RESUMEN
Biomechanical models predict that recruitment of gluteus maximus (GMax) will exert a compressive force across the sacroiliac joint (SIJ), yet this muscle requires morphologic assessment. The aims of this study were to document GMax's proximal attachments and assess their capacity to generate forces including compressive force at the SIJ. In 11 embalmed cadaver limbs, attachments of GMax crossing the SIJ were dissected and their fascicle orientation, length and attachment volume documented. The physiological cross-sectional area (PCSA) of each attachment was calculated along with its estimated maximum force at the SIJ and lumbar spine. GMax fascicles originated from the gluteus medius fascia, ilium, thoracolumbar fascia, erector spinae aponeurosis, sacrum, coccyx, dorsal sacroiliac and sacrotuberous ligaments in all specimens. Their mean fascicle orientation ranged from 32 to 45° below horizontal and mean length from 11 to 18 cm. The mean total PCSA of GMax was 26 cm(2) (range 16-36), of which 70% crossed the SIJ. The average maximum force predicted to be generated by GMax's total attachments crossing each SIJ was 891 N (range 572-1,215), of which 70% (702 N: range 450-1,009) could act perpendicular to the plane of the SIJ. The capacity of GMax to generate an extensor moment at lower lumbar segments was estimated at 4 Nm (range 2-9.5). GMax may generate compressive forces at the SIJ through its bony and fibrous attachments. These may assist effective load transfer between lower limbs and trunk.
Asunto(s)
Fascia/anatomía & histología , Cadera/anatomía & histología , Vértebras Lumbares/anatomía & histología , Músculo Esquelético/anatomía & histología , Articulación Sacroiliaca/anatomía & histología , Vértebras Torácicas/anatomía & histología , Anciano , Anciano de 80 o más Años , Fenómenos Biomecánicos/fisiología , Cadáver , Fascia/fisiología , Femenino , Cadera/fisiología , Humanos , Vértebras Lumbares/fisiología , Masculino , Persona de Mediana Edad , Modelos Anatómicos , Modelos Biológicos , Músculo Esquelético/fisiología , Articulación Sacroiliaca/fisiología , Vértebras Torácicas/fisiologíaRESUMEN
The emerging concept of recovery in mental health is often only loosely defined, but appears to be influenced more by specific human values and beliefs, than scientific research and 'evidence'. As a contribution to the further development of the philosophical basis of the concept of recovery, this paper reviews the discrete assumptions of the Tidal Model, describes the development of the Model's value base -- the 10 Commitments -- and illustrates the 20 Tidal Competencies, which aim to generate practice-based evidence for the process of recovery.
Asunto(s)
Convalecencia , Trastornos Mentales/terapia , Enfermería Psiquiátrica/métodos , Humanos , Trastornos Mentales/enfermeríaRESUMEN
Ig are multifunctional molecules with distinct properties assigned to individual domains. To assess the importance of IgM domain assembly in B cell development we generated two transgenic mouse lines with truncated muH chains by homologous integration of the neomycin resistance gene (neo(r)) into exons C(mu)1 and C(mu)2. Upon DNA rearrangement shortened muH chain transcripts, V(H)-D-J(H)-C(mu)3-C(mu)4, are produced independent of the transcriptional orientation and termination signals provided by neo(r). The truncated muH chain of approximately 52 kDa associates non-covalently with the L chain to form a monovalent HL heterodimer. Surface IgM is assembled into a defective BCR complex which has lost important signalling capacity. In immunizations with T-dependent and T-independent antigens, specific IgM antibodies cannot be detected, whilst IgG responses remain normal. B cell development in the bone marrow is characterized by an increase in early B cells, but a decrease of B220(+) cells from the stage when muH chain rearrangement is completed. The peritoneal lymphocyte population has elevated levels of CD5(+) B cells and their expansion may be the result of a negative feedback mechanism. The results show that antigenic stimulation is compromised by truncated monovalent IgM and that this deficit in stimulation leads to reduced levels of conventional B-2 lymphocytes, but dramatically increased levels of B-1 cells.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Antígenos CD5/biosíntesis , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas mu de Inmunoglobulina/genética , Receptores de Antígenos de Linfocitos B/fisiología , Eliminación de Secuencia/inmunología , Transducción de Señal/genética , Empalme Alternativo/inmunología , Animales , Especificidad de Anticuerpos/genética , Subgrupos de Linfocitos B/metabolismo , Subgrupos de Linfocitos B/patología , Secuencia de Bases , Células de la Médula Ósea/inmunología , Células de la Médula Ósea/patología , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Epítopos de Linfocito B/genética , Vectores Genéticos/síntesis química , Regiones Constantes de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/biosíntesis , Cadenas Ligeras de Inmunoglobulina/genética , Inmunoglobulina M/biosíntesis , Inmunoglobulina M/genética , Cadenas mu de Inmunoglobulina/biosíntesis , Activación de Linfocitos/genética , Recuento de Linfocitos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Ratones Transgénicos , Datos de Secuencia Molecular , Receptores de Antígenos de Linfocitos B/antagonistas & inhibidores , Transducción de Señal/fisiología , Bazo/inmunología , Bazo/patología , Regulación hacia Arriba/genética , Regulación hacia Arriba/inmunologíaRESUMEN
Gemifloxacin is a new fluoroquinolone that has been shown to possess a broad spectrum of antimicrobial activity against gram-positive and gram-negative microorganisms including methicillin-susceptible and methicillin-resistant staphylococci, Streptococcus pneumoniae, Haemophilus influenzae and most members of the family Enterobacteriaceae. The aim of the present study was to investigate the effect of gemifloxacin on the human intestinal microflora. Gemifloxacin was given in oral doses of 320 mg for 7 days to 10 healthy subjects and 5 subjects received a once-daily dose of matched placebo for 7 days. Faecal samples were collected prior to administration (days -8 and -6), during the administration period (days 2 and 4) and after withdrawal of administration (days 8, 11, 21, 28 and 56). In the aerobic intestinal microflora the numbers of enterobacteria were suppressed during the gemifloxacin administration and the numbers of enterococci and streptococci were also decreased. No other aerobic microorganisms were affected. In the anaerobic microflora the numbers of anaerobic cocci and lactobacilli were suppressed during the gemifloxacin administration while no other changes occurred. The microflora was normalized 49 days after the administration of gemifloxacin had stopped. No selection or overgrowth of resistant bacterial strains or yeasts occurred. The ecological impact of gemifloxacin was shown to be selective and similar to that of ciprofloxacin, levofloxacin and ofloxacin.
Asunto(s)
Antiinfecciosos/uso terapéutico , Bacterias/efectos de los fármacos , Fluoroquinolonas , Intestinos/microbiología , Naftiridinas/uso terapéutico , Adulto , Antiinfecciosos/farmacología , Enterobacteriaceae/efectos de los fármacos , Femenino , Gemifloxacina , Haemophilus influenzae/efectos de los fármacos , Humanos , Masculino , Naftiridinas/farmacología , Staphylococcus/efectos de los fármacos , Streptococcus pneumoniae/efectos de los fármacosRESUMEN
Senior lecturers/lecturers in mental health nursing (11 in round one, nine in round two, and eight in the final round) participated in a three-round Delphi study into the teaching of health care ethics (HCE) to students of nursing. The participants were drawn from six (round one) and four (round three) UK universities. Information was gathered on the organization, methods used and content of HCE modules. Questionnaire responses were transcribed and the content analysed for patterns of interest and areas of convergence or divergence. Findings include: the majority (72.8%) of the sample believed that insufficient time was allocated to the teaching of HCE; case studies were considered a popular, although problematic, teaching method; the 'four principles' approach was less than dominant in the teaching of HCE; and virtue ethics was taught by only 36.4% of the participants. The Delphi technique proved adequate and worth while for the purposes of this study. Further empirical research could aim to replicate or contradict these findings, using a larger sample and recruiting more university departments. Reflection is required on several issues, including the depth and breadth to which ethics theory and, more controversially, meta-ethics, are taught to nursing students.
Asunto(s)
Bioética/educación , Educación en Enfermería/normas , Ética en Enfermería/educación , Enfermería Psiquiátrica/educación , Adulto , Técnica Delphi , Docentes de Enfermería , Humanos , Investigación en Educación de Enfermería , Proyectos Piloto , Enfermería Psiquiátrica/ética , Estudiantes de Enfermería , Encuestas y Cuestionarios , Reino UnidoRESUMEN
Nephroblastoma overexpressed (NOV) is a member of the CCN family (connective tissue growth factor, CYR61, and NOV) of proteins that are involved in regulating the proliferation, differentiation, and adhesion of a variety of cell types. We have examined the expression of the NOV: gene and NOV protein by vascular smooth muscle cells (VSMCs), in vitro and in vivo, and the effects of recombinant NOV on VSMCs. Rat aortic VSMCs were found to express NOV: mRNA and NOV protein in vitro and in vivo. NOV: expression in adult rat tissues was very high in the aorta and was detected only weakly in the brain and lung by Northern analysis (relative levels 33:3:1). During postnatal development (3 days to 12 weeks), the expression of NOV: was correlated with markers of the differentiated smooth muscle cell phenotype (smooth muscle myosin heavy chain and SM22 alpha). In the rat carotid artery balloon injury model, NOV: was detectable by in situ hybridization and was downregulated in the media of the injured artery compared with the uninjured artery at 7 and 14 days after injury. Expression in the developing intima was barely detectable at 7 days after injury except for strong expression at the luminal surface. At 14 days after injury, NOV: expression was substantially increased throughout the intima. In vitro studies of the function of NOV protein showed that it promoted VSMC adhesion via a mechanism that was divalent cation and Arg-Gly-Asp independent but that it did not modulate VSMC proliferation or phenotype. The strong expression and dynamic regulation of NOV: in the arterial wall, together with its ability to promote VSMC adhesion, suggest that it may be involved in homeostasis and repair.
Asunto(s)
Moléculas de Adhesión Celular/genética , Proteínas Inmediatas-Precoces , Péptidos y Proteínas de Señalización Intercelular , Músculo Liso Vascular/metabolismo , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/fisiología , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/fisiología , Animales , Aorta/crecimiento & desarrollo , Enfermedades de las Arterias Carótidas/metabolismo , Adhesión Celular , Células Cultivadas , Clonación Molecular , Factor de Crecimiento del Tejido Conjuntivo , Ácido Edético/farmacología , Expresión Génica , Hibridación in Situ , Integrinas/fisiología , Oligopéptidos/farmacología , Reacción en Cadena de la Polimerasa , ARN Mensajero/análisis , Ratas , Ratas Wistar , Vitronectina/metabolismoRESUMEN
A three-round Delphi study was conducted to gather data on ethical reasoning among psychiatric nurses (N = 26 in round one (R1), decreasing to N = 14 in the final round (R3)). Transcripts of questionnaires were carefully read and compared. Responses were manually sorted into categories, themes and patterns of interest. Eight debates emerged from the data. This article discusses two in detail: the nature of moral virtues and the meaning of compassion in psychiatric nursing. A sympathetic overview of virtue ethics is also provided. The nurses' responses included a lot of virtue terms, such as, 'honest', 'fair', and 'care". However, 'nurses' moral virtues' was ranked low in importance as a notion invoked in ethical decision making in the round-one ranking exercise. Only half of the sample believed that the moral character of a psychiatric nurse is important in ethical decision-making. Further, most of the round-one sample thought the virtues could not be acquired. Compassion was identified as crucial to psychiatric nursing and the nurse-client relationship, though, as expected, many diverse meanings were attributed to this notion. While the Delphi method proved adequate for our purposes, problems with regard to accurately understanding the respondents' intended meanings highlighted a major weakness of this technique, in common with other methods relying on questionnaires. Further inquiry is needed regarding the role of moral virtues and virtue ethics in both psychiatric nursing and nurse education.
Asunto(s)
Empatía , Enfermería Psiquiátrica , Virtudes , Técnica Delphi , Ética , Humanos , Encuestas y CuestionariosRESUMEN
The findings of a pilot telephone survey of senior nurses' views of multidisciplinary teamwork (MDT) practice in 26 acute psychiatric admission wards within the ambit of one English health authority are reported. The survey was conducted in two stages. The first stage focused on obtaining the senior nurse's general perceptions of the organization of the work of the multidisciplinary team, and in the second stage more detailed responses to the original questions were obtained. The findings suggest that the composition and operations of the acute ward team are limited, and the relationship between the MDT and patients and their families do not entirely support the concept of increased collaboration with patients and their families. The findings are discussed within the context of the perceived crisis in acute psychiatric care in England and the ambitions for more egalitarian 'mental health' services.
Asunto(s)
Actitud Frente a la Salud , Trastornos Mentales/terapia , Grupo de Atención al Paciente , Enfermería Psiquiátrica , Enfermedad Aguda , Humanos , Servicios de Salud Mental , Defensa del PacienteRESUMEN
STUDY DESIGN: Superficial and deep laminae of the posterior layer of lumbar fascia were dissected. The lumbar portion was measured for evidence of segmental thickenings. Superior attachments were dissected, documented, and photographed. OBJECTIVES: To verify the existence of posterior accessory ligaments and establish the superior attachments and fiber angles of the posterior layer of lumbar fascia. SUMMARY OF BACKGROUND DATA: There have been two small dissection studies on the posterior layer. Their findings are conflicting in several areas of clinical significance. Thickenings in the lumbar region were described in one study, but have not been verified. The superior attachments of the posterior layer have not been formally documented. METHODS: Study 1: In 21 embalmed cadavers, the lumbar region of the posterior layer was dissected. The lumbar spinous processes and adjacent fascia were marked. The fascia was removed and examined, and its thickness measured with a manual micrometer. Results were statistically analyzed. Study 2: Superior attachments of the posterior layer in 20 cadavers were dissected and photographed. Capacity to transmit tension was estimated and documented photographically, and fiber angles measured in situ. RESULTS: Study 1: There was no evidence of macroscopic segmental thickening in the posterior layer. Study 2: The superficial lamina was continuous superiorly with the rhomboids, and the deep lamina with the tendons of splenius cervices and capitis. These previously undocumented attachments were of variable thickness and fibrosity, and capable of transmitting tension. CONCLUSIONS: Both superficial and deep laminae of the posterior layer are more extensive superiorly than previously thought. This may have implications for certain tests used in assessment and management of low back pain such as the slump and "nonorganic" tests. The thickness of the superior attachments is variable. Their capacity for load bearing is yet to be quantified.
Asunto(s)
Fascia Lata/anatomía & histología , Ligamentos Articulares/anatomía & histología , Vértebras Lumbares/anatomía & histología , Anciano , Anciano de 80 o más Años , Cadáver , Femenino , Humanos , Masculino , Músculo Esquelético/anatomía & histologíaRESUMEN
The plant virus cowpea mosaic virus (CPMV) has been developed as an epitope-presentation system. Numerous epitopes have been expressed in the betaB-betaC loop of the CPMV small coat protein, all of which undergo a cleavage reaction between their two carboxy-terminal residues. Although many peptides presented in this manner give an authentic immune response, this was not the case for the NIm-1A epitope from human rhinovirus-14. Crystallography revealed significant differences between the structure of NIm-1A on CPMV compared with its native configuration. The 3D structure of C PMV expressing NIm-1A was used to design alterations to the context of the NIm-1A graft.
Asunto(s)
Cápside/química , Comovirus , Vectores Genéticos , Procesamiento Proteico-Postraduccional , Rhinovirus/química , Secuencia de Aminoácidos , Secuencia de Bases , Cápside/biosíntesis , Cápside/genética , Proteínas de la Cápside , Humanos , Datos de Secuencia Molecular , Pisum sativum/virología , Péptidos , Rhinovirus/genéticaRESUMEN
Current mental health practice and legislation is influenced significantly by the ill-defined concept of serious mental illness (SMI). Although the concept of SMI originally appeared to include a consideration of a range of forms of mental illness, including dementias, by custom and practice the term has come to mean, often, only people with a psychotic diagnosis. This paper reviews the influence of the term SMI, considering the way older people, younger people and other subgroups within the 'adult' population have been marginalized by the narrow definition of the concept. The paper concludes with a consideration of some of the ethical implications of mental health nurses focusing on an ill-defined 'illness' grouping, and the impact that such a narrow focus might have on the ambition to establish 'health for all by the year 2000'.
Asunto(s)
Actitud Frente a la Salud , Trastornos Mentales/enfermería , Prejuicio , Enfermería Psiquiátrica , Adolescente , Adulto , Factores de Edad , Anciano , Ética en Enfermería , Humanos , Persona de Mediana Edad , Teoría de Enfermería , Distancia Psicológica , Índice de Severidad de la EnfermedadRESUMEN
Site-directed antisera have been developed against the two endothelin-converting enzyme-1 (ECE-1) isoforms cloned to date in humans, ECE-1 alpha and ECE-1 beta. Antisera were raised in rabbits against synthetic peptides corresponding to the deduced amino acid sequences that differ between ECE-1 alpha and ECE-1 beta. Antisera were highly selective for their corresponding antigen (titer 1 x 10(4)) and did not detect ET-1 or big ET-1. Furthermore, no detectable crossreactivity was observed between the different site-specific antisera and the other immunizing peptides, suggesting that the antisera would be selective for ECE-1 alpha and ECE-1 beta. Standard displacement curves have been developed to determine the levels of immunoreactive ECE-1 alpha and ECE-1 beta in solubilized microsomal fractions of human tissue. In conclusion, we have described the first production and characterization of site-directed antisera raised against ECE-1 alpha and ECE-1 beta capable of discriminating between the two ECE-1 isoforms. Furthermore, using these antisera, we have found that ECE-1 alpha appears to be the predominant isoform in human tissue.
Asunto(s)
Anticuerpos Bloqueadores/farmacología , Ácido Aspártico Endopeptidasas/inmunología , Isoenzimas/inmunología , Metaloendopeptidasas/inmunología , Secuencia de Aminoácidos , Animales , Especificidad de Anticuerpos , Ácido Aspártico Endopeptidasas/antagonistas & inhibidores , Reacciones Cruzadas , Enzimas Convertidoras de Endotelina , Humanos , Isoenzimas/antagonistas & inhibidores , Metaloendopeptidasas/antagonistas & inhibidores , Microsomas/metabolismo , Datos de Secuencia Molecular , ConejosRESUMEN
The use of synthetic random peptide libraries is a powerful technology for the study of many aspects of antigen presentation and peptide selection by major histocompatibility complex (MHC) molecules. Here we have used them in conjunction with a recombinant system to determine the peptide binding motifs of three classical class I MHC molecules of the laboratory rat: RT1-Aa, RT1-Au, and RT1-A1c. Described is a method for producing large amounts of soluble class I heavy and light chains in bacteria. Refolding RT1-Aa heavy chain (HC) with rat beta2-microglobulin (beta2m) in the presence of a specific peptide and the subsequent purification of the complex yielded conformationally correct material. This was assessed by gel chromatography, SDS-polyacrylamide gel electrophoresis, isoelectric focussing gel electrophoresis, enzyme-linked immunosorbent assay, and fluorescence-activated cell sorter analysis employing a previously unreported method utilizing a His-Tag affinity silica. By refolding RT1-Aa HC and rat beta2m around a random nonapeptide library and subjecting the resulting complex to acid elution of the bound peptides and pool sequencing, the peptide binding motif for this MHC class I molecule was determined. Results corresponded well with those previously determined from naturally bound peptides and in addition gave a clear and unambiguous signal for the C-terminal anchor residue. This method was then applied to determine the previously undescribed binding motifs for RT1-Au and RT1-A1c. For both molecules, the whole motif was confirmed from naturally bound peptides. We propose this method as an alternative way to obtain the whole class I MHC peptide motif, particularly when a specific antibody is unavailable and/or natural expression of the class I molecule of interest is low.
Asunto(s)
Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad/metabolismo , Oligopéptidos/metabolismo , Animales , Sitios de Unión , Simulación por Computador , Antígenos de Histocompatibilidad/genética , Antígenos de Histocompatibilidad Clase I/genética , Modelos Moleculares , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/metabolismo , Biblioteca de Péptidos , Unión Proteica , Conformación Proteica , Pliegue de Proteína , Ratas , Proteínas Recombinantes de Fusión/metabolismo , Microglobulina beta-2/genética , Microglobulina beta-2/metabolismoRESUMEN
An antigen complex unique for porcine gamma/delta T cells has previously been identified using the monoclonal antibodies MAC319 and MAC320. Here we use digestion with the proteolytic enzyme bromelain to selectively release the MAC319 antigen as a soluble fragment, for further characterisation. A cytofluorometric inhibition assay was developed to follow the purification of this fragment, as the conformation sensitivity of the MAC319 epitope prevented the use of immunoblotting techniques. The antigen has been purified using a combination of anion-exchange and hydrophobic-interaction columns, followed by separation on a size-exclusion column. Fractions from the size-exclusion column containing the antigen consisted of one major band at Mr 95,000. This species was shown to be specifically absorbed onto MAC319-coupled Sepharose, thereby identifying the MAC319 antigen. N-terminal amino acid sequencing of this band has revealed a previously unidentified sequence. This fragment was also shown to be glycosylated, most likely with a single sugar moiety. Enzymatic removal of the sugars showed that they did not appear to be necessary for binding of the polypeptide to the MAC319 antibody.
Asunto(s)
Mapeo Epitopo , Receptores de Antígenos de Linfocitos T gamma-delta/aislamiento & purificación , Animales , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Bromelaínas/farmacología , Femenino , Glicósido Hidrolasas/metabolismo , Humanos , Masculino , Peso Molecular , Receptores de Antígenos de Linfocitos T gamma-delta/química , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Porcinos , Linfocitos T/químicaRESUMEN
Psychiatric nurses in the United Kingdom (UK) have begun to reattend to people with 'serious and enduring mental illness'. At the same time research in the USA and UK has refocused much of its attention on neuroscientific theories and models of serious mental illness. Psychiatric nurses are being encouraged to consider the value of biomedical explanations of serious illness, such as schizophrenia, and to accommodate these theories and models in the practice of nursing. This paper will examine the challenge of the biomedical approach for the continued development of psychiatric nursing theory and practice. It is proposed that psychiatric nursing needs to develop further its own 'proper focus', if it is to survive as a key player in the health care field on the 21st century.
Asunto(s)
Teoría de Enfermería , Enfermería Psiquiátrica , Esquizofrenia/enfermería , Humanos , Modelos Psicológicos , Relaciones Enfermero-Paciente , Diagnóstico de Enfermería , Esquizofrenia/fisiopatologíaRESUMEN
In Saccharomyces cerevisiae the metabolite phosphoribosyl-pyrophosphate (PRPP) is required for purine, pyrimidine, tryptophan and histidine biosynthesis. Enzymes that can synthesize PRPP can be encoded by at least four genes. We have studied 5-phospho-ribosyl-1(alpha)-pyrophosphate synthetases (PRS) genetically and biochemically. Each of the four genes, all of which are transcribed, has been disrupted in haploid yeast strains of each mating type and although all disruptants are able to grow on complete medium, differences in growth rate and enzyme activity suggest that disruption of PRS1 or PRS3 has a significant effect on cell metabolism, whereas disruption of PRS2 or PRS4 has little measurable effect. Using Western blot analysis with antisera raised against peptides derived from the non-homology region (NHR) and the N-terminal half of the PRS1 gene product it has been shown that the NHR is not removed by protein splicing. However, the fact that disruption of this gene causes the most dramatic decrease in cell growth rate and enzyme activity suggests that Prs1p may have a key structural or regulatory role in the production of PRPP in the cell.
Asunto(s)
Genes Fúngicos , Genes del Tipo Sexual de los Hongos , Familia de Multigenes , Ribosa-Fosfato Pirofosfoquinasa/genética , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Cartilla de ADN/genética , Datos de Secuencia Molecular , Mutagénesis Insercional , Saccharomyces cerevisiae/crecimiento & desarrolloAsunto(s)
Epóxido Hidrolasas/aislamiento & purificación , Microsomas Hepáticos/enzimología , Secuencia de Aminoácidos , Animales , Cromatografía de Afinidad , Electroforesis en Gel de Poliacrilamida , Epóxido Hidrolasas/química , Epóxido Hidrolasas/metabolismo , Caballos , Humanos , Datos de Secuencia Molecular , Fragmentos de Péptidos/química , Conejos , Ratas , Homología de Secuencia de AminoácidoRESUMEN
It is known that dietary carbohydrates regulate the activity of the intestinal SGLT1. We have demonstrated that modifications in SGLT1 activity are due to alterations in SGLT1 expression in response to the sugar content of the diet. To assess the correlation between changes in the activity of SGLT1 and the abundance of SGLT1 protein, we have employed a method for the quantitative measurement of immunoreactive proteins. A calibration curve has been constructed using either a nonadecapeptide (amino acids 402-420), or a recombinant protein corresponding to amino acids 554-640 of the SGLT1 sequence. Immunoblotting the protein samples concurrently with specific quantities of either the peptide or recombinant standard, using antibodies raised against these antigens, enabled accurate quantification of the absolute amounts of immunoreactive protein in the samples. The amount of SGLT1 protein correlates well with measurements of SGLT1 activity. The modulation of the activity of SGLT1 in response to lumenal sugars is due to corresponding changes in the absolute levels of SGLT1 protein.
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Carbohidratos de la Dieta/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Mucosa Intestinal/metabolismo , Glicoproteínas de Membrana/efectos de los fármacos , Glicoproteínas de Membrana/genética , Proteínas de Transporte de Monosacáridos/efectos de los fármacos , Proteínas de Transporte de Monosacáridos/genética , Animales , Femenino , Mucosa Intestinal/química , Masculino , Glicoproteínas de Membrana/química , Microvellosidades/química , Microvellosidades/metabolismo , Proteínas de Transporte de Monosacáridos/química , Ovinos , Transportador 1 de Sodio-GlucosaRESUMEN
Mechanical injury in plants induces responses that are involved not only in healing but also in defense against a potential pathogen. To understand the intracellular signaling mechanism of wounding, we have investigated the involvement of protein kinases. Using specific antibodies, we showed that wounding alfalfa leaves specifically induces the transient activation of the p44MMK4 kinase, which belongs to the family of mitogen-activated protein kinases. Whereas activation of the MMK4 pathway is a post-translational process and was not blocked by [alpha]-amanitin and cycloheximide, inactivation depends on de novo transcription and translation of a protein factor(s). After wound-induced activation, the MMK4 pathway was subject to a refractory period of 25 min, during which time restimulation was not possible, indicating that the inactivation mechanism is only transiently active. After activation of the p44MMK4 kinase by wounding, transcript levels of the MMK4 gene increased, suggesting that the MMK4 gene may be a direct target of the MMK4 pathway. In contrast, transcripts of the wound-inducible MsWIP gene, encoding a putative proteinase inhibitor, were detected only several hours after wounding. Abscisic acid, methyl jasmonic acid, and electrical activity are known to mediate wound signaling in plants. However, none of these factors was able to activate the p44MMK4 kinase in the absence of wounding, suggesting that the MMK4 pathway acts independently of these signals.