Retinyl Ester

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Earl H. Harrison - One of the best experts on this subject based on the ideXlab platform.

  • Isolation and characterization of a microsomal acid Retinyl Ester hydrolase.
    The Journal of biological chemistry, 2005
    Co-Authors: Thomas Linke, Harry D. Dawson, Earl H. Harrison
    Abstract:

    Previous work demonstrated both acid and neutral, bile salt-independent Retinyl Ester hydrolase activities in rat liver homogenates. Here we present the purification, identification, and characterization of an acid Retinyl Ester hydrolase activity from solubilized rat liver microsomes. Purification to homogeneity was achieved by sequential chromatography using SP-Sepharose cation exchange, phenyl-Sepharose hydrophobic interaction, concanavalin A-Sepharose affinity and Superose 12 gel filtration chromatography. The isolated protein had a monomer molecular mass of approximately 62 kDa, as measured by mass spectrometry. Gel filtration chromatography of the purified protein revealed a native molecular mass of approximately 176 kDa, indicating that the protein exists as a homotrimeric complex in solution. The purified protein was identified as carboxylEsterase ES-10 (EC 3.1.1.1) by N-terminal Edman sequencing and extensive LC-MS/MS sequence analysis and cross-reaction with an anti-ES-10 antibody. Glycosylation analysis revealed that only one of two potential N-linked glycosylation sites is occupied by a high mannose-type carbohydrate structure. Using Retinyl palmitate in a micellar assay system the enzyme was active over a broad pH range and displayed Michaelis-Menten kinetics with a K(m) of 86 microm. Substrate specificity studies showed that ES-10 is also able to catalyze hydrolysis of triolein. CholEsteryl oleate was not a substrate for ES-10 under these assay conditions. Real time reverse transcriptase-PCR and WEstern blot analysis revealed that ES-10 is highly expressed in liver and lung. Lower levels of ES-10 mRNA were also found in kidney, testis, and heart. A comparison of mRNA expression levels in liver demonstrated that ES-10, ES-4, and ES-3 were expressed at significantly higher levels than ES-2, an enzyme previously thought to play a major role in Retinyl Ester metabolism in liver. Taken together these data indicate that carboxylEsterase ES-10 plays a major role in the hydrolysis of newly-endocytosed, chylomicron Retinyl Esters in both neutral and acidic membrane compartments of liver cells.

  • Retinyl Ester secretion by intestinal cells a specific and regulated process dependent on assembly and secretion of chylomicrons
    Journal of Lipid Research, 2001
    Co-Authors: Neeru Nayak, Earl H. Harrison, Mahmood M Hussain
    Abstract:

    Retinyl Esters (RE) have been used extensively as markers to study chylomicron (CM) catabolism because they are secreted in the postprandial state with CM and do not exchange with other lipoproteins in the plasma. To un- derstand the mechanism of secretion of RE by the intestine under the fasting and postprandial states, differentiated Caco-2 cells were supplemented with radiolabeled retinol under conditions that support or do not support CM secre- tion. We observed that these cells assimilate vitamin A by a rapid uptake mechanism. After uptake, cells store retinol in both Esterified and unEsterified forms. Under fasting con- ditions, cells do not secrete RE but secrete free retinol un- associated with lipoproteins. Under postprandial condi- tions, cells secrete significant amounts of RE only with CM. The secretion of RE with CM was independent of the rate of uptake of retinol and intracellular free and Esterified ret- inol levels, and was absolutely dependent on the assembly and secretion of CM. The secretion of RE was correlated with the secretion of CM and not with the secretion of total apolipoprotein B. Inhibition of CM secretion by Pluronic L81 decreased the secretion of RE and did not result in their increased secretion with smaller lipoproteins. These data strongly suggest that RE secretion by the intestinal cells is a specific and regulated process that occurs in the post- prandial state and is dependent on the assembly and secre- tion of CM. We propose that RE are added to CM during final stages of lipoprotein assembly and may serve as sign- posts for these steps. — Nayak, N., E. H. Harrison, and M. M. Hussain. Retinyl Ester secretion by intestinal cells: a specific and regulated process dependent on assembly and secretion of chylomicrons. J. Lipid Res. 2001. 42: 272-280.

  • lipoprotein lipase expression level influences tissue clearance of chylomicron Retinyl Ester
    Journal of Lipid Research, 1999
    Co-Authors: Arlette M Van Bennekum, Earl H. Harrison, Richard J. Deckelbaum, Ira J. Goldberg, Yuko Kako, Peter Weinstock, William S. Blaner
    Abstract:

    Approximately 25% of postprandial retinoid is cleared from the circulation by extrahepatic tissues. Little is known about physiologic factors important to this uptake. We hypothesized that lipoprotein lipase (LpL) contributes to extrahepatic clearance of chylomicron vitamin A. To in- vestigate this, ( 3 H)Retinyl Ester-containing rat mesenteric chylomicrons were injected intravenously into induced mu- tant mice and nutritionally manipulated rats. The tissue sites of uptake of 3 H label by wild type mice and LpL-null mice overexpressing human LpL in muscle indicate that LpL expression does influence accumulation of chylomi- cron retinoid. Skeletal muscle from mice overexpressing human LpL accumulated 1.7- to 2.4-fold more 3 H label than wild type. Moreover, heart tissue from mice overexpresss- ing human LpL, but lacking mouse LpL, accumulated less than half of the 3 H-label taken up by wild type heart. Fasting and heparin injection, two factors that increase LpL activity in skeletal muscle, increased uptake of chylomicron ( 3 H) retinoid by rat skeletal muscle. Using ( 3 H)Retinyl palmitate and its non-hydrolyzable analog Retinyl ( 14 C)hexadecyl ether incorporated into Intralipid emulsions, the importance of Retinyl Ester hydrolysis in this process was assessed. We ob- served that 3 H label was taken up to a greater extent than 14 C label by rat skeletal muscle, suggesting that retinoid up- take requires hydrolysis. In summary, for each of our ex- periments, the level of lipoprotein lipase expression in skel- etal muscle, heart, and/or adipose tissue influenced the amount of ( 3 H)retinoid taken up from chylomicrons and/

  • purification and characterization of a neutral bile salt independent Retinyl Ester hydrolase from rat liver microsomes relationship to rat carboxylEsterase es 2
    Journal of Biological Chemistry, 1997
    Co-Authors: Gwoshing Sun, Stefan E H Alexson, Earl H. Harrison
    Abstract:

    Abstract A neutral, bile salt-independent Retinyl Ester hydrolase (NREH) has been purified from a rat liver microsomal fraction. The purification procedure involved detergent extraction, DEAE-Sepharose ion exchange, Phenyl-Sepharose hydrophobic interaction, Sephadex G-100 and Sephacryl S-200 gel filtration chromatographies, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The isolated enzyme has an apparent molecular mass of approximately 66 kDa under denaturing conditions on SDS-PAGE. Analysis of the amino acid sequences of four peptides isolated after proteolytic digestion revealed that the enzyme is highly homologous with other rat liver carboxylEsterases. In particular, the sequences of the four peptides of the NREH (60 amino acids total) were identical to those of a rat carboxylEsterase expressed in the liver (Alexson, S. E. H., Finlay, T. H., Hellman, U., Svensson, L. T., Diczfalusy, U., and Eggertsen, G. (1994) J. Biol. Chem. 269, 17118–17124). Antibodies against this enzyme also react with the purified NREH. Purified NREH shows a substrate preference for Retinyl palmitate over triolein and did not catalyze the hydrolysis of cholEsteryl oleate. With Retinyl palmitate as substrate, the enzyme had a pH optimum of 7 and showed apparent saturation kinetics, with half-maximal activity achieved at substrate concentrations (K m) of approximately 70 μm.

  • lecithin retinol acyltransferase and Retinyl Ester hydrolase activities are differentially regulated by retinoids and have distinct distributions between hepatocyte and nonparenchymal cell fractions of rat liver
    Journal of Nutrition, 1997
    Co-Authors: Tomokazu Matsuura, Earl H. Harrison, Mohamed Z Gad, Catharine A Ross
    Abstract:

    The cellular distribution of enzymes that Esterify retinol and hydrolyze Retinyl Esters (RE) was studied in liver of vitamin A-sufficient, -deficient, and deficient rats treated with retinoic acid or N-(4-hydroxyphenyl)-retinamide. Livers were perfused and cell fractions enriched in hepatocytes, and nonparenchymal cells were obtained for assays of RE and enzyme activity. The specific activity of lecithin:retinol acyltransferase (LRAT) was approximately 10-fold greater in the nonparenchymal cell than the hepatocyte fraction from both vitamin A-sufficient and retinoid-treated rats. Total RE mass, newly synthesized [3H]RE and LRAT activity were positively correlated in liver and isolated cells of both normal (P < 0.0001) and retinoid-treated rats (P < 0.0002). In nonparenchymal cells, these three constituents were nearly equally enriched as evaluated by their relative specific activity values (RSA, defined as the percentage of recovered activity divided by the percentage of recovered protein), which were each significantly greater than 1.0, with values of 4.3 for total RE mass (P < 0.05), 3.6 for newly synthesized [3H]RE (P < 0.01) and 3.8 for LRAT activity (P < 0.01). In contrast, the specific activities of neutral and acid bile salt-independent Retinyl Ester hydrolases (REH) did not vary with vitamin A status, and their RSA values were close to 1.0 in both hepatocytes and nonparenchymal cells. These data show that LRAT and REH are differentially regulated by retinoids and that these enzymes also differ in their spacial distribution between liver parenchymal and nonparenchymal cells.

William S. Blaner - One of the best experts on this subject based on the ideXlab platform.

  • altered hepatic Retinyl Ester concentration and acyl composition in response to alcohol consumption
    Biochimica et Biophysica Acta, 2013
    Co-Authors: Robin D Clugston, Ira J. Goldberg, Hongfeng Jiang, Man Xia Lee, Paul D Berk, Lishin Huang, William S. Blaner
    Abstract:

    Retinoids (vitamin A and its metabolites) are essential micronutrients that regulate many cellular processes. Greater than 70% of the body's retinoid reserves are stored in the liver as Retinyl Ester (RE). Chronic alcohol consumption induces depletion of hepatic retinoid stores, and the extent of this has been correlated with advancing stages of alcoholic liver disease. The goal of this study was to analyze the mechanisms responsible for depletion of hepatic RE stores by alcohol consumption. A change in the fatty-acyl composition of RE in alcohol-fed mice was observed within two weeks after the start of alcohol consumption. Specifically, alcohol-feeding was associated with a significant decline in hepatic Retinyl palmitate levels; however, total RE levels were maintained by a compensatory increase in levels of usually minor RE species, particularly Retinyl oleate. Our data suggests that alcohol feeding initially stimulates a futile cycle of RE hydrolysis and synthesis, and that the change in RE acyl composition is associated with a change in the acyl composition of hepatic phosphatidylcholine. The alcohol-induced change in RE acyl composition was specific to the liver, and was not seen in lung or white adipose tissue. This shift in hepatic RE fatty acyl composition is a sensitive indicator of alcohol consumption and may be an early biomarker for events associated with the development of alcoholic liver disease.

  • the molecular basis of retinoid absorption a genetic dissection
    Journal of Biological Chemistry, 2008
    Co-Authors: Nuttaporn Wongsiriroj, Roseann Piantedosi, Ira J. Goldberg, Krzysztof Palczewski, Thomas P Johnston, Ellen Li, William S. Blaner
    Abstract:

    The intestine and other tissues are able to synthesize Retinyl Esters in an acyl-CoA-dependent manner involving an acyl-CoA:retinol acyltransferase (ARAT). However, the molecular identity of this ARAT has not been established. Recent studies of lecithin:retinol acyltransferase (LRAT)-deficient mice indicate that LRAT is responsible for the preponderance of Retinyl Ester synthesis in the body, aside from in the intestine and adipose tissue. Our present studies, employing a number of mutant mouse models, identify diacylglycerol acyltransferase 1 (DGAT1) as an important intestinal ARAT in vivo. The contribution that DGAT1 makes to intestinal Retinyl Ester synthesis becomes greater when a large pharmacologic dose of retinol is administered by gavage to mice. Moreover, when large retinol doses are administered another intestinal enzyme(s) with ARAT activity becomes apparent. Surprisingly, although DGAT1 is expressed in adipose tissue, DGAT1 does not catalyze Retinyl Ester synthesis in adipose tissue in vivo. Our data also establish that cellular retinol-binding protein, type II (CRBPII), which is expressed solely in the adult intestine, in vivo channels retinol to LRAT for Retinyl Ester synthesis. Contrary to what has been proposed in the literature based on in vitro studies, CRBPII does not directly prevent retinol from being acted upon by DGAT1 or other intestinal ARATs in vivo.

  • Retinyl Ester formation by lecithin retinol acyltransferase is a key regulator of retinoid homeostasis in mouse embryogenesis
    Journal of Biological Chemistry, 2008
    Co-Authors: Lesley Wassef, William S. Blaner, Roseann Piantedosi, Krzysztof Palczewski, Leora Hamberger, Loredana Quadro
    Abstract:

    The developing mammalian embryo is entirely dependent on the maternal circulation for its supply of retinoids (vitamin A and its metabolites). The mechanisms through which mammalian developing tissues maintain adequate retinoid levels in the face of suboptimal or excessive maternal dietary vitamin A intake have not been established. We investigated the role of Retinyl Ester formation catalyzed by lecithin:retinol acyltransferase (LRAT) in regulating retinoid homeostasis during embryogenesis. Dams lacking both LRAT and retinol-binding protein (RBP), the sole specific carrier for retinol in serum, were maintained on diets containing different amounts of vitamin A during pregnancy. We hypothesized that the lack of both proteins would make the embryo more vulnerable to changes in maternal dietary vitamin A intake. Our data demonstrate that maternal dietary vitamin A deprivation during pregnancy generates a severe retinoid-deficient phenotype of the embryo due to the severe retinoid-deficient status of the double mutant dams rather than to the lack of LRAT in the developing tissues. Moreover, in the case of excessive maternal dietary vitamin A intake, LRAT acts together with Cyp26A1, one of the enzymes that catalyze the degradation of retinoic acid, and possibly with STRA6, the recently identified cell surface receptor for retinol-RBP, in maintaining adequate levels of retinoids in embryonic and extraembryonic tissues. In contrast, the pathway of retinoic acid synthesis does not contribute significantly to regulating retinoid homeostasis during mammalian development except under conditions of severe maternal retinoid deficiency.

  • lipoprotein lipase expression level influences tissue clearance of chylomicron Retinyl Ester
    Journal of Lipid Research, 1999
    Co-Authors: Arlette M Van Bennekum, Earl H. Harrison, Richard J. Deckelbaum, Ira J. Goldberg, Yuko Kako, Peter Weinstock, William S. Blaner
    Abstract:

    Approximately 25% of postprandial retinoid is cleared from the circulation by extrahepatic tissues. Little is known about physiologic factors important to this uptake. We hypothesized that lipoprotein lipase (LpL) contributes to extrahepatic clearance of chylomicron vitamin A. To in- vestigate this, ( 3 H)Retinyl Ester-containing rat mesenteric chylomicrons were injected intravenously into induced mu- tant mice and nutritionally manipulated rats. The tissue sites of uptake of 3 H label by wild type mice and LpL-null mice overexpressing human LpL in muscle indicate that LpL expression does influence accumulation of chylomi- cron retinoid. Skeletal muscle from mice overexpressing human LpL accumulated 1.7- to 2.4-fold more 3 H label than wild type. Moreover, heart tissue from mice overexpresss- ing human LpL, but lacking mouse LpL, accumulated less than half of the 3 H-label taken up by wild type heart. Fasting and heparin injection, two factors that increase LpL activity in skeletal muscle, increased uptake of chylomicron ( 3 H) retinoid by rat skeletal muscle. Using ( 3 H)Retinyl palmitate and its non-hydrolyzable analog Retinyl ( 14 C)hexadecyl ether incorporated into Intralipid emulsions, the importance of Retinyl Ester hydrolysis in this process was assessed. We ob- served that 3 H label was taken up to a greater extent than 14 C label by rat skeletal muscle, suggesting that retinoid up- take requires hydrolysis. In summary, for each of our ex- periments, the level of lipoprotein lipase expression in skel- etal muscle, heart, and/or adipose tissue influenced the amount of ( 3 H)retinoid taken up from chylomicrons and/

  • Retinyl Ester hydrolysis and retinol efflux from bfc 1beta adipocytes
    Journal of Biological Chemistry, 1997
    Co-Authors: Shuanghong Wei, Roseann Piantedosi, Katherine Lai, Shailja Patel, Huan Shen, Vittorio Colantuoni, Fredric B Kraemer, William S. Blaner
    Abstract:

    Abstract Adipose tissue is an important storage depot for retinol, but there are no data regarding retinol mobilization from adipose stores. To address this, dibutyryl cAMP was provided to murine BFC-1β adipocytes and its effects on retinol efflux assessed. High performance liquid chromatography analysis of retinol and Retinyl Esters in adipocytes and media indicated that cAMP stimulated, in a time- and dose-dependent manner, retinol accumulation in the culture media and decreased cellular Retinyl Ester concentrations. Study of adipocyte retinol-binding protein synthesis and secretion indicated that cAMP-stimulated retinol efflux into the media did not result from increased retinol-retinol-binding protein secretion but was dependent on the presence of fetal bovine serum in the culture media. Since our data suggested that Retinyl Esters can be hydrolyzed by a cAMP-dependent enzyme like hormone-sensitive lipase (HSL), in separate studies, we purified a HSL-containing fraction from BFC-1β adipocytes and demonstrated that it catalyzed Retinyl palmitate hydrolysis. Homogenates of Chinese hamster ovary cells overexpressing HSL catalyzed Retinyl palmitate hydrolysis in a time-, protein-, and substrate-dependent manner, with an apparentK m for Retinyl palmitate of 161 μm, whereas homogenates from control Chinese hamster ovary cells did not.

Andrew T.c. Tsin - One of the best experts on this subject based on the ideXlab platform.

  • 11-cis ARAT Activity in Primary Culture of Chicken Muller Cells
    2015
    Co-Authors: Alberto Muniz, Elia T. Villazana-espinoza, Bridget Thackeray, Andrew T.c. Tsin
    Abstract:

    A novel retinoid cycle has recently been identified in the cone-dominated chicken retina and this cone cycle accumulates 11-cis Retinyl Esters upon light adaptation. The purpose of this study is to investigate how 11-cis Retinyl Esters are formed in the retina. Primary cultures of chicken Muller cells and cell membrane were incubated with all-trans or 11-cis retinol to study Retinyl Ester synthesis. In Muller cells, Esterification of 11-cis retinol was 4 times greater than Esterification of all-trans retinol. In the presence of palmitoyl CoA and CRALBP, Muller cell membranes synthesized 11-cis Retinyl Ester from 11-cis retinol at a rate which was 20 fold higher than that of all-trans Retinyl Ester. In the absence of CRALBP, 11-cis Retinyl Ester synthesis was greatly reduced (by 7 fold). In the absence of palmitoyl CoA, Retinyl Ester synthesis was not observed. Muller cell membranes incubated with radiolabeled palmitoyl CoA resulted in the transfer of the labeled acyl group to retinol. This acyl transfer was greatly reduced in the presence of progEsterone, a known ARAT inhibitor. 11-cis ARAT activity remained unchanged when assayed in the presence of all-trans retinol, suggesting a distinct catalytic activity from that of all-trans ARAT. Apparent kinetic rates for 11-cis ARAT were 0.135 nmol/min/mg (Vmax) and 11.25μM (KM); and for all-trans ARAT, 0.006

  • Colocalization of 11-cis Retinyl Esters and Retinyl Ester Hydrolase Activity in Retinal Pigment Epithelium Plasma Membrane
    2013
    Co-Authors: Nathan L. Mata, Elia T. Villazana, Andrew T.c. Tsin
    Abstract:

    PURPOSE. TO identify the subcellular locale of 11-cis Retinyl Esters in bovine retinal pigment epithelium (RPE) and to characterize the enzymic mechanism responsible for liberation of 11-cis retinoids in this compartment. METHODS. Endoplasmic reticulum (ER)-enriched and plasma membrane (PM)-enriched protein fractions were prepared from bovine RPE microsomes using sequential discontinuous sucrose and Percoll gradient fractionation. Enzyme markers for ER (such as carboxylEsterase), and PM (such as 5'-nucleotidase [5'-ND]; alkaline phosphatase [AP]; and ouabain-sensitive Na +,K +-ATPase [ATPase]) were used to identify the subfractions. Membrane-associated retinoids were quantified by high-performance liquid chromatography (HPLC) and Retinyl Ester hydrolase (REH) activities were determined by radiometric and chromatographic (HPLC) means. RESULTS. Chromatographic analyses of membrane-associated retinoids showed that 11-cis Retinyl Esters are localized mainly in PM-enriched fractions, whereas all-trans Retinyl Esters are associated predominantly with ER-enriched membranes; profiles of the distribution of 11-cis- and all-trans REH activities were consistent with the Retinyl Ester distribution. Further purification of the crude PM fraction yielded a fraction (P2) that was significantly enriched with 5'-ND (fivefold), ATPas

  • Effect of light exposure on the accumulation and depletion of Retinyl Ester in the chicken retina
    Experimental eye research, 2006
    Co-Authors: Elia T. Villazana-espinoza, Andrea L. Hatch, Andrew T.c. Tsin
    Abstract:

    A previous study has shown that in the cone-rich chicken retina, 11-cis Retinyl Ester increases with light exposure and decreases in subsequent dark adaptation. The purpose of this research is to study how light exposure (intensity and duration) determine the rate of accumulation and utilization as well as the size of this 11-cis Retinyl Ester pool in the chicken retina. Chickens were dark-adapted overnight before exposure to different light intensities and durations. Animals were sampled at regular time intervals. 11-cis retinal and 11-cis Retinyl Ester were extracted from the retina and analyzed by HPLC. An increase in light intensity from 1,000 and 2,000 Lux (for a 20 min exposure) increased the amount of 11-cis Retinyl Ester from 0.38 to 0.75 nmol/mg. An increase in the duration of light exposure from 10 to 20 min (at 2,000 Lux) also increased the amount of 11-cis Retinyl Ester in the chicken retina (from 0.37 to 0.75 nmol/mg). This 11-cis Retinyl Ester pool in the chicken retina was rapidly reduced to baseline level ( approximately 0.20 nmol/mg) upon dark adaptation. The rate of accumulation of 11-cis Retinyl Ester was dependent on light intensity and duration of exposure and the maximum rate was approximately 0.03 nmol/mg/min. In comparison, dark adaptation was associated with a significantly higher rate of 11-cis Retinyl Ester depletion (approximately 0.05), indicating that light exposure and dark adaptation were associated with different biochemical steps of retinoid storage and utilization. Results from this study are the first to show that the size of the 11-cis Retinyl Ester pool, as well as the rate of its accumulation and depletion in the cone-rich chicken retina, are determined by the intensity and duration of light exposure. These data support the suggestion that a light-driven cone cycle exists in the chicken retina.

  • The hepatic Retinyl Ester hydrolase activity is depressed at the onset of diabetes in BB rats.
    The British journal of nutrition, 2003
    Co-Authors: Min Chen, Andrew T.c. Tsin, Alan B. R. Thomson, Tapan K. Basu
    Abstract:

    Dietary vitamin A as Retinyl Ester is hydrolysed and re-Esterified with long-chain fatty acids in the small intestine. The Esterified vitamin A is subsequently stored in the liver, where it is hydrolysed to free retinol to be transported by carrier proteins to the target tissue. A decreased availability of retinol carrier proteins has been suggested to be responsible for affecting metabolic availability of vitamin A in type 1 diabetes. Using BB Wistar rats, the present study was undertaken to examine whether the presence of a hyperglycaemic state modifies Retinyl Ester hydrolase (REH) activity in the intestine and the liver. At the onset of diabetes, hepatic REH enzymatic activity was significantly ðP , 0·05Þ decreased. However, REH activity remained unaffected in the small intestine, including both ileum and jejunum. Diabetes also resulted in decreased plasma and liver concentrations of retinol. An in vitro study was conducted to examine the effect of diabetes on the intestinal uptake of Retinyl palmitate. Jejunum and ileum from diabetic and non-diabetic BB rats were incubated with labelled Retinyl palmitate at different concentrations ranging from 32 to 256 nmol/l. The uptake of Retinyl palmitate was increased in both diabetic and non-diabetic rats together with the increase of substrate concentration. However, no significant difference was observed in the uptake of Retinyl palmitate between diabetic and non-diabetic rats. These present results suggest that the depressed hepatic REH activities may contribute to the diabetes-associated metabolic derangement of vitamin A. BB rats: Diabetes: Vitamin A: Retinyl Ester hydrolases: Intestinal absorption

  • Substrate specificities of Retinyl Ester hydrolases in retinal pigment epithelium.
    Methods in enzymology, 2000
    Co-Authors: Andrew T.c. Tsin, Nathan L. Mata, Jennifer A. Ray, Elia T. Villazana
    Abstract:

    Publisher Summary Previous studies have shown that Retinyl Esters are located in the retinal pigment epithelium (RPE) of many vertebrate species from fishes, amphibians, reptilians, and birds to mammals including humans. To investigate the role of Retinyl Esters in the visual process, a simple hypothesis is porposed that in the RPE, there exists a substrate-specific Retinyl Ester hydrolase relevant to the visual cycle. This chapter presents biochemical methods employed in laboratory to investigate Retinyl Ester hydrolysis in the RPE, with an emphasis on substrate specificity. On the basis of results reported in the literature and on those from the investigation, the location and role of 11-cis-REH seem apparent. Starting from the branch point where 11-cis-Retinyl Esters are synthesized at the ER, they are somehow relocated to the plasma membrane, where they are stored. Hydrolysis of this pool of Retinyl Esters is carried out by a substrate-specific 11-cis-Retinyl Ester hydrolase and the product of the reaction, 11-cis- retinol, is then oxidized by the 11-cis-RD at the PM.

Ira J. Goldberg - One of the best experts on this subject based on the ideXlab platform.

  • altered hepatic Retinyl Ester concentration and acyl composition in response to alcohol consumption
    Biochimica et Biophysica Acta, 2013
    Co-Authors: Robin D Clugston, Ira J. Goldberg, Hongfeng Jiang, Man Xia Lee, Paul D Berk, Lishin Huang, William S. Blaner
    Abstract:

    Retinoids (vitamin A and its metabolites) are essential micronutrients that regulate many cellular processes. Greater than 70% of the body's retinoid reserves are stored in the liver as Retinyl Ester (RE). Chronic alcohol consumption induces depletion of hepatic retinoid stores, and the extent of this has been correlated with advancing stages of alcoholic liver disease. The goal of this study was to analyze the mechanisms responsible for depletion of hepatic RE stores by alcohol consumption. A change in the fatty-acyl composition of RE in alcohol-fed mice was observed within two weeks after the start of alcohol consumption. Specifically, alcohol-feeding was associated with a significant decline in hepatic Retinyl palmitate levels; however, total RE levels were maintained by a compensatory increase in levels of usually minor RE species, particularly Retinyl oleate. Our data suggests that alcohol feeding initially stimulates a futile cycle of RE hydrolysis and synthesis, and that the change in RE acyl composition is associated with a change in the acyl composition of hepatic phosphatidylcholine. The alcohol-induced change in RE acyl composition was specific to the liver, and was not seen in lung or white adipose tissue. This shift in hepatic RE fatty acyl composition is a sensitive indicator of alcohol consumption and may be an early biomarker for events associated with the development of alcoholic liver disease.

  • intestinal dgat1 deficiency reduces postprandial triglyceride and Retinyl Ester excursions by inhibiting chylomicron secretion and delaying gastric emptying
    Journal of Lipid Research, 2012
    Co-Authors: Gene P Ables, Kryscilla Jian Zhang Yang, Silke Vogel, Antonio Hernandezono, Shuiqing Yu, Jason J Yuen, Susan Birtles, Linda K Buckett, Andrew V Turnbull, Ira J. Goldberg
    Abstract:

    Acyl CoA:diacylglycerol acyltransferase (DGAT) 1 catalyzes the final step of triglyceride (TG) synthesis. We show that acute administration of a DGAT1 inhibitor (DGAT1i) by oral gavage or genetic deletion of intestinal Dgat1 (intestine-Dgat1−/−) markedly reduced postprandial plasma TG and Retinyl Ester excursions by inhibiting chylomicron secretion in mice. Loss of DGAT1 activity did not affect the efficiency of retinol Esterification, but it did reduce TG and retinoid accumulation in the small intestine. In contrast, inhibition of microsomal triglyceride transfer protein (MTP) reduced chylomicron secretion after oral fat/retinol loads, but with accumulation of dietary TG and retinoids in the small intestine. Lack of intestinal accumulation of TG and retinoids in DGAT1i-treated or intestine-Dgat1−/− mice resulted, in part, from delayed gastric emptying associated with increased plasma levels of glucagon-like peptide (GLP)-1. However, neither bypassing the stomach through duodenal oil injection nor inhibiting the receptor for GLP-1 normalized postprandial TG or Retinyl Esters excursions in the absence of DGAT1 activity. In summary, intestinal DGAT1 inhibition or deficiency acutely delayed gastric emptying and inhibited chylomicron secretion; however, the latter occurred when gastric emptying was normal or when lipid was administered directly into the small intestine. Long-term hepatic retinoid metabolism was not impacted by DGAT1 inhibition.

  • the molecular basis of retinoid absorption a genetic dissection
    Journal of Biological Chemistry, 2008
    Co-Authors: Nuttaporn Wongsiriroj, Roseann Piantedosi, Ira J. Goldberg, Krzysztof Palczewski, Thomas P Johnston, Ellen Li, William S. Blaner
    Abstract:

    The intestine and other tissues are able to synthesize Retinyl Esters in an acyl-CoA-dependent manner involving an acyl-CoA:retinol acyltransferase (ARAT). However, the molecular identity of this ARAT has not been established. Recent studies of lecithin:retinol acyltransferase (LRAT)-deficient mice indicate that LRAT is responsible for the preponderance of Retinyl Ester synthesis in the body, aside from in the intestine and adipose tissue. Our present studies, employing a number of mutant mouse models, identify diacylglycerol acyltransferase 1 (DGAT1) as an important intestinal ARAT in vivo. The contribution that DGAT1 makes to intestinal Retinyl Ester synthesis becomes greater when a large pharmacologic dose of retinol is administered by gavage to mice. Moreover, when large retinol doses are administered another intestinal enzyme(s) with ARAT activity becomes apparent. Surprisingly, although DGAT1 is expressed in adipose tissue, DGAT1 does not catalyze Retinyl Ester synthesis in adipose tissue in vivo. Our data also establish that cellular retinol-binding protein, type II (CRBPII), which is expressed solely in the adult intestine, in vivo channels retinol to LRAT for Retinyl Ester synthesis. Contrary to what has been proposed in the literature based on in vitro studies, CRBPII does not directly prevent retinol from being acted upon by DGAT1 or other intestinal ARATs in vivo.

  • lipoprotein lipase expression level influences tissue clearance of chylomicron Retinyl Ester
    Journal of Lipid Research, 1999
    Co-Authors: Arlette M Van Bennekum, Earl H. Harrison, Richard J. Deckelbaum, Ira J. Goldberg, Yuko Kako, Peter Weinstock, William S. Blaner
    Abstract:

    Approximately 25% of postprandial retinoid is cleared from the circulation by extrahepatic tissues. Little is known about physiologic factors important to this uptake. We hypothesized that lipoprotein lipase (LpL) contributes to extrahepatic clearance of chylomicron vitamin A. To in- vestigate this, ( 3 H)Retinyl Ester-containing rat mesenteric chylomicrons were injected intravenously into induced mu- tant mice and nutritionally manipulated rats. The tissue sites of uptake of 3 H label by wild type mice and LpL-null mice overexpressing human LpL in muscle indicate that LpL expression does influence accumulation of chylomi- cron retinoid. Skeletal muscle from mice overexpressing human LpL accumulated 1.7- to 2.4-fold more 3 H label than wild type. Moreover, heart tissue from mice overexpresss- ing human LpL, but lacking mouse LpL, accumulated less than half of the 3 H-label taken up by wild type heart. Fasting and heparin injection, two factors that increase LpL activity in skeletal muscle, increased uptake of chylomicron ( 3 H) retinoid by rat skeletal muscle. Using ( 3 H)Retinyl palmitate and its non-hydrolyzable analog Retinyl ( 14 C)hexadecyl ether incorporated into Intralipid emulsions, the importance of Retinyl Ester hydrolysis in this process was assessed. We ob- served that 3 H label was taken up to a greater extent than 14 C label by rat skeletal muscle, suggesting that retinoid up- take requires hydrolysis. In summary, for each of our ex- periments, the level of lipoprotein lipase expression in skel- etal muscle, heart, and/or adipose tissue influenced the amount of ( 3 H)retinoid taken up from chylomicrons and/

  • Lipoprotein lipase hydrolysis of Retinyl Ester. Possible implications for retinoid uptake by cells.
    The Journal of biological chemistry, 1994
    Co-Authors: William S. Blaner, Joseph C. Obunike, S B Kurlandsky, Maysoon Al-haideri, Roseann Piantedosi, Richard J. Deckelbaum, Ira J. Goldberg
    Abstract:

    Adipose tissue contains substantial stores of retinoid (retinol+Retinyl Ester) that, quantitatively, are second only to retinoid stores in the liver. Our studies show that retinoid levels in adipose tissue are markedly influenced by dietary retinoid intake. Because lipoprotein lipase (LPL) increases the uptake of lipoproteins and lipid emulsion particles by many cell types including adipocytes, we investigated whether LPL also increases retinoid uptake by adipocytes from lipid-containing particles. Addition of LPL (10 micrograms/ml) to BFC-1 beta adipocytes produced a 2-fold increase in cellular uptake of [3H]retinoid from a lipid emulsion containing [3H]Retinyl Ester. Heparin, which displaces LPL from binding sites on cell surface proteoglycans, increased [3H]retinoid uptake by an additional 2-fold. High performance liquid chromatography analyses showed that greater than 75% of the media and 85% of the cellular radioactivity was present as retinol. The conversion of Retinyl Ester to retinol by LPL was then assessed using model Retinyl Ester containing lipid emulsions. Although triglyceride appears to be the preferred substrate for LPL, after greater than 25% of the triglyceride was hydrolyzed, significant amounts of Retinyl Ester were hydrolyzed by LPL. Retinyl Ester hydrolysis was increased approximately 20-fold in the presence of a source of apolipoprotein C-II. The physiologically significant palmitate, stearate, oleate, and linoleate Esters of retinol were all hydrolyzed by LPL. When LPL was incubated with [3H]Retinyl Ester containing rabbit mesenteric chylomicrons and in the presence of heparin and apolipoprotein C-II, the LPL was able to completely hydrolyze the Retinyl Ester to retinol. Thus, LPL is able to catalyze the hydrolysis of Retinyl Esters and, through the process of hydrolysis, may facilitate uptake of retinoid by adipocytes.

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  • Specificity of binding of all-trans-Retinyl Ester to RPE65.
    Biochemistry, 2005
    Co-Authors: Pranab Maiti, Deviprasad R. Gollapalli, Robert R Rando
    Abstract:

    Membrane-bound RPE65 (mRPE65) is a binding protein for all-trans-Retinyl Esters, which are the substrates for the isomerization reaction that completes the visual cycle. RPE65 is essential for rhodopsin regeneration and, hence, for vision. As RPE65 appears to be part of the rate-limiting pathway in the visual cycle, specific antagonists of the molecule will be important in evaluating its full physiological role. The protein is known to stereoselectively bind all-trans-Retinyl Esters (tREs), with dissociation constants in the 50 nM range. This study explores the overall binding specificity of RPE65 with respect to both retinoids and other isoprenoids in an effort to define the specificity of binding, and to begin the process of designing specific antagonists for it. The nature of the specificity directed toward the three main structural elements (retinoid, linker, and acyl moieties) in the tRE molecule is reported. In the all-trans-Retinyl Ester series, binding affinity increased as a function of the hydrophobicity of the fatty acyl group. In the linker region, binding affinities were little affected by amide, ketone, and ether replacements for the carboxy Ester moiety of the naturally occurring tRE ligand. Finally, modifications in the all-trans-retinoid moiety are also tolerated. For example, E,E-farnesyl palmitate binds with approximately the same affinity as does all-trans-Retinyl palmitate. Other isoprenoid analogues also bind, as do truncated retinoids in the beta-ionone series. Therefore, mRPE65 is a moderately specific retinoid binding protein directed at long chain all-trans-Retinyl Esters.

  • All-trans-Retinyl Esters are the substrates for isomerization in the vertebrate visual cycle.
    Biochemistry, 2003
    Co-Authors: Deviprasad R. Gollapalli, Robert R Rando
    Abstract:

    The identification of the critical enzyme(s) that carries out the trans to cis isomerization producing 11-cis-retinol during the operation of the visual cycle remains elusive. Confusion exists in the literature as to the exact nature of the isomerization substrate. At issue is whether it is an all-trans-Retinyl Ester or all-trans-retinol (vitamin A). As both putative substrates interconvert rapidly in retinal pigment epithelial membranes, the choice of substrate can be ambiguous. The two enzymes that effect interconversion of all-trans-retinol and all-trans-Retinyl Esters are lecithin retinol acyl transferase (LRAT) and Retinyl Ester hydrolase (REH). The Retinyl Ester or all-trans-retinol pools are radioactively labeled separately in the presence of inhibitors of LRAT and REH, effectively preventing their interconversion. Pulse-chase experiments unambiguously demonstrate that all-trans-Retinyl Esters, and not all-trans-retinol, are the precursors of 11-cis-retinol. When the all-trans-Retinyl Ester pool is radioactively labeled, the resulting 11-cis-retinol is labeled with the same specific activity as the precursor Ester. The converse is true with vitamin A. These data unambiguously establish all-trans-Retinyl Esters as the precursors of 11-cis-retinol.