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1.
Nat Genet ; 25(3): 302-5, 2000 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-10888878

RESUMEN

Autosomal recessive juvenile parkinsonism (AR-JP), one of the most common familial forms of Parkinson disease, is characterized by selective dopaminergic neural cell death and the absence of the Lewy body, a cytoplasmic inclusion body consisting of aggregates of abnormally accumulated proteins. We previously cloned PARK2, mutations of which cause AR-JP (ref. 2), but the function of the gene product, parkin, remains unknown. We report here that parkin is involved in protein degradation as a ubiquitin-protein ligase collaborating with the ubiquitin-conjugating enzyme UbcH7, and that mutant parkins from AR-JP patients show loss of the ubiquitin-protein ligase activity. Our findings indicate that accumulation of proteins that have yet to be identified causes a selective neural cell death without formation of Lewy bodies. Our findings should enhance the exploration of the molecular mechanisms of neurodegeneration in Parkinson disease as well as in other neurodegenerative diseases that are characterized by involvement of abnormal protein ubiquitination, including Alzheimer disease, other tauopathies, CAG triplet repeat disorders and amyotrophic lateral sclerosis.


Asunto(s)
Ligasas/metabolismo , Enfermedad de Parkinson/metabolismo , Enzimas Ubiquitina-Conjugadoras , Humanos , Ligasas/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Células Tumorales Cultivadas , Ubiquitina-Proteína Ligasas , Ubiquitinas/genética , Ubiquitinas/metabolismo
2.
Nat Genet ; 27(2): 209-14, 2001 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-11175792

RESUMEN

In mammalian cells, regulation of the expression of proteins involved in iron metabolism is achieved through interactions of iron-sensing proteins known as iron regulatory proteins (IRPs), with transcripts that contain RNA stem-loop structures referred to as iron responsive elements (IREs). Two distinct but highly homologous proteins, IRP1 and IRP2, bind IREs with high affinity when cells are depleted of iron, inhibiting translation of some transcripts, such as ferritin, or turnover of others, such as the transferrin receptor (TFRC). IRPs sense cytosolic iron levels and modify expression of proteins involved in iron uptake, export and sequestration according to the needs of individual cells. Here we generate mice with a targeted disruption of the gene encoding Irp2 (Ireb2). These mutant mice misregulate iron metabolism in the intestinal mucosa and the central nervous system. In adulthood, Ireb2(-/-) mice develop a movement disorder characterized by ataxia, bradykinesia and tremor. Significant accumulations of iron in white matter tracts and nuclei throughout the brain precede the onset of neurodegeneration and movement disorder symptoms by many months. Ferric iron accumulates in the cytosol of neurons and oligodendrocytes in distinctive regions of the brain. Abnormal accumulations of ferritin colocalize with iron accumulations in populations of neurons that degenerate, and iron-laden oligodendrocytes accumulate ubiquitin-positive inclusions. Thus, misregulation of iron metabolism leads to neurodegenerative disease in Ireb2(-/-) mice and may contribute to the pathogenesis of comparable human neurodegenerative diseases.


Asunto(s)
Proteínas Hierro-Azufre/genética , Hierro/metabolismo , Trastornos del Movimiento/genética , Enfermedades Neurodegenerativas/genética , Proteínas de Unión al ARN/genética , Animales , Cerebelo/patología , Duodeno/patología , Ferritinas/metabolismo , Eliminación de Gen , Mucosa Intestinal/patología , Proteína 1 Reguladora de Hierro , Proteína 2 Reguladora de Hierro , Proteínas Reguladoras del Hierro , Ratones , Ratones Mutantes , Datos de Secuencia Molecular , Neuronas/patología , Oligodendroglía/patología , Células de Purkinje/patología , Putamen/patología , Elementos de Respuesta , Tálamo/patología , Ubiquitinas/metabolismo
3.
Br J Sports Med ; 44(12): 856-61, 2010 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-19042919

RESUMEN

BACKGROUND: Although high prevalence of anterior cruciate ligament injuries (ACL) in judokas has been reported, there has been very little research concerning events preceding the injury. OBJECTIVE: To determine the common situations and mechanisms of ACL injury in judo. METHODS: A total of 43 cases of ACL injuries that had occurred during judo competition or practice were investigated, using questionnaires with interviews conducted by a single certified athletic trainer who has 20 years of judo experience to obtain information regarding the situation and mechanism in which the ACL injury occurred. RESULTS: The number of ACL injuries when the participant's grip style was different from the style of the opponent (ie, kenka-yotsu style) (28 cases) was significantly greater than when the participant's grip style was the same as that of the opponent (ie, ai-yotsu style) (15 cases; p<0.001). The number of ACL injuries was significantly higher when the participant was attacked by the opponent than when counterattacked or when attempting the attack (p<0.001). In addition, being attacked with osoto-gari was revealed as the leading cause of ACL injury incidence among the participants (16.8%). CONCLUSIONS: Grip style may be associated with ACL injury occurrence in judo. In addition, direct contact due to the opponent's attack may be a common mechanism for ACL injuries in judo.


Asunto(s)
Lesiones del Ligamento Cruzado Anterior , Artes Marciales/lesiones , Adolescente , Femenino , Humanos , Masculino , Equilibrio Postural/fisiología , Postura/fisiología , Estudios Retrospectivos , Adulto Joven
4.
Science ; 243(4889): 375-8, 1989 Jan 20.
Artículo en Inglés | MEDLINE | ID: mdl-2911748

RESUMEN

DNA and nuclear proteins were transferred into cells simultaneously at more than 95% efficiency by means of vesicle complexes. The DNA was rapidly transported into the nuclei of cultured cells, and its expression reached a maximum within 6 to 8 hours after its introduction. Moreover, when the plasmid DNA and nuclear protein were cointroduced into nondividing cells in rat liver by injection into the portal veins of adult rats, the plasmid DNA was carried into liver cell nuclei efficiently by nuclear protein. The expression of the DNA in adult rat liver, on introduction of the DNA with nuclear protein, was more than five times as great as with nonnuclear protein.


Asunto(s)
ADN/metabolismo , Proteínas del Grupo de Alta Movilidad/metabolismo , Hígado/metabolismo , Animales , Northern Blotting , Compartimento Celular , Núcleo Celular/metabolismo , Células Cultivadas , ADN/farmacocinética , Ratones , Ratas , Transformación Genética
5.
Radiat Prot Dosimetry ; 128(3): 343-50, 2008.
Artículo en Inglés | MEDLINE | ID: mdl-18337296

RESUMEN

Surface dose monitoring in patients and physicians during 29 uterine artery embolisation (UAE) procedures was performed using photoluminescence dosemeters and thermo-luminescence dosemeters. Organ or tissue doses were measured with an anthropomorphic phantom using UAE exposure conditions averaged from the 29 cases, and effective doses were estimated for the patient. Entrance surface dose of the patients at the maximum dose position ranged from 121.5 to 1650 mGy. Estimated doses ranged from 3.16 to 43 mGy for the ovary and from 3.8 to 51.8 mGy for the uterus. The effective dose was 1.09-14.8 mSv. Monitored doses on the body surface of physicians were relatively high in the upper arm (5.41+/-1.52 to 163+/-17.25 microGy) and the hand and fingers (0.85+/-1.18 to 222+/-16.4 microGy).


Asunto(s)
Embolización Terapéutica , Fluoroscopía , Exposición Profesional , Ovario/efectos de la radiación , Médicos , Radiografía Intervencional , Dosimetría Termoluminiscente , Útero/irrigación sanguínea , Arterias , Femenino , Humanos , Dosis de Radiación , Útero/efectos de la radiación
6.
Mol Cell Biol ; 15(1): 552-60, 1995 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-7799964

RESUMEN

We have cloned a novel cDNA (Spa-1) which is little expressed in the quiescent state but induced in the interleukin 2-stimulated cycling state of an interleukin 2-responsive murine lymphoid cell line by differential hybridization. Spa-1 mRNA (3.5 kb) was induced in normal lymphocytes following various types of mitogenic stimulation. In normal organs it is preferentially expressed in both fetal and adult lymphohematopoietic tissues. A Spa-1-encoded protein of 68 kDa is localized mostly in the nucleus. Its N-terminal domain is highly homologous to a human Rap1 GTPase-activating protein (GAP), and a fusion protein of this domain (SpanN) indeed exhibited GAP activity for Rap1/Rsr1 but not for Ras or Rho in vitro. Unlike the human Rap1 GAP, however, SpanN also exhibited GAP activity for Ran, so far the only known Ras-related GTPase in the nucleus. In the presence of serum, stable Spa-1 cDNA transfectants of NIH 3T3 cells (NIH/Spa-1) hardly overexpressed Spa-1 (p68), and they grew as normally as did the parental cells. When NIH/Spa-1 cells were serum starved to be arrested in the G1/G0 phase of the cell cycle, however, they, unlike the control cells, exhibited progressive Spa-1 p68 accumulation, and following the addition of serum they showed cell death resembling mitotic catastrophes of the S phase during cell cycle progression. The results indicate that the novel nuclear protein Spa-1, with a potentially active Ran GAP domain, severely hampers the mitogen-induced cell cycle progression when abnormally and/or prematurely expressed. Functions of the Spa-1 protein and its regulation are discussed in the context of its possible interaction with the Ran/RCC-1 system, which is involved in the coordinated nuclear functions, including cell division.


Asunto(s)
Ciclo Celular , Proteínas Activadoras de GTPasa , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Células 3T3 , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Cartilla de ADN/química , Activación Enzimática , Proteínas de Unión al GTP/metabolismo , Expresión Génica , Linfocitos/citología , Linfocitos/metabolismo , Ratones , Mitógenos/farmacología , Datos de Secuencia Molecular , Proteínas Nucleares/fisiología , ARN Mensajero/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Proteína de Unión al GTP ran , Proteínas de Unión al GTP rap
7.
Mol Cell Biol ; 21(4): 1329-35, 2001 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-11158318

RESUMEN

Although chromosomal segregation at meiosis I is the critical process for genetic reassortment and inheritance, little is known about molecules involved in this process in metazoa. Here we show by utilizing double-stranded RNA (dsRNA)-mediated genetic interference that novel protein kinases (Ce-CDS-1 and Ce-CDS-2) related to Cds1 (Chk2) play an essential role in meiotic recombination in Caenorhabditis elegans. Injection of dsRNA into adult animals resulted in the inhibition of meiotic crossing over and induced the loss of chiasmata at diakinesis in oocytes of F(1) animals. However, electron microscopic analysis revealed that synaptonemal complex formation in pachytene nuclei of the same progeny of injected animals appeared to be normal. Thus, Ce-CDS-1 and Ce-CDS-2 are the first example of Cds1-related kinases that are required for meiotic recombination in multicellular organisms.


Asunto(s)
Caenorhabditis elegans/enzimología , Caenorhabditis elegans/genética , Meiosis/genética , Meiosis/fisiología , Proteínas Quinasas/genética , Proteínas Quinasas/fisiología , Proteínas Serina-Treonina Quinasas , Recombinación Genética , Secuencia de Aminoácidos , Aneuploidia , Animales , Secuencia de Bases , Quinasa de Punto de Control 2 , Cartilla de ADN/genética , ADN de Helmintos/genética , Femenino , Genes de Helminto , Masculino , Datos de Secuencia Molecular , Fenotipo , ARN Bicatenario/genética , ARN de Helminto/genética , Homología de Secuencia de Aminoácido
8.
J Sports Med Phys Fitness ; 46(3): 425-30, 2006 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-16998447

RESUMEN

AIM: The purpose of this study was to investigate the relationship between the influence of music on RPE during sub-maximal exercise and on the autonomic nervous system before and after sub-maximal exercise. METHODS: Heart rate (HR), HR variability (HRV) and rates of physical fatigue (RPE) during exercise at 60% and at 40% VO2max with and without music were measured. The exercise protocol consisted of a 30-min seated rest (control) period followed by a 30-min submaximal cycling exercise and a 35-min recovery period. Autonomic-nervous activity was measured before and after exercise. During exercise, RPE was recorded every 3 min and HR was recorded for every minute. RESULTS: Although RPE did not differ during exercise at 60% VO2max, this value was lower during exercise at 40% VO2max in the presence, than in the absence of a favorite piece music (P < 0.05). HR, HFA and LFA/HFA of HRV significantly differed with exercise intensity in the absence (P < 0.05), but not in the presence of music. CONCLUSIONS: These findings suggested that music evokes a ''distraction effect'' during low intensity exercise, but might not influence the autonomic nervous system. Therefore, when jogging or walking at comparatively low exercise intensity, listening to a favorite piece of music might decrease the influence of stress caused by fatigue, thus increasing the ''comfort'' level of performing the exercise.


Asunto(s)
Sistema Nervioso Autónomo/fisiología , Ejercicio Físico/fisiología , Frecuencia Cardíaca/fisiología , Fatiga Muscular/fisiología , Música , Esfuerzo Físico/fisiología , Adulto , Análisis de Varianza , Prueba de Esfuerzo , Tolerancia al Ejercicio/fisiología , Humanos , Masculino , Consumo de Oxígeno/fisiología , Valores de Referencia
9.
Cancer Res ; 44(5): 2115-21, 1984 May.
Artículo en Inglés | MEDLINE | ID: mdl-6324996

RESUMEN

Highly malignant rabbit tumor (VX-2) was implanted at the periphery of the liver in 63 rabbits. Selective delivery of the anticancer agent copoly(styrenemaleic acid) conjugated to neocarzinostatin (SMANCS), which was dissolved in an oil contrast medium (Lipiodol), was performed by injection via the proper hepatic artery. The anticancer effect was also evaluated by various parameters. By using low-kVp X-ray examination of the resected rabbit liver, Lipiodol was found to distribute throughout the entire liver arterial lumina and was retained there for about 24 hr, but disappeared from the normal liver arterial lumina gradually. However, Lipiodol was retained in the tumor tissue and vessels for at least 7 days, whereas it was undetectable in any other organs. A radioactive analogue of Lipiodol, a chloroiodinated fatty acid, was prepared by using [14C]linoleic acid. This analogue was used in the study of the distribution by low-kVp X-ray examination, Sudan III staining, and autoradiography. Lipiodol remained in the tumor vessels as well as the tumor cells. The use of the radioisotope yielded a quantitative profile of Lipiodol accumulation in tumor tissues; approximately 1000 times more at 15 min and 100 times more at 3 days after the injection than that of most other organs or plasma. Its major excretion route appeared to be through the bile and then the feces. The biological activity of SMANCS was also determined and was found to be significant in both tumor and liver even 7 days after injection. No activity was found in any other organ or tissue. The relatively high biological activity of SMANCS in the nontumorous liver adjacent to the tumor may be the result of continuous drug release from SMANCS-Lipiodol in the tumor tissue. By histological examination, massive tumor necrosis and infiltration of the inflammatory cells were found in the rabbits treated with SMANCS-Lipiodol. In the rabbits treated with Lipiodol alone, necrosis of the tumor was only minimal, and no infiltration of inflammatory cells was observed. Survival periods of the treated rabbits (n = 14) were significantly longer than those of controls (n = 10); 23.1 +/- 5.5 (S.D.) days versus 16.1 +/- 2.9 days (p less than 0.005), respectively, even though only one injection was used for this highly malignant tumor. Mean tumor size for both groups at laparotomy was 163.3 +/- 83.0 sq mm and 160.5 +/- 76.5 sq mm, respectively (not significant).(ABSTRACT TRUNCATED AT 400 WORDS)


Asunto(s)
Antibióticos Antineoplásicos/administración & dosificación , Furanos/administración & dosificación , Aceite Yodado/administración & dosificación , Anhídridos Maleicos/administración & dosificación , Neoplasias Experimentales/tratamiento farmacológico , Poliestirenos/administración & dosificación , Cinostatina/administración & dosificación , Animales , Autorradiografía , Radioisótopos de Carbono , Aceite Yodado/metabolismo , Cinética , Hígado/efectos de los fármacos , Hígado/patología , Anhídridos Maleicos/uso terapéutico , Neoplasias Experimentales/diagnóstico por imagen , Poliestirenos/uso terapéutico , Conejos , Cintigrafía , Distribución Tisular , Cinostatina/análogos & derivados , Cinostatina/uso terapéutico
10.
Lab Chip ; 16(4): 668-78, 2016 Feb 21.
Artículo en Inglés | MEDLINE | ID: mdl-26725379

RESUMEN

The miniaturization of integrated fluidic processors affords extensive benefits for chemical and biological fields, yet traditional, monolithic methods of microfabrication present numerous obstacles for the scaling of fluidic operators. Recently, researchers have investigated the use of additive manufacturing or "three-dimensional (3D) printing" technologies - predominantly stereolithography - as a promising alternative for the construction of submillimeter-scale fluidic components. One challenge, however, is that current stereolithography methods lack the ability to simultaneously print sacrificial support materials, which limits the geometric versatility of such approaches. In this work, we investigate the use of multijet modelling (alternatively, polyjet printing) - a layer-by-layer, multi-material inkjetting process - for 3D printing geometrically complex, yet functionally advantageous fluidic components comprised of both static and dynamic physical elements. We examine a fundamental class of 3D printed microfluidic operators, including fluidic capacitors, fluidic diodes, and fluidic transistors. In addition, we evaluate the potential to advance on-chip automation of integrated fluidic systems via geometric modification of component parameters. Theoretical and experimental results for 3D fluidic capacitors demonstrated that transitioning from planar to non-planar diaphragm architectures improved component performance. Flow rectification experiments for 3D printed fluidic diodes revealed a diodicity of 80.6 ± 1.8. Geometry-based gain enhancement for 3D printed fluidic transistors yielded pressure gain of 3.01 ± 0.78. Consistent with additional additive manufacturing methodologies, the use of digitally-transferrable 3D models of fluidic components combined with commercially-available 3D printers could extend the fluidic routing capabilities presented here to researchers in fields beyond the core engineering community.

11.
Oncogene ; 19(16): 1992-2001, 2000 Apr 13.
Artículo en Inglés | MEDLINE | ID: mdl-10803460

RESUMEN

Beta-catenin and plakoglobin are closely related armadillo family proteins with shared and distinct properties; Both are associated with cadherins in actin-containing adherens junctions. Plakoglobin is also found in desmosomes where it anchors intermediate filaments to the desmosomal plaques. Beta-catenin, on the other hand, is a component of the Wnt signaling pathway, which is involved in embryonic morphogenesis and tumorigenesis. A key step in the regulation of this pathway involves modulation of beta-catenin stability. A multiprotein complex, regulated by Wnt, directs the phosphorylation of beta-catenin and its degradation by the ubiquitin-proteasome system. Plakoglobin can also associate with members of this complex, but inhibition of proteasomal degradation has little effect on its levels while dramatically increasing the levels of beta-catenin. Beta-TrCP, an F-box protein of the SCF E3 ubiquitin ligase complex, was recently shown to play a role in the turnover of beta-catenin. To elucidate the basis for the apparent differences in the turnover of beta-catenin and plakoglobin we compared the handling of these two proteins by the ubiquitin-proteasome system. We show here that a deletion mutant of beta-TrCP, lacking the F-box, can stabilize the endogenous beta-catenin leading to its nuclear translocation and induction of beta-catenin/LEF-1-directed transcription, without affecting the levels of plakoglobin. However, when plakoglobin was overexpressed, it readily associated with beta-TrCP, efficiently competed with beta-catenin for binding to beta-TrCP and became polyubiquitinated. Fractionation studies revealed that about 85% of plakoglobin in 293 cells, is Triton X-100-insoluble compared to 50% of beta-catenin. These results suggest that while both plakoglobin and beta-catenin can comparably interact with beta-TrCP and the ubiquitination system, the sequestration of plakoglobin by the membrane-cytoskeleton system renders it inaccessible to the proteolytic machinery and stabilizes it.


Asunto(s)
Cisteína Endopeptidasas/metabolismo , Proteínas del Citoesqueleto/metabolismo , Complejos Multienzimáticos/metabolismo , Transactivadores , Ubiquitinas/metabolismo , Animales , Transporte Biológico , Células CHO/metabolismo , Compartimento Celular , Cricetinae , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Desmoplaquinas , Dexametasona/farmacología , Proteínas de Unión al GTP/genética , Proteínas de Unión al GTP/metabolismo , Humanos , Factor de Unión 1 al Potenciador Linfoide , Octoxinol/química , Complejo de la Endopetidasa Proteasomal , Proteínas Recombinantes/efectos de los fármacos , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , beta Catenina , Proteínas con Repetición de beta-Transducina , gamma Catenina
12.
Biochim Biophys Acta ; 422(1): 29-37, 1976 Jan 23.
Artículo en Inglés | MEDLINE | ID: mdl-1247595

RESUMEN

Amino acid analysis and chemical modification of the crystalline quinolinate phosphoribosyltransferase (EC 2.4.2.19) from hog liver were performed. The enzyme contained 29 residues of half cystine per mol. The enzyme activity was strongly inhibited by sulfhydryl reagents. The number of reactive (exposed) sulfhydryl group was determined to be 10.2 and total sulfhydryl group was to be 25.2 per mol by using 5,5'-dithiobis(2-nitrobenzoic acid). The enzyme activity was also inhibited by lysine residue-, histidine residue-, and arginine residue-modifying reagents. These results and the effect of preincubation with the substrates on chemical modifications suggest that the lysine residue, histidine residue and sulfhydryl group may be closely related to the binding site of quinolinic acid.


Asunto(s)
Hígado/enzimología , Pentosiltransferasa/metabolismo , Aminoácidos/análisis , Animales , Sitios de Unión , Cinética , Oxidación-Reducción , Fotoquímica , Unión Proteica , Ácidos Quinolínicos/metabolismo , Reactivos de Sulfhidrilo/farmacología , Porcinos
13.
Biochim Biophys Acta ; 611(2): 280-8, 1980 Feb 14.
Artículo en Inglés | MEDLINE | ID: mdl-7357010

RESUMEN

Crystalline quinolinate phosphoribosyltransferase (nicotinatenucleotide: pyrophosphate phosphoribosyltransferase (carboxylating), EC 2.4.2.19) was isolated from hog kidney and compared with the same enzyme prepared from hog liver. The enzyme preparation was homogeneous as shown by polyacrylamide gel electrophoresis and ultracentrifugation analysis. The enzyme had a molecular weight of 220 000 and the subunit 35 000. The physicochemical properties of the enzyme were: sedimentation coefficient (S200,W), 7.75 . 10(-13) s; difussion coefficient (D200,W), 5.04 . 10(-7) cm2/s; Stokes radius, 62.05 A, frictional ratio (f/f0), 1.62 and isoelectric point (pI), 4.5. The enzyme was stable at 37 degrees C for 30 min between pH 4.5 and 9.5. Enzyme activity was inhibited by various carboxylic acids; however, this inhibition was reversed by raising the Mg2+ concentration. Optimum pH was 5.5, and no detectable amounts of Mg2+, Mn2+, Fe2+, Cu2+, Zn2+ and Ca2+ were found by atomic absorption spectrophotometry. The enzyme was found to contain sugar. Mg2+ was completely replaceable by Mn2+. The reaction mechanism of this enzyme was suggested to be of the 'ping-pong' type. Km values of quinolinic acid and 5-phosphoribosyl 1-pyrophosphate were 4 . 10(-5) and 1.4 . 10(-4) M, respectively.


Asunto(s)
Riñón/enzimología , Pentosiltransferasa/aislamiento & purificación , Aminoácidos/análisis , Animales , Cationes Bivalentes , Fenómenos Químicos , Química Física , Cristalización , Cinética , Pentosiltransferasa/metabolismo , Porcinos
14.
Biochim Biophys Acta ; 757(3): 274-81, 1983 Jun 09.
Artículo en Inglés | MEDLINE | ID: mdl-6849975

RESUMEN

The absorption of [14C]pteroylglutamic acid (PteGlu) bound to the folate-binding protein of bovine milk was investigated in rat gastrointestinal tract in vivo and in situ. When bound [14C]PteGlu was given to rat intragastrically via oral intubation, a considerable amount of PteGlu was released from folate-binding protein under the acidic conditions in stomach, and it recombined with folate-binding protein in jejunum in vivo. Compared with free PteGlu, bound PteGlu was more gradually absorbed in small intestine, but finally the total amount of bound PteGlu absorbed was almost the same as that of free PteGlu. In all experiments in situ, bound PteGlu was only slightly absorbed in jejunum, where free PteGlu was rapidly absorbed under the same conditions. On the other hand, the absorption rates of the two forms of PteGlu were almost similar to each other in ileum. These results suggest that PteGlu bound to folate-binding protein is absorbed by a manner different from that of free PteGlu in rat gastrointestinal tract.


Asunto(s)
Proteínas Portadoras/farmacología , Ácido Fólico/metabolismo , Absorción Intestinal/efectos de los fármacos , Proteínas de la Leche/farmacología , Receptores de Superficie Celular , Animales , Proteínas Portadoras/aislamiento & purificación , Bovinos , Receptores de Folato Anclados a GPI , Concentración de Iones de Hidrógeno , Técnicas In Vitro , Masculino , Proteínas de la Leche/aislamiento & purificación , Ratas , Ratas Endogámicas
15.
Biochim Biophys Acta ; 434(1): 189-98, 1976 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-938662

RESUMEN

A method was devised to isolate N-terminal peptide fragments from the polypeptide chains constituting thyroglobulin even in the case when the terminal amino groups are naturally blocked, for instance, acylated. Reduced and carboxymethylated hog thyroglobulin was first acetylated and digested with thermolysin. The blocked N-terminal peptide fragments were separated from the unblocked N-terminal fragments by column chromatography on Dowex 50, then on Dowex 1 after dinitrophenylation, and finally fractionated into ten fractions by paper chromatography after gel filtration on Sephadex G-10. Structural analyses by enzymic or partial acid hydrolysis of these peptide fractions failed to detect N-terminal acetyl amino acid. Instead, pyroglutamyl peptides including pyroglutamylleucine were found. By the same method, acetylated lysine and glycine were identified for chicken lysozyme and horse myoglobin, respectively. The use of thermolysin because of its unique specificity, and the possible relevance of the present result to the previous data on the N-terminal analysis of thyroglobulin are discussed.


Asunto(s)
Tiroglobulina , Secuencia de Aminoácidos , Aminoácidos/análisis , Animales , Sitios de Unión , Carboxipeptidasas , Dinitrofenoles , Glutamatos/análisis , Fragmentos de Péptidos/análisis , Unión Proteica , Porcinos , Termolisina
16.
Biochim Biophys Acta ; 251(2): 208-16, 1971 Nov 19.
Artículo en Inglés | MEDLINE | ID: mdl-11452386

RESUMEN

In order to identify the number and types of peptide chains in thyroglobulin, noniodinated 19-S thyroglobulin obtained from goitrogen-treated hogs was exhaustively digested with trypsin (EC 3.4.4.4) after reduction and S-carboxymethylation. The digestion mixture was preliminarily separated into 30 fractions on Sephadex G-100 or G-15 and SE-Sephadex columns. The number of various tryptic peptides contained in each fraction was determined on peptide maps, where spots were detected with ninhydrin for total peptides and with each specific reagent for arginine, histidine or tyrosine-containing peptides. The number of total peptides observed in most of the fractions was estimated to be half the number of lysine plus arginine residues found in each fraction per mole of thyroglobulin, and the number of specific peptides was also close to half the number of each specific amino acid. These findings imply that thyroglobulin has 2-fold symmetry in the structure at the level of tryptic fragments and thus probably at the level of intact peptide chains.


Asunto(s)
Fragmentos de Péptidos/química , Tiroglobulina/química , Aminoácidos/análisis , Animales , Bocio/metabolismo , Bocio/veterinaria , Fragmentos de Péptidos/aislamiento & purificación , Mapeo Peptídico , Porcinos , Enfermedades de los Porcinos/metabolismo , Tripsina/metabolismo
17.
Biochim Biophys Acta ; 444(2): 618-22, 1976 Sep 24.
Artículo en Inglés | MEDLINE | ID: mdl-971422

RESUMEN

2-Amino-4-hydroxypteridine compounds were isolated as the Crithidia factor from spinach chloroplasts by DEAE-Sephadex A-50, DEAE-cellulose, Sephadex G-25, and thin-layer chromatography. One of the compounds was characterized as 6-hydroxymethylpterin by gas-liquid chromatography-mass spectrometry and by comparison with authentic specimen.


Asunto(s)
Cloroplastos/análisis , Plantas , Pterinas/aislamiento & purificación , Cromatografía de Gases , Eucariontes/crecimiento & desarrollo , Espectrometría de Masas , Pterinas/análisis
18.
Biochim Biophys Acta ; 884(1): 18-24, 1986 Oct 29.
Artículo en Inglés | MEDLINE | ID: mdl-2429706

RESUMEN

The intracellular localization of pancreatic enzyme secretion-stimulating activity in rat pancreas was investigated. We found and purified a pancreatic enzyme secretion-stimulating peptide from rat bile/pancreatic juice. The peptide is trypsin-sensitive (showing temporary trypsin inhibitory activity), and it is hypothesized that it acts as a trypsin-sensitive mediator in the feedback regulation of diet-induced pancreatic enzyme secretion. The zymogen granule fraction was purified 5-fold by ultracentrifugation by the Percoll density gradient method. The purity of the zymogen granule fraction was determined from the specific amylase activity and electron microscopic morphology. The specific enzyme activities of amylase and trypsin and the trypsin inhibitory activity increased in parallel during the purification, and the pancreatic enzyme secretion-stimulating activity was also localized in the zymogen granule fraction. These results suggest that the pancreatic enzyme secretion-stimulating peptide originates from the acinar cells, and that it is secreted through exocytosis of zymogen granules into the small intestine, its ratio to trypsin thus remaining constant. This idea supports our hypothesis that the stimulating peptide acts as a mediator for the feedback regulation of pancreatic enzyme secretion by trypsin.


Asunto(s)
Gránulos Citoplasmáticos/metabolismo , Precursores Enzimáticos/metabolismo , Páncreas/enzimología , Péptidos/aislamiento & purificación , Inhibidor de Tripsina Pancreática de Kazal/aislamiento & purificación , Inhibidores de Tripsina/aislamiento & purificación , Amilasas/aislamiento & purificación , Animales , Fraccionamiento Celular/métodos , Gránulos Citoplasmáticos/enzimología , Masculino , Páncreas/metabolismo , Ratas , Ratas Endogámicas , Tripsina/aislamiento & purificación
19.
Cell Death Differ ; 9(6): 682-92, 2002 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-12032677

RESUMEN

Insulin-like growth factor-1 (IGF-1) inhibited N-acetylsphingosine (C2-ceramide)-induced HL-60 cell apoptosis via relieving oxidative damage. This inhibitory action of IGF-1 was blocked by a phosphatidylinositol-3 (PI-3) kinase inhibitor wortmannin and enhanced by overexpression of the p110 catalytic subunit of PI-3 kinase. Either IGF-1 pretreatment or PI-3 kinase overexpression restored ceramide-depleted catalase function, and this restoration was inhibited by wortmannin. A catalase inhibitor 3-amino-1h-1, 2, 4-triazole (ATZ) blocked the inhibitory action of IGF-1 on ceramide-induced apoptosis, whereas exogenous purified catalase enhanced it. Ceramide-activated caspase-3 was inhibited by IGF-1/PI-3 kinase and enhanced by wortmannin, while the addition of a specific caspase-3 inhibitor DMQD-CHO significantly enhanced the restoration by IGF-1 of ceramide-depleted catalase function. Moreover, IGF-1 inhibited C2-ceramide-induced decrease of mitochondrial membrane potential, and increase of cytochrome c release, caspase-3 cleavage and caspase-3 activity as judged by PhiPhiLux cleaving method. In summary, these results suggest that IGF-1/PI-3 kinase inhibited C2-ceramide-induced apoptosis due to relieving oxidative damage, which resulted from the inhibition of catalase by activated caspase-3.


Asunto(s)
Apoptosis/fisiología , Catalasa/metabolismo , Factor I del Crecimiento Similar a la Insulina/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Esfingosina/análogos & derivados , Esfingosina/fisiología , Androstadienos/farmacología , Caspasa 3 , Inhibidores de Caspasas , Caspasas/metabolismo , Catalasa/antagonistas & inhibidores , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Células HL-60 , Humanos , Membranas Intracelulares/fisiología , Metabolismo de los Lípidos , Peroxidación de Lípido , Potenciales de la Membrana , Mitocondrias/fisiología , Oligopéptidos/farmacología , Oxidación-Reducción , Esfingosina/antagonistas & inhibidores , Wortmanina
20.
J Int Med Res ; 33(2): 207-14, 2005.
Artículo en Inglés | MEDLINE | ID: mdl-15790132

RESUMEN

The effects of adiponectin replacement therapy on myocardial damage were studied in leptin-deficient (OB) mice with acute viral myocarditis. Encephalomyocarditis virus was injected intraperitoneally into OB and wild-type (WT) mice. One subgroup of OB mice received no intervention and another subgroup received daily adiponectin replacement, simultaneously with viral inoculation. Differences in heart weight, cardiac histological score, numbers of infiltrating or apoptotic cells in the myocardium and the immunoreactivity of adiponectin receptors in myocytes were determined. The reactivity of adiponectin receptor 1 in myocytes from OB mice on day 4 and day 8 after viral inoculation was significantly decreased compared with that in myocytes from WT mice; the OB mice also had elevated cardiac weights and severe inflammatory myocardial damage. Adiponectin replacement in OB mice inhibited the development of severe myocarditis by augmenting myocyte adiponectin receptor 1 reactivity. Exogenously administered adiponectin may inhibit the progression of viral myocarditis through binding to the adiponectin receptor 1 in leptin-deficient conditions.


Asunto(s)
Virus de la Encefalomiocarditis/metabolismo , Péptidos y Proteínas de Señalización Intercelular/administración & dosificación , Leptina/metabolismo , Miocarditis/terapia , Miocarditis/virología , Miocardio/patología , Adiponectina , Animales , Apoptosis , Peso Corporal , Progresión de la Enfermedad , Terapia Genética , Corazón/fisiología , Corazón/virología , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Células Musculares/metabolismo , Miocardio/metabolismo , Tamaño de los Órganos , Unión Proteica , Receptores de Adiponectina , Receptores de Superficie Celular/metabolismo , Receptores de Leptina , Factores de Tiempo
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