RESUMEN
OBJECTIVE: The primary aim of this study was to investigate the association of polymorphisms of TRAF1-C5, a newly identified rheumatoid arthritis (RA) risk locus in Caucasians, with susceptibility to RA and systemic lupus erythematosus (SLE) in Japanese populations. Gene expression levels of TRAF1 and C5 to assess the functional significance of genotypes were also analysed. METHODS: A multicentre association study consisting of 4 RA case-control series (4397 cases and 2857 controls) and 3 SLE case-control series (591 cases and 2199 shared controls) was conducted. Genotyping was performed using TaqMan genotyping assay for two single nucleotide polymorphisms (SNPs) that showed the best evidence of association in the previous Caucasian studies. Quantifications of TRAF1 and C5 expression were performed with TaqMan expression assay. RESULTS: Significant differences in allele frequency for both SNPs were observed between RA and control subjects (combined odds ratio = 1.09), while no significant difference was detected between patients with SLE and controls. Interestingly, alleles rs3761847 A and rs10818488 G had increased the risk for RA in the present study, while they decreased the risk in the original studies. A significant difference was found between risk allele carriers and non-carriers of rs10818488 for the expression level of TRAF1 in phorbol myristate acetate-stimulated lymphoblastoid cell lines (p = 0.04). CONCLUSION: Association of TRAF1-C5 locus with RA susceptibility was detected in the Japanese populations with modest magnitude, while no significant association was observed for SLE. Significant positive effect of genotype on the expression of TRAF1 might support the genetic association between TRAF1 and RA.
Asunto(s)
Artritis Reumatoide/genética , Complemento C5/genética , Lupus Eritematoso Sistémico/genética , Polimorfismo de Nucleótido Simple , Factor 1 Asociado a Receptor de TNF/genética , Artritis Reumatoide/diagnóstico por imagen , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Pueblo Asiatico/genética , Autoanticuerpos/sangre , Estudios de Casos y Controles , Línea Celular , Complemento C5/metabolismo , Femenino , Predisposición Genética a la Enfermedad/genética , Estudio de Asociación del Genoma Completo/métodos , Genotipo , Articulaciones de la Mano/diagnóstico por imagen , Humanos , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Masculino , Persona de Mediana Edad , Radiografía , Factor 1 Asociado a Receptor de TNF/metabolismoRESUMEN
OBJECTIVES: Endoscopic spinal surgery has become increasingly common year for year because it is a minimally invasive procedure. In our hospital, we introduced endoscopic bilateral decompression with a unilateral approach (endo-BiDUA) in 2006. In this paper, we review clinically and radiographically the elderly patients who underwent the endo-BiDUA. METHODS: Fifteen patients aged over 65 years were included in this review. They had undergone endo-BiDUA between January 2006 and July 2008. Operation time, blood loss, complications, clinical outcome using the Japanese Orthopedic Association (JOA) score, and enlargement ratio of the dural tube on magnetic resonance imaging (MRI), were evaluated. RESULTS: The mean operation time per level of endo-BiDUA was 144 min. Blood loss was about 60.2 mL. One patient had a post-surgical hematoma and required an additional laminectomy and removal of the hematoma 2 days after the surgery. No other complications such as dural tear, nerve root injury, or infection were encountered. All patients but one, who had a post-surgical hematoma, could start walking within 2 days following the surgery. Before surgery, the mean JOA score was 17.0 and it improved to about 23.3 after the surgery. The area of the dural tube increased to 408.0% after the surgery (range: 211-774%). CONCLUSIONS: Endo-BiDUA facilitated the return of elderly patients with lumbar spinal canal stenosis to their original daily activities.
Asunto(s)
Descompresión Quirúrgica/métodos , Vértebras Lumbares/cirugía , Estenosis Espinal/cirugía , Anciano , Anciano de 80 o más Años , Endoscopía , Femenino , Humanos , Vértebras Lumbares/diagnóstico por imagen , Imagen por Resonancia Magnética , Masculino , Radiografía , Estenosis Espinal/diagnóstico por imagen , Resultado del TratamientoRESUMEN
BACKGROUND: Endoscopic lumbar decompression is useful for the treatment of various spinal conditions and is being performed in an increasing number of patients worldwide. We reviewed the surgery-related complications in patients who underwent endoscopic surgery and discuss the learning curve for this procedure. METHODS: Since the first case in August 2000, a total of 138 patients have undergone endoscopic posterior decompression surgery. Of these, there were 74 patients with Herniated Nucleus Pulposus (HNP), 57 with Lumbar Canal Stenosis (LCS), and 7 with other conditions. From 2003 to 2005, the senior surgeon took a sabbatical, and no endoscopic surgery was conducted. We divided the cases based on the date of surgery: there were 62 patients in the early (E) group (before September 2003), and 76 in the late (L) group (from January 2006 to April 2008). We compared the incidence of surgery-related complications between 2 disease types as well as between the E and L groups. RESULTS: We encountered 11 complications, which included 6 dural tears, 2 post-surgical hematomas, 2 neural complications and 1 fracture of the inferior articular process. The incidence of surgery-related complications was 8.6%. The incidences of complications were 8.1% and 9.3% for HNP and LCS, respectively, and 11.3%, and 5.3% in the E and L groups, respectively. The incidence was particularly high (16.7%) in the E group with LCS. CONCLUSION: There is a steep learning curve for endoscopic surgery. Based on the data, surgeons should start performing endoscopic techniques for LCS after gaining enough experience of endoscopic surgery for HNP.
Asunto(s)
Descompresión Quirúrgica/efectos adversos , Endoscopía/efectos adversos , Desplazamiento del Disco Intervertebral/cirugía , Vértebras Lumbares/cirugía , Estenosis Espinal/cirugía , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Descompresión Quirúrgica/métodos , Duramadre/lesiones , Endoscopía/métodos , Femenino , Hematoma/etiología , Humanos , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
Long-term follow-up of patients with adhesive capsulitis (AC) reveals that approximately half of them suffer from a limited range of shoulder motion, particularly external and/or internal rotation. We report the surgical technique and short-term clinical outcomes of ultrasound-guided release of the thickened coracohumeral (CH) ligament in 8 patients (9 shoulders) with AC. Passive external rotation with the arm by the side significantly increased from an average of 18° preoperatively to 47° immediately after CH ligament release. VAS and ASES scores were improved at 3months follow-up in all 9 shoulders, and maintained at 6months follow-up in 6 shoulders. No procedure-related adverse events developed over the 6-month follow-up period. Ultrasound-guided release for thickened CH ligament is a reliable and effective minimally invasive surgery for persistent limited external rotation due to AC of the shoulder.
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Bursitis/fisiopatología , Bursitis/cirugía , Ligamentos Articulares/diagnóstico por imagen , Ligamentos Articulares/cirugía , Articulación del Hombro/fisiopatología , Adulto , Anciano , Bursitis/complicaciones , Apófisis Coracoides , Femenino , Humanos , Húmero , Masculino , Persona de Mediana Edad , Rango del Movimiento Articular , Rotación , Articulación del Hombro/cirugía , Dolor de Hombro/etiología , Cirugía Asistida por Computador , UltrasonografíaRESUMEN
This study was undertaken to elucidate the mechanism of biological repair at the tendon-bone junction in a rat model. The stump of the toe flexor tendon was sutured to a drilled hole in the tibia (tendon suture group, n = 23) to investigate healing of the tendon-bone junction both radiologically and histologically. Radiological and histological findings were compared with those observed in a sham control group where the bone alone was drilled (n = 19). The biomechanical strength of the repaired junction was confirmed by pull-out testing six weeks after surgery in four rats in the tendon suture group. Callus formation was observed at the site of repair in the tendon suture group, whereas in the sham group callus formation was minimal. During the pull-out test, the repaired tendon-bone junction did not fail because the musculotendinous junction always disrupted first. In order to understand the factors that influenced callus formation at the site of repair, four further groups were evaluated. The nature of the sutured tendon itself was investigated by analysing healing of a tendon stump after necrosis had been induced with liquid nitrogen in 16 cases. A proximal suture group (n = 16) and a partial tenotomy group (n = 16) were prepared to investigate the effects of biomechanical loading on the site of repair. Finally, a group where the periosteum had been excised at the site of repair (n = 16) was examined to study the role of the periosteum. These four groups showed less callus formation radiologically and histologically than did the tendon suture group. In conclusion, the sutured tendon-bone junction healed and achieved mechanical strength at six weeks after suturing, showing good local callus formation. The viability of the tendon stump, mechanical loading and intact periosteum were all found to be important factors for better callus formation at a repaired tendon-bone junction.
Asunto(s)
Callo Óseo , Traumatismos de los Tendones/cirugía , Cicatrización de Heridas , Animales , Fenómenos Biomecánicos , Masculino , Modelos Teóricos , Periostio/irrigación sanguínea , Ratas , Ratas Endogámicas BB , Estrés MecánicoRESUMEN
A 54 year old man developed an unusual lipoma in the patellar tendon, consisting of a fibro-adipose component and a chondro-osseous component. The fibro-adipose component contained mature adipocytes, lipoblasts, and fibroblasts; the chondro-osseous component showed typical endochondral bone formation. Molecular analysis showed that the identical HMGA2-LPP fusion transcript-characteristic of lipoma, parosteal lipoma, and pulmonary chondroid hamartoma-was detectable in the both components.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Proteínas HMGA/análisis , Articulación de la Rodilla , Lipoma/química , Proteínas de Fusión Oncogénica/análisis , Neoplasias de los Tejidos Blandos/química , Tendones , Adipogénesis , Diferenciación Celular , Condrogénesis , Fibrosis , Humanos , Lipoma/patología , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Neoplasias de los Tejidos Blandos/patología , Coloración y EtiquetadoRESUMEN
Vascular leiomyomas, or angioleiomyomas, are benign tumours originating from smooth non-striated muscle. Leiomyomas in the hand are uncommon and their pre-operative diagnosis is difficult. We report a 65 year-old woman who developed a vascular leiomyoma arising from the deep palmar arterial arch.
Asunto(s)
Angiomioma/cirugía , Mano/cirugía , Neoplasias de los Tejidos Blandos/cirugía , Anciano , Angiomioma/irrigación sanguínea , Angiomioma/diagnóstico , Arterias/patología , Arterias/cirugía , Diagnóstico Diferencial , Femenino , Dedos/irrigación sanguínea , Mano/irrigación sanguínea , Mano/patología , Humanos , Procesamiento de Imagen Asistido por Computador , Imagenología Tridimensional , Imagen por Resonancia Magnética , Neoplasias de los Tejidos Blandos/irrigación sanguínea , Neoplasias de los Tejidos Blandos/diagnóstico , Tomografía Computarizada por Rayos XRESUMEN
Aberrant tyrosine phosphorylation of beta-catenin inactivates the E-cadherin-mediated cell adhesion and invasion suppressor system in cancer cells. Elucidation of the association between beta-catenin and c-erbB-2 protein prompted us to investigate whether interference with this interaction can change the invasive phenotype. In a human gastric cancer cell line, TMK-1, N-terminally deleted beta-catenin, which binds to c-erbB-2 but not to cadherin, inhibited the association between endogenous beta-catenin and c-erbB-2 protein, and suppressed the tyrosine phosphorylation of beta-catenin. Cells expressing truncated beta-catenin exhibited markedly reduced invasiveness in vitro and peritoneal metastasis in vivo, and developed an epithelial morphology. These results suggest that tyrosine phosphorylation of beta-catenin regulated by c-erbB-2 protein may play an important role in the invasion, metastasis and morphogenesis of cancer cells and that inhibition of the aberrant tyrosine phosphorylation of beta-catenin effectively prevents invasion and metastasis of cancer cells.
Asunto(s)
Proteínas del Citoesqueleto/metabolismo , Invasividad Neoplásica/genética , Metástasis de la Neoplasia/genética , Receptor ErbB-2/metabolismo , Neoplasias Gástricas/secundario , Transactivadores , Animales , Proteínas del Citoesqueleto/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Fosforilación/efectos de los fármacos , Receptor ErbB-2/efectos de los fármacos , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Eliminación de Secuencia , Transducción de Señal , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Transfección , Factor de Crecimiento Transformador alfa/farmacología , Trasplante Heterólogo , Tirosina/metabolismo , beta CateninaRESUMEN
Osteopontin (Opn) and polyoma enhancer-binding protein (PEBP) 2alphaA/core binding factor (CBFA) 1 have been suggested to play important roles in ossification. The overlapping localization of opn and PEBP2alphaA/CBFA1 mRNA, and the marked decrease of opn mRNA expression in PEBP2alphaA knockout mice, indicated that the transcription of opn gene was controlled by PEBP2alphaA. In the present study, we determined the direct regulation of PEBP2alphaA on the opn promoter activity. Opn promoter activity was markedly enhanced by PEBP2alphaA and ETS1 in a synergistic manner. The synergistic effect was diminished when either the PEBP2alphaA or ETS1 binding site was mutated, or the spatial arrangement of these sites was mutated by a 4-nt insertion. The distance between these sites was important for transactivation but not protein-DNA binding. The direct interaction between PEBP2alphaA and ETS1 was depended on protein-DNA binding. These results suggested that the specific spatial arrangement of both sites and direct interaction between PEBP2alphaA and ETS1, were essential for promoter function. Furthermore, endogenous opn mRNA was decreased with the introduction of dominant negative PEBP2alphaA to MC3T3/E1 cells expressing endogenous PEBP2alphaA, ETS1 and opn. These findings suggest that PEBP2alphaA and ETS1 cooperate in vivo to regulate expression of the opn gene in the skeletal tissue. Cell type-specific regulation of Opn gene expression will also be discussed.
Asunto(s)
Proteínas de Unión al ADN/fisiología , Regulación de la Expresión Génica , Proteínas de Neoplasias , Proteínas Proto-Oncogénicas/fisiología , Sialoglicoproteínas/genética , Factores de Transcripción/fisiología , Células 3T3 , Animales , Secuencia de Bases , Sitios de Unión , Secuencia Conservada , Subunidad alfa 1 del Factor de Unión al Sitio Principal , Humanos , Ratones , Datos de Secuencia Molecular , Osteogénesis/genética , Osteopontina , Regiones Promotoras Genéticas , Proteína Proto-Oncogénica c-ets-1 , Proteínas Proto-Oncogénicas c-ets , Alineación de Secuencia , Sialoglicoproteínas/metabolismo , Factor de Transcripción AP-2 , Transcripción Genética , Activación TranscripcionalRESUMEN
The prognosis of high-risk retinoblastoma (RB) with extraocular disease, relapse, or invasion of the cut end of the optic nerve is extremely poor. Following the discontinuation of thiotepa production in Japan, BU- and melphalan (Mel)-based regimens have been used, followed by the standard treatment for neuroblastoma. This study retrospectively analyzed 14 high-risk RB patients who underwent high-dose chemotherapy (HDC) and hematopoietic SCT; 8 received a BU/Mel conditioning regimen and 6 received other regimens. The disease status at HDC was relapse in 8 patients and extraocular involvement in 5. All patients received peripheral blood stem cell infusion >1.5 × 10(6)/kg. Engraftment occurred within a median of 11 days (BU/Mel: 10-13, others: 9-13). Primary toxicities included mucositis (⩾grade 3) in 9 patients (4 with BU/Mel, 5 with others). Veno-occlusive disease (VOD) occurred in two 1-year-old patients in the BU/Mel group. There were no treatment-related deaths. Of 4 (2 with BU/Mel, 2 with others) patients with central nervous system (CNS) relapse after HDC, 3 died. In conclusion, the BU/Mel regimen may be feasible for high-risk RB under careful monitoring for VOD, particularly in younger patients. CNS relapse associated with a lethal prognosis occurred after all regimens; therefore, further evaluation of HDC efficacy for high-risk RB is required.
Asunto(s)
Busulfano/administración & dosificación , Trasplante de Células Madre Hematopoyéticas , Melfalán/administración & dosificación , Agonistas Mieloablativos/administración & dosificación , Trasplante de Células Madre de Sangre Periférica , Neoplasias de la Retina/terapia , Retinoblastoma/terapia , Acondicionamiento Pretrasplante , Aloinjertos , Preescolar , Femenino , Estudios de Seguimiento , Humanos , Lactante , Japón , Masculino , Neoplasias de la Retina/diagnóstico , Retinoblastoma/diagnóstico , Estudios RetrospectivosRESUMEN
Bone lengthening with osteotomy and gradual distraction was achieved in 57 rats, and the effect of mechanical tension-stress on gene expression of bone morphogenetic proteins (BMPs) was investigated by in situ hybridization and Northern blot analysis using probes of BMP-2, BMP-4, BMP-6, BMP-7, and growth/differentiation factor (GDF)-5. There was a lag phase for 7 days after femoral osteotomy until gradual distraction was carried out for 21 days at a rate of 0. 25 mm/12 h using a small external fixator. The signals of the above BMPs mRNA were not detected in the intact rat bone but they were induced after osteotomy except those for BMP-7. By 4 days after osteotomy, BMP-2 and BMP-4 mRNAs were detected in chondrogenic precursor cells in the subperiosteal immature callus. BMP-6 and GDF-5 mRNA were detected in more differentiated cells in chondroid bone. By 7 days after osteotomy, cartilaginous external callus and bony endosteal callus were formed. Meanwhile, the signals of BMP-2 and BMP-4 mRNAs declined to preoperative levels, whereas the signals of BMP-6 and GDF-5 mRNAs were rather elevated. As distraction was started, the callus elongated and eventually separated into proximal and distal segments forming a fibrous interzone in the middle. Expression of BMP-2 and BMP-4 mRNAs was markedly induced at this stage. Their signals were detected widely among chondrogenic and osteogenic cells and their precursor cells sustaining mechanical tension-stress at the fibrous interzone. BMP-6 and GDF-5 mRNAs were detected exclusively in chondrogenic cells at both ends of the fibrous interzone, where endochondral ossification occurred. But neither mRNA was detected in terminally differentiated hypertrophic chondrocytes. As distraction advanced, the cartilage was progressively resorbed from both ends and new bone was formed directly by intramembranous ossification. There was no new cartilage formation in the advanced stage of distraction. The signals of BMP-6 and GDF-5 mRNA declined by this stage, while those of BMP-2 and BMP-4 were maintained at high level for as long as distraction was continued. After completion of distraction, the fibrous interzone fused and the lengthened segment was consolidated. BMP-2, BMP-4, BMP-6, nor GDF-5 was expressed at this stage. The signals of BMP-7 were not detected throughout the experiment. The present results suggest that excellent and uninterrupted bone formation during distraction osteogenesis owes to enhanced expression of BMP-2 and BMP-4 genes by mechanical tension-stress. Abundant gene products of BMP-2 and BMP-4 could induce in situ bone formation by paracrine and autocrine mechanisms.
Asunto(s)
Proteínas Morfogenéticas Óseas/biosíntesis , Fémur/metabolismo , Osteogénesis por Distracción , Factor de Crecimiento Transformador beta , Animales , Northern Blotting , Proteína Morfogenética Ósea 2 , Proteína Morfogenética Ósea 4 , Proteína Morfogenética Ósea 6 , Proteína Morfogenética Ósea 7 , Fémur/diagnóstico por imagen , Fémur/fisiología , Curación de Fractura , Factor 5 de Diferenciación de Crecimiento , Sustancias de Crecimiento/biosíntesis , Hibridación in Situ , Masculino , Osteotomía , ARN Mensajero/biosíntesis , Radiografía , Ratas , Ratas Sprague-Dawley , Estrés Mecánico , Factores de TiempoRESUMEN
Distraction osteogenesis is a recently advanced principle of bone lengthening in which a bone separated by osteotomy is subjected to slow progressive distraction using an external fixation device. Appropriate mechanical tension-stress is believed not to break the callus but rather to stimulate osteogenesis. To study the molecular features of this process, the expression and localization of the mRNAs encoding osteopontin (OPN), osteocalcin (OC), matrix Gla protein (MGP), osteonectin (ON), and collagen type I and I during distraction osteogenesis were examined by in situ hybridization and Northern blot analysis. The process can be divided into three distinct phases: the lag phase for 7 days between osteotomy and the beginning of distraction, the distraction phase for 21 days, and the consolidation phase for several weeks. The histologic and molecular events taking place during the lag phase were similar to those observed in fracture healing. The osteotomy site was surrounded by external callus consisting of hyaline cartilage. As distraction started at the rate of 0.25 mm/12 h, the cartilaginous callus was elongated, deformed, and eventually separated into proximal and distal segments. The chondrocytes were stretched along the tension vector and became fibroblast-like in shape. Although morphologically these cells were distinguishable from osteogenic cells, they expressed OPN, OC, and alkaline phosphatase mRNAs. As distraction advanced, the cartilaginous callus was progressively replaced by bony callus by endochondral ossification and thereafter new bone was formed directly by intramembranous ossification. OPN mRNA was detected in preosteoblasts and osteoblasts at the boundary between fibrous tissue and new bone. ON, MGP, and OC mRNAs appeared early in the differentiation stage. The variety of cell types expressing mRNA encoding bone matrix proteins in distraction osteogenesis was much greater than that detected in the embryonic bone formation and fracture healing process. Moreover, the levels of OPN, ON, MGP, and OC mRNA expression markedly increased during the distraction phase. These results suggested that mechanical tension-stress modulates cell shape and phenotype, and stimulates the expression of the mRNA for bone matrix proteins.
Asunto(s)
Proteínas Morfogenéticas Óseas/biosíntesis , Huesos/metabolismo , Proteínas de la Matriz Extracelular , Fémur/metabolismo , Osteogénesis por Distracción/métodos , ARN Mensajero/biosíntesis , Animales , Northern Blotting , Proteínas Morfogenéticas Óseas/genética , Proteínas de Unión al Calcio/biosíntesis , Proteínas de Unión al Calcio/genética , Condrogénesis , Colágeno/biosíntesis , Colágeno/genética , Colorantes , Fijadores Externos , Fémur/diagnóstico por imagen , Fémur/patología , Hibridación in Situ , Masculino , Osteocalcina/biosíntesis , Osteocalcina/genética , Osteogénesis , Osteonectina/biosíntesis , Osteonectina/genética , Osteopontina , Osteotomía , Radiografía , Ratas , Ratas Sprague-Dawley , Sialoglicoproteínas/biosíntesis , Sialoglicoproteínas/genética , Proteína Gla de la MatrizRESUMEN
In scorbutic patients, fractures are slow to heal because of impaired collagen synthesis. To investigate the influence of impaired collagen synthesis on the differentiation and proliferation of osteogenic and chondrogenic cells, we examined the expression of genes encoding bone matrix proteins, including osteonectin (ON), osteopontin (OPN), osteocalcin (OC), and matrix Gla protein (MGP), as differentiation markers for osteogenic and chondrogenic cells during fracture healing in Osteogenic Disorder Shionogi (ODS) rats, which have a hereditary defect in the ability to synthesize ascorbic acid (Asc). In ODS rats without Asc supplementation, intramembranous ossification was completely inhibited. Although a few fibroblast-like cells expressing ON mRNA were observed, no OPN mRNA-expressing cells were detected. During endochondral ossification, a small amount of metachromatic staining cartilage appeared at the fracture site, but there was no provisional calcification zone in the cartilage. Chondrocytes expressed ON and MGP mRNAs, but not OPN mRNA. When Asc was given to these rats, callus formation was soon detected around the fracture site, while OPN mRNA was expressed by differentiated osteoblasts and hypertrophic chondrocytes. Our data indicate that impaired collagen synthesis due to Asc deficiency inhibited the increase of ON and MGP mRNA-expressing cells as well as the appearance of OPN mRNA-expressing cells. Since OPN is considered to play an important role in normal and pathological mineralization, lack of OPN mRNA expression accompanying impaired collagen synthesis may have a role in defective mineralization and delayed fracture healing in scurvy.
Asunto(s)
Deficiencia de Ácido Ascórbico/metabolismo , Colágeno/biosíntesis , Proteínas de la Matriz Extracelular , Curación de Fractura/efectos de los fármacos , ARN Mensajero/análisis , Animales , Deficiencia de Ácido Ascórbico/genética , Proteínas de Unión al Calcio/biosíntesis , Diferenciación Celular/genética , Condrocitos/metabolismo , Curación de Fractura/genética , Osteocalcina/biosíntesis , Osteogénesis/genética , Osteonectina/biosíntesis , Osteopontina , Fosfoproteínas/biosíntesis , Ratas , Sialoglicoproteínas/biosíntesis , Factores de Tiempo , Proteína Gla de la MatrizRESUMEN
Pycnodysostosis is a rare autosomal recessive skeletal dysplasia characterized by short stature, osteosclerosis, acroosteolysis, bone fragility, and skull deformities. Recently, mutations in the gene encoding cathepsin K (CK), a lysosomal cysteine protease localized exclusively in osteoclasts, were found to be responsible for this disease. We analyzed genomic DNA from four unrelated Japanese patients with this disorder and identified three different mutations of their CK genes: a previously reported missense mutation (A277 V), a novel single base deletion mutation (531 del T) causing a frame shift from codon 142 that results in a premature termination codon, and a novel missense mutation (L9P) in the signal peptide region. To investigate whether the L9P mutation disrupts signal peptide function and decreases protein synthesis, mutant and wild-type CK complementary DNAs driven by the cytomegalovirus promoter were transfected into COS-7 cells, and their gene products were detected by immunohistochemistry and Western blotting. Expression of the mutant protein was markedly reduced, suggesting decreased mature CK production in this patient, which may have been due to dysfunction of the signal peptide. These results provide evidence that a structural change in the signal peptide of the CK protein was involved in the pathogenesis of pycnodysostosis.
Asunto(s)
Huesos/anomalías , Catepsinas/genética , Osteosclerosis/genética , Adulto , Sustitución de Aminoácidos , Animales , Western Blotting , Células COS , Catepsina K , ADN/genética , ADN/aislamiento & purificación , Epítopos/genética , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Mutación/genética , Osteosclerosis/congénito , Linaje , Señales de Clasificación de Proteína/genética , ARN Mensajero/genética , ARN Mensajero/aislamiento & purificación , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Hypophosphatasia is associated with a defect of the tissue-non-specific alkaline phosphatase gene. We performed a mutational analysis in a surviving patient diagnosed at birth as having hypophosphatasia, on the basis of a low level of serum alkaline phosphatase (ALP) activity and characteristic radiographical findings. She had two sisters, one of whom died of respiratory failure complicated by perinatal hypophosphatasia; the other seemed healthy, with a relatively low activity level of ALP. The patient's parents also had low ALP activity. Sequence analysis of the tissue-nonspecific alkaline phosphatase gene was performed, using genomic DNA and total RNA from the skin fibroblasts of the patient and the peripheral mononuclear cells of her parents. The conversion of Phe to Leu at codon 310 (F310L) and Gly to Arg at 439 (G439R) were identified in the patient. Interestingly, the reconstructive experiments demonstrated that the F310L mutant exhibited an ALP activity level 65% of the normal level, whereas the mutant G439R had no activity. Moreover, the digestion by StuI, after a PCR using complementary DNA extracted from fibroblasts of the patient and lymphocytes of her father, revealed a relatively low messenger RNA level of F310L. These findings suggest that the neonatal case of hypophosphatasia was associated with compound mutations, one of which caused the loss of ALP activity and the other of which caused a slight reduction of the ALP activity, with a relatively low level of messenger RNA.
Asunto(s)
Hipofosfatasia/genética , Mutación , Adulto , Fosfatasa Alcalina/sangre , Fosfatasa Alcalina/genética , ADN Complementario/análisis , Femenino , Humanos , Recién Nacido , Masculino , Polimorfismo Genético , EmbarazoRESUMEN
Hypophosphatasia is associated with a defect of the tissue-nonspecific alkaline phosphatase (TNSALP) gene. The onset and clinical severity are usually correlated in hypophosphatasia; patients with perinatal hypophosphatasia die approximately at the time of birth. In contrast, we describe a male neonatal patient with hypophosphatasia who had no respiratory problems and survived. He was compound heterozygous for the conversion of Phe to Leu at codon 310 (F310L) and the deletion of a nucleotide T at 1735 (delT1735), causing the frame shift with the result of the addition of 80 amino acids at the C-terminal of the protein. Because the C-terminal portion of TNSALP is known to be important for TNSALP to bind to the plasma membrane, the localization of wild-type and mutated TNSALP proteins was analyzed using green fluorescent protein chimeras. The expression vectors containing the complementary DNA of fusion proteins consisting of signal peptide, green fluorescent protein, and wild-type or mutated TNSALP, caused by delT1735 or F310L mutation, were introduced transiently or stably in Saos-2 cells. The delT1735 mutant failed to localize at the cell surface membrane, whereas the wild-type and the F310L mutants were located in the plasma membrane and cytoplasm. The assay for enzymatic activity of TNSALP revealed that the delT1735 mutant lost the activity and that the F310L mutant exhibited an enzymatic activity level that was 72% of the normal level. The F310L mutation was also detected in another neonatal patient with relatively mild (nonlethal) hypophosphatasia (reported in J Clin Endocrinol Metab, 81:4458-4461, 1996), suggesting that residual ALP activity of the F310L mutant contributes to the less severe phenotype. The patient is unique, with respect to a discrepancy between onset and clinical severity in hypophosphatasia.
Asunto(s)
Fosfatasa Alcalina/genética , Mutación del Sistema de Lectura , Hipofosfatasia/genética , Fosfatasa Alcalina/análisis , Sustitución de Aminoácidos , Femenino , Eliminación de Gen , Proteínas Fluorescentes Verdes , Humanos , Indicadores y Reactivos , Recién Nacido , Leucina , Proteínas Luminiscentes , Masculino , Especificidad de Órganos , Fenilalanina , Polimorfismo Genético , Proteínas Recombinantes de FusiónRESUMEN
The chemical structure of a lipid A, which was obtained as a minor component from lipopolysaccharide of Helicobacter pylori strain 206-1, was determined to be a glucosamine beta-(1 -6) disaccharide 1-(2-aminoethyl)phosphate acylated by (R)-3-hydroxyoctadecanoic acid, (R)-3- hydroxyhexadecanoic acid, and (R)-3-(octadecanoyloxy)octadecanoic acid at the 2-, 3- and 2'-positions, respectively. Compared with the other major lipid A from the same strain, which was previously reported [Suda Y, Ogawa T, Kashihara W et al. Chemical structure of lipid A from Helicobacter pylori strain 206-1 lipopolysaccharide. J Biochem 1997; 121: 1129--1133], the structure was very similar with one exception. An (R)-3-hydroxyhexadecanoic acid was present at the 3-position of the novel lipid A component. The structure is apparently identical to one of the proposals by Moran et al. [Moran AP, Lindner B, Walsh EJ. Structural characterization of the lipid A component of Helicobacter pylori rough- and smooth-form lipopolysaccharides. J Bacteriol 1997; 179: 6453--6463], who concluded the same structure as the so-called major lipid A from the H. pylori strain NCTC 11637 but without isolating a homogeneous component. The endotoxic properties and pro-inflammatory cytokine-inducing activities of this novel tetra-acyl type lipid A were lower than those of previously reported major tri-acyl type lipid A.
Asunto(s)
Helicobacter pylori/química , Lípido A/química , Lípido A/fisiología , Animales , Células Cultivadas , Citocinas/metabolismo , Relación Dosis-Respuesta a Droga , Galactosamina/toxicidad , Humanos , Dosificación Letal Mediana , Prueba de Limulus , Activación de Linfocitos , Espectroscopía de Resonancia Magnética , Masculino , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
The antidepressant mianserin is administered as a racemate of the S(+)- and R(-)-enantiomers. Previous in-vitro studies have suggested that CYP2D6 is involved in the stereoselective metabolism of mianserin and its active metabolite, desmethylmianserin. To determine a role for CYP2D6 in vivo, the effects of thioridazine, an inhibitor of CYP2D6, on the steady-state plasma concentrations of the enantiomers of mianserin and desmethylmianserin were examined in 13 depressed Japanese patients. All patients were taking 30 mg of racemic mianserin at bedtime for 8-50 days. Thioridazine (40 mg/day) was coadministered for 1 week, and blood samplings were performed before and after thioridazine coadministration, 12 h after bedtime dosing. Plasma concentrations of the enantiomers of mianserin and desmethylmianserin were measured by HPLC, and the CYP2D6 genotype was determined by allele-specific PCR analysis. Thioridazine significantly increased plasma concentration of S(+)-mianserin (mean SD: 78.2 +/- 35.0 vs. 150.8 +/- 48.7 nM, P < 0.001), but not R(-)-mianserin (39.8 +/- 21.2 vs. 39.5 +/- 20.6 nM, NS). Thioridazine also significantly increased plasma concentrations of both S-desmethylmianserin (11.9 +/- 2.8 vs. 24.4 +/- 10.7 nM, P < 0.01) and R-desmethylmianserin (42.6 +/- 28.4 vs. 115.6 +/- 36.9 nM, P < 0.001). One patient homozygous for the defective allele CYP2D6*5 had the second highest and highest plasma concentrations of S(+)-mianserin and R-desmethylmianserin, respectively, before thioridazine coadministration, and exhibited little increase in plasma concentration of the drugs after thioridazine coadministration. These results suggest that thioridazine specifically inhibits the metabolism of S(+)-mianserin and R-desmethylmianserin, probably through inhibition of CYP2D6, but not R(-)-mianserin.
Asunto(s)
Antidepresivos de Segunda Generación/sangre , Inhibidores del Citocromo P-450 CYP2D6 , Depresión/sangre , Inhibidores Enzimáticos/farmacología , Mianserina/análogos & derivados , Mianserina/sangre , Tioridazina/farmacología , Antidepresivos de Segunda Generación/uso terapéutico , Depresión/tratamiento farmacológico , Femenino , Homocigoto , Humanos , Japón , Masculino , Mianserina/farmacocinética , Mianserina/uso terapéutico , EstereoisomerismoRESUMEN
OBJECTIVE: To assess the possible involvement of CYP3A4 in the metabolism of alprazolam in vivo. METHOD: Twelve healthy male volunteers were randomly allocated to one of the two different treatment sequences, placebo-erythromycin or erythromycin-placebo, with an at least 6-week washout period between the two trial phases. Each volunteer received 400 mg erythromycin or matched placebo given orally three times a day for 10 days and an oral dose (0.8 mg) of alprazolam on the posttreatment day 8. Plasma concentration of alprazolam was measured up to 48 hours after the administration, and psychomotor function was assessed at each time of blood samplings with use of the Digit Symbol Substitution Test, visual analog scale, and Udvalg for kliniske undersøgelser side effect rating scale. RESULTS: Erythromycin significantly (p < 0.001) increased the area under the plasma concentration-time curves (200 +/- 43 versus 322 +/- 49 ng . hr/ml from 0 to 48 hours and 229 +/- 52 versus 566 +/- 161 ng . hr/ml from 0 hour to infinity), decreased the apparent oral clearance (1.02 +/- 0.31 versus 0.41 +/- 0.12 ml/min/kg), and prolonged the elimination half-life (16.0 +/- 4.5 versus 40.3 +/- 14.4 hours) of alprazolam. However, any psychomotor function variables did not differ significantly between the erythromycin and placebo trial phases. CONCLUSION: This study suggests that erythromycin, an inhibitor of CYP3A4, inhibits the metabolism of alprazolam, providing an in vivo evidence for the involvement of CYP3A4 in its metabolism. However, the kinetic change of alprazolam by erythromycin does not result in the pharmacodynamic change of this triazolobenzodiazepine, at least after single dosing.
Asunto(s)
Alprazolam/farmacocinética , Ansiolíticos/farmacocinética , Sistema Enzimático del Citocromo P-450/metabolismo , Eritromicina/farmacología , Oxigenasas de Función Mixta/metabolismo , Inhibidores de la Síntesis de la Proteína/farmacología , Administración Oral , Adulto , Alprazolam/administración & dosificación , Alprazolam/sangre , Alprazolam/orina , Ansiolíticos/sangre , Ansiolíticos/orina , Cromatografía Líquida de Alta Presión , Estudios Cruzados , Citocromo P-450 CYP3A , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Interacciones Farmacológicas , Eritromicina/administración & dosificación , Semivida , Humanos , Masculino , Inhibidores de la Síntesis de la Proteína/administración & dosificación , Estadística como AsuntoRESUMEN
BACKGROUND: The CYP2D6*10 (*10) allele that causes decreased CYP2D6 activity is present in Asians with a high frequency of about 50%. In this study we studied the effects of the *10 allele on the steady-state plasma concentrations (Css) of haloperidol and reduced haloperidol. METHODS: The subjects were 67 Japanese inpatients with schizophrenia who had only the wild-type or *10 alleles. Thirty-four patients were homozygous for the wild-type allele, and 26 were heterozygous and 7 were homozygous for the *10 allele. All patients had been receiving 12 mg/day haloperidol for at least 2 weeks. Plasma concentrations of haloperidol and reduced haloperidol were measured by HPLC. RESULTS: The mean +/- SD values of haloperidol Css in the patients with 0, 1, and 2 *10 alleles were 22.8+/-11.0, 30.1+/-10.6, and 31.2+/-21.2 nmol/L, respectively, and those values for reduced haloperidol were 6.1+/-2.9, 9.5+/-3.7, and 9.9+/-6.2 nmol/L, respectively. The mean haloperidol Css was significantly (P < .05) higher in the patients with 1 *10 allele than in those with no *10 alleles. The mean Css of reduced haloperidol was significantly (P < .05) higher in the patients with 1 and 2 *10 alleles than in those with no *10 alleles. CONCLUSION: This study suggests that the *10 allele plays an important role in controlling the Css of both haloperidol and reduced haloperidol, especially in Asian subjects.