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R R Brenner

Publications and source records attributed to R R Brenner.

At least 73 records · Page 4Linked to original sources
PubMed1986

Cholesterol-dependent modification of microsomal dynamics and UDPglucuronyltransferase kinetics.

The effect of both in vitro incorporation and removal of cholesterol in guinea pig liver microsomes on the lipid composition, dynamic properties of the membrane, and kinetic constants of UDPglucuronyltransferase was studied. No significant changes either in the fatty acid composition or in the distribution of phospholipid classes were observed upon cholesterol incorporation and removal. Lateral and rotational mobility measured by the efficiency of pyrene excimer formation and fluorescence of 1,6-diphenylhexatriene decreased with cholesterol incorporation and increased in parallel to cholesterol removal. These changes were associated with alterations in the kinetic properties of UDPglucuronyltransferase. Whereas Vmax increased, the Km of the different steps of the reaction decreased with cholesterol incorporation. The negative homotropic effect and apparent cooperativity of UDP-glucuronic acid decreased when cholesterol was incorporated and increased after cholesterol removal. Moreover, the UDP-N-acetylglucosamine-dependent activation of the enzyme decreased in correlation with an increase of cholesterol concentration in microsomes. It has been demonstrated that both the shift of the non-Michaelian kinetics of the enzyme to Michaelian and the decrease of the UDP-N-acetylglucosamine-dependent activation of the enzyme are evoked by a change of the physical state of the UDPglucuronyltransferase milieu from a gel phase to a liquid-crystalline phase. Therefore, we must admit that cholesterol incorporation in the microsomes while producing an increased packing of the bulk lipids would also cause the separation of more fluid phospholipids, which increase the proportion of molecules in the liquid-crystalline state within the enzyme environment.

Animals↗
PubMed1986

Regulation of linoleic acid delta 6-desaturation by a cytosolic lipoprotein-like fraction in isolated rat liver microsomes.

A peripheral component of the delta 6-fatty acid-desaturase system of rat liver microsomes has been isolated from the cytosol by ultracentrifugation at a saline density of 1.26 g/ml. It exhibited lipoprotein characteristics with an approximate protein/lipid ratio of 1.22 and free fatty acids and phosphatidylcholine as its main lipid components. Linoleic acid desaturation activity diminished in washed microsomes, since they lost the adsorbed cytosolic fraction. Addition of the factor reactivated the reaction and the recovery was dependent on the concentration of the factor in the medium. Linoleic acid and linoleyl-CoA were bound by the cytosolic fraction. However, the transport of substrate to the desaturase was not apparently a main function of the cytosolic fraction, since transport occurred equally in the absence of the factor. Moreover, the solubilization of linoleyl-CoA was not enhanced and the free monomeric concentration was not altered by the presence of the cytosolic fraction. In addition, the factor did not divert delta 6-desaturase substrate to or from other metabolic pathways such as esterification to phospholipids. gamma-Linolenic acid produced by delta 6-desaturation of linoleic acid in the microsomes inhibited the desaturase, but it was removed by the factor from the membrane towards the cytosol, preventing the inhibition. The anti-inhibitory effect of the cytosolic factor was blockaded by addition of columbinic acid or gamma-linolenic acid to the factor. Moreover, the inhibitory effect of arachidonic acid was not prevented by addition of the cytosolic fraction. These results suggest that the cytosolic fraction studied would optimize the delta 6-desaturation of linoleic acid in vitro in rat liver microsomes by removal of the product, gamma-linolenic acid, as it is formed.

Animals↗
PubMed1986

Effect of dietary cholesterol on microsomal membrane composition, dynamics and kinetic properties of UDPglucuronyl transferase.

The effect of cholesterol administration in vivo on the lipid composition, dynamic properties of the microsomal membrane of guinea pig livers and the kinetic properties of UDPglucuronyl transferase were studied. Cholesterol administration in the diet evoked an increase of microsomal cholesterol, but no significant changes in the fatty-acid composition of total lipids or of each phospholipid class. Instead, the phosphatidylethanolamine, phosphatidylcholine molar ratio of the membrane was markedly decreased from 0.57 to 0.38. This decline was not enough to counterbalance the overall 'ordering' effect of cholesterol and consequently, the fluorescence anisotropy of the membranes labeled with 1,6-diphenylhexatriene was increased. The lateral diffusion evaluated by measuring the pyrene excimer formation was decreased by the cholesterol incorporation. These physical changes were associated with changes in the kinetic properties of UDPglucuronyl transferase: Vmax increased, while the Km of the different steps of the reaction decreased in the modified microsomes. Furthermore, a shift of the non-michaelian kinetics to michaelian, equivalent to a decrease of a negative homotropic effect and apparent cooperativity of UDPglucuronic acid was observed since the Hill coefficient changed, approaching 1. A non-michaelian kinetics of this enzyme is an indication of boundary lipids in the gel phase and a shift to michaelian, a change of the surrounding lipids to a liquid-crystalline structure. In consequence, our results suggest that cholesterol incorporation in the microsomal membrane while producing a condensing effect of bulk lipids would produce an opposite effect on the UDPglucuronyl transferase boundary lipids.

Animals↗
PubMed1986

Dexamethasone blocks arachidonate biosynthesis in isolated hepatocytes and cultured hepatoma cells.

The effect of dexamethasone on the incorporation and conversion of [1-14C]eicosa-8,11,14-trienoic acid to arachidonic acid in isolated hepatocytes and in hepatoma tissue culture (HTC) cells was studied. In both kinds of cells, no changes in the exogenous acid incorporation were found when the hormone was added to the incubation media at 0.1 or 0.2 mM concentration, while the biosynthesis of arachidonic acid was significantly depressed. The effect on the biosynthesis was faster in isolated normal liver cells (60 min) than in tumoral cells (120 min) and reached an inhibition of ca. 50% after 3 hr of treatment. The addition of cycloheximide (10(-6) M) also caused a marked decrease in the biosynthesis of this polyunsaturated fatty acid, but when dexamethasone was added to the media simultaneously with cycloheximide, a synergistic action was not observed. The results obtained show that protein synthesis would be involved in the modulation of the biosynthesis of arachidonic acid by glucocorticoids. The changes in the delta 5 desaturation of labeled 20:3 omega 6 to arachidonic acid correlated with changes in the fatty acid composition in isolated cells.

8,11,14-Eicosatrienoic Acid↗
PubMed1986

Effect of epinephrine on the oxidative desaturation of fatty acids in the rat adrenal gland.

Delta-6 and delta 5 desaturation activity of rat adrenal gland microsomes was studied to determine the effect of microsomal protein and the substrate saturation curves. This tissue has a very active delta 6 desaturase for linoleic and alpha-linolenic acids. The administration of epinephrine (1 mg/kg body weight) 12 hr before killing, produced approximately a 50% decrease in desaturation of [1-14C]linoleic acid to gamma-linolenic acid, [1-14C]alpha-linolenic acid to octadeca-6,9,12,15-tetraenoic acid and [1-14C]eicosa-8,11,14-trienoic acid to arachidonic acid. A 30% decrease in delta 5 desaturation activity was also shown after 7 hr of epinephrine treatment. The changes on the oxidative desaturation of the same fatty acids in liver microsomes were similar. No changes were observed in the total fatty acid composition of adrenal microsomes 12 hr after epinephrine treatment. Mechanisms of action of the hormone on the biosynthesis of polyunsaturated fatty acids in the adrenal gland are discussed.

Adrenal Glands↗
PubMed1986

Effect of different acids with delta 9,12-dienoic structures on delta 9 desaturation activity in rat liver microsomes.

The effect of oral administration, for 24 or 48 hr, of different octadeca fatty acids containing a 9,12-dienoic structure on the fatty acid composition and delta 9 desaturation activity of liver microsomes of rat fed a fat-free diet was studied. The ethyl esters of linoelaidic and gamma-linolenic acids, the methyl ester of linoleic acid and free columbinic acid were administered to rats maintained on a fat-free diet. The supplementation of the fat-free diet with linoelaidate produced no relevant changes in the fatty acid composition pattern of liver microsomes and did not modify the percentage of conversion of palmitic to palmitoleic acid. The addition of linoleate or gamma-linolenate to the fat-free diet returned liver microsome delta 9 desaturation activity toward the control and partially restored the liver microsome fatty acid spectrum found in the fat-free diet. Columbinic acid (5-trans-9-cis,12-cis-18:3), which cannot be transformed into arachidonic acid, also decreased the delta 9 desaturation activity enhanced by the fat-free diet and evoked changes in the microsomal fatty acid composition similar to those produced by the omega 6 fatty acids. These results suggest that the modulation of delta 9 desaturase activity evoked by dietary administration of unsaturated acids of omega 6 series would depend on the cis double bond configuration of these acids.

Animals↗
PubMed1986

Effect of malonyl-CoA on delta 6 desaturation activity of rat liver microsomes.

The effect of malonyl-CoA on linoleic acid desaturation and elongation reactions of rat liver microsomes was studied. Under strict desaturation conditions, the in vitro microsomal conversion of linoleic acid to gamma-linolenic acid is time-dependent. When malonyl-CoA was added to the aforementioned incubation medium, linoleic acid was desaturated to gamma-linolenic acid and elongated to its higher homologues. Under these conditions, delta 6 desaturation activity, calculated by adding gamma-18:3, 20:3 and 20:4 acids, was neither inhibited nor activated by malonyl-CoA. These results indicate that the elongation of gamma-linolenyl-CoA coupled to the desaturation of linoleic acid did not modify delta 6 desaturase activity.

Acyl Coenzyme A↗
PubMed1986

[Biochemistry of the evolutive cycle of Triatoma infestans. IX. Composition of cuticular hydrocarbons compared to other Triatominae].

The hydrocarbon composition of epicuticular lipids of different species of Triatomini were determined and compared by gas-liquid chromatography. T. infestans, T. delpontei, T. rubrovaria, T. sordida, T. guasayana and T. vitticeps from infestans subgroup and T. pallidipennis from rubrofasciata, were studied. All these species had in common the same kind of hydrocarbons. The major components were n C29; n C31; 12+14+15+18 methyl C38; 15, 19+14, 18 dimethyl C39; 12+14+17+19 methyl C36 and 4 methyl C34. However, the relative proportions of selective hydrocarbons were significantly different among the species studied. These differences may be used with a chemotaxonomic criterion. Either the gas-liquid chromatographic curve of the cuticular hydrocarbons or the ratio of selective hydrocarbons may be used as fingerprints of the species considered.

Animals↗
PubMed1986

[Biochemistry of the development cycle of Triatoma infestans (vinchuca). X. Hemolymph lipoproteins of females].

A lipoprotein of high density (HDL) and three of very high density (VHDL-I, VHDL-f and VHDL-II) were separated by ultracentrifugation from T. infestans female hemolymph. Lipids were mainly transported by HDL, whereas VHDL-II presented the highest protein content. In all the lipoproteins, 1,2 and 1,3-diacylglycerols, triacylglycerols, free fatty acids, cholesterol, cholesterol esters and hydrocarbons, were present. Phosphatidylcholine and phosphatidylethanolamine were also identified. The main lipids were represented by phospholipids, 1,2 and 1,3-diacylglycerols and hydrocarbons. All lipoproteins were delipidated to study the corresponding apolipoproteins. They were examined by SDS-polyacrylamide gel electrophoresis in the presence of 2-mercaptoethanol. Apo-HDL was separated into four intense polypeptide bands of approximate MW 19,000; 45,000; 86,000 and 200,000. Apo-VHDL-I was fractionated into three polypeptide bands--a major one of MW 18,500 and two minor ones of 28,000 and 45,000. The apolipoproteins belonging to VHDL-f that only appeared in females, were separated into 10 bands; the major ones corresponded to MW of 58,000, 86,000 and 160,000. In these apolipoproteins, the specimens of 58,000 and 160,000 showed positive carbohydrate reaction. Yet, in the apo-VHDL-II the most intense protein subunits primarily corresponded to bands of MW 160,000 and 86,000. Apparently, the four lipoproteins would share two polypeptide chains.

Animals↗
PubMed1986

Effect of dietary columbinic acid on the fatty acid composition and physical membrane properties of different tissues of EFA-deficient rats.

The effect of columbinic acid (5 trans, 9 cis, 12 cis, octadeca-trienoic acid) supplemented to a fat-free diet on the fatty acid composition and its correlation to the physical properties of several tissues of rats, was studied. The absence of lipids in the diet produced the typical changes in the fatty acid composition characteristic of essential fatty acid (EFA) deficiency, namely a significant increase in the relative percentage of monoenoic fatty acids with a concomitant decrease in linoleic and arachidonic acids and a rise in eicosa-5,8,11-trienoic acid in liver, kidney, lung and spleen homogenates. Columbinic acid supplemented to a fat-free diet for 24 or 48 hr was incorporated into the different tissues and was partially elongated to 7 trans, 11 cis, 14 cis eicosatrienoic acid, but it was not desaturated. It modified the fatty acid spectrum of the lipids in the different tissues returning it to a similar composition of non-EFA deficient animals, except for a decrease of linoleic acid. The absence of lipids in the diet produced an increase in the 1-6 diphenyl-1,3,5-hexatriene (DPH) steady-state fluorescence anisotropy (rs) in liver microsomes, that was corrected by the administration of columbinic acid for 24 hr. It is concluded that columbinic acid produced a change in the pattern of total fatty acid composition of the different tissues studied which induced a favorable effect on the physical properties of the liver microsomal membranes (rs), leading to an improvement on the fatty acid deficiency in those membranes. Besides, columbinic acid would also exert a favorable effect in the short term, but not in the long-term eicosanoids production.

Animals↗
PubMed1985

In vitro modification of cholesterol content of rat liver microsomes. Effects upon membrane 'fluidity' and activities of glucose-6-phosphatase and fatty acid desaturation systems.

The cholesterol content of rat liver microsomal membranes was modified in vitro by incubating microsomes and cytosol with liposomes prepared by sonication of microsomal lipids and cholesterol. In this way, the cholesterol to phospholipid molar ratio was increased from 0.11-0.13 in untreated microsomes to a maximal of 0.8 in treated ones. Cholesterol incorporation in microsomes produced an increase in the diphenyl-hexatriene steady-state fluorescence anisotropy and a decrease in the efficiency of pyrene-excimer formation which indicated a decrease in the rotational and translational mobility, respectively, of these probes in the membranes lipid phase. Cholesterol incorporation in microsomes did not affect significantly the glucose-6-phosphatase activity in 0.1% Triton X-100 totally disrupted microsomes, but diminished the glucose-6-phosphatase activity of 'intact' microsomes. This indicates that possibly the glucose 6-phosphate translocation across the microsomal membrane is impeded by an increase in the membrane apparent 'microviscosity'. Cholesterol incorporation in microsomes decreased NADH-cytochrome c reductase without affecting NADH-ferricyanide reductase activity. The delta 9 desaturation reaction rate was enhanced by cholesterol incorporation at low but not at high palmitic acid substrate concentration. delta 5 and delta 6 desaturase reaction-rates were increased both at low and high fatty acid substrate concentrations. These results suggest that a mechanism involving fatty acid desaturase enzymes, might exist to self-regulate the microsomal membrane lipid phase 'fluidity' in the rat liver.

Animals↗
PubMed1985

Uptake and metabolism of eicosa-8, 11, 14-trienoic acid in normal hepatocytes and HTC cells.

The incorporation and conversion of eicosa-8,11,14-trienoic acid to arachidonic acid were studied in HTC cells (7288 c hepatoma cells) and isolated hepatocytes of rat. The cells were incubated at different concentrations of the acid during 0 to 6 hours. Both kinds of cells were able to take up the acid. However, the HTC cells have a greater avidity for the eicosatrienoic acid than normal liver cells. The incorporation of the acid modified the fatty acid composition of the cells and caused a decrease in the amount of saturated and monoenoic acid. In both cells eicosatrienoic acid was converted to arachidonic acid. However, in HTC cells arachidonic acid level was low and not correlated with the amount of the eicosatrienoic acid incorporated. This fact is apparently not due to an impairment in delta 5 desaturation activity since this enzyme is active in both cells. It would be possible that arachidonic acid production in malignant cells would be also interrupted in another metabolic pathway after delta 6 desaturation step. The strikingly low amount of arachidonic acid biosynthesized in HTC cells compared to normal hepatocytes could be interpreted as a consequence of a lower availability of eicosatrienoic acid for the microsomal desaturation system in malignant cells, in addition to the low delta 6 desaturase activity.

8,11,14-Eicosatrienoic Acid↗
PubMed1985

Effect of insulin on the oxidative desaturation of fatty acids in non-diabetic rats and in isolated liver cells.

The effect of insulin on the oxidative desaturation of 1-14C palmitic acid to palmitoleic acid (delta 9 desaturase) 1-14C linoleic acid to alpha-linolenic acid (delta 6 desaturase) on rat liver microsomes and 1-14C eicosa-8,11,14-trienoic acid to arachidonic acid (delta 5 desaturase) on rat liver microsomes and hepatoma tissue culture (HTC) cells was studied. After 12 h of insulin injection, at a dose of 2.5-12.5 U/kg no change was found in delta 9 desaturation activity while delta 5 desaturation activity decreased. The conversion of linoleic to alpha-linolenic acid decreased when the amount of insulin injected was 5 U/kg or more. The effect of insulin (5 U/kg) on delta 9, delta 6 or delta 5 desaturation activity was tested from 1 to 12 h after the injection. The conversion of palmitic acid to palmitoleic acid showed no important changes along the time, while delta 5 desaturation activity decreased at all the times tested. delta 6 Desaturation activity showed a slight increase after 1 h of insulin treatment and then decreased significantly up to the end of the experiment. The addition of 400 mU/ml or more of insulin to the incubation medium of HTC cells produced a significant decrease on the conversion of eicosatrienoic acid to arachidonic acid. The effect of insulin on fatty acid desaturation activity of liver microsomes of normal rats differs from that of diabetic rats. The role of this hormone in relation to other hormones, carbohydrate metabolism and lipid biosynthesis on the activity of the desaturases was discussed.

8,11,14-Eicosatrienoic Acid↗
PubMed1985

[Biochemistry of the developmental cycle of Triatoma infestans. VII. Lipid composition of the cuticle surface extracted with hexane].

The lipid composition of Triatoma infestans (vinchuca) epicuticle was studied. The insect lipids were extracted with hexane and fractionated by chromatography. Long chain fatty alcohols, hydrocarbons, waxes, triacylglycerols, diacylglycerols, free fatty acids, cholesterol, cholesterol esters and monoacylglycerols were detected. The composition of fatty acids, hydrocarbons and alcohols was studied by gas-liquid chromatography and mass spectrography. Linear and branched hydrocarbons were separated. Both types had an uneven number of up to 41 carbons. The predominant straight-chain hydrocarbons were in the range of 27 to 33 carbons. The branched hydrocarbons identified were 2-methyl, 3-methyl and internally branched monomethyl-alkanes and dimethyl-alkanes. The fatty alcohols identified had a straight chain of 16 to 34 carbons with predominance of 28 to 34 specimens. In the different fractions of esterified fatty acids and in the free acids two groups were detected: one formed by acids of 16 to 18 carbons saturated and non-saturated, and another by very long chain saturated acids of 24 to 32 carbons. The presence of alcohols, hydrocarbons and waxes, constituted mainly by long and saturated chains, conveys to the cuticle high resistance to physical and chemical aggressors of the environment.

Animals↗
PubMed1985

[Biochemistry of the growth cycle of Triatoma infestans (Vinchuca). VIII. Preliminary study of hemo-lymphatic apolipoproteins in the adult male].

The three lipoproteins of high (HDL) and very high density (VHDL-I and VHDL-II) were separated from the hemolymph of adult males of T. infestans. They were delipidated and the corresponding apolipoproteins examined by sodium dodecyl sulphate (SDS) gel electrophoresis in polyacrylamide in the presence of 2-mercaptoethanol. They were also iso-electro focused in a pH gradient of 5 to 8. All three groups of apolipoproteins were separated in several polypeptide chains, but all of them had in common a glycoproteic unit of PM 86 000. The VHDL-II presented the simplest composition with predominance of the 86 000 band. Apo-HDL was the most complicated and showed intense bands of PM as high as 210 000 and as low as 17 000 together with 44 000 and 86 000 and less intense bands. The apo-VHDL-I was separated in polypeptides chains in the range of 86 000 to 17 000. The different apoproteins are apparently formed by the association at least in part of common polypeptides.

Animals↗
PubMed1984

Short-chain aliphatic alcohols increase rat-liver microsomal membrane fluidity and affect the activities of some microsomal membrane-bound enzymes.

n-Butyl and isoamyl alcohols decrease the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene and enhance the efficiency of pyrene excimer formation when these probes are incorporated in rat-liver microsomal membrane, suggesting an increase in rotational and translational mobilities. Neither alcohol modifies NADH-ferricyanide reductase activity but both increase NADH-cytochrome c reductase activity. This was interpreted as an increase in the rate of lateral diffusion of the proteins cytochrome b5 and cytochrome b5 reductase as a consequence of the enhanced membrane lipid phase fluidity. Microsomal delta 9 and delta 6 desaturase activities in the presence of isoamyl alcohol were also studied. This alcohol decreases delta 9 desaturation when it is measured at a low substrate concentration (13 microM palmitic acid), but it is not modified when it is measured at a high substrate concentration (66 microM palmitic acid). delta 6 desaturation is diminished by isoamyl alcohol when it is measured with both 13 microM and 66 microM linoleic acid. The influence of isoamyl alcohol on the glucose-6-phosphatase system activity was also studied. In non-detergent-treated microsomes, isoamyl alcohol enhances glucose-6-phosphatase activity. However, if microsomes are previously treated with 0.1% Triton X-100 isoamyl alcohol does not modify this activity. The enhancement of the glucose 6-phosphate transport rate is not due to membrane permeability barrier disruption, since isoamyl alcohol does not modify mannose-6-phosphohydrolase latency. This would suggest that an increase in membrane lipid phase fluidity specifically activates glucose 6-phosphate transport across the membrane.

Animals↗
PubMed1984

Effect of fatty acids of omega 6 series on the biosynthesis of arachidonic acid in HTC cells.

The influence of the preincubation of HTC cells with fatty acids of omega 6 series and columbinic acid (5t, 9c, 12c 18:3) on the biosynthesis of arachidonic acid was studied. The cells were incubated on a chemically defined medium with or without the addition of unlabeled linoleic, gamma-linolenic, eicosatrienoic, arachidonic, docosatetraenoic, docosapentaenoic and columbinic acids. After 24 hr of preincubation in the presence of the aforementioned fatty acids, [1-14C]eicosa-8,11,14-trienoic acid was added to the culture medium as the only lipidic source. Twenty-four hours later the synthesis of arachidonic acid and the fatty acid composition of the cells were determined. At 20 microM concentration the omega 6 fatty acids studied except docosapentaenoic acid produced an increase on the biosynthesis of arachidonic acid compared to the cells incubated in the absence of unlabeled fatty acids in the medium. The fatty acids added to the culture medium were incorporated into the cells and modified their fatty acid composition. Columbinic acid, with a similar structure to linoleic acid, also produced a significant increase on the conversion of eicosatrienoic acid to arachidonic acid. These results would suggest that the effect of both, linoleic and columbinic acids, may be adscribed to their configuration and not necessarily to their transformation in higher homologs, since columbinic acid is unable to be desaturated.

8,11,14-Eicosatrienoic Acid↗