of the separated proteins revealed exclusive photolabeling of a 30-kDa protein in the absence of inhibitors of the carrier. This specific photolabeling of the 30-kDa protein was prevented in a concentration-dependent manner by either carboxyatractylate or palmitoyl-CoA (0.1-5 microM), two known inhibitors of the ADP/ATP carrier. ADP reduced the extent of photolabeling of the 30-kDa protein, but palmitic acid, free CoASH, and dinitrophenol were ineffective, indicating the specificity of the reaction. CoA photolabels may be useful in probing ligand and/or substrate binding sites and in determining the structure-function relationship of the ADP/ATP carrier.

Acyl Coenzyme A↗
PubMed1989

Impact of lemongrass oil, an essential oil, on serum cholesterol.

To test the hypothesis that non-sterol mevalonate pathway end products lower serum cholesterol levels, we asked 22 hypercholesterolemic subjects (315 +/- 9 mg cholesterol/dl) to take a daily capsule containing 140 mg of lemongrass oil, an essential oil rich in geraniol and citral. The paired difference in serum cholesterol levels of subjects completing the 90-day study approached significance (P less than 0.06, 2-tailed t-test). The subjects segregated into two groups, one consisting of 14 subjects resistant to the protocol and the other consisting of 8 subjects who responded. Paired differences in cholesterol level at 30, 60 and 90 d for resistant subjects were +2 +/- 6, +2 +/- 7 and -1 +/- 6 mg/dl; paired differences for the responding subjects were -25 +/- 10 (p less than 0.05), -33 +/- 8 (p less than 0.01) and -38 +/- 10 (p less than 0.025), respectively. The paired difference (+8 +/- 4) in the cholesterol levels of six responders 90 days after the discontinuation of lemongrass oil was not significant.

Cholesterol↗
PubMed1989

Effect of dietary fat on individual long-chain fatty acyl-CoA esters in rat liver and skeletal muscle.

The effect of dietary fat on the long-chain acyl-CoA ester profile of liver and skeletal muscle was investigated by feeding weanling rats 12%-fat diets composed of high-linoleic safflower oil (73% 18:2n-6), high-oleic safflower oil (70% 18:1n-9) or olive oil (70% 18:1n-9) for six and ten weeks. Approximately 50% of both hepatic and skeletal muscle acyl-CoA esters comprised linoleoyl-CoA or oleoyl-CoA with high-linoleic or oleic feeding, respectively. Total hepatic acyl-CoA ester concentration was 40% higher (p less than 0.05) in rats fed 12% fat compared with controls fed a 4%-fat diet. These data demonstrate that the long-chain acyl-CoA ester profile of liver and skeletal muscle reflects the dietary fatty acid profile.

Acyl Coenzyme A↗
PubMed1989

Reconstitution of purified brown adipose tissue mitochondria uncoupling protein: demonstration of separate identity of nucleotide binding and proton translocation sites by chemical probes.

The reaction of two arginine-modifying reagents, phenylglyoxal and 2,3-butanedione, with a highly purified uncoupling protein of brown adipose tissue mitochondria decreased the GDP binding to the uncoupling protein. This inhibition was irreversible and dependent on time and concentration of the reagent. Complete inhibition of GDP binding by both reagents establishes that arginine is one of the critical amino acid residues involved in the binding of GDP. Reconstitution of the uncoupling protein (both unmodified and modified) into phospholipid vesicles by our procedure showed no effect of phenylglyoxal modification on the H+ conductance, thus demonstrating that the proton translocation site is structurally different and distinct from the GDP binding site.

Adipose Tissue, Brown↗
PubMed1988

Energy substrate utilization in freshly isolated Morris Hepatoma 7777 cells.

Freshly isolated Morris Hepatoma 7777 cells were prepared for a study of the utilization of palmitate and beta-hydroxybutyrate as metabolic fuels compared to other major energy substrates (glucose and glutamine). These cells were capable of oxidizing both [U-14C]palmitate and beta-[3-14C]hydroxybutyrate although the rates accounted for only 10 +/- 3 (SD) and 9 +/- 4% of total oxygen consumed, respectively; n = 4. Incorporation of [U-14C]glutamine and [U-14C]glucose carbon into 14CO2 made up for 38 +/- 13 and 9 +/- 2% of oxygen consumed by these cells, respectively. The conversion of glucose carbon into lactate was estimated to supply 26 +/- 6% of ATP. Thus, glutamine oxidation and lactate formation from glucose were the major contributors to estimated ATP needs. Conversion of these substrates into lipids was also studied and compared with oxidized products. Incorporation of glucose, glutamine, and beta-hydroxybutyrate into nonsaponifiable lipids and fatty acids was only 6.0 +/- 2.9, 0.8 +/- 0.2 and 17.7 +/- 6.65% (n = 3) of their respective rates of CO2 formation. This suggests that in freshly isolated Morris Hepatoma 7777 cells, citrate export from the mitochondria for cholesterogenesis and lipogenesis is a minor fate of substrate carbon entering the mitochondria for oxidation.

Adenine Nucleotides↗
PubMed1988

Determination of individual long-chain fatty acyl-CoA esters in heart and skeletal muscle.

A method has been developed for determination of individual long-chain fatty acyl-CoA esters from heart and skeletal muscle using high performance liquid chromatography (HPLC). The esters were extracted from freeze-clamped tissue of pig and rat hearts and rat skeletal muscle for analysis on a radially compressed C18 5mu reverse-phase column. Nine peaks in the extract with carbon chain lengths from C12 to C20 that subsequently disappeared on alkaline hydrolysis were identified. The major acyl-CoA peaks were 14:1, 18:2, 16:0 and 18:1 and additionally in rat heart 18:0. Total long-chain acyl-CoA esters obtained by summation of the individual molecular species was 11.34 +/- 1.48 nmol/g wet wt. pig heart; 14.51 +/- 2.11 nmol/g wet wt. in rat heart, and 4.35 +/- 0.71 nmol/g wet wt. in rat skeletal muscle. These values were approximately 132% of those obtained using a separate procedure that measured total CoA by HPLC after alkaline hydrolysis of the esters. The described method demonstrates the quantitation of individual acyl-CoA species in muscle tissue. Therefore, it has a number of advantages in that it permits information to be obtained on the individual molecular species under various nutritional and metabolic conditions.

Acyl Coenzyme A↗
PubMed1988

Age-related changes in respiration coupled to phosphorylation. I. Hepatic mitochondria.

Age-related changes affecting mitochondrial adenine nucleotide metabolism may underlie age-related decreases in hepatic metabolic activities. Oxidative activity coupled with phosphorylation, the apparent Km and Vmax of the adenine nucleotide translocase (AdNT), the adenine nucleotide pool size and membrane lipid composition were determined for hepatic mitochondria from young (3 months), mature (12 months) and aged (24 months) Fischer 344 male rats which had been fed NIH-31 diet. The age-related decreases in state 3 respiration supported by NAD-linked substrates were 2-4-fold greater than that of an FAD-linked substrate. The 32% (P less than 0.05) decrease in the AdNT Vmax calculated for the aged rats was accompanied by a 17% decrease in the AdNT Km. The exchangeable pool of adenine nucleotides in mitochondria from aged rats was 72% (P less than 0.05) that in the young rats. While the age-related increase in the cholesterol/phospholipid-Pi ratio and changes in the phospholipid head group pattern were not significant, the overall change in the fatty acid pattern effected a 20% (P less than 0.05) decrease in the n-6/n-3 fatty acid ratio. These data suggest that the reduced Vmax of the AdNT is a consequence of a diminished pool of exchangeable adenine nucleotides. The lower AdNT velocity may reflect the effect of changes in the lipid environment of the membrane in which it is embedded. The major shifts in these parameters occurred during the second year of life.

Adenine Nucleotides↗
PubMed1988

Age-related changes in respiration coupled to phosphorylation. II. Cardiac mitochondria.

Age-related changes in mitochondrial adenine nucleotide metabolism may underlie the progressive decline in cardiac function. Oxidase activity coupled with phosphorylation, adenine nucleotide translocase (AdNT) activity, adenine nucleotide pool size and membrane lipid composition were determined using cardiac mitochondria from young (3 months), mature (12 months) and aged (24 months) Fischer 344 male rats which had been fed NIH-31 diet. While an age-associated 15% decrease in respiratory activity was not significant, AdNT activity of the aged rat was 20% lower (P less than 0.05) than that of the young rat. The exchangeable matrix adenine nucleotide pool (ATP + ADP) tended to decrease with age. In comparison to the young, membrane lipids of cardiac mitochondria from aged rat had a 43% higher (P less than 0.01) cholesterol/phospholipid-Pi ratio and a significantly lower (P less than 0.01) phosphatidyl ethanolamine/phosphatidyl choline ratio. The overall change in the fatty acid pattern of mitochondrial membrane lipids resulted in a significant (P less than 0.01) decrease in the n-6/n-3 fatty acid ratio. All values obtained for the mature rat fell between those of the young and aged rats. These data suggest that the reduced cardiac AdNT activity in the aged rat is a consequence of both a diminished pool of exchangeable adenine nucleotides and a lower AdNT velocity. Age-related changes in the lipid components of the membrane matrix in which the AdNT is embedded may underlie the decrease in respiratory activity.

Adenine Nucleotides↗
PubMed1988

Host and tumor growth and energy substrates in blood of hepatoma-bearing rats receiving high-fat parenteral infusions.

We examined the effects of isocaloric carbohydrate-based TPN vs fat-based TPN on plasma levels of energy substrates and insulin and on growth of Morris Hepatoma 7777 and host animals. Hepatoma-bearing rats fed similar diets ad libitum served as controls. Although no differences in tumor growth were observed, host weight loss was less in parenterally than in orally fed animals. Liver lipid and plasma free fatty acids and triglycerides in tumor-bearing and normal rats infused high-fat TPN were markedly higher than in all other groups, suggesting that this level of lipid infusion exceeds the rate of utilization. Plasma glucose and insulin in tumor-bearing rats infused high-fat TPN were higher than in all other groups, which may indicate insulin resistance. Replacing glucose energy in TPN with lipid does not appear to reduce glucose availability to tumors but does have potentially deleterious effects on the host.

Adipose Tissue↗
PubMed1988

Adaptive changes in individual acyl-CoA esters from hamster BAT during cold acclimation.

Long-chain fatty acyl-CoA esters (LCFACoAE) were extracted from freeze-clamped powdered brown adipose tissue (BAT) obtained from thermoneutral control and cold-acclimated hamsters and the CoA esters individually separated by high-performance liquid chromatography. LCFACoAE of carbon chain length C12 to C20 were identified by increasing column retention time in the following order: C12:0, C14:1, C14:0, C16:1, C18:2, C16:0, C18:1, C18:0, and C20:4. The mean total LCFACoAE concentrations were 235 +/- 40 nmol/g protein for the control hamsters and 648 +/- 105 nmol/g protein for the 22-day cold-acclimated hamsters. A rapid fourfold increase in the levels of C16:0, C18:0, and C18:1 occurred within hours after initiation of the cold temperature, whereas the concentrations of the other six LCFACoAE either increased only slightly or remained unchanged. Almost 50% of the total LCFACoAE in the BAT of cold-acclimated hamsters was made up of C16:0, C18:0, and C18:1. These results, which demonstrate some dynamic changes in adipose tissue LCFACoAE, are consistent with their proposed role in the initiation and maintenance of BAT thermogenesis.

Acclimatization↗
PubMed1987

Activity of long chain acyl-CoA synthetase in isolated alveolar type II cells.

Fatty-acyl-CoA synthetase activity was determined in rat alveolar type II cells. Compared to whole-lung homogenate, the enzyme specific activity with palmitic acid was 3.6-fold higher in isolated type II alveolar cells. The enzyme in rat alveolar type II cells did not discriminate among various fatty acids, suggesting that supply of fatty acids rather than specificity might be an important factor for their activation in these cells.

Animals↗
PubMed1987

Relation of lung fatty acid binding protein to the biosynthesis of pulmonary phosphatidic acid and phosphatidylcholine.

The activities of glycerophosphate and lysophosphatidylcholine (LPC) acyltransferases were determined using lung microsomes in the presence of lung fatty acid binding protein (FABP). The synthesis of phosphatidic acid (PA) was increased two- to fourfold in the presence of FABP as compared to albumin. Lung FABP did not increase the incorporation of palmitoyl CoA into phosphatidylcholine. The results indicate that FABP-bound fatty acyl CoA may be a preferred substrate for glycerophosphate acyltransferase.

1-Acylglycerophosphocholine O-Acyltransferase↗
PubMed1986

Carnitine status, plasma lipid profiles, and exercise capacity of dialysis patients: effects of a submaximal exercise program.

Carnitine status, blood lipid profiles, and exercise capacity were evaluated in a combined group of hemodialysis (N = 4) and continuous ambulatory peritoneal dialysis (N = 6) patients before and after an 8-week submaximal exercise program. Maximal aerobic capacity (VO2max) was only 18.5 +/- 5.9 (mean +/- SD) mL O2/kg/min, well below the expected 30 to 35 mL O2/kg/min for age-matched sedentary controls. Plasma short-chain acylated carnitine levels, which were two to three times normal values, were reduced after the exercise program, but the long-chain acylcarnitines were significantly reduced during acute exercise. Muscle biopsies of the vastus lateralis were performed at rest in five patients prior to and after the 8-week exercise program. Total carnitine in skeletal muscle was 3.09 (.076 SD) mumol/g ww, with only 11.3% acylated prior to the exercise program, which was much lower than the 4.25 +/- 1.27 mumol/g ww, with 28.5% acylated in a group of healthy athletic subjects (N = 28). Muscle free carnitine concentrations decreased significantly following the 8-week training period, with only a slight reduction in total carnitine. The percent of acylated carnitine was therefore significantly increased (P less than 0.05) from 11.3% to 25.2% after the experimental period. Pretraining carnitine palmitoyl transferase activity at rest was 0.57 +/- 0.28 nmol palmitoyl carnitine formed/5 min/mg mitochondrial protein, which was not changed by exercise training v 1.80 +/- 0.51 nmol/5 min/mg protein in 28 healthy normals (P less than 0.001). Free fatty acid concentrations were reduced significantly during acute exercise as a result of the exercise training program whereas other plasma lipids were not altered. (ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1986

Effects of recombinant interferon-alpha 2 treatment upon lipid concentrations and lipoprotein composition.

Interferon-alpha 2 (IFN-alpha 2) produced by recombinant DNA technology and purified to homogeneity, was assessed for effects on plasma lipids and lipoprotein composition in 10 patients with metastatic malignant melanoma. Patients received 30 X 10(6) U/m2 IFN intravenously for 5 consecutive days every 3 weeks. Plasma cholesterol concentrations were significantly decreased after 3 days of IFN administration (171 +/- 37 mg/dl, mean +/- SD) when compared with pretreatment concentrations (211 +/- 28 mg/dl, p less than 0.05). Approximately 34% of the decrease in plasma cholesterol concentration was contributed by high-density lipoprotein (HDL). Significant decreases in plasma HDL cholesterol (44 +/- 8 to 34 +/- 7 mg/dl, p less than 0.05) and apolipoprotein A-1 concentrations (124 +/- 14 to 95 +/- 17 mg/dl, p less than 0.05) occurred. Although decreases in low-density lipoprotein (LDL) cholesterol concentrations contributed to the majority of the total decrease observed in plasma cholesterol, IFN did not alter the cholesterol-to-protein ratio of the LDL. When IFN-alpha 2 was discontinued, alterations in plasma lipoproteins returned to pretreatment levels. Plasma triglyceride concentrations were not influenced by IFN treatment nor did administration of IFN decrease very-low-density lipoprotein (VLDL). The overall effect of recombinant (r) IFN-alpha 2 on lipid composition was to reduce LDL and HDL without alteration of VLDL or triglycerides.

Adult↗
PubMed1985

Adenine nucleotide translocase-dependent anion transport in pea chloroplasts.

Pea chloroplasts were found to take up actively ATP and ADP and exchange the external nucleotides for internal ones. Using carrier-free [14C]ATP, the rate of nucleotide transport in chloroplasts prepared from 12-14-day-old plants was calculated to be 330 mumol ATP/g chlorophyll/min, and the transport was not affected by light or temperature between 4 and 22 degrees C. Adenine nucleotide uptake was inhibited only slightly by carboxyatractylate, whereas bongkrekic acid was nearly as effective an inhibitor of the translocator in pea chloroplasts as it was in mammalian mitochondria. There was no counter-transport of adenine nucleotides with substrates carried on the phosphate translocator including inorganic phosphate, 3-phosphoglycerate and dihydroxyacetone phosphate. However, internal or external phosphoenolpyruvate, normally considered to be transported on the phosphate carrier in chloroplasts, was able to exchange readily with adenine nucleotides. Furthermore, inorganic pyrophosphate which is not transported by the phosphate carrier initiated efflux of phosphoenolpyruvate as well as ATP from the chloroplast. These findings illustrate some interesting similarities as well as differences between the various plant phosphate and nucleotide transport systems which may relate to their role in photosynthesis.

Adenine Nucleotides↗
PubMed1985

Substrate inhibition of carnitine palmitoyltransferase by palmitoyl-CoA and activation by phospholipids and proteins.

When carnitine palmitoyltransferase is purified it shows increasing substrate inhibition by palmitoyl-CoA as the protein content of the assay mixture is decreased. The purified enzyme is stimulated by addition of phospholipids (phosphatidylcholine, cardiolipin) and proteins (albumin, fatty acid-binding protein, lambda-globulin) to the reaction mixture. The effects of phospholipid and protein are more than additive, particularly with relatively high concentrations of palmitoyl-CoA. It is suggested that the enzyme contains hydrophobic sites which require phospholipid to prevent spurious binding of palmitoyl-CoA and which normally anchor the enzyme to the mitochondrial membrane.

Acyl Coenzyme A↗