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BACKGROUND: Higher frequency of Smad4 inactivation or loss of expression is observed in metastasis of colorectal cancer (CRC) leading to unfavourable survival and contributes to chemoresistance. However, the molecular mechanism of how Smad4 regulates chemosensitivity of CRC is unknown. METHODS: We evaluated how the loss of Smad4 in CRC enhanced chemoresistance to 5-fluorouracil (5-FU) using two CRC cell lines in vitro and in vivo. Immunoblotting with cell and tumour lysates and immunohistochemical analyses with tissue microarray were performed. RESULTS: Knockdown or loss of Smad4 induced tumorigenicity, migration, invasion, angiogenesis, metastasis, and 5-FU resistance. Smad4 expression in mouse tumours regulated cell-cycle regulatory proteins leading to Rb phosphorylation. Loss of Smad4 activated Akt pathway that resulted in upregulation of anti-apoptotic proteins, Bcl-2 and Bcl-w, and Survivin. Suppression of phosphatidylinositol-3-kinase (PI3K)/Akt pathway by LY294002 restored chemosensitivity of Smad4-deficient cells to 5-FU. Vascular endothelial growth factor-induced angiogenesis in Smad4-deficient cells might also lead to chemoresistance. Low levels of Smad4 expression in CRC tissues correlated with higher levels of Bcl-2 and Bcl-w and with poor overall survival as observed in immunohistochemical staining of tissue microarrays. CONCLUSION: Loss of Smad4 in CRC patients induces resistance to 5-FU-based therapy through activation of Akt pathway and inhibitors of this pathway may sensitise these patients to 5-FU.
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Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Fluorouracilo/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteína Smad4/genética , Animales , Antimetabolitos Antineoplásicos/farmacología , Proteínas Reguladoras de la Apoptosis , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Supervivencia Celular/genética , Cromonas/farmacología , Neoplasias Colorrectales/genética , Resistencia a Antineoplásicos/genética , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Humanos , Proteínas Inhibidoras de la Apoptosis/biosíntesis , Ratones , Ratones Endogámicos BALB C , Morfolinas/farmacología , Invasividad Neoplásica , Metástasis de la Neoplasia/genética , Trasplante de Neoplasias , Neovascularización Patológica/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Fosforilación , Proteínas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Proteínas Represoras/biosíntesis , Proteína de Retinoblastoma/metabolismo , Proteína Smad4/deficiencia , Survivin , Regulación hacia Arriba , Factor A de Crecimiento Endotelial Vascular/metabolismo , Cicatrización de Heridas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Patients presented with the supraclavicular lymphadenopathy in the medicine department have a strong suspicion of serious illness like tuberculosis, sarcoidosis, toxoplasmosis and malignancy of lymphnode, blood, lung, upper GIT, breast, ovary, testes, and other sites of body. This prospective type of observational study carried out in the indoor and out patient department of medicine of Mymensingh Medical College Hospital over a period of 6 month from April 2011 to September 2011 to diagnose the causes of supraclavicular lymphadenopathy. Patient of either sex, 18 years or above presented with supraclavicular lymphadenopathy were included. Biopsy or FNAC were done. The study showed that mean age of the patient of supraclavicular lymphadenopathy that finally diagnosed as malignant was 49.7 years and that of non malignant was 33.7 years. Male patient have suffered more (60%) from malignant disease than that of female patient (40%). Discrete, hard, non tender either fixed or non fixed supraclavicular lymphadenopathy was found malignant (18 of 18 cases, 100%) and discrete, firm, tender lymphnode were found non malignant (5 of 5 cases, 100%). Increased frequency (11 of 28, 39.3%) of granulomatous inflammation from the tuberculoid lymphadenitis were found among the patient undergone supraclavicular lymphnode biopsy. FNAC result was also of simillar type and finally it was found that frequency of tuberculosis (20 of 53, 37.7%) was highest and bronchial carcima was the second most frequent diagnosis (14 of 53, 26.4%). This study showed that supraclavicular lymphadenopathy is associated mostly with serious disease like tuberculosis and malignancy.
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Enfermedades Linfáticas/etiología , Neoplasias/patología , Adenocarcinoma/secundario , Adulto , Distribución por Edad , Biopsia , Carcinoma Broncogénico/secundario , Carcinoma de Células Escamosas/secundario , Femenino , Hospitales Universitarios , Humanos , Ganglios Linfáticos/patología , Linfadenitis/etiología , Enfermedades Linfáticas/patología , Metástasis Linfática , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Distribución por Sexo , Hombro , Tuberculosis Ganglionar/patologíaRESUMEN
This cross sectional descriptive study was done to find out common clinical presentations, etiologies and laboratory investigation abnormalities in patients of periodic paralysis. Study was carried out in 30 patients with an age range from 8 to 70 years who were enrolled from July 2008 to June 2009 in Mymensingh Medical College Hospital (MMCH) medicine unit. Individuals who were admitted with sudden onset generalized muscle weakness, had history of previous attack and serum potassium level <3mmol/l or >5.5mmol/l were included in this study. In this series, majority of the patients were male (66.67%). Male: female ratio was approximately 2:1. The mean age of the patients was 27.4±4.5 years. Majority (26.67%) of them were in age range of 31-40 years. About 30% of the patients experienced the first attack of paralysis at the age of 20-24 years. Majority of patients (53%) were from middle class family with occupation of private service (26.66%) and farmer (20%). Positive family history was reported in 20% of patients. Regarding the precipitating factors, majority of patients (83.3%) were related to high carbohydrate meal, 56.67% related to temperature, 41.67% to exercise. Flaccid muscle weakness with variables muscle power (MRC grade 4/5 to 2/5 in 60% and 1/5 to 0/5 in 40%) was found. Cerebellar functions, all modalities of sensations and functions of cranial nerves were intact in all patients. In this series, laboratory investigations revealed reduced serum potassium level (<3mmol/l) in 90% of patients. Serum potassium value >5.5mmol/l was found in only 3.33% of patients. Creatine kinase (MM) was raised in 23% of the patients and Thyroid stimulating hormone (TSH) level was 0.8-2mmol/l in 6% of the patients. More than half of the patients (56%) showed variable ECG changes. Impaired nerve conduction function was found in 28.00%. So, careful history taking, meticulous clinical examination and simple laboratory investigations is sufficient to make a prompt diagnosis and rapid management of patients with periodic paralysis.
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Parálisis Periódicas Familiares/diagnóstico , Adolescente , Adulto , Anciano , Niño , Creatina Quinasa/sangre , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana Edad , Parálisis Periódicas Familiares/sangre , Potasio/sangre , Tirotropina/sangre , Adulto JovenRESUMEN
A 15 years old Bangladeshi boy presented with hepatosplenomegaly, anaemia, multiple fractures (symptomatic and asymptomatic) without jaundice was investigated. Laboratory findings revealed leukoerythroblastic blood picture with reduced haemoglobin (7.7 gm/dl). Skeletal survey showed generalized increased bone density, sclerosed medulary space, Rugger-Jersey spine and diploic space filled with dense materials. Overlapping clinical features of both intermediate autosomal recessive and adult autosomal dominant variety of osteopetrosis were found in this patient but diagnosis were made on the basis of typical radiological finding which was mostly consistent with the adult autosomal dominant variety. The patient was treated conservatively and specialist consultation was taken in managing bony abnormalities. This patient was discharged with advised of subsequent follow-up.
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Osteopetrosis/diagnóstico , Adolescente , Humanos , MasculinoRESUMEN
The ß-carboline moiety, substituted at the C1 and C3 benzylic positions with a leaving group, has been demonstrated for the first time as a photoremovable protecting group for time-dependent sequential release of two (same or different) carboxylic acids upon one- and two-photon light irradiation. Density functional theory calculations suggest that the electronic environment of the ß-carboline moiety at C1 and C3 positions plays a key role in the rate of photorelease.
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Understanding SARS-CoV-2 associated immune pathology is crucial to develop pan-effective vaccines and treatments. Here we investigate the immune events from the acute state up to four weeks post SARS-CoV-2 infection, in non-human primates (NHP) with heterogeneous pulmonary pathology. We show a robust migration of CD16 expressing monocytes to the lungs occurring during the acute phase, and we describe two subsets of interstitial macrophages (HLA-DR+CD206-): a transitional CD11c+CD16+ cell population directly associated with IL-6 levels in plasma, and a long-lasting CD11b+CD16+ cell population. Trafficking of monocytes is mediated by TARC (CCL17) and associates with viral load measured in bronchial brushes. We also describe associations between disease outcomes and high levels of cell infiltration in lungs including CD11b+CD16hi macrophages and CD11b+ neutrophils. Accumulation of macrophages is long-lasting and detectable even in animals with mild or no signs of disease. Interestingly, animals with anti-inflammatory responses including high IL-10:IL-6 and kynurenine to tryptophan ratios show less severe illness. Our results unravel cellular mechanisms of COVID-19 and suggest that NHP may be appropriate models to test immune therapies.
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COVID-19/inmunología , Modelos Animales de Enfermedad , Pulmón/inmunología , SARS-CoV-2/inmunología , Enfermedad Aguda , Animales , COVID-19/diagnóstico , COVID-19/patología , COVID-19/virología , Citocinas/metabolismo , Progresión de la Enfermedad , Femenino , Humanos , Pulmón/citología , Pulmón/virología , Macaca mulatta/inmunología , Macaca mulatta/virología , Macrófagos/inmunología , Masculino , Monocitos/inmunología , Monocitos/metabolismo , Neutrófilos/inmunología , Neutrófilos/metabolismo , SARS-CoV-2/aislamiento & purificación , Índice de Severidad de la Enfermedad , Carga Viral/inmunología , Replicación Viral/inmunologíaRESUMEN
We report spin-wave excitations in annular antidot lattice fabricated from 15 nm-thin Ni80Fe20 film. The nanodots of 170 nm diameters are embedded in the 350 nm (diameter) antidot lattice to form the annular antidot lattice, which is arranged in a square lattice with edge-to-edge separation of 120 nm. A strong anisotropy in the spin-wave modes are observed with the change in orientation angle (Ï) of the in-plane bias magnetic field by using Time-resolved Magneto-optic Kerr microscope. A flattened four-fold rotational symmetry, mode hopping and mode conversion leading to mode quenching for three prominent spin-wave modes are observed in this lattice with the variation of the bias field orientation. Micromagnetic simulations enable us to successfully reproduce the measured evolution of frequencies with the orientation of bias magnetic field, as well as to identify the spatial profiles of the modes. The magnetostatic field analysis, suggest the existence of magnetostatic coupling between the dot and antidot in annular antidot sample. Further local excitations of some selective spin-wave modes using numerical simulations showed the anisotropic spin-wave propagation through the lattice.
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In spite of achieving high power conversion efficiency (PCE), organo-halide perovskites suffer from long term stability issues. Especially the grain boundaries of polycrystalline perovskite films are considered as giant trapping sites for photo-generated carriers and therefore play an important role in charge transportation dynamics. Surface engineering via grain boundary modification is the most promising way to resolve this issue. A unique antisolvent-cum-quantum dot (QD) assisted grain boundary modification approach has been employed for creating monolithically grained, pin-hole free perovskite films, wherein the choice of all-inorganic CsPbBr x I3-x (x = 1-2) QDs is significant. The grain boundary filling by QDs facilitates the formation of compact films with 1-2 µm perovskite grains as compared to 300-500 nm grains in the unmodified films. The solar cells fabricated by CsPbBr1.5I1.5 QD modification yield a PCE of â¼16.5% as compared to â¼13% for the unmodified devices. X-ray photoelectron spectral analyses reveal that the sharing of electrons between the PbI6 - framework in the bulk perovskite and Br- ions in CsPbBr1.5I1.5 QDs facilitates the charge transfer process while femtosecond transient absorption spectroscopy (fs-TAS) suggests quicker trap filling and enhanced charge carrier recombination lifetime. Considerable ambient stability up to â¼720 h with <20% PCE degradation firmly establishes the strategic QD modification of bulk perovskite films.
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A one- and two-photon activated photoremovable protecting group (PRPG) was designed based on a carbazole fused o-hydroxycinnamate platform for the dual (same or different) release of alcohols. The mechanism for the dual release proceeds through a stepwise pathway and also monitors the first and second photorelease in real time by an increase in fluorescence intensity and color change, respectively. Further, its application in staining live neurons and ex vivo imaging with two-photon excitation is shown.
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The retinoblastoma-related protein p107 has been shown to be a regulator of the transcription factor E2F. p107 associates with E2F via its pocket region and represses E2F-dependent transcription. In this study, we provide evidence for a novel interaction between p107 and the transcription factor Sp1. We show that p107 can be found endogenously associated with Sp1 in the extracts of several different cell lines. Moreover, in transient transfection assays, expression of p107 represses Sp1-dependent transcription. This repression of Sp1-dependent transcription does not require the DNA-binding domain of Sp1. Transcription driven by a chimeric protein containing the Ga14 DNA-binding domain and the Sp1 activation domains is inhibited by p107. Interestingly, unlike the repression of E2F-dependent transcription, the repression of Sp1-dependent transcription does not depend on an intact pocket region. We show that distinct regions of p107 are involved in the control of Sp1 and E2F.
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Proteínas Portadoras , Proteínas de Ciclo Celular , Proteínas de Unión al ADN , Proteínas Nucleares/metabolismo , Proteínas Represoras/metabolismo , Factor de Transcripción Sp1/metabolismo , Factores de Transcripción/fisiología , Animales , Anticuerpos Monoclonales , Secuencia de Bases , Reacciones Cruzadas , ADN/metabolismo , Factores de Transcripción E2F , Células L , Ratones , Datos de Secuencia Molecular , Proteínas Nucleares/genética , Fragmentos de Péptidos , Pruebas de Precipitina , Regiones Promotoras Genéticas/genética , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/metabolismo , Proteína 1 de Unión a Retinoblastoma , Proteína p107 Similar a la del Retinoblastoma , Factor de Transcripción Sp1/genética , Factor de Transcripción DP1 , Transcripción Genética/fisiologíaRESUMEN
Smad proteins play a key role in the intracellular signaling of the transforming growth factor beta (TGF-beta) superfamily of extracellular polypeptides that initiate signaling from the cell surface through serine/threonine kinase receptors. A subclass of Smad proteins, including Smad6 and Smad7, has been shown to function as intracellular antagonists of TGF-beta family signaling. We have previously reported the identification of a WD40 repeat protein, STRAP, that associates with both type I and type II TGF-beta receptors and that is involved in TGF-beta signaling. Here we demonstrate that STRAP synergizes specifically with Smad7, but not with Smad6, in the inhibition of TGF-beta-induced transcriptional responses. STRAP does not show cooperation with a C-terminal deletion mutant of Smad7 that does not bind with the receptor and consequently has no inhibitory activity. STRAP associates stably with Smad7, but not with the Smad7 mutant. STRAP recruits Smad7 to the activated type I receptor and forms a complex. Moreover, STRAP stabilizes the association between Smad7 and the activated receptor, thus assisting Smad7 in preventing Smad2 and Smad3 access to the receptor. STRAP interacts with Smad2 and Smad3 but does not cooperate functionally with these Smads to transactivate TGF-beta-dependent transcription. The C terminus of STRAP is required for its phosphorylation in vivo, which is dependent on the TGF-beta receptor kinases. Thus, we describe a mechanism to explain how STRAP and Smad7 function synergistically to block TGF-beta-induced transcriptional activation.
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Proteínas de Unión al ADN/metabolismo , Proteínas , Transducción de Señal , Transactivadores/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Células COS , Línea Celular , Humanos , Immunoblotting , Fosforilación , Plásmidos , Pruebas de Precipitina , Unión Proteica , Proteínas de Unión al ARN , Receptores de Factores de Crecimiento/metabolismo , Proteína Smad2 , Proteína smad3 , Proteína smad6 , Proteína smad7 , Transcripción Genética , Activación TranscripcionalRESUMEN
The complement system is a central player of the innate immune system. Activation of the complement system protects the host against pathogens. However, uncontrolled synthesis can be detrimental to host. This concise review summarizes the current understanding of the mechanism(s) of complement activation, the mechanism of C3 regulation, and the role of complement in human immunodeficiency virus (HIV) pathogenesis with emphasis on the cross-talk between HIV and complement system in NeuroAIDS and HIV-associated nephropathy (HIVAN).
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Proteínas del Sistema Complemento/fisiología , Infecciones por VIH/inmunología , Complejo SIDA Demencia/inmunología , Nefropatía Asociada a SIDA/inmunología , Terapia Antirretroviral Altamente Activa , Quimiotaxis , Activación de Complemento , VIH/patogenicidad , VIH/fisiología , Infecciones por VIH/tratamiento farmacológico , Humanos , Receptores de Complemento/fisiología , Replicación ViralRESUMEN
We have previously demonstrated that the core promoter of rat ribosomal RNA gene (rDNA) contains an E-box-like sequence to which the core promoter binding factor CPBF binds and that the 44 kDa subunit of this protein is immunologically related to USF1, the helix--loop--helix-zipper DNA binding protein. Further, we showed that RNA polymerase I (pol I) transcription in vitro is competed by oligonucleotides containing USF-binding site, which suggested a key role for USF in rDNA transcription. To prove the potential role of USF in pol I transcription in vivo, USF1 and USF2 homodimers and USF1/USF2 heterodimer were overexpressed in CHO cells by transfection of the respective cDNAs. Co-transfection of a plasmid containing rDNA followed by primer extension analysis showed that overexpression of USF1 and USF2 as homodimers resulted in inhibition of rDNA transcription by as much as 85-90% whereas overexpression of USF1/USF2 in the heterodimeric form activated transcription approximately 3.5-fold. Transfection of mutant USF2 cDNA that is devoid of the basic DNA-binding domain produced only minimal inhibition of rDNA transcription. These data show that USF can modulate transcription of rRNA gene in vivo by functioning as a repressor (homodimer) or activator (heterodimer) of pol I transcription in vivo and suggest that inhibition of rDNA transcription may be responsible for the antiproliferative action of USF homodimers.
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ADN Ribosómico/biosíntesis , ADN Ribosómico/metabolismo , Proteínas de Unión al ADN , Secuencias Hélice-Asa-Hélice , Factores de Transcripción/metabolismo , Transcripción Genética , Animales , Células CHO , Cricetinae , Dimerización , Genes Reporteros , Ratas , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/metabolismo , Factores de Transcripción/biosíntesis , Factores de Transcripción/química , Transfección , Factores Estimuladores hacia 5' , beta-Galactosidasa/biosíntesisRESUMEN
The erythroagglutinating activity of purified Vicia faba lectin was enhanced in the presence of gums; gum guar caused the highest enhancement. Circular dichroism probe demonstrated 40-57% beta-conformation and 4-23% alpha-conformation of the lectin at pH 7.2 depending upon the analytical methods used. The beta-conformations of untreated and modified V. faba lectins were increased in the presence of gums. The mixing of gum guar with lectin and with modified lectin, respectively, led to the highest values of beta-conformational change in the protein molecule, thereby increasing the number of receptor sites of the lectin molecule. The enhancement of the activity of V. faba lectin in the presence of gum guar might be due to the conformational change of the protein molecule.
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Goma Arábiga/farmacología , Hemaglutinación/efectos de los fármacos , Lectinas/farmacología , Lectinas de Plantas , Polisacáridos/farmacología , Tragacanto/farmacología , Animales , Dicroismo Circular , Técnicas In Vitro , Conformación Proteica/efectos de los fármacos , ConejosRESUMEN
Increased expression of cyclooxygenase-2 (COX-2) expression has been observed in several human tumor types and in selected animal and cell culture models of carcinogenesis, including lung cancer. Increased expression of COX-2 and production of prostaglandins appear to provide a survival advantage to transformed cells through the inhibition of apoptosis, increased attachment to extracellular matrix, increased invasiveness, and the stimulation of angiogenesis. In the present studies, we found that transforming growth factor beta1 (TGF-beta1) and epidermal growth factor (EGF) synergistically induced the expression of COX-2 and prostaglandin E2 (PGE2) production in mink lung epithelial (Mv1Lu) cells. EGF, but not PDGF or IGF-1, was able to inhibit TGF-beta1-induced apoptosis in Mv1Lu cells and this effect was blocked by NS-398, a selective inhibitor of COX-2 activity, suggesting a possible role for COX-2 in the anti-apoptotic effect of EGF receptor ligands. The combination of TGF-beta1 and EGF also significantly induced COX-2 expression in rat intestinal epithelial (RIE-1) cells and completely prevented sodium butyrate (NaBu)-induced apoptosis. The synergistic induction of COX-2 by TGF-beta1 and EGF was not observed in R1B-L17 cells, a line derived from Mv1Lu cells that lacks the TGF-beta type-I receptor. AG1478, a selective inhibitor of EGF receptor tyrosine kinase activity, completely suppressed the induction of COX-2 expression by either EGF or TGF-beta1+EGF. Also, PD98059, a specific inhibitor of MEK/ERK pathway, and SB203580, a specific inhibitor of p38 MAPK activity, significantly inhibited the induction of COX-2 in response to combined EGF and TGF-beta1. These results suggest an important collaborative interaction of TGF-beta1 and EGF signaling in the induction of COX-2 and prostaglandin production in Mv1Lu cells.
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Apoptosis/efectos de los fármacos , Factor de Crecimiento Epidérmico/farmacología , Isoenzimas/biosíntesis , Prostaglandina-Endoperóxido Sintasas/biosíntesis , Factor de Crecimiento Transformador beta/farmacología , Animales , Células Cultivadas , Ciclooxigenasa 2 , Dinoprostona/biosíntesis , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Inducción Enzimática/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Humanos , Proteínas de la Membrana , Visón , Proteínas Quinasas Activadas por Mitógenos/fisiología , Ratas , Factores de TiempoRESUMEN
Increased Cdk4 expression occurs coincident with over-expression of cyclin D1 in many human tumours and tumourigenic mouse models. Here, we investigate both in vivo and in vitro the mechanism by which Cdk4 expression is regulated in the context of cyclin D1 over-expression. Cdk4 mRNA levels in cyclin D1-over-expressing tissue and cultured cells were unchanged compared with controls. In contrast, Cdk4 protein levels were increased in cyclin D1-over-expressing tissue and cells versus their respective controls. This increase was not due to altered protein stability, but appeared to be due to an increase in Cdk4 protein synthesis. We also performed immunoprecipitation and in vitro kinase assays to demonstrate an increase in cyclin D1-Cdk4 complex formation and associated kinase activity. Blocking cyclin D1 expression resulted in diminished Cdk4 protein but not mRNA levels. These findings suggest a mechanism by which Cdk4 expression is increased in the context of cyclin D1 over-expression during tumourigenesis.
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Ciclina D1/metabolismo , Quinasas Ciclina-Dependientes/biosíntesis , Regulación Enzimológica de la Expresión Génica , Proteínas Proto-Oncogénicas , Animales , Línea Celular Transformada , Quinasa 4 Dependiente de la Ciclina , Hepatocitos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , ARN Mensajero/metabolismo , Transducción GenéticaRESUMEN
An erythroagglutinin from the hemolymph of the scorpion, Heterometrus bengalensis, has been purified by gel filtration and ion-exchange chromatography. Its homogeneity has been demonstrated by polyacrylamide gel electrophoresis. The purified agglutinin appears to be a monomeric protein having a possible molecular weight between 146,000 and 148,000. It has no divalent cation requirement for erythroagglutination. The erythroagglutination is not inhibited by saccharides, glycoproteins and mucin. Identical erythroagglutination pattern is obtained with normal as well as neuraminidase treated erythrocytes.
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Hemaglutininas/aislamiento & purificación , Hemolinfa/análisis , Escorpiones , Animales , Carbohidratos/análisis , Cationes Bivalentes , Hemaglutinación , Hemaglutininas/toxicidad , Peso Molecular , Neuraminidasa , ConejosRESUMEN
A protease inhibitor has been purified by ultracentrifugation, affinity chromatography on trypsin-sepharose 4B, and chromatofocusing on PBE-94 from hemolymph of the scorpion Heterometrus bengalensis. Homogeneity of the protease inhibitor was demonstrated by high performance liquid chromatography (HPLC). The protease inhibitor is a monomeric glycoprotein with a molecular weight of 120,000 dalton, which is stable between pH 4 and pH 8. The molecule inhibits serine proteases like trypsin and alpha-chymotrypsin and shows a noncompetitive mode of inhibition towards trypsin, with a Ki value of 6.1 x 10(-6) mM. Amino acid analysis shows a preponderance of aspartic acid, glutamic acid, serine, and glycine. The protease inhibitor is efficient in inhibiting phenoloxidase activity in both the hemolymph and the isolated phenoloxidase. Melanin synthesis by phenoloxidase may be influenced by this protease inhibitor.
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Glicoproteínas/sangre , Hemolinfa/química , Escorpiones/metabolismo , Inhibidores de Serina Proteinasa/sangre , Aminoácidos/análisis , Animales , Quimotripsina/antagonistas & inhibidores , Glicoproteínas/aislamiento & purificación , Glicoproteínas/farmacología , Melaninas/biosíntesis , Peso Molecular , Monofenol Monooxigenasa/antagonistas & inhibidores , Inhibidores de Serina Proteinasa/aislamiento & purificación , Inhibidores de Tripsina/sangre , Inhibidores de Tripsina/aislamiento & purificaciónRESUMEN
Acetylcholinesterase (AchE: EC 3.1.1.7) was identified and purified from the hemolymph of the scorpion Heterometrus bengalensis. The purity of the enzyme was determined by polyacrylamide gel electrophoresis (PAGE). The molecular weight of the enzyme, determined by sodium dodecyl sulfate-PAGE, was 80,000. The purified AchE hydrolysed acetylthiocholine iodide, but it did not react with butyrylthiocholine iodide. BW284C51, a specific inhibitor of AchE, strongly inhibited the enzyme. The known inhibitor (tetramonoisopropylpyrophosphortetramide) of pseudocholinesterase did not produce any inhibition of the enzyme activity. The purified AchE of scorpion hemolymph was vulnerable to high substrate concentration. The presence of Cu2+ and Ni2+ reduced the enzyme activity, whereas the metal ion, Sn2+, enhanced AchE activity. Ca2+ produced neither inhibition nor activation. (Na+, K+)-ATPase and Mg2+-ATPase activities were greatly enhanced by the purified AchE.
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Acetilcolinesterasa/aislamiento & purificación , Hemolinfa/enzimología , Acetiltiocolina/metabolismo , Adenosina Trifosfatasas/metabolismo , Animales , Bencenamina, 4,4'-(3-oxo-1,5-pentanodiil)bis(N,N-dimetil-N-2-propenil-), Dibromuro , Butiriltiocolina/metabolismo , Cobre/farmacología , Electroforesis en Gel de Poliacrilamida , Cinética , Peso Molecular , Níquel/farmacología , EscorpionesRESUMEN
Datta, P. K., Moulder, J. E., Fish, B. L., Cohen, E. P. and Lianos, E. A. Induction of Heme Oxygenase 1 in Radiation Nephropathy: Role of Angiotensin II. Radiat. Res. 155, 734-739 (2001). In a rat model of radiation-induced nephropathy, we investigated changes in expression of heme oxygenase 1 (Hmox1, also known as HO-1), an enzyme that catalyzes conversion of heme into biliverdin, carbon monoxide and iron. The study explored whether radiation induces Hmox1 expression in the irradiated kidney and whether angiotensin II (AII) mediates Hmox1 expression in glomeruli isolated from irradiated kidneys. To assess the effects of radiation on Hmox1 expression, rats received 20 Gy bilateral renal irradiation and were randomized to groups receiving an AII type 1 (AT(1)) receptor antagonist (L-158,809) or no treatment. Drug treatment began 9 days prior to bilateral renal irradiation and continued for the duration of the study. Estimation of Hmox1 levels in glomerular protein lysates assessed by Western blot analysis revealed a significant increase in Hmox1 protein at 50 and 65 days postirradiation. In animals treated with the AT(1) receptor antagonist, there was no induction of Hmox1, suggesting that AII may be a mediator of Hmox1 induction. To confirm that AII stimulates Hmox1 expression, animals were infused with 200, 400 or 800 ng/kg min(-1) of AII for 18-19 days, and Hmox1 protein levels in glomeruli were assessed. There was a significant induction of Hmox1 in glomeruli of animals infused with 800 ng/kg min(-1) of AII. These studies demonstrate that glomerular Hmox1 expression is elevated in the middle phase of radiation nephropathy and that AII can increase glomerular Hmox1 levels.