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1.
Semergen ; 47(8): 508-514, 2021.
Article in Spanish | MEDLINE | ID: mdl-34531125

ABSTRACT

OBJECTIVE: COVID-19 infection requires early diagnosis, with PCR being the gold standard test. The protocols advocate the use of rapid antigenic tests that require evaluation in actual clinical practice. The objective was to evaluate the diagnostic test for rapid antigen detection, Panbio Covid rapid test, compared with PCR, in patients with symptoms of 5 or less days of evolution and with a high-suspicion of infection by COVID-19 in a health center. MATERIALS AND METHODS: 103 patients over 14 years of age who attended an urban health center located in the Usera District of Madrid, with high-suspicion of COVID-19 infection, in the first 5 days of evolution from the onset of symptoms during the month of November 2020. INTERVENTIONS: diagnostic tests for COVID-19 are performed: antigen and PCR. RESULTS: The prevalence of the disease was 24.3% according to the PCR test and 17.5% according to the rapid antigenic test. The sensitivity was 72% (95% CI: 54.3-89.6%). The specificity was 100%. The positive and negative predictive values were 100% and 91.8% respectively. In the bivariate analysis, there was no relationship between symptoms and the presence of disease, except for myalgias (p=0.030). The multivariate analysis found a relationship between cough, dyspnea, fever, myalgia, anosmia/ageusia, and ocular symptoms and the presence of disease. CONCLUSIONS: The sensitivity and specificity for the Panbio rapid antigen test are similar to other studies performed in primary care. In high-prevalence of disease and with highly suspected symptoms, positive test results can be considered definitive, but negative results will require confirmation. Myalgia, fever, dyspnea, anosmia/ageusia, and ocular symptoms may be more related to the presence of COVID-19.


Subject(s)
COVID-19 , Diagnostic Tests, Routine , Humans , Primary Health Care , SARS-CoV-2 , Sensitivity and Specificity
2.
Med. intensiva (Madr., Ed. impr.) ; 44(9): 566-576, dic. 2020. tab
Article in Spanish | IBECS | ID: ibc-189899

ABSTRACT

La pandemia por SARS-CoV-2 ha generado nuevos escenarios que requieren modificaciones de los protocolos habituales de reanimación cardiopulmonar. Las guías clínicas vigentes sobre el manejo de la parada cardiorrespiratoria no incluyen recomendaciones para situaciones aplicables a este contexto. Por ello, el Plan Nacional de Reanimación Cardiopulmonar de la Sociedad Española de Medicina Intensiva, Crítica y Unidades Coronarias, en colaboración con el Grupo Español de RCP Pediátrica y Neonatal y con el programa de Enseñanza de Soporte Vital en Atención Primaria de la Sociedad Española de Medicina Familiar y Comunitaria, ha redactado las siguientes recomendaciones, que están divididas en 5 partes que tratan los principales aspectos para cada entorno asistencial. En este artículo se presenta un resumen ejecutivo de las mismas


The SARS-CoV-2 pandemic has created new scenarios that require modifications to the usual cardiopulmonary resuscitation protocols. The current clinical guidelines on the management of cardiorespiratory arrest do not include recommendations for situations that apply to this context. Therefore, the National Cardiopulmonary Resuscitation Plan of the Spanish Society of Intensive and Critical Care Medicine and Coronary Units (SEMICYUC), in collaboration with the Spanish Group of Pediatric and Neonatal CPR and with the Teaching Life Support in Primary Care program of the Spanish Society of Family and Community Medicine (SEMFyC), have written these recommendations, which are divided into 5 parts that address the main aspects for each healthcare setting. This article consists of an executive summary of them


Subject(s)
Humans , Coronavirus Infections/diagnosis , Cardiopulmonary Resuscitation/standards , Cardiopulmonary Resuscitation/instrumentation , Heart Arrest/complications , Polymerase Chain Reaction , Societies, Medical/standards , Patient Safety , Respiratory Tract Infections/prevention & control , Respiratory Tract Infections/transmission
3.
Med Intensiva (Engl Ed) ; 44(9): 566-576, 2020 Dec.
Article in Spanish | MEDLINE | ID: mdl-32425289

ABSTRACT

The SARS-CoV-2 pandemic has created new scenarios that require modifications to the usual cardiopulmonary resuscitation protocols. The current clinical guidelines on the management of cardiorespiratory arrest do not include recommendations for situations that apply to this context. Therefore, the National Cardiopulmonary Resuscitation Plan of the Spanish Society of Intensive and Critical Care Medicine and Coronary Units (SEMICYUC), in collaboration with the Spanish Group of Pediatric and Neonatal CPR and with the Teaching Life Support in Primary Care program of the Spanish Society of Family and Community Medicine (SEMFyC), have written these recommendations, which are divided into 5 parts that address the main aspects for each healthcare setting. This article consists of an executive summary of them.


Subject(s)
COVID-19/complications , Cardiopulmonary Resuscitation/standards , SARS-CoV-2 , Adult , Advanced Cardiac Life Support/methods , Advanced Cardiac Life Support/standards , Age Factors , Airway Management/methods , Airway Management/standards , COVID-19/epidemiology , COVID-19/prevention & control , COVID-19/transmission , Cardiopulmonary Resuscitation/methods , Child , Disease Progression , Electric Countershock , Heart Arrest/therapy , Humans , Pandemics , Patient Positioning/methods , Personal Protective Equipment , Protective Clothing , Societies, Medical , Spain
4.
J Chemother ; 21(5): 550-7, 2009 Nov.
Article in English | MEDLINE | ID: mdl-19933047

ABSTRACT

Plitidepsin (Aplidin) is a novel antitumor agent, derived from the mediterranean tunicate Aplidium albicans, and is currently in phase ii clinical trials with evidence of activity in heavily pretreated multiple myeloma, renal cell carcinoma, melanoma and neuroblastoma patients. As compared to its parental compound didemnin B, plitidepsin has shown a better therapeutic index with less bone marrow toxicity, cardiotoxicity and neurotoxicity in patients and a more potent cytotoxic effect in several tumor cell lines. As sensitivity to the drug varies between cell lines and fresh leukemia samples, we performed studies on transport of plitidepsin in leukemia and lymphoma cell lines to determine the mechanism of uptake. The drug is taken up by an active transport process, i.e. the process is temperature and energy dependent, and has a high-affinity binding site with Kt =212 nM and Vmax = 15 pmoles/min. Importantly, once inside the cell, efflux of plitidepsin is minimum, suggesting that the drug is bound to intracellular macromolecules. Further work showed that plitidepsin binds to G-Protein Coupled Receptors (GPCRs), since GPCR and GRK (GPCR kinases) inhibitors suramin and heparin respectively, markedly reduce the drug uptake and its cytotoxic activity. Signaling via Jak/Stat pathway is inhibited by pharmacological concentrations of plitidepsin, further confirming the relationship between plitidepsin and GPCRs.


Subject(s)
Depsipeptides/metabolism , Heparin/pharmacology , Membrane Microdomains/metabolism , Receptors, G-Protein-Coupled/antagonists & inhibitors , Suramin/pharmacology , 4-Aminopyridine/pharmacology , Adenosine Triphosphate/metabolism , Antimetabolites, Antineoplastic/pharmacology , Biological Transport, Active , Cell Proliferation , Cytarabine/pharmacology , Humans , JNK Mitogen-Activated Protein Kinases/antagonists & inhibitors , JNK Mitogen-Activated Protein Kinases/metabolism , Peptides, Cyclic , Potassium Channel Blockers/pharmacology , Potassium Channels/drug effects , Receptors, G-Protein-Coupled/metabolism , STAT Transcription Factors/antagonists & inhibitors , STAT Transcription Factors/metabolism , Subcellular Fractions , Time Factors , Tumor Cells, Cultured/metabolism
5.
Leukemia ; 21(12): 2399-405, 2007 Dec.
Article in English | MEDLINE | ID: mdl-17713546

ABSTRACT

Aplidin (plitidepsin) is a novel marine-derived antitumor agent presently undergoing phase II clinical trials in hematological malignancies and solid tumors. Lack of bone marrow toxicity has encouraged further development of this drug for treatment of leukemia and lymphoma. Multiple signaling pathways have been shown to be involved in Aplidin-induced apoptosis and cell cycle arrest in G1 and G2 phase. However, the exact mechanism(s) of Aplidin action remains to be elucidated. Here we demonstrate that mitochondria-associated or -localized processes are the potential cellular targets of Aplidin. Whole genome gene-expression profiling (GEP) revealed that fatty acid metabolism, sterol biosynthesis and energy metabolism, including the tricarboxylic acid cycle and ATP synthesis are affected by Aplidin treatment. Moreover, mutant MOLT-4, human leukemia cells lacking functional mitochondria, were found to be resistant to Aplidin. Cytosine arabinoside (araC), which also generates oxidative stress but does not affect the ATP pool, showed synergism with Aplidin in our leukemia and lymphoma models in vitro and in vivo. These studies provide new insights into the mechanism of action of Aplidin. The efficacy of the combination of Aplidin and araC is currently being evaluated in clinical phase I/II program for the treatment of patients with relapsed leukemia and high-grade lymphoma.


Subject(s)
Antineoplastic Agents/pharmacology , Cytarabine/pharmacology , Depsipeptides/pharmacology , Mitochondria/drug effects , Adenosine Triphosphate/biosynthesis , Animals , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Cell Cycle/drug effects , Cell Line, Tumor/drug effects , Cell Line, Tumor/transplantation , Cytarabine/administration & dosage , Depsipeptides/administration & dosage , Doxorubicin/pharmacology , Drug Screening Assays, Antitumor , Drug Synergism , Gene Expression Profiling , Gene Expression Regulation, Leukemic/drug effects , Humans , K562 Cells/drug effects , Leukemia-Lymphoma, Adult T-Cell/drug therapy , Leukemia-Lymphoma, Adult T-Cell/pathology , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/pathology , Methylprednisolone/pharmacology , Mice , Mice, SCID , Mitochondria/physiology , Mitoxantrone/pharmacology , Oxidative Stress/drug effects , Peptides, Cyclic , Specific Pathogen-Free Organisms , Xenograft Model Antitumor Assays
8.
Rev Neurol ; 35(10): 994-7, 2002.
Article in Spanish | MEDLINE | ID: mdl-12436405

ABSTRACT

In vitro studies have shown that the cerebrospinal fluid of patients with multiple sclerosis transports compounds which may affect the function or viability of central nervous system cell function. The presence of nerve ion channel blockers and other molecules, as yet unidentified, which may cause the death of nerve cells or oligodendrocytes has been shown, although their relevance and clinical correlation is still not clear. If their usefulness is proved, these methods may be useful in the search for inhibitors of the noxious effects of the substances mentioned.


Subject(s)
Apoptosis , Multiple Sclerosis/cerebrospinal fluid , Neurons/physiology , Retrograde Degeneration , Humans , Ion Channels/physiology , Neurons/cytology
9.
Rev. neurol. (Ed. impr.) ; 35(10): 994-997, 16 nov., 2002.
Article in Es | IBECS | ID: ibc-22329

ABSTRACT

Los estudios in vitro han permitido demostrar que el líquido cefalorraquídeo de los pacientes con esclerosis múltiple transporta compuestos que pueden modificar la funcionalidad de células del sistema nervioso central en cultivo o afectar su viabilidad. Así, se ha comprobado la existencia de bloqueadores de los canales iónicos neuronales y de otras moléculas, aún por identificar, que pueden originar muerte celular, bien de los oligodendrocitos o de las neuronas. Está por aclarar la relevancia de estos hallazgos y su correlación clínica. Si se comprueba su utilidad, estos métodos podrían emplearse en la búsqueda de inhibidores de los efectos nocivos de los compuestos comentados (AU)


Subject(s)
Humans , Apoptosis , Retrograde Degeneration , Multiple Sclerosis , Neurons , Ion Channels
10.
Aten Primaria ; 25(3): 172-5, 2000 Feb 28.
Article in Spanish | MEDLINE | ID: mdl-10730441

ABSTRACT

OBJECTIVE: To analyse the causes of overuse of hospital emergency services (HES). DESIGN: Cross sectional, descriptive study. SETTING: Emergency service at a general hospital. PATIENTS: Patients who attended the HES on their own initiative. MEASUREMENTS AND MAIN RESULTS: Telephone poll to a representative sample of patients attending on their own initiative the emergency department of the 12 de Octubre Hospital in Madrid between October 5th and 12th. Average age: 46.95 (SD, 20.81); 52.2% women and 47.8% men, 50% were ignorant of the existence of primary care emergencies. 77% were ignorant of the existence of ongoing care points. Main motives for attendance at HES were: ignorance of non-hospital emergency services (32%), better technical means (25.6%), quicker care (21%), sensation of vital urgency (11.4%), poor quality of care in PC (8%). The care received at HES was evaluated as positive in 90% of cases, though 33% thought the information provided insufficient, and 34% the waiting-time excessive. Although 40% believed afterwards that their problem could have been resolved in PC, as many as 75% would return to the hospital. CONCLUSIONS: Widespread ignorance of the existence of non-hospital emergency services affects the over-use of HES. Most users use the HES as a service of immediate PC, a rapid way of obtaining health care. Users have a very favourable opinion of HES care, which is not stated in the case of non-hospital emergency services. In order to improve use of the HES, the population needs to receive better health education.


Subject(s)
Emergency Service, Hospital/statistics & numerical data , Cross-Sectional Studies , Female , Humans , Interviews as Topic/methods , Male , Middle Aged , Primary Health Care/statistics & numerical data , Spain , Surveys and Questionnaires
11.
Aten. prim. (Barc., Ed. impr.) ; 25(3): 172-175, feb. 2000.
Article in Es | IBECS | ID: ibc-4074

ABSTRACT

Objetivo. Analizar las causas de sobreutilización de los servicios de urgencias hospitalarios (SUH). Diseño. Estudio descriptivo transversal. Emplazamiento. Servicio de urgencias de un hospital general. Pacientes. Pacientes que acudieron al SUH por iniciativa propia. Mediciones y resultados principales. Encuesta telefónica a una muestra representativa de pacientes que acudieron a urgencias del Hospital 12 de Octubre de Madrid del 5 al 12 de octubre de 1998 por iniciativa propia. Edad media, 46,95 años (DE, 20,81), un 52,2 por ciento mujeres y el 47,8 por ciento varones. Un 50 por ciento desconoce la urgencia en atención primaria (AP), el 77 por ciento desconoce la existencia de puntos de atención continuada. Principales motivos por los que acuden al SUH: desconocimiento de urgencias extrahospitalarias (SUEH) (32 por ciento), empleo de mejores medios técnicos (25,6 por ciento), asistencia más rápida (21 por ciento), sensación de urgencia vital (11,4 por ciento), mala calidad asistencial en AP (8 por ciento). La valoración de la atención recibida en SUH es positiva en un 90 por ciento, el 33 por ciento considera insuficiente la información facilitada y un 34 por ciento el tiempo de espera excesivo. Aunque un 40 por ciento cree a posteriori que su problema era solucionable en AP, hasta el 75 por ciento retornaría al hospital. Conclusiones. El gran desconocimiento del SUEH influye en la sobreutilización de los SUH. La mayoría de los usuarios utiliza los SUH como servicio de AP inmediata, siendo una forma rápida de conseguir atención. La opinión de los usuarios sobre la asistencia en un SUH es muy favorable, hecho que no se objetiva con los SUHE. Debería mejorarse la educación sanitaria de la población para mejorar el uso de los SUH (AU)


Subject(s)
Middle Aged , Child, Preschool , Child , Male , Infant , Female , Humans , Disease Outbreaks , Spain , Urban Health , Erythema Infectiosum , Surveys and Questionnaires , Primary Health Care , Cross-Sectional Studies , Hospitals, Urban , Interviews as Topic , Emergency Service, Hospital
12.
Cir. Esp. (Ed. impr.) ; 67(1): 14-17, ene. 2000. graf
Article in Es | IBECS | ID: ibc-3687

ABSTRACT

Introducción. La pancreatitis postoperatoria es una complicación temida tras la intervención quirúrgica sobre la región biliopancreática. Entre los mecanismos patogénicos de la pancreatitis sigue especulándose alrededor de los secretagogos y, en concreto, sobre el papel de las hormonas estimuladoras de la secreción pancreática. Pacientes y método. Se realizó un estudio prospectivo en pacientes que iban a ser sometidos a cirugía biliar electiva por procesos benignos, determinándose la gastrina y secretina en el preoperatorio, a las 24 y 48 h del postoperatorio por técnicas de radioinmunoanálisis. Resultados. Quedaron incluidos 98 pacientes, 72 mujeres y 26 varones, con una edad media de 59 años. El perfil de la gastrinemia fue de 177 pg/ml en el preoperatorio, 186 pg/ml a las 24 h del postoperatorio y 172 pg/ml a las 48 h de éste; el perfil de la secretinemia proporcionó esta secuencia: 87, 86 y 84 pg/ml. Conclusiones. La cirugía sobre la vía biliar no modifica significativamente los valores hormonales de gastrina y secretina; aunque su liberación se sigue produciendo en concentraciones similares a las preoperatorias (AU)


Subject(s)
Adult , Aged , Female , Male , Middle Aged , Humans , Liver/surgery , Liver , Gastrins , Secretin , Bile Duct Diseases/surgery , Gastrointestinal Hormones , Biliary Tract Surgical Procedures , Receptors, Gastrointestinal Hormone , Prospective Studies , Pancreatitis/surgery
13.
Protein Expr Purif ; 13(2): 243-50, 1998 Jul.
Article in English | MEDLINE | ID: mdl-9675069

ABSTRACT

Stromelysin 1 (ST1) is a member of the matrix metalloproteinase (MMP) family probably involved in extracellular matrix degradation. Stromelysin 3 (ST3), considered by sequence homology to be a member of the MMP family of proteases, is specifically expressed in the stroma adjacent to the invasive tumoral cells, but its role in cancer progression remains to be elucidated. Genes encoding ST1 and ST3 were expressed in lepidopteran insect cells using the baculovirus expression vector system. Recombinant baculoviruses were obtained after cloning the full-length cDNA of ST1 and ST3 in plasmids pBacPAK1 and pBacPAK9, respectively. Sf9 insect cells infected with the recombinant baculovirus overexpressed the zymogen proST1 (60 kDa) in an insoluble form, a peak of expression being reached from 24 h postinfection. After solubilization in 8 M urea, and further refolding, activation, and purification, 0.3 mg of mature ST1 (30 kDa), purified to 90% homogeneity, was obtained per 5 x 10(8) infected cells. Recombinant ST1 exhibited proteolytic activity on alpha2-macroglobulin, casein, fibronectin, alpha1-antitrypsin, and laminin. The recombinant zymogen proST3 (55 kDa) was expressed as a soluble form in insect cells, maximal expression occurring at 72 h postinfection. After purification to 95% homogeneity, 2.5 mg of proST3 was obtained per 5 x 10(8) infected cells. A number of proteases including plasmin, urokinase, and ST1 were shown to be able to cleave proST3 giving rise to defined bands of 50-30 kDa. The ST3 mature form of 45 kDa (mST3) was also expressed in the baculovirus system and the obtained protein, 2. 5 mg per 5 x 10(8) infected cells purified to 80% homogeneity, was shown to be active on both casein degradation and alpha2-macroglobulin entrapment assays. Our results suggest that the baculovirus system offers a convenient and efficient means to produce ST1 and ST3 in order to carry out further biochemical studies.


Subject(s)
Matrix Metalloproteinase 3/biosynthesis , Metalloendopeptidases/biosynthesis , Recombinant Proteins/biosynthesis , Animals , Baculoviridae/genetics , Caseins/metabolism , Cloning, Molecular , DNA, Complementary/genetics , Gene Expression , Humans , Matrix Metalloproteinase 11 , Matrix Metalloproteinase 3/genetics , Matrix Metalloproteinase 3/metabolism , Metalloendopeptidases/genetics , Metalloendopeptidases/metabolism , Protein Precursors/biosynthesis , Protein Precursors/genetics , Spodoptera/cytology , Spodoptera/virology , alpha-Macroglobulins/metabolism
14.
J Biol Chem ; 272(41): 25706-12, 1997 Oct 10.
Article in English | MEDLINE | ID: mdl-9325295

ABSTRACT

To elucidate the physiological role of human stromelysin-3 (hST-3) in tumor progression and/or wound healing, insulin-like growth factor-binding protein-1 (IGFBP-1) was analyzed as a potential physiological substrate. hST-3 proteolysis generates two fragments of 16 and 9 kDa that react with IGFBP-1 monoclonal antibody, although they do not bind insulin-like growth factor-I (IGF-I) in ligand blot. N-terminal sequencing shows that hST-3 cleaves IGFBP-1 at the His140-Val141 bond located in the IGFBP-1 midregion. We show that IGFBP-1 inhibits IGF-I-induced survival and proliferation of BAF/3 cells, as well as IGF-I-mediated activation of phosphatidylinositol 3-kinase (PI 3-K). Co-incubation of the IGF-I. IGFBP-1 complex with hST-3 restores IGF-I-induced proliferation and PI 3-K kinase activity in these cells. BAF/3 proliferation is significantly increased with the hST-3-treated IGF-I.IGFBP-1 complex compared with that obtained using IGF-I alone. To produce this enhanced proliferation, IGF-I must bind to IGFBP-1 before hST-3 proteolysis, demonstrated using an IGF-I variant that does not bind IGFBP. IGFBP-1 also inhibits IGF-I-induced proliferation of the MCF-7 breast adenocarcinoma, and this inhibition was not seen in hST-3-transfected MCF-7 cells. Such proteolysis may thus play a role in in vivo tumor progression. These results indicate that hST-3 may regulate IGF-I bioavailability by proteolyzing IGFBP, thus favoring cell survival and proliferation.


Subject(s)
Insulin-Like Growth Factor Binding Protein 1/metabolism , Metalloendopeptidases/metabolism , Humans , Hydrolysis , Insulin-Like Growth Factor I/metabolism , Matrix Metalloproteinase 11 , Metalloendopeptidases/genetics , Molecular Weight , Peptide Mapping , Phosphatidylinositol 3-Kinases/metabolism , Signal Transduction , Transfection , Tumor Cells, Cultured
15.
Gene ; 195(2): 151-9, 1997 Aug 22.
Article in English | MEDLINE | ID: mdl-9305759

ABSTRACT

We report the molecular cloning of a novel gene family. The first member of this family was cloned from a mouse lambda gt11 cDNA library using the B92 monoclonal antibody (mAb) raised against stromal cell extracts. This was followed by RACE-PCR using mRNA from the stromal cell line. A 4 kb cDNA was obtained encoding a unique protein sequence of 1258 aa, that we designate stromal antigen (SA)-1. The human SA-1 gene was cloned by homology from a thymus cDNA library and the sequence of the predicted protein was found to be highly homologous to the murine SA-1 (>98.9%). Another cDNA was cloned and the deduced protein (SA-2) was 71% homologous to SA-1. Northern blot and PCR analysis indicated that on the mRNA level the SA-1 gene is expressed in all tissues analyzed and probably encodes a single transcript. The identification of SA-1 protein in tissues and cells required combined immunoprecipitation and Western blotting using a polyclonal antiserum raised against a predicted peptide of SA-1 and the B92 mAb. Using this assay we identified a protein of about 120 kDa in hemopoietic organs. Subcellular fractionation indicated that SA-1 is a nuclear protein. Thus, despite the ubiquitous expression on the mRNA level, the protein was predominantly detected in hemopoietic organs and may therefore be controlled on a post-transcriptional level. The SA-1 gene described in this study is highly conserved between mouse and man. This implies a crucial function for this protein.


Subject(s)
Gene Expression , Nuclear Proteins/genetics , Amino Acid Sequence , Animals , Base Sequence , Blotting, Western , Bone Marrow Cells/cytology , Bone Marrow Cells/metabolism , Cells, Cultured , Cloning, Molecular , Female , Gene Library , Humans , Lymphocytes/metabolism , Male , Mice , Mice, Inbred BALB C , Molecular Sequence Data , Neoplasms, Experimental/genetics , Neoplasms, Experimental/metabolism , Nuclear Proteins/chemical synthesis , Nuclear Proteins/metabolism , Peptides/chemical synthesis , Peptides/immunology , Polymerase Chain Reaction , Precipitin Tests , RNA, Messenger/genetics , RNA, Messenger/metabolism , Recombination, Genetic , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Stromal Cells , Thymus Gland/metabolism , Transcription, Genetic
16.
Clin Exp Metastasis ; 15(1): 26-32, 1997 Jan.
Article in English | MEDLINE | ID: mdl-9009103

ABSTRACT

To measure matrix metalloproteinase (MMP) activity in a large number of samples it is advisable to use easily automated methods. We have evaluated and compared the activity of stromelysin-1 (MMP-3), matrilysin (MMP-7), 72 kDa gelatinase A (MMP-2) and 92 kDa gelatinase B (MMP-9) by zymogram analysis and fluorescent substrate degradation assays. FITC-casein and the fluorogenic peptide Dnp-Pro-beta-cyclo-hexyl-Ala-Gly-Cys(Me)-His-Ala-Lys-(N-Me-Abz)-NH 2 were used as fluorescent substrates. FITC-casein was more efficiently degraded than the fluorogenic peptide by all MMPs tested except MMP-9. MMP-2 was not significantly able to degrade the fluorogenic peptide. Gelatin zymography was the most sensitive method to detect the activity of both gelatinases but quantitation problems compromise its use. The degradation of fluorogenic substrates by MMPs could be inhibited by the chelating agent EDTA and by the tissue inhibitor of metalloproteinases 2 (TIMP-2), an MMP-specific inhibitor. Fluorometric methods represent a good alternative for MMP activity measurement, especially when a large number of samples must be processed.


Subject(s)
Collagenases/analysis , Electrophoresis, Polyacrylamide Gel , Fluorometry , Gelatinases/analysis , Matrix Metalloproteinase 3/analysis , Metalloendopeptidases/analysis , Recombinant Fusion Proteins/analysis , Calcium/physiology , Caseins/metabolism , Chelating Agents/pharmacology , Collagenases/metabolism , Edetic Acid/pharmacology , Enzyme Inhibitors/pharmacology , Fluorescein-5-isothiocyanate , Fluorescent Dyes , Gelatinases/antagonists & inhibitors , Gelatinases/metabolism , Humans , Matrix Metalloproteinase 3/metabolism , Matrix Metalloproteinase 7 , Matrix Metalloproteinase 9 , Matrix Metalloproteinase Inhibitors , Metalloendopeptidases/antagonists & inhibitors , Metalloendopeptidases/metabolism , Proteins/pharmacology , Recombinant Fusion Proteins/antagonists & inhibitors , Tissue Inhibitor of Metalloproteinase-2
17.
An Med Interna ; 14(9): 478-84, 1997 Sep.
Article in Spanish | MEDLINE | ID: mdl-9453760

ABSTRACT

The aim of this work is to review the role of the main risks factors in relation to cardiovascular diseases, and to evaluate the relative effectiveness of different preventive strategies. In the first part of the paper we summarize the historical background that allows us to understand the magnitude of this problem and its evolution. Secondly, we evaluate the significance of these strategies in the primary prevention of stroke and myocardial infarction, and how an intervention in one risk factor can have a greater significance for one of these events. But in any case, we must consider the whole benefit in each patient's health in our final evaluation of this intervention. Finally, we describe the strategies for hypertension and diabetes in order to reduce the incidence of end renal disease.


Subject(s)
Cerebrovascular Disorders/prevention & control , Myocardial Infarction/prevention & control , Alcohol Drinking/adverse effects , Aspirin/therapeutic use , Cerebrovascular Disorders/etiology , Diabetes Complications , Diabetes Mellitus/prevention & control , Estrogen Replacement Therapy , Exercise , Female , Humans , Hypertension/complications , Hypertension/prevention & control , Kidney Failure, Chronic/complications , Kidney Failure, Chronic/prevention & control , Male , Myocardial Infarction/etiology , Obesity/complications , Obesity/prevention & control , Platelet Aggregation Inhibitors/therapeutic use , Risk Factors , Smoking Cessation
18.
J Biotechnol ; 46(3): 235-41, 1996 May 15.
Article in English | MEDLINE | ID: mdl-8672291

ABSTRACT

The baculovirus expression system was used to produce recombinant human matrilysin. Expression of promatrilysin reached a peak at 72 h post-infection. Most of the recombinant protein remained in the intracellular fraction in an insoluble form, which after renaturation was purified by S-Sepharose and Green A Dyematrex chromatography in order to remove host proteases. Active recombinant matrilysin degraded casein, type I and type IV collagens and fibronectin. Expression of recombinant human matrilysin using the baculovirus system represents a useful tool for obtaining large amounts of this metalloproteinase in order to carry out further biochemical studies and to screen for inhibitors.


Subject(s)
Baculoviridae/genetics , Gene Expression , Metalloendopeptidases/genetics , Metalloendopeptidases/isolation & purification , Animals , Base Sequence , Cell Line , Colonic Neoplasms , DNA, Complementary/chemistry , Humans , Matrix Metalloproteinase 7 , Metalloendopeptidases/biosynthesis , Metalloendopeptidases/metabolism , Molecular Sequence Data , Recombinant Proteins/biosynthesis , Recombinant Proteins/isolation & purification , Spodoptera , Transfection , Tumor Cells, Cultured
19.
Exp Hematol ; 24(6): 728-37, 1996 May.
Article in English | MEDLINE | ID: mdl-8635529

ABSTRACT

Leukemia cell lines that do not proliferate in the absence of serum grow well when cultured with stromal cells. To study this growth dependence on stroma, we selected the M1 myeloblast clone, since its stroma dependence is reminiscent of that exhibited by hematopoietic stem cells. Conditioned medium form a stromal cell line, prepared under serum-free conditions, contained an activity that induced the proliferation of M1 cells and was therefore designated M1 myeloid activity (MMA). Among the various cytokines tested for MMA-like activity, only transforming growth factor-beta (TGF-beta) and macrophage colony-stimulating factor (M-CSF) were found to affect M1 cell survival, and the two cytokines acted synergistically to induce M1 cell growth. Antibodies to both TGF-beta and M-CSF abolished most, but not all, of the MMA in the medium conditioned by stromal cells, indicating that additional factors contribute to MMA. A subclone of M1 cells, M1/M2, selected in medium conditioned by stroma, was found to respond to stromal stimulation but was unable to proliferate in fetal calf serum (FCS). Neutralization experiments indicated that M1/2 cell growth depended mainly on M-CSF and also partially on TGF-beta. By contrast, the same neutralizing antibodies did not affect the ability of serum to support M1 cell growth. The molecules that promoted leukemia cell growth in serum seemed therefore to differ from those provided by stroma. This model system may offer novel information on the interactions of normal and leukemic hematopoietic cells with their stromal microenvironment.


Subject(s)
Bone Marrow/pathology , Leukemia, Experimental/pathology , Macrophage Colony-Stimulating Factor/physiology , Transforming Growth Factor beta/physiology , Animals , Cell Adhesion , Cell Division/drug effects , Culture Media , Cytokines/pharmacology , Drug Synergism , Growth Substances/blood , Macrophage Colony-Stimulating Factor/administration & dosage , Mice , Transforming Growth Factor beta/administration & dosage , Tumor Cells, Cultured
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