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1.
The kinetics and metabolism of physiological doses of all-trans-retinoic acid were examined in blood and small intestinal mucosa of vitamin A-depleted rats. A major portion of intrajugularly injected retinoic acid is rapidly (within 2 min) sequestered by tissues; subsequently 13-cis-retinoic acid and polar metabolites are released into circulation. All-trans-retinoic acid appears in small intestinal epithelium within 2 min after dosing and is the major radioactive compound there for at least 2 h. Retinoyl glucuronide and 13-cis-retinoic acid are early metabolites of all-trans-retinoic acid in the small intestine of bile duct-cannulated rats. Retinoyl glucuronide, the major metabolite of retinoic acid intestinal epithelium, in contrast to other polar metabolites, was not detected in circulation. An examination of [3H]retinyl acetate metabolites under steady state conditions in vitamin A-repleted rats demonstrates the occurrence of all-trans-retinoic acid and 13-cis-retinoic acid in circulation and in intestinal epithelium, in a pattern similar to that found after injection of retinoic acid into vitamin A-depleted rats. Our data establish that all-trans-retinoic acid, 13-cis-retinoic acid, and retinoyl glucuronide are physiological metabolites of vitamin A in target tissues, and therefore are important candidates as mediators of the biological effect of the vitamin. 相似文献
2.
Immunological memory to tetanus toxoid is established and maintained in the vitamin A-depleted rat 总被引:2,自引:0,他引:2
We have previously shown that vitamin A deficiency severely impairs the young rat's ability to produce specific antibodies after primary immunization with tetanus toxoid (TT). In the present studies, we asked whether immunologic memory to TT is established even in the vitamin A-depleted animal, and if so, whether such memory can be elicited after subsequent repletion with retinol. Vitamin A-depleted rats produced very low concentrations of TT-specific IgM and IgG antibodies in both the primary and secondary responses; however, the ratios of secondary to primary IgM anti-TT and of IgG anti-TT were normal. When rats were repleted with retinol 1 day after immunization, IgM and IgG anti-TT concentrations in both the primary and secondary responses were at least as great as those of control rats. For rats repleted with retinol 2 days before the booster immunization, secondary IgM and IgG anti-TT concentrations were equal in magnitude to those of vitamin A-sufficient controls. For all groups, the kinetics of the antibody response were similar. We conclude that immunological memory is intact in the vitamin A-depleted animal, as shown by 1) the normal ratio of its secondary to primary antibody responses, 2) the restoration of a quantitatively normal secondary antibody response in previously vitamin A-depleted animals repleted with retinol just before boosting with TT, and 3) a normal class switch from IgM to IgG. Retinol deficiency is also characterized by an abnormal elevation of total plasma IgG, despite the inability of the vitamin A-depleted animal to produce normal quantities of specific antibodies after challenge with antigen. 相似文献
3.
The effects of retinoid addition of vitamin A-depleted (UV-irradiated) culture medium on uridine metabolism in cultured Sertoli cells have been studied. After vitamin A depletion, a consistent 2- to 4-fold enhancement of [3H]uridine incorporation into RNA was observed. Several lines of evidence indicate that this enhancement is the result of an increase in the specific activity of the uridine-labeled precursors of RNA. Although vitamin A depletion did not affect either uridine uptake or alter cellular RNA content, a 5-fold increase in the specific activity of UMP was found in vitamin A-depleted cells. This increase results because the cellular content of uracil nucleosides plus nucleotides is selectively lowered in vitamin A-depleted cells. The decreased content of uridine derivatives could be accounted for by a 45-57% decrease in the activity of glutamine-dependent carbamylphosphate synthetase in vitamin A-depleted cells. The effects of vitamin A deprivation on uridine incorporation, as well as carbamylphosphate synthetase activity, could be completely restored to or above control values by supplementing vitamin A-depleted cell culture medium with either retinol or retinoic acid. This effect of vitamin A depletion appears to be highly specific. Under the same conditions, no gross alteration in either the pattern or extent of synthesis of cellular or secreted proteins, glycoproteins, glycosaminoglycans, and lipids was observed. In addition, vitamin A depletion/repletion had no effect on the growth rate or morphology of the cells. 相似文献
4.
Polyadenosine 5′-diphosphate ribose (poly-ADPR) was isolated from animal tissue nuclei which had been extracted by the citric acid method. The isolation of poly-ADPR involved four steps; acidic precipitation of the homogenized nuclei; alkaline hydrolysis of the RNA and proteins; separation of poly-ADPR and DNA by cesium chloride equilibrium centrifugation; and chromatography on Sephadex G-10, enabling us to obtain poly-ADPR without DNA or RNA contamination and in water solution. The isolated poly-ADPR was broken down into its subunit, the ADPR, and then quantitatively determined by a fluorometric method modified for the determination of picomole quantities of ADPR. 相似文献
5.
P A Peterson L Rask T Helting L Ostberg Y Fernstedt 《The Journal of biological chemistry》1976,251(16):4986-4995
Crude cell membrane fractions from a number of tissues can form acidic glycolipids. The formation of acidic galactose lipid and mannose lipid was greatly reduced in vitamin A deficiency, primarily in tissues known to be mucus-producing. Mouse mastocytoma tissue was active in forming acidic galactose lipids with UDP-galactose as substrate. One of the products was identified as retinylphosphate galactose. The synthetase reaction producing this compound exhibited an apparent pH optimum at 6.3. The presence of detergent and retinol stimulated the synthetase reaction, which exhibited an absolute requirement for Mn2+ or Mg2+. The synthetase reaction was readily reversible. Incubation of particulate enzyme with retinylphosphate galactose and UDP yielded UDP-galactose and a compound tentatively identified as retinylphosphate. The galactose lipid was isolated by column chromatography on DEAE-cellulose and silica gel. The retinylphosphate galactose was homogeneous when examined by thin layer chromatography. Mild acid hydrolysis of labeled retinylphosphate galactose yields [14C]galactose, whereas alkaline hydrolysis and hydrogenolysis produced [14C]galactose 1-phosphate. Retinylphosphate galactose bound to vitamin A-depleted, retinol-binding protein. 相似文献
6.
A A Rutten J P Bruyntjes F C Ramaekers 《Virchows Archiv. B, Cell pathology including molecular pathology》1988,56(2):111-117
Keratin expression in hamster tracheal epithelium was investigated during organ culture in serum-free, hormone-supplemented medium using monospecific monoclonal antibodies. Generally, tracheal basal cells expressed keratins detected by antibodies RCK102 and RCK103, while columnar epithelial cells were stained positively by RGE53, RCK103, RCK105 and HCK19. Metaplastic squamous cell foci reacted with antibodies RKSE60, RCK103 and HCK19. Early metaplastic alterations were more clearly RKSE60-positive than the mature lesions. In the vitamin A-depleted tracheas basal cells were clearly RCK102-positive. Superficial cells in the central part of areas of squamous metaplasia induced by cigarette smoke condensate expressed the basal cell keratins, and were negative for the columnar cell keratin 18 detected by the RGE53 antibody. This finding suggests that in cigarette smoke condensate-induced squamous metaplasia basal cells play an important role. The mucus-producing cells at the edges of metaplastic squamous cell foci expressed the keratins specific to columnar cells. Cigarette smoke condensate exposure accelerated epithelial keratinization compared to the vitamin A-depleted epithelium. It was concluded that not only small mucous granule cells, but also basal cells are involved in the development and maintenance of induced squamous metaplasia in tracheal epithelium. Furthermore, in vitro vitamin A-depleted epithelium did not coexpress vimentin in addition to the different keratins. 相似文献
7.
Thompson CL Selby CP Van Gelder RN Blaner WS Lee J Quadro L Lai K Gottesman ME Sancar A 《Journal of biological rhythms》2004,19(6):504-517
Mice exhibit multiple nonvisual responses to light, including 1) photoentrainment of circadian rhythm; 2) "masking," which refers to the acute effect of light on behavior, either negative (activity suppressing) or positive (activity inducing); and 3) pupillary constriction. In mammals, the eye is the sole photosensory organ for these responses, and it contains only 2 known classes of pigments: opsins and cryptochromes. No individual opsin or cryptochrome gene is essential for circadian photoreception, gene photoinduction, or masking. Previously, the authors found that mice lacking retinol-binding protein, in which dietary depletion of ocular retinaldehyde can be achieved, had normal light signaling to the SCN, as determined by per gene photoinduction. In the present study, the authors analyzed phototransduction to the SCN in vitamin A-replete and vitamin A-depleted rbp-/- and rbp-/-cry1-/-cry2-/- mice using molecular and behavioral end points. They found that vitamin A-depleted rbp-/- mice exhibit either normal photoentrainment or become diurnal. In contrast, while vitamin A-replete rbp-/-cry1-/-cry2-/- mice are light responsive (with reduced sensitivity), vitamin A-depleted rbp-/-cry1-/-cry2-/- mice, which presumably lack functional opsins and cryptochromes, lose most behavioral and molecular responses to light. These data demonstrate that both cryptochromes and opsins regulate nonvisual photoresponses. 相似文献
8.
A comparative study of glucose-6-phosphatase, alcaline RNase, ATPase, inosine diphosphatase and 5'-nucleotidase activities in isolated rat liver and hepatoma-27 nuclei and nuclear envelopes was performed. The tumor nuclear membranes were shown to be free from G-6-Pase activity in contrast to the liver nuclear membranes. The nuclear RNase activity was strongly inhibited in the hepatoma and could be unmasked in the presence of 3-10(-4) M pCMB. Hepatoma nuclear and nuclear envelopes ATP-ase activity was found to be moderately decreased as compared to those of the normal tissue. The values of inosine diphosphatase activity in hepatoma were similar to those in liver. The role of the nuclear envelope in nuclear-cytoplasmic interactions as well as nuclear location of G-6-Pase are discussed. 相似文献
9.
Reductions in the concentration of retinol (vitamin A) in serum, lung and kidney were observed in rats subjected to inflammation-inducing treatments (turpentine oil injection of thermal injury). At the same time, the liver showed an almost normal vitamin A content. Feeding of retinol to vitamin A-depleted rats with inflammation revealed that intestinal absorption of retinol was still active in the inflamed state, and the livers of these rats showed good incorporation of retinol. The livers of normal and vitamin A-depleted rats subjected to the inflammatory treatments showed a normal RBP content (retinol-binding protein) and hepatic release of holo-RBP into the serum was not impaired functionally. These results suggest the possibility that the decreases of vitamin A in the lung, serum and kidney may be due primarily to enhanced local consumption of vitamin A related to the inflammation, rather than to a reduced supply of vitamin A from the liver or to decreased intestinal absorption. In bovine serum albumin (BSA)-sensitized rats produced by direct intubation of BSA into the lungs, the level of vitamin A in the lung decreased prior to that in the liver or serum, supporting the hypothesis that the decrease in vitamin A in the inflamed lungs of these rats may be due mainly to the consumption of vitamin A in the lung in response to inflammation. 相似文献
10.
11.
Steroid-receptor complexes (SRC) of estrogen and progestin were isolated from rat liver and purified 1500-2000-fold. The SRC within the composition of cytosol and purified 2000-fold were characterized by gel filtration of Sephadex G-100 and by DEAE-cellulose chromatography. The purified SRC from rat liver were bound to isolated liver cell nuclei of rats of various age (1.5, 6, 12 and 24 month-old). The maximal binding of progestin and estrogen SRC from rat liver was observed in homologous nuclei of 1.5-month-old animals. The binding of SRC by the nuclei decreased progressively with age, reaching its minimum in 24-month-old rats. The observed differences in the SRC binding by cell nuclei of experimental animals may be the cause of functional changes at various stages of ontogenesis. 相似文献
12.
Nuclear membranes from mammalian liver. VI. Glucose-6-phosphatase in rat liver, a cytochemical and biochemical study 总被引:6,自引:0,他引:6
Activities of glucose-6-phosphatase (G-6-Pase) and other phosphatases were determined in nuclei, nuclear membrane and microsomal fractions and subfractions, and condensed chromatin isolated from the liver of adult, newly born and prenatal rats. The purity of the fractions was controlled by electron microscopic morphometry and by measurement of various marker enzymes. The specific G-6-Pase activity of the nuclear membranes was found to be about 60% that of the microsomes. However, when calculated on the basis of the phospholipid content, all fractions had similar activities. Determinations of G-6-Pase enrichments and recoveries were also made. The correspondence of the hydrolysing activities of glucose-6-phosphate, mannose-6-phosphate, and inorganic pyrophosphate, together with various phosphotransferases, showed the same association of the G-6-Pase with these enzymes in the nuclear envelope as in the microsomal membranes. G-6-Pase was also demonstrated in the fractions by cytochemistry, and the activity was localized alongside the cisternal surfaces of both, inner and outer, nuclear membrane. ‘Free’ inner nuclear membrane fragments contained also G-6-Pase. No activity was observed at the nuclear pore complexes. Both, nuclear and microsomal membranes revealed a parallel rapid perinatal increase of G-6-Pase activity climaxing at 23 to 28 h after birth. Triton-X-100 treatment of isolated nuclei, which was found not to selectively release outer nuclear membranes, resulted in a great decrease of G-6-Pase activity as well as in losses of membrane phospholipids. The results clarify the divergence of earlier reports concerning the presence of G-6-Pase in the perinuclear cisterna and add biochemical evidence to the morphologically derived view of the nuclear envelope as being a special form of the ER system. 相似文献
13.
The effect of alpha-tocopherol on the RNA-polymerase activity in isolated rat nuclei and chromatin from normal and E-deficient rats and the possible role of tocopherol-binding proteins in this process were studied. Some differences in the RNA-polymerase activities of the nuclei were found; however, in vitro added alpha-tocopherol had no effect on the level of the label incorporation into RNA. No effect of alpha-tocopherol on this process was observed after addition of cytosol either. Analysis of chromatins from normal and E-deficient rats revealed no differences in their RNA-polymerase activities. In vitro added alpha-tocopherol increased the RNA-polymerase activity of normal (but not of vitamin E-deficient) rats. Some differences in the RNA-polymerase activities were noted after addition to the incubation medium of the Triton X-100-solubilized nuclear fraction specifically binding alpha-tocopherol. This effect was enhanced in the presence of exogenous alpha-tocopherol. The susceptibility of chromatin from normal and E-deficient rats to DNAse I hydrolysis was also found to be different. It was concluded that vitamin E can influence the RNA-polymerase activity of the nuclei and chromatin as well as the chromatin structure and that alpha-tocopherol-binding proteins are necessary for the vitamin E effect on the RNA-polymerase activity to be manifested. 相似文献
14.
A M Spielvogel M I Mednieks U Eppenberger R A Jungmann 《European journal of biochemistry》1977,73(1):199-212
Protein phosphokinase activity from a 0.5 M NaCl extract of purified porcine ovary nuclei has been resolved by Sephadex G-200 gel filtration into three forms of kinase, protein kinase I and III, both independent of adenosine 3':5'-monophosphate (cyclic AMP), and cyclic-AMP-dependent protein kinase II. Cyclic AMP-binding activity was associated with protein kinase II but not with protein kinases I and III. Protein kinases I, II, and III exhibited different cyclic nucleotide dependency and substrate specificity. Protein kinase II was inhibited by a heat-stable protein from rabbit skeletal muscle, whereas protein kinases I and III were not inhibited. According to previously established criteria [Traugh, J.A., Ashby, C.D. and Walsh, D.A. (1974) nuclear protein kinase II can be classified as cyclic-AMP-dependent protein kinase consisting of regulatory and catalytic subunits. Nuclear protein kinases I and III are cyclic-AMP-independent enzymes. Evidence for the identity of nuclear cyclic-AMP-dependent protein kinase II with cytosol (105 000 X g supernatant fraction) cyclic-AMP-dependent protein kinase was obtained in several ways. Nuclear and cytosol cyclic-AMP-dependent protein kinases exhibited identical elution characteristics on DEAE-cellulose and Sephadex G-200 indicating that both kinases are of similar molecular size and possess similar ionic charge. Both kinases exhibited an identical Km for ATP of 8 muM, showed similar substrate specificity, and revealed similar antigenic properties. Cyclic-AMP-dependent protein kinase II was also identified in nuclei isolated in nonaqueous media, eliminating the possibility that the cyclic-AMP-dependent protein kinase activity identified in nuclei isolated in aqueous media may have arisen as the result of cytoplasmic contamination. After incubation of neonatal porcine ovaries which lack nuclear cyclic-AMP-dependent protein kinase with 0.1 muM 8-p-chlorophenylthio cyclic AMP, considerable cyclic-AMP-dependent protein kinase II activity was identified in nuclei isolated in nonaqueous media. From these data it is concluded that the nuclear cyclic-AMP-dependent protein kinase II is related to or identical with the ovary cytoplasmic cyclic-AMP-dependent protein kinase, supporting the concept that nuclear cyclic-AMP-dependent protein kinase is of cytoplasmic origin. 相似文献
15.
Intrinsic factor or cobalophilin were removed by incubating human gastric juice and pig pyloric extract with purified anti-intrinsic factor and anti-cobalophilin immunoglobulin-G, respectively, covalently coupled to Sepharose. Cobalophilin (transcobalamin I) was also removed from pig serum either by using anti-cobalophilin immunoglobulin-G Sepharose or by Sephadex G-200 chromatography. The one remaining semipurified vitamin B-12-binding protein (intrinsic factor, cobalophilin or transcobalamin II) was then isolated by vitamin B-12-Sepharose affinity chromatography. Intrinsic factors, cobalophilins and transcobalamin II isolated by this two-step procedure were compared by double isotope techniques with the corresponding protein not subjected to affinity chromatography and found to be identical in reaction to antiserum, gel filtration and electrofocusing. The avidity of the isolated and unisolated intrinsic factors for the ileal intrinsic factor receptor was also the same. 相似文献
16.
Isolation of chick intestinal nuclei. Effect of vitamin D3 on nuclear metabolism 总被引:10,自引:6,他引:4
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1. Chick intestinal nuclei were isolated, with practically no contamination from other organelles and whole cells, by centrifugation through 2.4m-sucrose. 2. The proportions of RNA, DNA and protein of the isolated nuclei were unaffected by the vitamin D status of the birds. The RNA/DNA ratio was 0.15. 3. The incorporation of [5-(3)H]orotic acid into the rapidly labelled intestinal nuclear RNA, after a 10min. pulse of the orotic acid, was increased in vitamin D-deficient chicks only 10min. after a 125mug. dose of cholecalciferol. 4. There was no stimulation of the DNA-dependent RNA polymerase activity of the isolated nuclei from birds treated with cholecalciferol. 5. The results are discussed in relation to the changes occurring during the lag period, after administration of cholecalciferol and before Ca(2+) transport is detected, and the function of the vitamin. 相似文献
17.
SYNOPSIS. The basic proteins of Paramecium aurelia nucleus were extracted from isolated nuclei and deoxyribonucleoprotein (DNP) of such nuclei. About 35–40% of the nuclear protein, predominantly a lysine-rich histone, is acid soluble. Five major components of the histone can be distinguished by polyacrylamide gel electrophoresis. Some components of Paramecium histone are similar to those of mammalian histones in their electrophoretic mobility, but they differ from the latter in the electrophoretic velocity and relative levels. The basic to acidic amino acid ratio of the histone from the ciliate is ~1.1–1.5, and its amino acid composition resembles closely that of yeast histone. Through the use of Sephadex G-200 gel filtration for purification of the histones extracted directly from isolated nuclei 2 basic proteins were resolved—component I, with an elution volume of 1.4, constitutes ~20% and component II, with an elution volume of 1.9, ~80%. 相似文献
18.
Synthesis of lipids was studied in isolated nuclei from rat thymus and liver cells. On incubation of the isolated nuclei with [2-14C]acetate and [1-14C]glycerol, the label was intensively incorporated into phospholipids and with a significantly lower intensity into fatty acids and cholesterol. Only trace amounts of radioactivity were detected in the lipids of chromatin prepared from isolated thymus nuclei after their incubation, and this suggested that lipids were mainly synthesized on the nuclear membrane. On the preincubation of thymus tissue homogenate with [2-14C]acetate and the subsequent isolation of the nuclei and chromatin, the radioactivity of chromatin lipids was comparable to the radioactivity of nuclear lipids. The findings suggested that in the isolated nuclei the newly synthesized lipids were not transported into chromatin from the nuclear membrane. The specific radioactivities of individual phospholipids and fatty acids were different in the isolated nuclei and in nuclei obtained from preincubated homogenate. Mechanisms of lipid synthesis in isolated nuclei and causes of the different radioactivities of lipids in the isolated nuclei and in the nuclei obtained from the preincubated homogenate are discussed. 相似文献
19.
B A Tsudzevich N V Roginets Ia B Blium N E Kucherenko 《Ukrainski? biokhimicheski? zhurnal》1984,56(5):515-519
There forms of histone-specific acetyltransferases--A, B and C are obtained from the rat liver nuclei. The isolation process included nuclei generation, ammonium sulphate salting-out of proteins, DEAE-cellulose, hydroxyl-apatite, phosphocellulose chromatography and Sephadex C-200 gel-filtration. Acetyltransferases A, B and C from the nuclei were purified 56.8, 144.1 and 42.3 times, respectively. Histones were preferential substrates of the obtained enzymes. Molecular mass of acetyltransferases was determined by Sephadex G-150 and G-200 gel-filtration. It was 120 for enzyme A, about 90 for B and above 200 kDa for C. 相似文献
20.
Interaction of the retinol/cellular retinol-binding protein complex with isolated nuclei and nuclear components 总被引:12,自引:4,他引:8
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《The Journal of cell biology》1981,91(1):63-68
Retinol (vitamin A alcohol) is involved in the proper differentiation of epithelia. The mechanism of this involvement is unknown. We have previously reported that purified cellular retinol-binding (CRBP) will mediate specific binding of retinol to nuclei isolated from rat liver. We now report that pure CRBP delivers retinol to the specific nuclear binding sites without itself remaining bound. Triton X-100-treated nuclei retain the majority of these binding sites. CRBP is also capable of delivering retinol specifically to isolated chromatin with no apparent loss of binding sites, as compared to whole nuclei. CRBP again does not remain bound after transferring retinol to the chromatin binding sites. When isolated nuclei are incubated with [3H]retinol- CRBP, sectioned, and autoradiographed, specifically bound retinol is found distributed throughout the nuclei. Thus, CRBP delivers retinol to the interior of the nucleus, to specific binding sites which are primarily, if not solely, on the chromatin. The binding of retinol to these sites may affect gene expression. 相似文献