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2.
Eur Cell Mater ; 31: 95-106, 2016 Jan 27.
Artículo en Inglés | MEDLINE | ID: mdl-26815642

RESUMEN

Transplantation of activated nucleus pulposus (NP) cells obtained by coculturing NP cells and bone marrow mesenchymal stromal cells having cell-to-cell contact has been shown to be effective in animal models and, more recently, in human clinical trials. If the NP cells can be cryopreserved, then autologous cell transplantation could be offered to patients as and when required. In a previous study, we confirmed that activated NP cells can be obtained by coculturing with mesenchymal cells after cryopreservation. However, the in vivo effects of cell transplantation therapy using activated NP cells prepared from cryopreserved cells are not known. In this in vivo canine model, we compared indicators of disc degeneration in animals that received transplanted activated normal NP cells, transplanted cryopreserved NP cells, and no cell transplantation after induction of disc degeneration. The intervertebral disc height on radiographs and T2-weighted magnetic resonance imaging were significantly higher in both cell transplantation groups compared with the degenerated disc group. Macroscopic and histological findings demonstrated attenuated disc degeneration in the two transplanted groups. Intense staining of proteoglycan and collagen type II was seen in green fluorescent protein-labelled transplanted cells, which suggested that the cells had survived and were functioning after transplantation. No significant differences were observed between the two transplanted groups. Transplanted activated cryopreserved NP cells induced a similar attenuation of intervertebral disc degeneration as that of conventionally activated NP cells. These findings suggest that the use of cryopreserved cells specific to a patient's condition has potential in transplantation therapy.


Asunto(s)
Trasplante de Células/métodos , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Criopreservación , Degeneración del Disco Intervertebral/terapia , Disco Intervertebral/citología , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/citología , Animales , Células Cultivadas , Técnicas de Cocultivo , Perros , Femenino , Dolor de la Región Lumbar/terapia , Vértebras Lumbares/metabolismo
3.
J Thromb Haemost ; 11(9): 1742-50, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23848403

RESUMEN

BACKGROUND: Familial platelet disorder (FPD) is a rare autosomal dominant disease characterized by thrombocytopenia and abnormal platelet function. Causal mutations have been identified in the gene encoding runt-related transcription factor 1 (RUNX1) of FPD patients. OBJECTIVES: To elucidate the role of RUNX1 in the regulation of expression of platelet factor 4 (PF4) and to propose a plausible mechanism underlying RUNX1-mediated induction of the FPD phenotype. METHODS: We assessed whether RUNX1 and its mutants, in combination with E26 transformation-specific-1 (ETS-1), Core-binding factor subunit beta (CBFß), and Friend leukemia virus integration 1 (FLI-1), cooperatively regulate PF4 expression during megakaryocytic differentiation. In an embryonic stem cell differentiation system, expression levels of endogenous and exogenous RUNX1 and PF4 were determined by real-time RT-PCR. Promoter activation by the transcription factors were evaluated by reporter gene assays with HepG2 cells. DNA binding activity and protein interaction were analyzed by electrophoretic mobility shift assay and immunoprecipitation assay with Cos-7 cells, respectively. Protein localization was analyzed by immunocytochemistry and Western blotting with Cos-7 cells. RESULTS: We demonstrated that RUNX1 activates endogenous PF4 expression in megakaryocytic differentiation. RUNX1, but not its mutants, in combination with ETS-1 and CBFß, or FLI-1, synergistically activated the PF4 promoter. Each RUNX1 mutant harbors various functional abnormalities, including loss of DNA-binding activity, abnormal subcellular localization, and/or alterations of binding affinities for ETS-1, CBFß, and FLI-1. CONCLUSIONS: RUNX1, but not its mutants, strongly and synergistically activates PF4 expression along with ETS family proteins. Furthermore, loss of the RUNX1 transcriptional activation function is induced by various functional abnormalities.


Asunto(s)
Trastornos de las Plaquetas Sanguíneas/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Regulación de la Expresión Génica/genética , Mutación , Factor Plaquetario 4/genética , Proteínas Proto-Oncogénicas c-ets/metabolismo , Línea Celular , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Fracciones Subcelulares/metabolismo
4.
Med Phys ; 39(6Part8): 3685, 2012 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-28518900

RESUMEN

PURPOSE: We have recently developed a dynamic tumor tracking irradiation system using Vero4DRT (MHI-Tm2 000). It is needed to create a 4D correlation model between a fiducial marker implanted near a tumor and an external surrogate as a function of time by continuously acquiring both fluoroscopy images and external surrogate signals. The purpose of this study was to propose a new dosimetry method using Gafchromic XR-SP2 films to measure surface dose by fluoroscopy imaging. METHODS: First, half-value layers (HVLs) were measured using aluminum (Al) thicknesses (15 mm) at 40125 kVp. Subsequently, several films were irradiated using various milliampere second values on a solid water phantom. The surface air kerma were also measured using the chamber to calculate the surface doses under the same condition. Then, the calibration curve of dose vs. pixel values was calculated. Finally, surface dose by fluoroscopy imaging was measured using several pieces of film taped on the chest phantom. Orthogonal X-ray fluoroscopy imaging was simultaneously performed until completion of data acquisition for creating a 4D correlation model. Those films were scanned after irradiation using a flat-bed scanner and converted to dose by calibration curve. RESULTS: The HVLs for tube voltage within 40125 kVp ranged from 2.35 to 5.98 mm Al. The calibration curve between surface dose and pixel values was reasonably smooth. The differences between the measured and the calibrated doses were less than 3%. The hot spots with the maximum dose of 37.12 mGy were observed around the area overlapped by both fluoroscopic fields. CONCLUSIONS: We have proposed a new dosimetry method using Gafchromic XR-SP2 films to measure surface dose by fluoroscopy imaging. This phantom study has demonstrated that it may be feasible to assess surface dose to patients during dynamic tumor tracking irradiation in clinic with ease after further investigation. This research was supported by the Japan Society for the Promotion of Science (JSPS) through its Funding Program for World-Leading Innovation R&D on Science and Technology (FIRST Program). Research sponsored in part by Mitsubishi Heavy Industries, Ltd.

5.
Osteoporos Int ; 18(7): 1009-15, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17554581

RESUMEN

UNLABELLED: Forty-three patients who had undergone cementless THA were randomly assigned to receive no osteoactive drug or oral risedronate for 6 months. Postoperative decrease of BMD in the risedronate group was significantly lower than that seen in the control group in zones 1, 2, 3, 6, and 7. INTRODUCTION: Proximal bone resorption around the femoral stem often has been observed after total hip arthroplasty (THA), could lead to late stem loosening. We previously reported the efficacy of etidronate on periprosthetic bone resorption after cementless THA. Recently risedronate is suggested to be effective for the prevention and treatment of for osteoporosis. The purpose of the present study was to evaluate the effects of risedronate on periprosthetic bone loss after cementless THA. METHODS: Forty-three patients who had undergone cementless THA were randomly assigned to receive no osteoactive drug (21 patients) or oral risedronate 2.5 mg/day (22 patients) for 6 months. Three patients were eliminated from the risedronate group because of dyspepsia. Periprosthetic bone mineral density (BMD) in seven regions of interest based on the zones of Gruen et al. was measured with dual energy X-ray absorptiometry at 3 weeks and 6 months postoperatively. RESULTS: At 6 months after surgery, postoperative decrease of BMD in the risedronate group was significantly lower than that seen in the control group in zones 1, 2, 3, 6, and 7 (p < 0.05, p < 0.01, p < 0.01, p < 0.05, and p < 0.05, respectively). CONCLUSION: These outcomes suggested that risedronate might reduce the periprosthetic bone resorption after cementless THA.


Asunto(s)
Artroplastia de Reemplazo de Cadera/métodos , Conservadores de la Densidad Ósea/administración & dosificación , Resorción Ósea/tratamiento farmacológico , Ácido Etidrónico/análogos & derivados , Osteoporosis/cirugía , Absorciometría de Fotón , Administración Oral , Anciano , Densidad Ósea/efectos de los fármacos , Ácido Etidrónico/administración & dosificación , Femenino , Articulación de la Cadera/diagnóstico por imagen , Articulación de la Cadera/cirugía , Humanos , Masculino , Persona de Mediana Edad , Osteoporosis/tratamiento farmacológico , Complicaciones Posoperatorias/tratamiento farmacológico , Ácido Risedrónico , Resultado del Tratamiento
6.
Gut ; 54(9): 1254-62, 2005 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-16099792

RESUMEN

BACKGROUND AND AIMS: The pathogenesis of Crohn's disease (CD), a chronic inflammatory bowel disease characterised by a Th1 immune response, remains unclear. Osteopontin (OPN) is a phosphoprotein known as an adhesive bone matrix protein. Recent studies have shown that OPN plays an important role in lymphocyte migration, granuloma formation, and interleukin 12 (IL-12) production. The present study investigated expression and the pathophysiological role of OPN in CD. METHODS: Plasma OPN concentration was measured by enzyme linked immunosorbent assay. Expression of OPN in human intestinal mucosa was determined using reverse transcription-polymerase chain reaction and western blot, and localisation of OPN was examined by immunohistochemistry. Expression of integrin beta3, an OPN receptor, on lamina propria mononuclear cells (LPMC) was assessed by flow cytometry. Functional activation of OPN in LPMC was investigated by measuring the production of cytokines. RESULTS: Plasma OPN concentration was significantly higher in patients with CD compared with normal controls or patients with ulcerative colitis (UC). OPN was upregulated in intestinal mucosa from UC and CD patients. OPN producing cells were epithelial or IgG producing plasma cells, or partial macrophages. OPN was detected in areas surrounding granuloma from mucosa in CD. Integrin beta3 expressing macrophages infiltrated inflamed mucosa in UC and CD; in contrast, there was no expression of integrin beta3 on intestinal macrophages in normal mucosa. OPN induced production of IL-12 from LPMC in CD but not in normal controls or UC. CONCLUSIONS: Increased OPN expression facilitates cytokine production and is closely involved in the Th1 immune response associated with CD.


Asunto(s)
Enfermedad de Crohn/inmunología , Sialoglicoproteínas/sangre , Células TH1/inmunología , Biomarcadores/análisis , Biomarcadores/sangre , Estudios de Casos y Controles , Colitis Ulcerosa/inmunología , Células Epiteliales/química , Humanos , Integrina beta3/análisis , Interleucina-12/análisis , Interleucina-12/inmunología , Mucosa Intestinal/química , Mucosa Intestinal/inmunología , Activación de Linfocitos , Macrófagos/química , Osteopontina , Células Plasmáticas/química , ARN Mensajero/análisis , Sialoglicoproteínas/análisis , Sialoglicoproteínas/genética
7.
Br J Radiol ; 77(918): 516-8, 2004 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-15151975

RESUMEN

This report describes a 58-year-old female with adenosquamous carcinoma (ASC) of the liver presenting with right lower abdominal pain. In most ASC of the liver, surgery is the first choice of treatment. However, surgery often seems to be ineffective because of the aggressive behaviour of this disease. At surgical laparotomy in the present case, there was a large tumour occupying the entire right lobe of the liver and invading the diaphragm. Thus, partial hepatectomy was performed. For the residual tumour, we performed repeated transcatheter arterial infusion chemotherapy with degradable starch microspheres (DSM). After four sessions of transcatheter arterial infusion chemotherapy, tumour size decreased remarkably and tumour markers had also decreased. Despite the poor prognosis, the patient remains alive and well 12 months after laparotomy. It is suggested that minimally invasive transcatheter infusion chemotherapy with DSM can be an effective treatment preserving a high quality of life.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Carcinoma Adenoescamoso/terapia , Quimioembolización Terapéutica/métodos , Neoplasias Hepáticas/terapia , Almidón/administración & dosificación , Carcinoma Adenoescamoso/diagnóstico por imagen , Femenino , Humanos , Infusiones Intraarteriales , Neoplasias Hepáticas/diagnóstico por imagen , Persona de Mediana Edad , Tomografía Computarizada por Rayos X
8.
Int J Clin Oncol ; 6(3): 149-52, 2001 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-11706785

RESUMEN

We report findings in a 76-year-old man who underwent a lateral segmentectomy of the liver for hepatocellular carcinoma in July 1996. In July 1997, transarterial embolization (TAE) was performed for recurrent tumors in the remnant liver. Augmentation of the tumors and an increase in protein induced by vitamin K absence or antagonist (PIVKA)-II level were noted in October 1997, and, although we recommended TAE again, the patient and his family refused further treatment. Subsequently, the patient was only observed, and, except for a small lesion that was probably a scar, no tumors were noted on image examinations in November 1998, and the PIVKA-II level had returned to a normal value at this time. Two years after the regression, the tumors appeared to be in complete spontaneous remission. This patient had no history indicative of ischemic necrosis, and levels of cellular surface markers for natural killer (NK) cells and NK cell activity showed high values, which suggested that tumor immunity was activated by some, unknown, mechanism.


Asunto(s)
Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/terapia , Embolización Terapéutica/métodos , Neoplasias Hepáticas/terapia , Recurrencia Local de Neoplasia/patología , Recurrencia Local de Neoplasia/terapia , Anciano , Humanos , Células Asesinas Naturales , Neoplasias Hepáticas/patología , Masculino , Remisión Espontánea , Negativa del Paciente al Tratamiento
9.
Arch Orthop Trauma Surg ; 121(7): 365-7, 2001 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-11510898

RESUMEN

There were 56 consecutive patients enrolled in our study from the outpatient clinic of our institution, mean age of 59 years, who had been on hemodialysis for 2-26 years. The duration of hemodialysis was less than 10 years for 36 patients (short-term group) and 10 years or more for 20 patients (long-term group) with a mean duration of 11 years at time of the investigation. In the short-term group, plain radiography revealed bone cysts in 9 hips (17%) and joint space narrowing in 10 hips (19%), while in the long-term group, there were bone cysts in 19 hips (48%) and joint space narrowing in 6 hips (15%). Bone mineral density as determined by dual-energy X-ray absorptiometry was 0.621 g/cm2 for patients in the short-term group and 0.503 g/cm2 for patients in the long-term group. Hip arthroplasties were performed in 3 patients suffering from femoral neck fracture due to bone cysts. All of them showed marked bone loss (mean 0.380 g/cm2). In conclusion, plain radiography and dual-energy X-ray absorptiometry are useful for evaluating the hip arthropathy of hemodialysis patients, especially those who have undergone long-term hemodialysis.


Asunto(s)
Articulación de la Cadera/diagnóstico por imagen , Artropatías/diagnóstico por imagen , Artropatías/etiología , Diálisis Renal/efectos adversos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Radiografía , Estudios Retrospectivos , Factores de Tiempo
10.
Biochemistry ; 40(15): 4633-44, 2001 Apr 17.
Artículo en Inglés | MEDLINE | ID: mdl-11294630

RESUMEN

Histidinol-phosphate aminotransferase (HspAT) is a key enzyme on the histidine biosynthetic pathway. HspAT catalyzes the transfer of the amino group of L-histidinol phosphate (Hsp) to 2-oxoglutarate to form imidazole acetol phosphate (IAP) and glutamate. Thus, HspAT recognizes two kinds of substrates, Hsp and glutamate (double substrate recognition). The crystal structures of native HspAT and its complexes with Hsp and N-(5'-phosphopyridoxyl)-L-glutamate have been solved and refined to R-factors of 19.7, 19.1, and 17.8% at 2.0, 2.2, and 2.3 A resolution, respectively. The enzyme is a homodimer, and the polypeptide chain of the subunit is folded into one arm, one small domain, and one large domain. Aspartate aminotransferases (AspATs) from many species were classified into aminotransferase subgroups Ia and Ib. The primary sequence of HspAT is less than 18% identical to those of Escherichia coli AspAT of subgroup Ia and Thermus thermophilus HB8 AspAT of subgroup Ib. The X-ray analysis of HspAT showed that the overall structure is significantly similar to that of AspAT of subgroup Ib rather than subgroup Ia, and the N-terminal region moves close to the active site like that of subgroup Ib AspAT upon binding of Hsp. The folding of the main-chain atoms in the active site is conserved between HspAT and the AspATs, and more than 40% of the active-site residues is also conserved. The eHspAT recognizes both Hsp and glutamate by utilizing essentially the same active-site folding as that of AspAT, conserving the essential residues for transamination reaction, and replacing and relocating some of the active-site residues. The binding sites for the phosphate and the alpha-carboxylate groups of the substrates are roughly located at the same position and those for the imidazole and gamma-carboxylate groups at the different positions. The mechanism for the double substrate recognition observed in eHspAT is in contrast to that in aromatic amino acid aminotransferase, where the recognition site for the side chain of the acidic amino acid is formed at the same position as that for the side chain of aromatic amino acids by large-scale rearrangements of the hydrogen bond networks.


Asunto(s)
Escherichia coli/enzimología , Glutamatos/química , Ácido Glutámico/análogos & derivados , Ácido Glutámico/química , Histidinol/química , Piridinas/química , Transaminasas/química , Secuencia de Aminoácidos , Animales , Sitios de Unión , Cristalografía por Rayos X , Enlace de Hidrógeno , Sustancias Macromoleculares , Modelos Moleculares , Datos de Secuencia Molecular , Conformación Proteica , Estereoisomerismo , Especificidad por Sustrato , Porcinos
11.
J Surg Oncol ; 75(3): 197-202, 2000 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-11088052

RESUMEN

A case of erythrocytosis caused by a hepatocellular carcinoma (HCC) that produced erythropoietin (Epo) is described. A 64-year-old man, with a huge HCC tumor in the right lobe of the liver, showed a high concentration of hemoglobin and increased levels of serum Epo, alpha-fetoprotein (AFP), and protein induced by vitamin K absence II (PIVKA-II). Right lobectomy of the liver was performed. Histological findings of the specimen showed a moderately differentiated HCC. The existence of Epo was confirmed immunohistochemically only in the tumor tissue and not in the normal liver tissue. Erythrocytosis disappeared and the serum levels of Epo, AFP, and PIVKA-II returned to the normal range after the operation. Within 2 months after the operation, recurrent tumors appeared in the remnant liver, and the patient died 13 months after the operation.


Asunto(s)
Biomarcadores , Carcinoma Hepatocelular/complicaciones , Eritropoyetina/biosíntesis , Neoplasias Hepáticas/complicaciones , Policitemia/etiología , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Humanos , Inmunohistoquímica , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Precursores de Proteínas/sangre , Protrombina , alfa-Fetoproteínas/metabolismo
12.
J Bone Joint Surg Am ; 82(10): 1421-5, 2000 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-11057470

RESUMEN

BACKGROUND: The purpose of this study was to assess the usefulness of bone scintigraphy in predicting progressive collapse of the femoral head after transtrochanteric rotational osteotomy for the treatment of osteonecrosis of the femoral head. METHODS: We studied thirty-three hips in thirty patients with osteonecrosis of the femoral head who had undergone transtrochanteric rotational osteotomy. There were twenty male and ten female patients, with a mean age of 34.4 years at the time of the operation. The mean duration of follow-up was 10.0 years. According to the staging system of Ficat and Arlet, there were nineteen stage-2 hips and fourteen stage-3 hips at the time of the operation. Conventional anteroposterior and lateral radiographs were assessed. In addition, bone scans were performed at three weeks after the operation to predict the outcome with regard to the rotated femoral head. On the basis of the location of low scan activity within the femoral head, the scintigraphic findings were classified into one of two categories: type A if there was no low scan activity in the weight-bearing area of the femoral head or type B if low scan activity occupied the entire weight-bearing area. Six hips with collapse were studied histologically. RESULTS: Postoperative scintiscans revealed sixteen type-A hips and seventeen type-B hips. Of the type-A hips, only three exhibited progressive collapse of the femoral head after the osteotomy, whereas fourteen of the type-B hips exhibited progressive collapse. A significant association was found between the postoperative scintigraphic findings and the final radiographic result (p < 0.01). CONCLUSIONS: Bone scintiscans made three weeks after transtrochanteric rotational osteotomy were useful for predicting the final clinical result.


Asunto(s)
Necrosis de la Cabeza Femoral/cirugía , Cabeza Femoral/diagnóstico por imagen , Osteotomía , Adulto , Femenino , Necrosis de la Cabeza Femoral/diagnóstico por imagen , Estudios de Seguimiento , Humanos , Masculino , Complicaciones Posoperatorias/diagnóstico por imagen , Valor Predictivo de las Pruebas , Cintigrafía , Radiofármacos , Estudios Retrospectivos , Medronato de Tecnecio Tc 99m , Factores de Tiempo , Soporte de Peso
13.
Arch Orthop Trauma Surg ; 120(9): 489-92, 2000.
Artículo en Inglés | MEDLINE | ID: mdl-11011665

RESUMEN

We investigated the pathology of femoral head collapse following transtrochanteric anterior rotational osteotomy. Six femoral heads were obtained during total hip arthroplasty some 2-12 years after osteotomy. In all cases, the preoperatively necrotic lesions exhibited mostly osteonecrosis with accumulation of bone marrow cell debris and trabecular bone with empty lacunae, although repair tissue such as granulation tissue and appositional bone formation were observed in limited areas in some cases. In the transposed intact articular surface of the femoral head, osteoarthritic changes such as fissure penetration to the subchondral bone and osteophyte formation were commonly observed. In newly created subchondral areas at weight-bearing sites, trabecular thickness and the number of trabecular bones had decreased, with few osteoblasts, osteoclasts, and osteocytes being present, resulting in a coarse lamellar structure of the trabecular bone. These findings suggest that transposed areas in cases of failure consist mostly of low-turnover osteoporotic lesions which could cause collapse of the femoral head.


Asunto(s)
Necrosis de la Cabeza Femoral/cirugía , Fémur/patología , Osteotomía/métodos , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad
14.
FEBS Lett ; 477(3): 273-7, 2000 Jul 21.
Artículo en Inglés | MEDLINE | ID: mdl-10908734

RESUMEN

It was reported that nicotine-induced dopamine release in the rat pheochromocytoma cell line, PC12 cells, was inhibited by kappa-opioid. However, it is not known whether inhibition of catecholamine biosynthesis is involved in the inhibitory mechanisms of kappa-opioids in PC12 cells. U-69593 (a kappa-opioid agonist: >/=100 nM) significantly inhibited the nicotine-induced increase of tyrosine hydroxylase (TH, a rate-limiting enzyme in biosynthesis of catecholamine) enzyme activity and TH mRNA levels. These inhibitory effects were completely reversed by naloxone and nor-binaltorphimine dihydrochloride (nor-BNI), a specific kappa-antagonist, whereas pertussis toxin (PTX) only partially reversed this inhibitory effect. Also, U-69593 (>/=100 nM) significantly inhibited the nicotine-induced increase of cAMP production. This inhibitory effect was completely reversed by naloxone and nor-BNI, whilst only partially reversed by PTX. Moreover, U-69593 (>/=100 nM) significantly inhibited the nicotine-induced increase of both the TH protein level and intracellular catecholamine levels. These results indicate that the anti-cholinergic actions of kappa-opioid can be explained partially by its inhibition of both TH enzyme activity and TH synthesis, through suppression of the cAMP/protein kinase A pathway. It would also appear that the PTX-sensitive G-protein mediates the inhibitory effect of this pathway, at least in part.


Asunto(s)
Bencenoacetamidas , Catecolaminas/antagonistas & inhibidores , Péptidos Opioides/farmacología , Animales , Catecolaminas/biosíntesis , AMP Cíclico/biosíntesis , Nicotina/farmacología , Células PC12 , Pirrolidinas/farmacología , ARN Mensajero/genética , Ratas , Receptores Opioides kappa/agonistas , Tirosina 3-Monooxigenasa/antagonistas & inhibidores , Tirosina 3-Monooxigenasa/genética
15.
Biochem Biophys Res Commun ; 274(2): 310-5, 2000 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-10913336

RESUMEN

New orexigenic peptides called orexin-A and -B have recently been described in neurons of the lateral hypothalamus and perifornical area. No orexins have been found in adipose tissues or visceral organs, including the adrenal gland. However, expression of the orexin-receptor 2 (OX2R) in the rat adrenal gland has been reported. To test the effects of orexins on peripheral organs, we investigated their effects on catecholamine synthesis and secretion in the rat pheochromocytoma cell line PC12. Orexin-A and -B (100 nM) significantly reduced basal and PACAP-induced tyrosine hydroxylase (TH) (the rate-limiting enzyme in the biosynthesis of catecholamines) mRNA levels. Orexin-A and -B (100 nM) also significantly inhibited the PACAP-induced increase in the cAMP level, suggesting that the suppressive effect on TH mRNA is mediated, at least in part, by the cAMP/protein kinase A pathway. Furthermore, orexin-A and -B (100 nM) significantly suppressed basal and PACAP-induced dopamine secretion from PC12 cells. Next, we examined whether orexin receptors (OX1R, OX2R) were present in the rat adrenal gland and PC12 cells. In the adrenal glands, OX2R was as strongly expressed as in the hypothalamus, but OX1R was not detected. On the other hand, neither OX1R nor OX2R was expressed in PC12 cells. However, binding assays showed equal binding of orexin-A and -B to PC12 cells, suggesting the existence in these cells of some receptors for orexins. These results indicate that orexins suppress catecholamine release and synthesis, and that the inhibitory effect is mediated by the cAMP/protein kinase A pathway.


Asunto(s)
Neoplasias de las Glándulas Suprarrenales/metabolismo , Proteínas Portadoras/metabolismo , Dopamina/biosíntesis , Dopamina/metabolismo , Péptidos y Proteínas de Señalización Intracelular , Neuropéptidos/metabolismo , Feocromocitoma/metabolismo , Neoplasias de las Glándulas Suprarrenales/patología , Animales , Unión Competitiva/efectos de los fármacos , Calcio/metabolismo , Proteínas Portadoras/farmacología , AMP Cíclico/biosíntesis , Relación Dosis-Respuesta a Droga , Neuropéptidos/farmacología , Neurotransmisores/metabolismo , Neurotransmisores/farmacología , Receptores de Orexina , Orexinas , Células PC12 , Feocromocitoma/patología , Polipéptido Hipofisario Activador de la Adenilato-Ciclasa , ARN Mensajero/biosíntesis , Ratas , Receptores Acoplados a Proteínas G , Receptores de Neuropéptido/biosíntesis , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismo
17.
J Biochem ; 128(1): 29-38, 2000 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-10876155

RESUMEN

4-Amino-4-deoxychorismate lyase (ADCL) is a member of the fold-type IV of PLP dependent enzymes that converts 4-amino-4-deoxychorismate (ADC) to p-aminobenzoate and pyruvate. The crystal structure of ADCL from Escherichia coli has been solved using MIR phases in combination with density modification. The structure has been refined to an R-factor of 20.6% at 2.2 A resolution. The enzyme is a homo dimer with a crystallographic twofold axis, and the polypeptide chain is folded into small and large domains with an interdomain loop. The coenzyme, pyridoxal 5'-phosphate, resides at the domain interface, its re-face facing toward the protein. Although the main chain folding of the active site is homologous to those of D-amino acid and L-branched-chain amino acid aminotransferases, no residues in the active site are conserved among them except for Arg59, Lys159, and Glu193, which directly interact with the coenzyme and play critical roles in the catalytic functions. ADC was modeled into the active site of the unliganded enzyme on the basis of the X-ray structures of the unliganded and liganded forms in the D-amino acid and L-branched-chain amino acid aminotransferases. According to this model, the carboxylates of ADC are recognized by Asn256, Arg107, and Lys97, and the cyclohexadiene moiety makes van der Waals contact with the side chain of Leu258. ADC forms a Schiff base with PLP to release the catalytic residue Lys159, which forms a hydrogen bond with Thr38. The neutral amino group of Lys159 eliminates the a-proton of ADC to give a quinonoid intermediate to release a pyruvate in accord with the proton transfer from Thr38 to the olefin moiety of ADC.


Asunto(s)
Escherichia coli/enzimología , Oxo-Ácido-Liasas/química , Secuencia de Aminoácidos , Sitios de Unión , Cristalografía por Rayos X , Ligandos , Modelos Moleculares , Datos de Secuencia Molecular , Oxo-Ácido-Liasas/metabolismo , Conformación Proteica , Fosfato de Piridoxal/metabolismo , Homología de Secuencia de Aminoácido
18.
Biochem Biophys Res Commun ; 272(2): 544-50, 2000 Jun 07.
Artículo en Inglés | MEDLINE | ID: mdl-10833449

RESUMEN

Angiotensin II subtype 2 receptor (AT(2)-R) is abundantly expressed in adrenal medullary chromaffin cells. However, the physiological roles of AT(2)-R in chromaffin cells remain to be clarified. Therefore, we investigated the effects of CGP42112 (AT(2)-R agonist) on catecholamine biosynthesis in cultured porcine adrenal medullary cells. We initially confirmed AT(2)-R was predominantly expressed in porcine adrenal medullary cells by [(125)I]-Ang II binding studies. CGP42112 (>==1 nM) significantly inhibited cGMP production from the basal value. Tyrosine hydroxylase (TH) is a rate-limiting enzyme in the biosynthesis of catecholamine, and its activity is regulated by both TH-enzyme activity and TH-synthesis. CGP42112 (>==1 nM) significantly inhibited TH-enzyme activity from the basal value. These inhibitory effects of CGP42112 on TH-enzyme activity and-cGMP production were abolished by PD123319 (AT(2)-R antagonist) while CV-11974 (AT(1)-R antagonist) was ineffective. We also tested whether decrease of cGMP is involved in the inhibitory effect of CGP42112 on TH-enzyme activity. Pretreatment of 8-Br-cGMP (membrane-permeable cGMP analogue) prevented the inhibitory effect of CGP 42112 on TH-enzyme activity. Similar to that of TH-enzyme activity, CGP42112 (>==1 nM) significantly reduced TH-mRNA and TH-protein level from the basal value, and these inhibitory effects were abolished by PD123319 but not CV-11974. These findings demonstrate that CGP 42112 reduces both TH-enzyme activity and TH-synthesis and that these inhibitory effects could be mediated by decrease of cGMP production.


Asunto(s)
Médula Suprarrenal/efectos de los fármacos , Catecolaminas/biosíntesis , Células Cromafines/efectos de los fármacos , Oligopéptidos/farmacología , Receptores de Angiotensina/agonistas , 8-Bromo Monofosfato de Adenosina Cíclica/farmacología , Médula Suprarrenal/citología , Médula Suprarrenal/enzimología , Médula Suprarrenal/metabolismo , Angiotensina II/antagonistas & inhibidores , Angiotensina II/metabolismo , Angiotensina II/farmacología , Antagonistas de Receptores de Angiotensina , Animales , Bencimidazoles/metabolismo , Bencimidazoles/farmacología , Unión Competitiva , Compuestos de Bifenilo , Células Cultivadas , Células Cromafines/citología , Células Cromafines/enzimología , Células Cromafines/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , GMP Cíclico/farmacología , Relación Dosis-Respuesta a Droga , Activación Enzimática/efectos de los fármacos , Inducción Enzimática/efectos de los fármacos , Imidazoles/metabolismo , Imidazoles/farmacología , Oligopéptidos/antagonistas & inhibidores , Oligopéptidos/metabolismo , Piridinas/metabolismo , Piridinas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptor de Angiotensina Tipo 1 , Receptor de Angiotensina Tipo 2 , Receptores de Angiotensina/metabolismo , Porcinos , Tetrazoles/metabolismo , Tetrazoles/farmacología , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismo
19.
Life Sci ; 66(22): PL303-11, 2000 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-10834306

RESUMEN

Atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) are present in adrenal chromaffin cells, and are co-secreted with catecholamines suggesting that these natriuretic peptides (NPs) may modulate functions of chromaffin cells in an autocrine and/or paracrine manner. Therefore, we investigated the effects of NPs on tyrosine hydroxylase (TH: a rate-limiting enzyme in biosynthesis of catecholamine) mRNA in rat pheochromocytoma PC12 cells. It was also determined whether the cyclic GMP/cGMP-dependent protein kinase (cGMP/PKG) pathway was involved in theses effects. Finally, we examined the effects of NPs on intracellular catecholamine content to confirm increase of catecholamine synthesis following TH mRNA induction. NPs (0.1 microM) induced significant increases of the TH mRNA (ANP= BNP> CNP). Also, the effects of NPs on TH mRNA were mimicked by 8-bromo cyclic GMP (1mM), and were blocked by KT5823 (1 microM) (inhibitor PKG) or LY83583 (1 microM) (guanylate cyclase inhibitor). Moreover, NPs were shown to induce significant increases of intracellular catecholamine contents (ANP= BNP> CNP). These findings suggest that NPs induced increases of TH mRNA through cGMP/PKG dependent mechanisms, which, in turn, resulted in stimulation of catecholamine synthesis in PC12 cells.


Asunto(s)
Factor Natriurético Atrial/farmacología , Catecolaminas/biosíntesis , Péptido Natriurético Encefálico/farmacología , Péptido Natriurético Tipo-C/farmacología , Tirosina 3-Monooxigenasa/metabolismo , Animales , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , GMP Cíclico/farmacología , Proteínas Quinasas Dependientes de GMP Cíclico , Inhibidores Enzimáticos/farmacología , Guanilato Ciclasa/antagonistas & inhibidores , Células PC12 , Inhibidores de Proteínas Quinasas , Proteínas Quinasas/metabolismo , ARN Mensajero/metabolismo , Ratas , Tirosina 3-Monooxigenasa/genética
20.
Arch Orthop Trauma Surg ; 120(5-6): 252-4, 2000.
Artículo en Inglés | MEDLINE | ID: mdl-10853889

RESUMEN

We reviewed 37 patients with avascular necrosis of the femoral head (ANF). There were 23 men and 14 women with a mean age of 36 years at the time of the operation. The duration of follow-up was 9 years. Twenty patients had undergone transtrochanteric rotational osteotomy (TRO) and 17, hip arthroplasties. Assessment of their quality of life (QoL) was performed using the Rosser Index Matrix for disability and distress. Concerning TRO, the mean preoperative and postoperative QoL scores were 0.944 and 0.957, respectively. Twelve patients exhibited increases and 7 patients decreases in their scores. Regarding the arthroplasty, the mean preoperative and postoperative QoL scores were 0.949 and 0.998, respectively. All patients showed increases in QoL scores after arthroplastic surgery. Concerning heavy manual work, all five of those patients returned to their preoperative occupations. These findings suggest that hip arthroplasty has more reliable therapeutic effects than TRO on QoL improvement for patients with ANF.


Asunto(s)
Artroplastia de Reemplazo de Cadera , Necrosis de la Cabeza Femoral/cirugía , Osteotomía , Complicaciones Posoperatorias/diagnóstico , Calidad de Vida , Adulto , Artroplastia de Reemplazo de Cadera/rehabilitación , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Osteotomía/rehabilitación , Complicaciones Posoperatorias/rehabilitación , Rehabilitación Vocacional
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