RESUMO
Differential cross sections and photon-beam asymmetries for the γ[over â]pâπ^{-}Δ^{++}(1232) reaction have been measured for 0.7
RESUMO
The Ï-Λ(1520) interference effect in the γpâK^{+}K^{-}p reaction has been measured for the first time in the energy range from 1.673 to 2.173 GeV. The relative phases between Ï and Λ(1520) production amplitudes were obtained in the kinematic region where the two resonances overlap. The measurement results support strong constructive interference when K^{+}K^{-} pairs are observed at forward angles but destructive interference for proton emission at forward angles. Furthermore, the observed interference effect does not account for the sqrt[s]=2.1 GeV bump structure in forward differential cross sections for Ï photoproduction. This fact suggests possible exotic structures such as a hidden-strangeness pentaquark state, a new Pomeron exchange, or rescattering processes via other hyperon states.
RESUMO
The exclusive reaction γpâK(+)π(-)Σ(+) was measured for the first time using linearly polarized photons at beam energies from 1.85 to 2.96 GeV. Angular distributions in the rest frame of the K(+)π(-) system were fitted to extract spin-density matrix elements of the K(*0) decay. The measured parity spin asymmetry shows that natural-parity exchange is dominant in this reaction. This result clearly indicates the need for t-channel exchange of the κ(800) scalar meson.
RESUMO
Immunocytochemical analyses revealed the presence and distribution of apolipoprotein E (apo E) in normal human brain tissue as well as in 77 human intracranial neoplasms. In normal brain tissues, the perikarya of astrocytes exhibited a strong positive reaction, whereas the Bergmann glia were stained to a moderate degree. However, no immunoreactivity was observed with neurons, oligodendrocytes, ependymal cells, and choroidal epithelium. Among the intracranial neoplasms, oligodendroglioma, choroid plexus papilloma, hemangioblastoma, primary malignant lymphoma, neurinoma, meningioma, pituitary adenoma, and craniopharyngioma were all negative. Immunoreactivity in the peripheral neuroblastoma was nil. However, the perikarya of astrocytomas and glioblastomas showed a positive reaction. Analyses on the degree of anaplasia and the amount of apo-E as an intensity of immunostaining showed a negative correlation. The astrocytic elements were stained in mixed oligoastrocytomas and medulloblastomas with glial differentiation. A few cases of ependymomas showed weak perikaryal immunostaining. Western blot analyses with anti-apo E antibody of a freshly prepared surgical specimen with astrocytomas revealed a single band with a molecular weight of approximately 37,000. The well differentiated cultured human astrocytoma cells secreted apo E into the medium. These lines of evidence suggest that apo E may serve as a potential marker specific for astrocytomas and glioblastomas, as well as an indicator of astrocytic tumor cell differentiation. The apo E localization in human brain tumors could be clinically relevant and diagnostically useful.
Assuntos
Apolipoproteínas E/análise , Biomarcadores Tumorais/análise , Glioma/análise , Imuno-Histoquímica , Apolipoproteínas E/metabolismo , Astrocitoma/análise , Química Encefálica , Neoplasias Encefálicas/análise , Diferenciação Celular , Transformação Celular Neoplásica/análise , Glioma/metabolismo , Glioma/patologia , Humanos , Células Tumorais CultivadasRESUMO
A 360 residue region encoded by the neurofibromatosis type 1 (NF1) gene shows significant homology to the catalytic domains of both mammalian GTPase-activating proteins (GAP) and yeast IRA proteins. This GAP-related domain of the NF1 gene (NF1-GRD), like the GAP and IRA protein, has been reported to mediate hydrolysis of Ras-bound GTP to GDP, resulting in inactivation of Ras protein. In the present study, we identified two different types of NF1-GRD cDNA. One (type I) is identical to the previously reported sequence, and the other (type II) contained an additional 63 bp insertion that encodes for a region of 21 amino acids in the center of the NF1-GRD molecule. Alternative splicing is the most likely mechanism by which these two types of transcripts arise. Our observations reveal that the type I transcript is predominantly expressed in undifferentiated cells, whereas the type II transcript predominates in differentiated cells. Furthermore, the expression pattern of type I and type II NF1-GRD mRNA immediately changed in SH-SY5Y neuroblastoma cells when neuronal differentiation programs were induced by retinoic acid treatment. We propose that the differential expression of type I and type II NF1-GRD transcripts might be an 'on/off' switch that regulates the catalytic activity of the NF1 gene product, which plays an important role in the regulation of neuronal differentiation.
Assuntos
Regulação Neoplásica da Expressão Gênica , Genes da Neurofibromatose 1 , MAP Quinase Quinase Quinases , Neurônios/citologia , Proteínas Oncogênicas , Proteínas Quinases/genética , Proteínas Proto-Oncogênicas , Transcrição Gênica , Sequência de Aminoácidos , Astrocitoma , Sequência de Bases , Diferenciação Celular , Linhagem Celular , Clonagem Molecular , DNA de Neoplasias/genética , Humanos , Dados de Sequência Molecular , Neurofibromina 1 , Oligodesoxirribonucleotídeos , Reação em Cadeia da Polimerase , Conformação Proteica , Proteínas Quinases/análise , Proteínas Quinases/metabolismo , Proteínas/genética , RNA Mensageiro/genética , Homologia de Sequência do Ácido NucleicoRESUMO
To enhance degradation of unconjugated bilirubin in hyperbilirubinemic subjects, we synthesized a bilirubin oxidase (EC 1.3.3.5) (BO) derivative (PEGBO) by covalently linking (2,4-bis[O-methoxy(polyethyleneglycol)]-6-chloro-s-triazine) (PEG) to the enzyme. Intravenously injected BO in rats disappeared from the circulation with a half-life of 2.5 min; the half-life of PEGBO was 190 min. Intravenously injected BO minimally and transiently decreased plasma bilirubin levels in jaundiced Gunn rats and in bile-duct-ligated jaundiced rats. In contrast, PEGBO rapidly and substantially decreased plasma bilirubin levels and the effect persisted for longer than 3 h. Renal dysfunction often occurs in patients with liver diseases. To study the role of bilirubin toxicity for the kidney, functions of transtubular transport for organic anions was measured in bile-duct-ligated jaundiced animals before and after treatment with PEGBO. Bile duct ligation decreased urinary excretion of phenolsulfophthalein (PSP), an organic anion used for renal function test. Treatment of the jaundiced animals with PEGBO increased the rate of PSP disappearance from the circulation and normalized its urinary excretion. Thus, PEGBO might be useful for the study of bilirubin toxicity in jaundiced animals.
Assuntos
Bilirrubina/sangue , Oxirredutases atuantes sobre Doadores de Grupo CH-CH , Polietilenoglicóis/sangue , Triazinas/sangue , Animais , Ductos Biliares , Meia-Vida , Hiperbilirrubinemia/tratamento farmacológico , Imunodifusão , Rim/efeitos dos fármacos , Ligadura , Fígado/metabolismo , Oxirredutases/sangue , Oxirredutases/imunologia , Polietilenoglicóis/síntese química , Polietilenoglicóis/imunologia , Polietilenoglicóis/farmacologia , Ratos , Ratos Gunn , Triazinas/síntese química , Triazinas/imunologia , Triazinas/farmacologiaRESUMO
Rat sinusoidal liver cells take up maleylated bovine serum albumin (maleyl-BSA) and its demaleylated form (demaleyl-BSA) by scavenger receptor-mediated endocytosis. Cellular binding of maleyl-BSA and demaleyl-BSA and its quantitative relation to subsequent intracellular degradation were investigated. The binding affinities of these ligands were almost equal whereas the number of binding sites for maleyl-BSA was more than twice as large than that for demaleyl-BSA. However, no difference was observed in their endocytic degradation. The amounts of maleyl-BSA degraded were proportional to those bound to the cell surface up to a certain level. However, a further increase in cell-bound ligands did not affect the degradation of maleyl-BSA. Several polyanions such as fucoidin and dextran sulfate of Mr = 5000 inhibited the binding of maleyl-BSA but did not affect its degradation. In contrast, acetylated or oxidized low density lipoprotein had virtually no effect on cellular binding of maleyl-BSA but exhibited profound effects on its intracellular degradation. Similar results were obtained with rat peritoneal macrophages. Based on these data, we would propose that two binding sites are involved in the receptor-mediated ligand recognition; one is coupled to subsequent endocytic degradation, and the other serves as a binding site for polyanionic compounds.
Assuntos
Albuminas/metabolismo , Fígado/metabolismo , Proteínas de Membrana , Receptores Imunológicos/metabolismo , Receptores de Lipoproteínas , Soroalbumina Bovina/metabolismo , Animais , Sítios de Ligação , Endocitose/efeitos dos fármacos , Técnicas In Vitro , Lipoproteínas LDL/metabolismo , Fígado/citologia , Macrófagos/metabolismo , Polieletrólitos , Polímeros/farmacologia , Ratos , Receptores Imunológicos/efeitos dos fármacos , Receptores Depuradores , Receptores Depuradores Classe B , Relação Estrutura-AtividadeRESUMO
When S-benzyl-N-acetyl-L-[U-14C]cysteine, a mercapturic acid, was administered to rats intravenously, the plasma level of radioactivity decreased very rapidly with a concomitant increase in the renal level of radioactivity. The renal radioactivity reached its maximum within 2 min and then decreased rapidly with concomitant appearance of the radioactive mercapturic acid in the urine. Bilateral ligation of the ureters resulted in only a slight decrease in the rate of disappearance of mercapturic acid from the plasma, while bilateral nephrectomy caused a marked retardation of its clearance from the plasma. Intravenous administration of probenecid, a well known inhibitor of a renal transtubular transport system for organic acids, caused a significant retardation of mercapturate clearance from the plasma in both of the control and ureter-ligated animals. The renal accumulation of this mercapturic acid as well as its excretion into urine was inhibited by probenecid. All these data suggested that a mercapturic acid in the plasma was preferentially taken up by renal tubule cells from the basolateral side of plasma membranes via the probenecid-sensitive transtubular transport system and then excreted rapidly into the lumenal space. This transtubular transport of a mercapturic acid seems to constitute an important process in the hepato-renal cooperation in the mercapturic acid biosynthesis in vivo.
Assuntos
Acetilcisteína/metabolismo , Túbulos Renais/metabolismo , Acetilcisteína/análogos & derivados , Acetilcisteína/sangue , Animais , Transporte Biológico/efeitos dos fármacos , Cinética , Fígado/metabolismo , Masculino , Probenecid/farmacologia , RatosRESUMO
Topological features and some properties of the membrane-bound peptidase(s) participating in the metabolism of a glutathione S-conjugate in the kidney were studied. S-Carbamidomethyl glutathione, a model compound for glutathione S-conjugate, was demonstrated to be sequentially hydrolyzed by gamma-glutamyltransferase (5-glutamyl)-peptide:amino-acid 5-glutamyltransferase; EC 2.3.2.2) and peptidase(s) bound to rat renal brush border membrane vesicles. Hydrolysis of S-carbamidomethyl cysteinylglycine was found to be inhibited by 1,10-o-phenanthroline, suggesting a participation of a metal-requiring peptidase in this process. The hydrolytic activity of the membranous peptidase was markedly depressed by cysteinylglycine S-acetyldextran polymer (molecular weight, 500 000), a nonpermeating derivative for cysteinylglycine. Papain treatment of brush border membrane vesicles resulted in the solubilization of most hydrolytic activity toward S-carbamidomethyl cysteinylglycine. Amino-peptidase M was also solubilized from the membrane and the increase in the specific activity of this enzyme in the papain-soluble fraction was in parallel within that of the peptidase activity for hydrolysis of S-carbamidomethyl cysteinylglycine. The hydrolytic activity of purified brush border membrane vesicles toward S-carbamidomethyl glutathione was fully reconstituted by the combined use of purified gamma-glutamyltransferase and aminopeptidase M. These findings indicated that, as in the case of the cleavage of gamma-glutamyl linkage of glutathione and related compounds, hydrolysis of the S-substituted cysteinylglycine occurred exclusively on the lumenal surface of renal brush border membrane as catalyzed mainly by aminopeptidase M.
Assuntos
Glutationa/análogos & derivados , Rim/enzimologia , Peptídeo Hidrolases/metabolismo , Aminopeptidases/metabolismo , Antígenos CD13 , Fracionamento Celular , Diazo-Oxo-Norleucina/farmacologia , Dipeptidases/metabolismo , Glutationa/metabolismo , Membranas/enzimologia , Microssomos/enzimologia , Microvilosidades/enzimologia , Fenantrolinas/farmacologia , gama-Glutamiltransferase/metabolismoRESUMO
This study was performed to establish the structural abnormality of a new hemoglobin variant discover-d in a Japanese patient with angina pectoris. The hybridization of the separated hemoglobin with canine hemoglobin revealed a beta-chain anomaly. Peptide betaTp-6 was found to be abnormally located on the peptide map of tryptic digests of the S-carboxymethylated beta-chain from the variant hemoglobin. A structural study on the abnormal betaTp-6 revealed that the variant hemoglobin differs from hemoglobin A by substitution of leucine for valine at residue 60 of the beta-chain. This new variant hemoglobin is designated as hemoglobin Yatsushiro after the name of the city where the propositus lived. The patient is hematologically healthy and his clinical history has nothing to do with this abnormal hemoglobin.
Assuntos
Hemoglobinas Anormais , Código Genético , Hemoglobina A , Hemoglobinas Anormais/isolamento & purificação , Humanos , Leucina/análise , Lisina/análise , Valina/análiseRESUMO
To elucidate a possible function of plasma albumin in vectorial transport of various cholephilic organic anions, such as bile acids, plasma clearance and transhepatic transport of radioactive taurocholate were studied in vivo in normal and mutant analbuminemic rats. Intravenous administration of taurocholate was followed by its rapid disappearance from the circulation in both animal groups. However, plasma clearance of taurocholate was significantly larger in analbuminemic (68.3 ml/min per kg of body weight) than in normal rats (29.8 ml/min per kg of body weight) at a dose of 8 mumol/kg of body weight. The increased plasma clearance in analbuminemic rats was accompanied by a more prompt biliary secretion of the ligand than occurred in normal animals; 79 and 42% of the injected dose was recovered in analbuminemic and normal rat bile, respectively, within 10 min after administration. Ultrafiltration analysis revealed that the binding of taurocholate to serum protein(s) was significantly lower in analbuminemic rats as compared with that in normal rat serum; 24 and 76% of taurocholate bound to protein fractions of analbuminemic and normal rat serum, respectively, at 0.5-mM ligand concentration. Binding of taurocholate to cytosolic proteins of normal and analbuminemic liver were similar; 23 and 28% of taurocholate bound to protein fractions from analbuminemic and normal rat, respectively, at 10 mg protein/ml and 20-microM ligand concentration. These results indicate that plasma albumin does not play a role in directing circulating taurocholate to the liver and that transhepatic transport of the bile acid increases with the increase in concentration of unbound ligand in the circulation.
Assuntos
Fígado/metabolismo , Albumina Sérica/deficiência , Ácido Taurocólico/metabolismo , Animais , Bile/metabolismo , Proteínas Sanguíneas/metabolismo , Proteínas de Transporte/metabolismo , Citosol/metabolismo , Proteínas de Ligação a Ácido Graxo , Glutationa Transferase/metabolismo , Masculino , Taxa de Depuração Metabólica , Ratos , Ratos EndogâmicosRESUMO
Renal transtubular transport of phenolsulfophthalein (PSP), a nephrophilic organic anion that circulates bound to albumin, was studied in normal and bile-duct-ligated rats. Intravenously injected PSP disappeared from the circulation more rapidly in bile-duct-ligated jaundiced rats than in intact animals. However, urinary excretion of PSP was significantly lower in the former than in the latter. Kinetic analysis revealed that binding of PSP to plasma protein(s) was significantly lower with jaundiced rats than with intact animals. Addition of albumin to plasma samples from bile-duct-ligated rats markedly increased PSP binding. The decreased PSP binding returned to normal levels after treating the jaundiced plasma with bilirubin oxidase, an enzyme that degrades amphiphilic bilirubin to water soluble metabolites. These results suggest that bilirubin might be the major metabolite that occupied the PSP binding site(s) on albumin in jaundiced rats. When PSP was injected bound to equimolar amount of albumin, the rate of PSP disappearance from the circulation decreased and urinary excretion of the ligand increased markedly; urinary excretion of PSP was significantly larger in bile-duct-ligated rats than in intact animals. These results suggest that the renal transport capacity for amphiphilic organic anions, such as PSP, might be increased compensatively in bile-duct-ligated animals, and that the apparent decrease in renal secretory transport for PSP might result from, at least in part, random distribution of the ligand to extrarenal tissues due to decrease in the binding activity of albumin.
Assuntos
Colestase/metabolismo , Túbulos Renais/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH , Fenolftaleínas/metabolismo , Fenolsulfonaftaleína/metabolismo , Animais , Transporte Biológico , Proteínas Sanguíneas/metabolismo , Masculino , Oxirredutases/metabolismo , Ratos , Ratos Endogâmicos , Albumina Sérica/metabolismoRESUMO
Although the possible involvement of superoxide radical and its metabolite(s) in the pathogenesis of various types of edema have been suggested, direct evidence supporting this concept is lacking. Since intravenously administered Cu2+Zn2(+)-type superoxide dismutase (SOD) rapidly disappeared from the circulation with a half-life of 4 min, the enzyme could not be used to test whether superoxide radicals played a critical role in the modulation of vascular permeability. We previously synthesized a SOD derivative (SM-SOD) by linking poly(styrene co-maleic acid butyl ester) (SM) to the enzyme (Ogino, T., Inoue, M., Ando, Y., Awai, M., Maeda, H. and Morino Y. (1988) Int. J. Pept. Protein Res. 32, 1583-1588); SM-SOD circulates bound to albumin with a half-life of 6 h. To test whether superoxide radicals play an important role in the regulation of vascular permeability, the effect of SM-SOD on experimental paw edema was studied in the rat. Subcutaneous injections of carrageenin to the paw rapidly induced local edema by increasing vascular permeability. Intravenous administration of SM-SOD markedly inhibited the carrageenin-induced increase in vascular permeability and suppressed the development of paw edema. In contrast, the same dose of SOD showed no such inhibitory effect. These results suggest that superoxide radical and/or its metabolite(s) might play a critical role in the pathogenesis of carrageenin-induced vasogenic edema.
Assuntos
Carragenina , Edema/prevenção & controle , Maleatos , Poliestirenos , Poliestirenos/farmacologia , Superóxido Dismutase/farmacologia , Animais , Permeabilidade Capilar/efeitos dos fármacos , Edema/induzido quimicamente , Edema/patologia , Azul Evans/metabolismo , Radicais Livres , Masculino , Neutrófilos/patologia , Poliestirenos/metabolismo , Poliestirenos/farmacocinética , Ratos , Ratos Endogâmicos , Albumina Sérica/metabolismo , Superóxido Dismutase/metabolismo , Superóxido Dismutase/farmacocinética , Superóxidos/metabolismoRESUMO
Long-term incubation of proteins with glucose undergo a series of nonenzymatic reactions to form advanced glycosylation end product (AGE) with fluorescence and brown color. The receptor for AGE-proteins was demonstrated in murine macrophages (Vlassara et al. (1985) Proc. Natl. Acad. Sci. USA 82. 5588). Our recent study with rat macrophages revealed that the receptor also recognized proteins modified with aliphatic aldehydes such as formaldehyde or glycolaldehyde, indicating its close identity to a scavenger receptor for aldehyde-modified proteins (Takata, K. et al. (1988) J. Biol. Chem. 263. 14819). This notion was tested in the present study with human monocytic leukemia cell line (THP-1 cells), human monocyte macrophages and murine peritoneal macrophages. Endocytic uptake of AGE-proteins and aldehyde-modified proteins was inhibited in a cross-competitive fashion. The receptor activities of THP-1 cells for AGE-albumin and aldehyde-modified proteins were induced synchronously by phorbol 12-myristate 13-acetate. Furthermore, upon reduction by NaBH4 of the Schiff base formed between proteins and glucose or aldehydes, no ligand activity was generated. However, once the ligand activity was generated, NaBH4 was no longer effective for the ligand activity. Thus, a structure in common between AGE-proteins and aldehyde-modified proteins may be crucial for recognition by the human macrophage receptor.
Assuntos
Proteínas Sanguíneas/metabolismo , Endocitose , Glicoproteínas , Proteínas de Membrana , Receptores Imunológicos/metabolismo , Receptores de Lipoproteínas , Células Tumorais Cultivadas/metabolismo , Animais , Linhagem Celular , Glicosilação , Humanos , Cinética , Leucemia , Camundongos , Receptores Depuradores , Receptores Depuradores Classe B , Acetato de Tetradecanoilforbol/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacos , Proteínas Séricas GlicadasRESUMO
Rat liver glutathione S-transferases (RX: glutathione R-transferase, EC 2.5.1.18) were found to adsorb S-carbamidomethyl glutathione linked to Sepharose CL-4B via lysyl or aliphatic diamine spacers of various carbon chain lengths (-NH-(CH2)n-NH-, n = 2, 4, 5, 6, 8 and 10). Proteins were eluted specifically by reduced glutathione. The affinity of the enzymes for the adsorbent increased with increase in the carbon chain length of aliphatic diamine spacers used. Adsorbent having a free carboxyl group within the spacer moiety had high capacity and was specific for glutathione S-transferases. The transferases were specifically eluted from the column in high yield by low concentrations of glutathione. Enzymes purified by the lysyl spacer adsorbent were homogeneous in sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and contained most of the hepatic glutathione S-transferase isozymes in isoelectric focusing. Oxidized glutathione and S-methyl glutathione were equally effective as reduced glutathione in eluting glutathione S-transferases from the adsorbent, but gamma-glutamylcysteinylglycineamide or gamma-glutamylcysteinylglycine-1-methyl ester were not effective. These data suggested that the free carboxyl group of glycyl moiety of glutathione might also be important for the specific binding of the transferases to this adsorbent.
Assuntos
Glutationa Transferase/isolamento & purificação , Fígado/enzimologia , Polissacarídeos/metabolismo , Sefarose/metabolismo , Animais , Sítios de Ligação , Cromatografia de Afinidade/métodos , Eletroforese em Gel de Poliacrilamida , Glutationa/análogos & derivados , Isoenzimas/isolamento & purificação , Masculino , Ratos , Sefarose/análogos & derivadosRESUMO
To investigate the post-binding events of high-density lipoprotein (HDL), rat peritoneal macrophages were enriched by collagen gel-coated plates and incubated in a cell-suspension system with fluorescein isothiocyanate-labeled HDL (FITC-HDL), followed by fluorescence spectroscopic analyses. Upon incubation with FITC-HDL at 37 degrees C for 30 min, the microenvironmental pH of the cell-associated FITC-HDL was 6.50, whereas a 0 degree C-incubation gave a corresponding pH of 7.15. Carbonyl cyanide m-chlorophenylhydrazone, a protonophore known to dissipate the proton gradient, restored the former acidic pH (pH 6.40) to pH 7.20, but had no effect on the latter. This indicates that cell surface-bound HDL is internalized and exposed to an acidic compartment. When cells were incubated with FITC-HDL at 37 degrees C for 30 min and the cell-associated FITC-HDL was chased at 37 degrees C, the fluorescence intensity at 490 nm showed a time-dependent increase. This increase was explained by a release of endocytosed FITC-HDL into the extracellular medium but not by a simple outward dissociation of the cell-associated FITC-HDL. The microenvironmental acidic pH of the cell-associated FITC-HDL changed to a less acidic pH during the chase whereas that of FITC-HDL became constant after released into the medium, indicating that endocytosed HDL was resected back into the extracellular medium. This resection process was temperature-dependent and accelerated by HDL itself. These results provide the biochemical evidence for the presence of an endocytic-exocytic pathway for HDL in rat peritoneal macrophages.
Assuntos
Endocitose , Lipoproteínas HDL/metabolismo , Macrófagos/fisiologia , Peritônio , Animais , Fluoresceína-5-Isotiocianato , Fluoresceínas , Corantes Fluorescentes , Concentração de Íons de Hidrogênio , Macrófagos/metabolismo , Ratos , Análise Espectral , Temperatura , Tiocianatos , Fatores de TempoRESUMO
Rat peritoneal macrophages possess a surface receptor for high-density lipoprotein (HDL). To obtain the functional aspect of the HDL receptor, the present study was undertaken to modify HDL with three different cross-linkers; dimethylsuberimidate, disuccinimidylsuberate and dithiobissuccinimidylpropionate (DSP) and determine their effect on the ligand activity for the HDL receptor. Upon modification at a low reagent concentration, DSP was found to be most effective in cross-linking of HDL apolipoproteins. The ligand activity of DSP-HDL for the HDL receptor was reduced by greater than 60%. Experiments with these macrophages at 37 degrees C showed; (i) the amounts of the cell-associated [125I]DSP-HDL as 3.5-fold higher than [125I]HDL; (ii) the cell-association of [125I]DSP-HDL was effectively (greater than 70%) inhibited by unlabeled DSP-HDL, whereas HDL showed a partial inhibition (30%); (iii) [125I]DSP-HDL underwent chloroquine-sensitive intracellular degradation; and (iv) DSP-HDL induced a 3-fold increase in the incorporation of [14C]oleic acid into cholesteryl oleate when compared with unmodified HDL. Experiments at 0 degrees C showed that the cellular binding of [125I]DSP-HDL was competed by acetylated low-density lipoprotein and dextran sulfate. These findings indicate that DSP-HDL is recognized as a ligand by a scavenger receptor of rat peritoneal macrophages, a notion consistent with HDL modified with tetranitromethane (Kleinherenbrink-Stins, M.F. et al. (1989) J. Lipid Res. 39, 511-520).
Assuntos
Proteínas de Transporte , Lipoproteínas HDL/química , Macrófagos/metabolismo , Proteínas de Ligação a RNA , Receptores de Superfície Celular/química , Receptores de Lipoproteínas , Animais , Cloroquina/farmacologia , Reagentes de Ligações Cruzadas , Eletroforese , Ligantes , Lipoproteínas HDL/efeitos dos fármacos , Lipoproteínas HDL/metabolismo , Peritônio , Ratos , Receptores de Superfície Celular/metabolismoRESUMO
Alterations in the molecular form of gamma-glutamyl transferase (5-glutamyl)-peptide:amino-acid 5-glutamyltransferase, EC 2.3.2.2.) were studied in the bile and serum of rats under surgical ligation of the bile duct. Polyacrylamide gel electrophoresis of the bile, followed by the enzyme stain, revealed a major, slowly migrating broad band and a minor, faster migrating band. The former was converted to the latter upon limited proteolysis of the bile with a very small amount of papain. This conversion was accompanied by a decrease in molecular size of the enzyme. Both enzyme forms were specifically adsorbed to a concanavalin A-Sepharose column. Most of the papain-treated enzyme preparation could be eluted from the column by alpha-methyl-D-glucoside, a haptenic sugar of this lectin. On the other hand, the predominant form of the enzyme in the untreated bile was eluted only in the presence both of the sugar and Triton X-100. Based on the chromatographic behavior of the two enzyme forms (detergent-solubilized and protease-solubilized form) purified from rat renal brush border membrane on concanavalin A-Sepharose column, it was concluded that the predominant form of the enzyme in the bile was the detergent-solubilized form and that the minor component represents the protease-solubilized enzyme. The serum from icteric rats was also found to contain both types of the enzyme. However, the relative amount of the protease-solubilized form to the detergent-solubilized form in the serum was much greater than that in the bile. These findings suggested that gamma-glutamyl transferase in the hepatobiliary membrane systems was solubilized into the bile mainly as the detergent-solubilized form, and that, during the process of translocation into the blood circulation, the enzyme was partly converted to the protease-solubilized form by some protease-like action.
Assuntos
Bile/enzimologia , Icterícia/enzimologia , Rim/enzimologia , gama-Glutamiltransferase/análise , Animais , Cromatografia em Gel , Masculino , Papaína , Polietilenoglicóis , Ratos , Solubilidade , gama-Glutamiltransferase/sangueRESUMO
To elucidate an anti-atherogenic nature of high density lipoprotein (HDL) at cellular level, its in vitro effect on macrophage foam cells was examined. Rat peritoneal macrophages were converted to foam cells by incubation with [3H]cholesterol-labeled acetylated LDL. HDL addition to these foam cells resulted in a reduction in cellular radioactive cholesteryl esters (CE) as well as its CE mass. The radioactive free cholesterol (FC) was similarly reduced with time, whereas its FC mass level was unaltered. Other lipoproteins such as very low density lipoprotein and low density lipoprotein also reduced the radioactive FC. However, their CE-reducing capacity was negligibly weak. These results suggest that (i) CE reduction is selective to HDL, (ii) FC transfer from plasma membrane to lipoprotein (cholesterol efflux) expressed by reduction in radioactive FC is not selective to HDL but occurs to other lipoproteins, (iii) the CE-reducing capacity of HDL became weaker when cellular binding of HDL was reduced by chemical modification with tetranitromethane or a chemical cross-linker, dithiobis-succinimidylpropionate, suggesting an importance of the specific binding in the HDL-mediated CE reduction. These in vitro results gave an experimental support to a definite role of HDL as an anti-atherogenic lipoprotein in vivo.
Assuntos
Ésteres do Colesterol/metabolismo , Células Espumosas/metabolismo , Lipoproteínas HDL/farmacologia , Animais , Colesterol/metabolismo , LDL-Colesterol/farmacologia , Lipoproteínas HDL/química , Masculino , Oxirredução , Ratos , Ratos Endogâmicos , Succinimidas , TetranitrometanoRESUMO
OBJECTIVES: The study was done to elucidate the relationship between baseline arterial remodeling and clinical outcome following stenting. BACKGROUND: The impact of preintervention arterial remodeling on subsequent vessel response and clinical outcome has been reported following nonstent coronary interventions. However, in stented segments, the impact of preintervention remodeling on clinical outcome has not been clarified. METHODS: Preintervention remodeling was assessed in 108 native coronary lesions by using intravascular ultrasound (IVUS). Positive remodeling (PR) was defined as vessel area (VA) at the target lesion greater than that of average reference segments. Intermediate or negative remodeling (IR/NR) was defined as VA at the target lesion less than or equal to that of average reference segment. Remodeling index expressed as a continuous variable was defined as VA at the target lesion site divided by that of average reference segments. RESULTS: Positive remodeling was present in 59 (55%) and IR/NR in 49 (45%) lesions. Although final minimal stent areas were similar (7.76 +/- 1.80 vs. 8.09 +/- 1.90 mm2, p = 0.36), target vessel revascularization (TVR) rate at nine-month follow-up was significantly higher in the PR group (22.0% vs. 4.1%, p = 0.01). By multivariate logistic regression analysis, higher remodeling index was the only independent predictor of TVR (p = 0.02). CONCLUSIONS: Lesions with PR before intervention appear to have a worse clinical outcome following IVUS-guided stenting. Intravascular ultrasound imaging before stenting may be helpful to stratify lesions at high risk for accelerated intimal proliferation.