RESUMEN
Objective: To establish an animal model of modified cuff tear arthropathy (mCTA) in order to better replicate the pathophysiology associated with rotator cuff tear-induced humeral head collapse. Design: mCTA was induced by transection of the rotator cuff, the long head of the biceps brachii (LHB), and superior half of the joint capsule in the right shoulder of 12-week-old rats; the left shoulder underwent sham surgery. The severity of CTA was quantitated using the Murine Shoulder Arthritis Score (MSAS). The trabecular bone of the humeral head and metaphysis was analyzed using bone histomorphometry. The expression of proinflammatory cytokines and catabolic enzymes was evaluated immunohistochemically. Results: In the mCTA model, the MSAS increased starting from 2 weeks after induction, and there was notable subchondral bone collapse with fibrous cells at 4 weeks. The mCTA cartilage exhibited positive staining for TNF-α, IL-1ß/6, MMP-3/13, and ADAMTS5. The trabecular bone volume was reduced not only in the subchondral bone but also in the metaphysis of the humeri, and bone resorption was enhanced in these areas. In the collapsed subchondral bone, both bone formation and resorption were increased. The fibrous cells showed expression of TNF-α, IL-6, and MMP-13, along with specific markers of mesenchymal stem cells. Furthermore, the fibrous cells showed osteoblastic characteristics (RUNX2-positive) and expressed RANKL. Conclusions: The LHB and the capsuloligamentous complex are critical stabilizers of the glenohumeral joint, serving to prevent the advancement of CTA following massive rotator cuff tears. Fibrous cells appear to play a role in the humeral head bone resorption.
RESUMEN
Down syndrome (DS) is the most frequent genetic disorder with mental retardation and caused by trisomy 21. Although the gene dosage effect hypothesis has been proposed to explain the impact of extra chromosome 21 on the pathology of DS, a series of evidence that challenge this hypothesis has been reported. The availability of the complete sequences of genes on chromosome 21 serves now as starting point to find functional information of the gene products, but information on gene products is limited so far. We therefore evaluated expression levels of six proteins whose genes are encoded on chromosome 21 (synaptojanin-1, chromosome 21 open reading frame 2, oligomycin sensitivity confering protein, peptide 19, cystatin B and adenosine deaminase RNA-specific 2) in fetal cerebral cortex from DS and controls at 18-19 weeks of gestational age using Western blot analysis. Synaptojanin-1 and C21orf2 were increased in DS, but others were comparable between DS and controls, suggesting that the DS phenotype cannot be simply explained by gene dosage effects. We are systematically quantifying all proteins whose genes are encoded on chromosome 21 in order to provide a better understanding of the pathobiochemistry of DS at the protein level. These studies are of significance as they show for the first time protein levels that are carrying out specific function in human fetal brain with DS.
Asunto(s)
Encéfalo/metabolismo , Cromosomas Humanos Par 21 , Síndrome de Down/genética , Dosificación de Gen , Western Blotting , Encéfalo/embriología , Estudios de Casos y Controles , Femenino , HumanosRESUMEN
Synaptojanin is a highly abundant polyphosphoinositide phosphatase in nerve terminals, and has been thought to play roles in clathrin-mediated synaptic vesicle endocytosis and signaling. In order to determine the broader role of synaptojanin in the central nervous system, we examined synaptojanin expression in the cerebrum and cerebellum from the fetal to the adult period by means of immunohistochemical and Western blot analyses. Immunohistochemistry consistently revealed the localization of synaptojanin in Cajal--Retzius cells, cortical plate neurons, subplate neurons, intermediate neurons, germinal matrix cells and the ventricular neuroepithelium of the fetal cerebrum. In the fetal cerebellum, synaptojanin immunoreactivity was localized in the external granular cell layer, Purkinje cell layer neuropil, cytoplasm of Purkinje cells and internal granular cells. The immunoreactivity in these structures was decreased around birth. After birth, the synaptojanin immunoreactivity of cortical neurons in the cerebrum, Purkinje cell layer neuropil, and internal granular cells and Purkinje cells in the cerebellum increased and reached a plateau after 11 years of age. These results were consistent with the intensity observed on Western blot analysis. These developmental changes of synaptojanin suggest a broader role in not only synaptic vesicle recycling, but also the regulation of neuronal migration and synaptogenesis in the fetal cerebrum and cerebellum.
Asunto(s)
Cerebelo/crecimiento & desarrollo , Cerebelo/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Telencéfalo/crecimiento & desarrollo , Telencéfalo/metabolismo , Adulto , Anciano , Western Blotting , Cerebelo/citología , Niño , Preescolar , Feto/química , Feto/metabolismo , Humanos , Inmunohistoquímica , Lactante , Recién Nacido , Proteínas del Tejido Nervioso/análisis , Neuronas/química , Neuronas/metabolismo , Monoéster Fosfórico Hidrolasas/análisis , Terminales Presinápticos/química , Terminales Presinápticos/metabolismo , Telencéfalo/citologíaRESUMEN
We have characterized a novel Sac domain-containing inositol phosphatase, hSac2. It was ubiquitously expressed but especially abundant in the brain, heart, skeletal muscle, and kidney. Unlike other Sac domain-containing proteins, hSac2 protein exhibited 5-phosphatase activity specific for phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 3,4,5-trisphosphate. This is the first time that the Sac domain has been reported to possess 5-phosphatase activity. Its 5-phosphatase activity for phosphatidylinositol 4,5-bisphosphate (K(m) = 14.3 microm) was comparable with those of Type II 5-phosphatases. These results imply that hSac2 functions as an inositol polyphosphate 5-phosphatase.
Asunto(s)
Fosfatidilinositoles/metabolismo , Fosfoproteínas Fosfatasas/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , ADN Complementario , Humanos , Inositol Polifosfato 5-Fosfatasas , Datos de Secuencia Molecular , Monoéster Fosfórico Hidrolasas/química , Monoéster Fosfórico Hidrolasas/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homología de Secuencia de AminoácidoRESUMEN
We have identified a cDNA encoding a novel inositol polyphosphate 5-phosphatase. It contains two highly conserved catalytic motifs for 5-phosphatase, has a molecular mass of 51 kDa, and is ubiquitously expressed and especially abundant in skeletal muscle, heart, and kidney. We designated this 5-phosphatase as SKIP (Skeletal muscle and Kidney enriched Inositol Phosphatase). SKIP is a simple 5-phosphatase with no other motifs. Baculovirus-expressed recombinant SKIP protein exhibited 5-phosphatase activities toward inositol 1,4,5-trisphosphate, inositol 1,3,4,5-tetrakisphosphate, phosphatidylinositol (PtdIns) 4,5-bisphosphate, and PtdIns 3,4, 5-trisphosphate but has 6-fold more substrate specificity for PtdIns 4,5-bisphosphate (K(m) = 180 microM) than for inositol 1,4, 5-trisphosphate (K(m) = 1.15 mM). The ectopic expression of SKIP protein in COS-7 cells and immunostaining of neuroblastoma N1E-115 cells revealed that SKIP is expressed in cytosol and that loss of actin stress fibers occurs where the SKIP protein is concentrated. These results imply that SKIP plays a negative role in regulating the actin cytoskeleton through hydrolyzing PtdIns 4,5-bisphosphate.
Asunto(s)
Monoéster Fosfórico Hidrolasas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Northern Blotting , Células COS , Clonación Molecular , ADN Complementario/aislamiento & purificación , Humanos , Inositol Polifosfato 5-Fosfatasas , Datos de Secuencia Molecular , Monoéster Fosfórico Hidrolasas/análisis , Monoéster Fosfórico Hidrolasas/metabolismo , Conejos , Especificidad por SustratoRESUMEN
OBJECTIVES: To investigate the effect of menopause on regional and total body lean mass. METHODS: Evaluation of 123 healthy premenopausal women (40.6 +/- 10.8 years) and 123 healthy postmenopausal women (61.8 +/- 7.5 years). All subjects were right side dominant. Regional (head, bilateral arms, trunk, and bilateral legs) and total body lean mass were measured using whole-body scanning by dual-energy X-ray absorptiometry. Baseline characteristics including age, height, weight, and menopausal state were recorded. These variables were compared between pre- and postmenopausal women. In all subjects, correlations between regional or total body lean mass and baseline characteristics were investigated using univariate and multiple regression analyses. RESULTS: Height, and lean mass of the trunk, bilateral legs and total body were significantly lower in postmenopausal women than in premenopausal women, while lean mass of the bilateral arms did not differ between the two groups. On univariate regression analysis, bilateral arms lean mass was positively correlated with height (P < 0.001). Trunk, bilateral legs, and total body lean mass were inversely correlated with age and menopausal state (P < 0.001), but were positively correlated with height (P < 0.001). After adjusting for age and height, trunk lean mass was still correlated with menopausal state (P < 0.01). CONCLUSIONS: Menopause induces lean mass loss, independent of aging and height. Trunk lean mass is more prone to decline with menopause than lean mass of other sites.
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Composición Corporal/fisiología , Índice de Masa Corporal , Menopausia/fisiología , Adulto , Femenino , Humanos , Persona de Mediana Edad , Estándares de Referencia , Valores de ReferenciaRESUMEN
Here, we identify a novel rat phosphatidylinositol-5-phosphate 4-kinase, phosphatidylinositol-phosphate kinase IIgamma (PIPKIIgamma). PIPKIIgamma comprises 420 amino acids with a molecular mass of 47,048 Da, showing greater homology to the type IIalpha and IIbeta isoforms (61.1 and 63.7% amino acid identities, respectively) of phosphatidylinositol-phosphate kinase than to the type I isoforms. It is predominantly expressed in kidney, with low expression in almost all other tissues. PIPKIIgamma was found to have phosphatidylinositol-5-phosphate 4-kinase activity as demonstrated in other type II kinases such as PIPKIIalpha. The PIPKIIgamma that is present endogenously in rat fibroblasts, PC12 cells, and rat whole brain lysate or that is exogenously overexpressed in COS-7 cells shows a doublet migrating pattern on SDS-polyacrylamide gel electrophoresis. Alkaline phosphatase treatment and metabolic labeling in [32P]orthophosphate experiments revealed that PIPKIIgamma is phosphorylated in vivo, resulting in a shift in its electrophoretic mobility. Phosphorylation is induced by treatment of mitogens such as serum and epidermal growth factor. Immunostaining experiments and subcellular fractionation revealed that PIPKIIgamma localizes dominantly in the endoplasmic reticulum (ER). Phosphorylation also occurs in the ER. Thus, PIPKIIgamma may have an important role in the synthesis of phosphatidylinositol bisphosphate in the ER.
Asunto(s)
1-Fosfatidilinositol 4-Quinasa/química , Fosfatidilinositol 4,5-Difosfato/biosíntesis , Fosfatos de Fosfatidilinositol/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/química , Fosfatasa Alcalina/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Fraccionamiento Celular , Células Cultivadas , Clonación Molecular , Retículo Endoplásmico/enzimología , Técnica del Anticuerpo Fluorescente , Riñón/enzimología , Mitógenos/farmacología , Datos de Secuencia Molecular , Fosfoproteínas/química , Fosforilación , Fosfotransferasas/química , ARN Mensajero/metabolismo , Ratas , Análisis de Secuencia de ADN , Homología de Secuencia de AminoácidoRESUMEN
OBJECTIVE: To investigate whether there is evidence of thyroid dysfunction in women with maternity blues. METHODS: Twenty women with maternity blues and 20 age-matched normal controls were enrolled in our study. The serum levels of 6 kinds of thyroid hormones, cortisol, and prolactin (PRL) of the 2 groups were compared, and obstetric variables were recorded for each subject. In addition, significant variables correlating with the development of maternity blues were determined by stepwise regression analysis. RESULTS: The serum-free triiodothyronine (FT3) level at 5 days puerperium was lower in the maternity blues group (p < 0.05) than in the control group. In the maternity blues group, the FT3 level at 5 days puerperium was lower than that at 37 weeks of pregnancy and at 1 month puerperium (p < 0.05). The reverse T3 levels and TSH levels at 5 days postpartum were higher in women with maternity blues than among the controls (p < 0.05). The proportion of primiparous women was higher in the maternity blues group (p < 0.01). Stepwise regression analysis revealed that the low FT3 level and primiparity were significantly correlated with the development of maternity blues (R2 = 0.281, p < 0.001). CONCLUSION: Thyroid dysfunction might be associated with the development of maternity blues.
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Depresión Posparto/etiología , Periodo Posparto/metabolismo , Embarazo/metabolismo , Glándula Tiroides/fisiopatología , Hormonas Tiroideas/sangre , Adulto , Estudios de Cohortes , Depresión Posparto/sangre , Depresión Posparto/psicología , Femenino , Humanos , Hidrocortisona/sangre , Paridad , Periodo Posparto/sangre , Periodo Posparto/psicología , Embarazo/sangre , Embarazo/psicología , Prolactina/sangre , Escalas de Valoración Psiquiátrica , Valores de Referencia , Análisis de Regresión , Estrés Fisiológico/complicaciones , Estrés Fisiológico/etiología , Estrés Fisiológico/fisiopatología , Hormonas Tiroideas/metabolismoAsunto(s)
Fosfatidilinositoles/metabolismo , 1-Fosfatidilinositol 4-Quinasa , Animales , Calcio/metabolismo , Proteínas de Unión al GTP/fisiología , Humanos , Inositol Polifosfato 5-Fosfatasas , Fosfatidilinositol 3-Quinasas , Monoéster Fosfórico Hidrolasas/fisiología , Fosfotransferasas (Aceptor de Grupo Alcohol)/fisiología , Transducción de Señal , Fosfolipasas de Tipo C/fisiologíaRESUMEN
Novel sulfated and nonsulfated oligosialylglycosphingolipids were isolated from sperm of the sea urchin, Hemicentrotus pulcherrimus, and their structures were established as follows: [formula: see text] This provides the first evidence for the natural occurrence of a tetrasialic acid structure in glycosphingolipids. The finding of sulfated oligosialyl chains is especially noteworthy in that the sulfate group exclusively resides on the C-8 of the nonreducing terminal residues of oligo/polysialyl chains and that sulfation appears to be a termination signal for elongation of oligosialyl chains. Sulfation at the nonreducing terminal Neu5Ac residues of oligosialyl chains was also found to facilitate the formation of an inter-residue lactone between the carboxyl group at the nonreducing terminal sulfated Neu5Ac and the hydroxyl group at C-9 of the penultimate Neu5Ac residue. The long chain base was 4-hydroxysphinganine (t18:0) and the major fatty acid species were identified as C20:1, C21:1, and C22:1.
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Glicoesfingolípidos/química , Ácidos Siálicos/análisis , Espermatozoides/química , Animales , Secuencia de Carbohidratos , Cromatografía Líquida de Alta Presión , Cromatografía en Capa Delgada , Glicoesfingolípidos/aislamiento & purificación , Masculino , Metilación , Datos de Secuencia Molecular , Neuraminidasa , Oligosacáridos/química , Oligosacáridos/aislamiento & purificación , Erizos de Mar , Espectrometría de Masa Bombardeada por Átomos VelocesRESUMEN
The effect of 14 types of nonionic surfactants on the permeation of Tenoxicam (TEN) through guinea pigs from propylene glycol solution was investigated in vitro. The flux of TEN was significantly enhanced by polyoxyethylene alkyl ether and polyoxyethylene monoalkyl carboxylate as hydrophilic surfactants, and by polyglyceryl trialkylate, glyceryl monoalkylate, sorbitan monoalkyl carboxylate and sorbitan trialkyl carboxylate as hydrophobic surfactants. In the presence of these surfactants, the flux was increased about 2 to 4 times, as compared to the flux without them. The mechanism of nonionic surfactant action was analyzed and classified by recent methods. the including HLB of surfactants, the solubility of the drugs in vehicle and the hemolysis of erythrocytes. The results suggested that vital factors in the enhancement of skin permeation by nonionic surfactants include the affinity of the surfactants to the stratum corneum, the solubility of the drugs in vehicle and the penetration of surfactants to the stratum corneum.
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Antiinflamatorios no Esteroideos/farmacocinética , Piroxicam/análogos & derivados , Absorción Cutánea/efectos de los fármacos , Tensoactivos/farmacología , Animales , Fenómenos Químicos , Química Física , Eritrocitos/efectos de los fármacos , Cobayas , Hemólisis/efectos de los fármacos , Técnicas In Vitro , Masculino , Piroxicam/farmacocinética , Conejos , Solubilidad , Tensoactivos/químicaRESUMEN
Amylases (I and II) extracellularly produced by an imperfect fungus, Fusidium sp. BX-1 in a medium containing glycerol as a carbon source, were purified as electrophoretically and isoelectrophoretically homogeneous proteins. The electrophoretical mobilities of amylase I and II on native and SDS-polyacrylamide gels exactly coincided with each other. Their molecular weights were estimated to be about 52,000. The sugar contents of amylase I and II were 3.4 and 4.7%, and the pIs were 8.70 and 8.55, respectively. The Kms of the enzymes for soluble starch were 0.053 and 0.044%. The actions of the enzymes on soluble starch, short chain amyloses, and maltose were examined. Amylase I and II are identified as being to an α-amylase and a glucoamylase, respectively.