Retinoids are produced by glia in the lateral ganglionic eminence and regulate striatal neuron differentiation.
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In order to identify molecular mechanisms involved in striatal development, we employed a subtraction cloning strategy to enrich for genes expressed in the lateral versus the medial ganglionic eminence. Using this approach, the homeobox gene Meis2 was found highly expressed in the lateral ganglionic eminence and developing striatum. Since Meis2 has recently been shown to be upregulated by retinoic acid in P19 EC cells (Oulad-Abdelghani, M., Chazaud, C., Bouillet, P., Sapin, V., Chambon, P. and Dolle, P. (1997) Dev. Dyn. 210, 173-183), we examined a potential role for retinoids in striatal development. Our results demonstrate that the lateral ganglionic eminence, unlike its medial counterpart or the adjacent cerebral cortex, is a localized source of retinoids. Interestingly, glia (likely radial glia) in the lateral ganglionic eminence appear to be a major source of retinoids. Thus, as lateral ganglionic eminence cells migrate along radial glial fibers into the developing striatum, retinoids from these glial cells could exert an effect on striatal neuron differentiation. Indeed, the treatment of lateral ganglionic eminence cells with retinoic acid or agonists for the retinoic acid receptors or retinoid X receptors, specifically enhances their striatal neuron characteristics. These findings, therefore, strongly support the notion that local retinoid signalling within the lateral ganglionic eminence regulates striatal neuron differentiation. (+info)
Differential mechanisms of retinoid transfer from cellular retinol binding proteins types I and II to phospholipid membranes.
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Cellular retinol-binding proteins types I and II (CRBP-I and CRBP-II) are known to differentially facilitate retinoid metabolism by several membrane-associated enzymes. The mechanism of ligand transfer to phospholipid small unilamellar vesicles was compared in order to determine whether differences in ligand trafficking properties could underlie these functional differences. Unidirectional transfer of retinol from the CRBPs to membranes was monitored by following the increase in intrinsic protein fluorescence that occurs upon ligand dissociation. The results showed that ligand transfer of retinol from CRBP-I was >5-fold faster than transfer from CRBP-II. For both proteins, transfer of the other naturally occurring retinoid, retinaldehyde, was 4-5-fold faster than transfer of retinol. Rates of ligand transfer from CRBP-I to small unilamellar vesicles increased with increasing concentration of acceptor membrane and with the incorporation of the anionic lipids cardiolipin or phosphatidylserine into membranes. In contrast, transfer from CRBP-II was unaffected by either membrane concentration or composition. Preincubation of anionic vesicles with CRBP-I was able to prevent cytochrome c, a peripheral membrane protein, from binding, whereas CRBP-II was ineffective. In addition, monolayer exclusion experiments demonstrated differences in the rate and magnitude of the CRBP interactions with phospholipid membranes. These results suggest that the mechanisms of ligand transfer from CRBP-I and CRBP-II to membranes are markedly different as follows: transfer from CRBP-I may involve and require effective collisional interactions with membranes, whereas a diffusional process primarily mediates transfer from CRBP-II. These differences may help account for their distinct functional roles in the modulation of intracellular retinoid metabolism. (+info)
Regulation of cellular retinol-binding protein type II gene expression by arachidonic acid analogue and 9-cis retinoic acid in caco-2 cells.
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We previously showed that unsaturated fatty acids induced gene expression of cellular retinol-binding protein type II (CRBPII) in rat jejunum [Suruga, K., Suzuki, R., Goda, T. and Takase, S. (1995) J. Nutr. 125, 2039-2044]. In the present study, we investigated this induction mechanism(s) using the human intestinal Caco-2 cell line. The postconfluent mature Caco-2 cells were maintained in serum-free medium containing arachidonic acid or its analogue, 5,8,11, 14-eicosatetraynoic acid (ETYA). Northern blot analysis showed that these compounds induced CRBPII mRNA levels to rise and that this induction was more effective when combined with 9-cis retinoic acid. This effect was independent of cycloheximide and inhibited by actinomycin D. Nuclear run-on assays confirmed that the ETYA and 9-cis retinoic acid-induced increase of CRBPII mRNA levels was due to an increased rate of transcription of its gene. In Caco-2 cells, the transcripts of peroxisome proliferator-activated receptor alpha (PPARalpha) and retinoid X receptor alpha (RXRalpha), which were activated by their ligands ETYA and 9-cis retinoic acid, respectively, were coexpressed. The gel shift study using rat CRBPII gene nuclear receptor response elements (RXRE, RE2, RE3) revealed that several forms of nuclear proteins from Caco-2 cells specifically bound to these elements. Some of these protein/DNA complexes reacted to both anti-RXRalpha and anti-PPAR antibodies. In addition, in-vitro synthesized RXRalpha and PPARalpha cooperatively bound to these elements as a heterodimer and these binding activities were enhanced by addition of ETYA or arachidonic acid but not by addition of 9-cis retinoic acid. These studies suggest that fatty acid or its analogue may regulate CRBPII gene expression through PPAR/RXR heterodimer bound to the nuclear receptor response element(s) of the CRBPII genes. (+info)
Vitamin A-sensitive tissues in transgenic mice expressing high levels of human cellular retinol-binding protein type I are not altered phenotypically.
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The suggested function of cellular retinol-binding protein type I [CRBP(I)] is to carry retinol to esterifying or oxidizing enzymes. The retinyl esters are used in storage or transport, whereas oxidized forms such as all-trans or 9-cis retinoic acid are metabolites used in the mechanism of action of vitamin A. Thus, high expression of human CRBP(I) [hCRBP(I)] in transgenic mice might be expected to increase the production of retinoic acid in tissues, thereby inducing a phenotype resembling vitamin A toxicity. Alternatively, a vitamin A-deficient phenotype could also be envisioned as a result of an increased accumulation of vitamin A in storage cells induced by a high hCRBP(I) level. Signs of vitamin A toxicity or deficiency were therefore examined in tissues from transgenic mice with ectopic expression of hCRBP(I). Testis and intestine, the tissues with the highest expression of the transgene, showed normal gross morphology. Similarly, no abnormalities were observed in other tissues known to be sensitive to vitamin A status such as cornea and retina, and the epithelia in the cervix, trachea and skin. Furthermore, hematologic variables known to be influenced by vitamin A status such as the hemoglobin concentration, hematocrits and the number of red blood cells were within normal ranges in the transgenic mice. In conclusion, these transgenic mice have normal function of vitamin A despite high expression of hCRBP(I) in several tissues. (+info)
Cellular retinol-binding protein I is essential for vitamin A homeostasis.
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The gene encoding cellular retinol (ROL, vitA)-binding protein type I (CRBPI) has been inactivated. Mutant mice fed a vitA-enriched diet are healthy and fertile. They do not present any of the congenital abnormalities related to retinoic acid (RA) deficiency, indicating that CRBPI is not indispensable for RA synthesis. However, CRBPI deficiency results in an approximately 50% reduction of retinyl ester (RE) accumulation in hepatic stellate cells. This reduction is due to a decreased synthesis and a 6-fold faster turnover, which are not related to changes in the levels of RE metabolizing enzymes, but probably reflect an impaired delivery of ROL to lecithin:retinol acyltransferase. CRBPI-null mice fed a vitA-deficient diet for 5 months fully exhaust their RE stores. Thus, CRBPI is indispensable for efficient RE synthesis and storage, and its absence results in a waste of ROL that is asymptomatic in vitA-sufficient animals, but leads to a severe syndrome of vitA deficiency in animals fed a vitA-deficient diet. (+info)
Distribution of vitamin A, retinol-binding protein, cellular retinoic acid-binding protein I, and retinoid X receptor beta in the porcine uterus during early gestation.
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Retinol and retinol-binding protein (RBP), among the major secretory products of the uterine endometrium in the uterine fluid of pigs, are assumed to be of importance for early embryonic development. While uterine RBP has been widely characterized, little information is available on the metabolism of vitamin A itself or other specific binding proteins or nuclear receptors in the uterus of pigs. In the present study, the content and distribution of vitamin A in uterine tissue of pigs during early gestation (Days 14-30) were examined macroscopically and microscopically via autofluorescence and HPLC. In addition, the distribution of specific proteins involved in vitamin A metabolism at the cellular and nuclear level was investigated. Macroscopically, the yellowish-greenish autofluorescence characteristic of vitamin A was observed in uterine endometrium. Microscopy showed that the autofluorescence was associated with glandular and surface epithelium of the endometrium. In these structures, immunoreactive RBP was localized, as was cellular retinoic acid-binding protein I. Retinoid X receptor beta was observed in the nucleus of myometrium and endometrium. The intensity of fluorescence decreased with the progress of gestation. This decrease was paralleled by a decrease in vitamin A content of endometrium and myometrium. In general, vitamin A concentration in the endometrium was higher than in the myometrium (P < 0.01). In the myometrium, if present at all, vitamin A was found almost exclusively as retinyl esters. In the endometrium, the dominant fraction was retinol, representing more than 90% of total vitamin A. These results show for the first time that the yellowish-greenish autofluorescence in the pig uterus can be attributed to vitamin A. Differences in the form of vitamin A present in endometrium and myometrium might point to differences in metabolism. In the myometrium, vitamin A might be stored, and in the endometrium, vitamin A is present primarily as retinol-the form in which it is secreted into the uterine fluid. (+info)
Metabolism of retinaldehyde and other aldehydes in soluble extracts of human liver and kidney.
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Purification and characterization of enzymes metabolizing retinaldehyde, propionaldehyde, and octanaldehyde from four human livers and three kidneys were done to identify enzymes metabolizing retinaldehyde and their relationship to enzymes metabolizing other aldehydes. The tissue fractionation patterns from human liver and kidney were the same, indicating presence of the same enzymes in human liver and kidney. Moreover, in both organs the major NAD(+)-dependent retinaldehyde activity copurified with the propionaldehyde and octanaldehyde activities; in both organs the major NAD(+)-dependent retinaldehyde activity was associated with the E1 isozyme (coded for by aldh1 gene) of human aldehyde dehydrogenase. A small amount of NAD(+)-dependent retinaldehyde activity was associated with the E2 isozyme (product of aldh2 gene) of aldehyde dehydrogenase. Some NAD(+)-independent retinaldehyde activity in both organs was associated with aldehyde oxidase, which could be easily separated from dehydrogenases. Employing cellular retinoid-binding protein (CRBP), purified from human liver, demonstrated that E1 isozyme (but not E2 isozyme) could utilize CRBP-bound retinaldehyde as substrate, a feature thought to be specific to retinaldehyde dehydrogenases. This is the first report of CRBP-bound retinaldehyde functioning as substrate for aldehyde dehydrogenase of broad substrate specificity. Thus, it is concluded that in the human organism, retinaldehyde dehydrogenase (coded for by raldH1 gene) and broad substrate specificity E1 (a member of EC 1. 2.1.3 aldehyde dehydrogenase family) are the same enzyme. These results suggest that the E1 isozyme may be more important to alcoholism than the acetaldehyde-metabolizing enzyme, E2, because competition between acetaldehyde and retinaldehyde could result in abnormalities associated with vitamin A metabolism and alcoholism. (+info)
Changes in levels of cellular retinol- and retinoic-acid-binding proteins of liver and lung during perinatal development of rat.
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Cellular retinol-binding protein and cellular retinoic-acid-binding protein, canditates for mediating the action of vitamin A, were found to be present in tissues of the fetal rat. Cellular retinol-binding proteins were still present in most tissues of the adult, but the retinoic-acid-binding protein was not detected in some, including lung, liver, intestine, and kidney. During perinatal development of lung the level of cellular retinol-binding protein remained relatively constant while the level of the cellular retinoic-acid-binding protein peaked at 10 days postnatally, then declined. It was not detectable in lung tissue from 21-day-old rats. In liver, however, the retinoic-acid-binding protein was not detectable later than 5 days postnatally, while the level of the cellular retinol-binding proteinrose sharply near birth, declining only after 21 days to the lower adult levels. The variations observed in the levels of the two binding proteins suggest different and changing requirements for retinol and retinoic acid in organ development and maturation. (+info)