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1.
Blockade of intestinal lipoprotein clearance in rabbits injected with Triton WR 1339-ethyl oleate 总被引:1,自引:0,他引:1
Although Triton WR 1339 has been used to block triglyceride or cholesterol removal from plasma, no data are available on the extent to which Triton WR 1339 administered to rabbits blocks clearance of newly absorbed dietary lipids. In the present study, we have measured the efficiency of this blockade during a 24-hr interval. After the Triton WR 1339 administration, plasma Sf greater than 400 and d less than 1.019 g/ml lipoprotein lipid concentrations increased greatly, but the concentration of d greater than 1.019 g/ml lipids decreased. In the rabbits fed 0.5% cholesterol for 1 week, the increase in d less than 1.019 g/ml and the decrease in 1.019 less than d less than 1.063 g/ml lipoprotein fractions 24 hr after the Triton WR 1339 injection were much greater than in the chow-fed Tritonized rabbits. After the Triton treatment, 50% of intravenously injected LDL-125I-labeled apoB disappeared in 24 hr, but little or no apoB appeared in other lipoprotein fractions and no VLDL apoB was converted to LDL. Labeled cholesterol and retinol were fed to rabbits and 24-hr increments in plasma cholesteryl- and retinyl-ester label and mass were measured. In chow-fed Tritonized rabbits about one-half of the absorbed oral doses of both labeled lipids was recovered in plasma, indicating that Triton WR 1339 does not completely inhibit the clearance of intestinal lipoproteins. When rabbits were injected with Triton and an ethyl oleate emulsion, the blockade of dietary lipid removal from plasma was substantially improved and chylomicron cholesterol uptake by extra-hepatic tissues was completely abolished. 相似文献
2.
The participation of endocytosis in the formation of Triton-filled lysosomes was followed after injecting Triton WR 1339 simultaneously with colloidal gold or horseradish peroxidase. According to cytomorphometric measurements Triton WR 1339 also significantly enhances autophagy. The analysis of the influence of Triton WR 1339 subfractions shows that autophagy is only augmented by the polymer component (‘macrotriton’) but not by the low molecular component (‘microtriton’) alone. When the latter was injected combined with either macrotriton or colloidal gold particles, autophagy increased to a level observed with Triton WR 1339 and beyond the levels determined for either component alone. Thus, autophagy appears to be stimulated by endocytosis. Autolysosomes are transformed within a few hours to peribiliary ‘dense bodies’. These do not display any significant turnover rate within a period of several days; therefore, dense bodies could be prelabelled with colloidal gold in order to show that they are the target organelles ingesting (macro-)Triton. According to difference spectra autophagy leads to a considerable concentration of microsomal and mitochondria! cytochromes in (macro-)Triton-filled lysosomes far beyond the level detected in ‘normal’ lysosomes. Because of the participation of both hetero- and autophagy in the formation of Triton WR 1339-filled lysosomes they have to be classified as telolysosomes. 相似文献
3.
Triton WR 1339 was found to contain a high molecular weight fraction with globular polymers of ˜ 105 D and a diameter of ˜80 Å (‘macrotriton’) and a low molecular weight fraction (‘microtriton’). The intracellular distribution of subfractions of [3H]Triton WR 1339 was followed by cell fractionation and by gel chromatography techniques in parallel with electron microscopy and autoradiography. Macrotriton is selectively stored in lysosomes and all evidence supports a slowly working endocytotic uptake mechanism. Microtriton permeates quickly into the cells and is rapidly and efficiently released into the bile. The data presented suggest some intracellular leaking of lysosomal contents due to the action of Triton WR 1339. 相似文献
4.
Omega-3 triglycerides modify blood clearance and tissue targeting pathways of lipid emulsions 总被引:4,自引:0,他引:4
Qi K Seo T Al-Haideri M Worgall TS Vogel T Carpentier YA Deckelbaum RJ 《Biochemistry》2002,41(9):3119-3127
Omega-3-rich (n-3) triglycerides (TG) are increasingly recognized as having modulating roles in many physiological and pathological conditions. We questioned whether the catabolism of lipid emulsions would be changed after enrichment with fish oil (n-3) TG as compared to enrichment with omega-6-rich soy oil (n-6) TG. Phospholipid-stabilized emulsions of n-3 TG and n-6 TG were labeled with [(3)H]cholesteryl oleoyl ether and administered by bolus injection to wild-type (WT) mice, mice lacking the low-density lipoprotein receptor (LDL-R) (LDL-R -/-), and apolipoprotein E (apoE) knockout mice (apoE -/-). The effects of exogenous apoE, heparin, Triton WR 1339, and lactoferrin on catabolism of emulsions were also assayed. n-3 TG emulsions were cleared faster from blood and had different extrahepatic tissue targeting compared to n-6 TG emulsions. In apoE -/- and LDL-R -/- mice, blood clearance of n-6 TG emulsions slowed with decreased liver uptake, but no changes were observed in n-3 TG emulsion clearance and tissue uptake compared to WT mice. In WT mice, addition of exogenous apoE to the emulsion increased liver uptake of n-6 TG emulsions but had no impact on n-3 TG emulsions. Pre-injection of heparin increased and Triton WR 1339 and lactoferrin decreased blood clearance of n-6 TG emulsions with little or no effect on n-3 TG emulsions. Liver uptake of n-6 TG emulsions increased after heparin injection and decreased after Triton WR 1339 injection, but uptake of n-3 TG emulsions was not changed. These data show that the catabolism of n-3 TG emulsions and the catabolism of n-6 TG emulsions occur via very different mechanisms. Removal of chylomicron-sized n-6 TG emulsions is modulated by lipoprotein lipase (LPL), apoE, LDL-R, and lactoferrin-sensitive pathways. In contrast, clearance of chylomicron-sized n-3 TG emulsions relies on LPL to a very minor extent and is independent of apoE, LDL-R, and lactoferrin-sensitive pathways. 相似文献
5.
E D Poliakova E B Dizhe T A Klimova T V Denisenko L E Vasil'eva 《Biokhimii?a (Moscow, Russia)》1981,46(2):296-305
The effects of Triton WR 1339, starvation and cholesterol diet on the activities of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) and acetyl-CoA carboxylase and on the rates of mevalonic acid (MVA) biosynthesis from acetyl-CoA and malonyl-CoA in the soluble (140 000 g) and microsomal fractions of rat liver, on the rate of incorporation of these substrates into squalene, cholesterol and lanosterol in the rat liver postmitochondrial fraction and on the rate of fatty acid biosynthesis was studied. The administration of Triton WR 1339 (200 mg per 100 g of body weight twice) stimulated the activity of HMG-CoA reductase and MVA biosynthesis from acetyl-CoA and malonyl-CoA in the intact and solubilized microsomal fractions and had no effect on these parameters in the soluble fraction. Starvation for 36 hrs did not cause inhibition of the reductase activity or MVA biosynthesis from both substrates in the soluble fraction. Alimentary cholesterol significantly increased the activity of HMG-CoA reductase, had no effect on the rate of MVA biosynthesis from acetyl-CoA and stimulated the malonyl-CoA incorporation in to MVA in the soluble fraction. Starvation an alimentary cholesterol inhibited the HMG-CoA reductase activity and MVA biosynthesis from both substrates in the solubilized microsomal fraction. Triton WR 1339 stimulated 4--19-fold the lipid formation in the total unsaponified fraction and its components i.e. squalene, lanosterol, cholesterol, from acetyl-CoA and only insignificantly (1,2--1,7-fold) increased malonyl-CoA incorporation into these compounds. Starvation and alimentary cholesterol repressed lanosterol and cholesterol biosynthesis from acetyl-CoA, decreased malonyl-CoA incorporation into these sterols and had no influence on squalene biosynthesis from the two substrates. Triton WR 1339 and starvation inhibited the acetyl-CoA carboxylase activity, unaffected by alimentary cholesterol. No significant changes in the rate of fatty acid biosynthesis from the substrates were observed. The data obtained provide evidence for the existence of autonomic pathways of MVA biosynthesis localized in the soluble and microsomal fractions of rat liver. The pathway of MVA biosynthesis in the soluble fraction is less sensitive to regulatory factors. Sterol biosynthesis from malonyl-CoA is also more resistant to regulatory effects than sterol biosynthesis from acetyl-CoA. This suggests that HMG-CoA reductase localized in the soluble fraction takes part in MVA and sterol biosynthesis from malonyl-CoA. 相似文献
6.
Colaço M Misquith S Bapat MM Wattiaux-De Coninck S Wattiaux R 《Biochemical and biophysical research communications》2004,319(4):1299-1306
Intravenous injection of gelonin and deglycosylated gelonin led to rapid clearance from the blood. Both molecules distributed similarly in liver and kidney suggesting that they followed the same pathway. Deglycosylation reduced the uptake by a third in liver, but did not affect uptake by kidney. Studies with Triton WR1339 showed a classical lysosomal pathway for both molecules. The deglycosylated molecule was degraded to a greater extent than native gelonin as seen by the presence of acid soluble radioactivity. Cell separation showed that while endothelial cells mainly took up native gelonin, Kupffer cells took up the deglycosylated molecule. 相似文献
7.
Fate of asialofetuin endocytosed by rat liver 总被引:1,自引:0,他引:1
R Wattiaux S Misquith S Wattiaux-De Coninck F Dubois 《Biochemical and biophysical research communications》1989,158(1):313-318
We have investigated the endocytosis by rat liver of asialofetuin coupled to [125I] tyramine cellobiose: [125I] TCASF. Subcellular distribution of radioactive compounds was established after differential and isopycnic centrifugation and by analysing the fractions by SDS electrophoresis. Labelling secondary lysosomes was performed by injecting rats with Triton WR 1339 four days before injecting the protein. Results show that after being associated with endosomes [125I] TCASF is recovered in organelles where they are subjected to a first degradation, the density of these organelles is practically not affected by Triton WR 1339 injection. Later the degradation products are associated with lysosomes whose density is markedly lowered by Triton WR 1339 treatment. These observations suggest that the first intracellular organelles where [125I] TCASF is subjected to digestion are distinct from the secondary lysosome population. This could be in agreement with the hypothesis that supposes that endosomes acquire enzymes from primary lysosomes before fusion with secondary lysosomes. 相似文献
8.
Victor Lorian 《Applied microbiology》1967,15(5):1202-1205
In a comparison of Lowenstein's medium with two agar media, 7H10 without and with the addition of Triton WR 1339, 4,100 sputa were planted on all three, yielding 803 (19.6%) positive cultures. The addition of Triton WR 1339 to the 7H10 medium made it possible to observe the cord formation directly on the isolation plates and did not alter, in any respect, the results obtained with the basic 7H10 medium. The agar media produced much earlier and more positive cultures than Lowenstein's medium. A greater number of cultures showed drug resistance on Lowenstein's medium as compared with the agar media. 相似文献
9.
Takahashi Y Inaba N Kuwahara S Kuki W 《Bioscience, biotechnology, and biochemistry》2003,67(11):2448-2450
We studied lipid metabolism to evaluate the effects of gamma-terpinene on suppression of increases in serum lipid concentrations using Triton WR1339-treated rats. At 6 hr after Triton WR1339 injection, the total cholesterol and triglyceride concentrations in the gamma-terpinene group underwent statistically significant decreases (18.3 and 30.3%, respectively) compared with those of the Triton-treated group. 相似文献
10.
Identification of a defence mechanism in vivo against the leakage of enterokinase into the blood.
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D A Grant A I Magee D Meeks C Regan D R Bainbridge J Hermon-Taylor 《The Biochemical journal》1979,184(3):619-626
1. The serum proteinase inhibitors alpha 1-antitrypsin, alpha 2-macroglobulin, inter-alpha-trypsin inhibitor and C1-esterase inhibitor were found not to affect the catalytic activity of human enterokinase, whereas bovine trypsin activity was modified essentially as expected. Enterokinase was also not inhibited by Trasylol (trypsin inhibitor from bovine lung) or bovine pancreatic trypsin inhibitor. No other component in human or mouse serum complexing with enterokinase was identified. 2. Human enterokinase administered intravenously into mice was rapidly cleared from the circulation with a half-life of 2.5 min. This removal was not the result of the difference in species, since partially purified mouse enterokinase was cleared at the same rate as the human enzyme. Clearance was mediated by recognition of the carbohydrate portion of enterokinase and not through specific recognition of its catalytic site. Immunofluorescent staining showed that the enzyme accumulated in the liver. Attempts to block the clearance by the simultaneous infusion of competing glycoproteins suggested that enterokinase was taken up by hepatocytes. Of the glycoproteins tested only two, human lactoferrin (terminal fucosyl alpha 1 leads to 3 N-acetylglucosamine) and bovine asialo-fetuin (terminal galactosyl beta 1 leads to 4 N-acetylglucosamine) were weakly competitive. Two inhibitors of endocytosis, Intralipid and Triton WR1339, failed to delay the removal of enterokinase. It is proposed that enterokinase is cleared from the circulation by an as yet uncharacterized hepatocyte receptor. 相似文献
11.
New Zealand white rabbits exhibited as much as a threefold increase in plasma cholesterol but no change in hepatic cholesterol when fasted for 7-9 days. Agarose electrophoresis and ultracentrifugation of plasma samples showed that only low density lipoprotein increased during fasting. Fasting changed the composition of the low density lipoprotein by increasing the percentage of cholesterol and decreasing the percentage of triglyceride while protein and phospholipid remained the same. Rates of cholesterol secretion into plasma, measured by Triton WR 1339 injection, and rates of plasma cholesteryl ester synthesis, determined by [2-(14)C]mevalonate injection, were similar for fed and fasted rabbits. These findings suggest that fasting hypercholesterolemia in rabbits did not result from increased production of low density lipoproteins. Triton WR 1339 was shown to inhibit plasma cholesterol esterification in vitro. 相似文献
12.
Rat liver lysosomes were isolated by free-flow electrophoresis and were examined morphologically and enzymatically for purity. Their membrane fraction was prepared by osmotic shock and analyzed for cholesterol, phospholipids and fatty acids. The results were compared with the membrane fraction of Triton WR 1339-filled lysosomes and with mitochondria. The cholesterol content (0.269 M cholesterol per M lipid phosphorus), the sphingomyelin concentration (7.9% of total lipid phosphorus) and the degree of unsaturation of fatty acids (38–45%) were found to be intermediate between those of membranes of Triton WR 1339-filled lysosomes (“plasma membrane-like”) and mitochondria (“endoplasmic reticulum-like”). The similarity of these results with corresponding data for the Golgi apparatus support the present view concerning the formation of primary lysosomes via the Golgi apparatus. The drastic changes in the lipid composition found after overloading with Triton WR 1339 confirm that the plasma membrane participates in the formation of the secondary lysosomal membrane. The data presented here underline the significance of the analysis of membrane lipids in evaluating correlations between morphologically different but functionally closely related membrane types. 相似文献
13.
Moen CJ Tholens AP Voshol PJ de Haan W Havekes LM Gargalovic P Lusis AJ van Dyk KW Frants RR Hofker MH Rensen PC 《Journal of lipid research》2007,48(10):2182-2192
The Hyplip2 congenic mouse strain contains part of chromosome 15 from MRL/MpJ on the BALB/cJ background. Hyplip2 mice show increased plasma levels of cholesterol and predominantly triglycerides (TGs) and are susceptible to diet-induced atherosclerosis. This study aimed at elucidation of the mechanism(s) explaining the hypertriglyceridemia. Hypertriglyceridemia can result from increased intestinal or hepatic TG production and/or by decreased LPL-mediated TG clearance. The intestinal TG absorption and chylomicron formation were studied after intravenous injection of Triton WR1339 and an intragastric load of olive oil containing glycerol tri[(3)H]oleate. No difference was found in intestinal TG absorption. Moreover, the hepatic VLDL-TG production rate and VLDL particle production, after injection of Triton WR1339, were also not affected. To investigate the LPL-mediated TG clearance, mice were injected intravenously with glycerol tri[(3)H]oleate-labeled VLDL-like emulsion particles. In Hyplip2 mice, the particles were cleared at a decreased rate (half-life of 25 +/- 6 vs. 11 +/- 2 min; P < 0.05) concomitant with a decreased uptake of emulsion TG-derived (3)H-labeled fatty acids by the liver and white adipose tissue. The increased plasma TG levels in Hyplip2 mice do not result from an enhanced intestinal absorption or increased hepatic VLDL production but are caused by decreased LPL-mediated TG clearance. 相似文献
14.
Uptake and degradation of 125I-labelled asialo-fetuin by isolated rat hepatocytes. 总被引:16,自引:0,他引:16
125I-Labelled asialo-fetuin was taken up by isolated rat hepatocytes by a saturable process. Half maximum uptake was seen at about 3 - 10(-8) M asialo-fetuin. Non-parenchymal liver cells did not take up asialo-fetuin in vitro. Rate of uptake of asialo-fetuin exceeded rate of degradation at all concentrations of asialo-fetuin tested. Asialo-fetuin consequently accumulated in the cells until the extracellular supply was exhausted. Asialo-fetuin degradation could be studied without concurrent uptake by incubating cells, previously exposed to asialo-fetuin, in asialo-fetuin-free medium. Degradation, as evidenced by increase in acid-soluble radioactivity, was inhibited by NH4Cl and chloroquine. The change with time in the intracellular distribution pattern of radioactivity in cells that had been exposed to 125I-labelled asialo-fetuin for 10 min was examined by means of differential centrifugation. Initially, the radioactivity was found mostly in the microsomal fraction. 60 min after the exposure to labelled protein, the distribution pattern of radioactivity resembled that of the lysosomal enzyme beta-acetylglucosaminidase. The possibility that asialo-fetuin digestion takes place in lysosomes is discussed. 相似文献
15.
125I-labelled asialo-fetuin was taken up by isolated rat hepatocytes by a saturable process. Half maximum uptake was seen at about 3 . 10(-8) M asialo-fetuin. Rate of uptake of asialo-fetuin exceeded rate of degradation at all concentrations of asialo-fetuin tested. Degradation of asialo-fetuin, as indicated by release of acid-soluble radioactivity from the cells, was inhibited by NH4Cl and chloroquine. The intracellular distribution of labelled asialo-fetuin was studied by differential and density gradient centrifuging. The distribution curves for radioactivity indicated that asialo-fetuin was present in lysosomes about 1 h after the uptake had started. Chloroquine and ammonium ions seemed to inhibit the uptake of asialo-fetuin into the lysosomes, possibly by interfering with the fusion between phagosomes and lysosomes. 相似文献
16.
Inhibition of proteolytic degradation of low density lipoprotein in human fibroblasts by chloroquine, concanavalin A, and Triton WR 1339. 总被引:23,自引:0,他引:23
The proteolytic degradation of 125I-labeled low density lipoprotein by monolayers of cultured human fibroblasts was prevented by exposure of the cells to chloroquine, an agent that has been reported previously to inhibit lysosomal degradative processes. Chloroquine did not inhibit the binding of low density lipoprotein to its cell surface receptor. However, the two regulatory actions that normally follow low density lipoprotein binding to its receptor, namely suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and stimulation of cholesteryl ester formation, were both prevented when degradation of the lipoprotein was inhibited by chloroquine. Two other agents affecting lysosomal function, Triton WR 1339 and concanavalin A, also inhibited the proteolytic degradation of low density lipoprotein in intact fibroblasts and simultaneously prevented low density lipoprotein-mediated suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and stimulation of cholesteryl ester formation. Unlike chloroquine, however, these two agents also affect the binding of low density lipoprotein to the cells. The inhibitory action of chloropuine, concanavalin A, and Triton WR 1339 could each be reversed by removal of the agent from the culture medium. These in vivo culture data, together with the observation that cell-free extracts of fibroblasts maximally degrade 125I-labeled low density lipoprotein at pH 4 and do not form acid-soluble material above pH 6, are consistent with the hypothesis that the proteolytic degradation of low density lipoprotein by monolayers of fibroblasts occurs within lysosomes. The data also suggest that normal lysosomal function is required in order for low density lipoprotein to regulate cholesterol synthesis and cholesteryl ester formation in the fibroblast system. 相似文献
17.
Carcinoembryonic antigen binding to Kupffer cells is via a peptide located at the junction of the N-terminal and first loop domains 总被引:1,自引:0,他引:1
A 11kD glycopeptide has been isolated by pepsin digestion of carcinoembryonic antigen (CEA) that is rapidly endocytosed by isolated rat Kupffer cells and lung alveolar macrophages. Uptake of this glycopeptide by the isolated cells can be inhibited by excess unmodified CEA. Removal of the N-linked oligosaccharide chains by N-glycanase did not alter cellular uptake but reduced the MW to approximately 5500. A seventeen amino acid N-terminal sequence locates this peptide at the junction of the N-terminal and first loop domain of CEA. It is suggested that the recognition of a peptide sequence in this area of CEA is responsible for its clearance from the circulation. 相似文献
18.
Isolation and characterization of Kupffer and endothelial cells from the rat liver 总被引:20,自引:0,他引:20
Non-parenchymal cell suspensions were prepared from rat livers by three different methods based on a collagenase, a pronase and a combined collagenase-pronase treatment. The highest yield of Kupffer and endothelial cells was obtained with the pronase treatment. Attempts were made for a further purification of these cells by Metrizamide density gradient centrifugation after preferentially loading lysosomal structures in Kupffer cells with Triton WR 1339, Jectofer®, Neosilvol®, Zymosan or colloidal carbon. After loading with Triton WR 1339 or Jectofer®, highly purified endothelial cell suspensions were obtained, but the final Kupffer cell preparations were contaminated with about 20% of endothelial cells. Kupffer and endothelial cells purified in this way showed an altered ultrastructure and contained increased activities of the lysosomal enzymes acid phosphatase, arylsulphatase B and cathepsin D. As an alternative procedure for the purification of Kupffer and endothelial cells, a method based on centrifugal elutriation was employed. With this procedure, highly purified preparations of Kupffer or endothelial cells with a well preserved ultrastructure were obtained. Compared with endothelial cells, purified Kupffer cells had a three times higher cathepsin D activity, whereas the arylsulphatase B activity was three times higher in endothelial cells. The high cathepsin D activity in Kupffer cells could be nearly completely inhibited by the specific cathepsin D inhibitor pepstatin, which excludes a possible contribution to this activity by proteases endocytosed during the isolation of the cells. 相似文献
19.
(1) When lysosomes isolated from the livers of Triton WR 1339-treated rats were incubated for 30 min in the presence of 100 mM KCl and 14CH3NH2, a stimulation by MgATP of the calculated accumulation of the base was observed, in agreement with previous results (Schneider, D.L. (1979) Biochem. Biophys. Res. Commun. 87, 559-565). A similar stimulation was seen with MgITP. Excess EDTA had very little effect on the stimulation by MgATP. (2) There was little effect of MgATP or MgITP on the calculated accumulation of 14CH3NH2 if the base was added to the incubation medium 1, 3, 4 or 5 min before terminating the incubation instead of being present for the total incubation period of 30 min. (3) The accumulation of the basic dye, acridine orange, by a crude lysosomal preparation isolated from the livers of untreated rats was found to be stimulated by MgATP, in agreement with earlier results (Dell'Antone, P. (1979) Biochem. Biophys. Res. Commun. 86, 180-189). Similar results were obtained with a crude lysosomal preparation isolated from the livers of Triton WR 1339-treated rats. In both cases, the stimulation was partly oligomycin-sensitive. (4) There was very little or no effect of MgATP on the accumulation of acridine orange by preparations of pure lysosomes isolated from the livers of Triton WR 1339-treated rats. (5) Our data do not acquire us to postulate the existence of an MgATP-dependent proton pump in lysosomes. 相似文献
20.
Uptake of glutamate into vesicles of Halobacterium salinarium has been studied during respiration and in the nonrespiring state. Uptake requires respiration or a minimum gradient in NaCl, which is consistent with an Na+ symport mechanism for uptake, as proposed for H. halobium. By replacing KCl or NaCl by choline chloride, it has been possible to distinguish between the effects of gradients and/or absolute concentration effects of NaCl and KCl. Uptake depends on the concentration of KCl on the inside, but not on a gradient in KCl. This points to a role for K+ as a regulator of uptake rate, but not of total uptake. The uptake of glutamate is not inhibited by a number of acids with similar chemical groups. Inhibition is, however, caused by D-glutamate. This indicates a specific transport site for glutamate. Parallel results are obtained for binding of glutamate to a Triton extract of the vesicle membrane. The variation in binding and uptake properties with the salt concentration is discussed with reference to transport kinetics. 相似文献