RESUMEN
The objective of this study was to determine the effect of size-fractionation by centrifugation on the film structure of casein micelles. Fractionated casein micelles in solution were asymmetrically distributed with a small distribution width as measured by dynamic light scattering. Films prepared from the size-fractionated samples showed a smooth surface in optical microscopy images and a homogeneous microstructure in atomic force micrographs. The nano- and microstructure of casein films was probed by micro-beam grazing incidence small angle x-ray scattering (µGISAXS). Compared to the solution measurements, the sizes determined in the film were larger and broadly distributed. The measured GISAXS patterns clearly deviate from those simulated for a sphere and suggest a deformation of the casein micelles in the film.
Asunto(s)
Caseínas/análisis , Quelantes/análisis , Micelas , Nanoestructuras/análisis , Animales , Caseínas/química , Quelantes/química , Microscopía de Fuerza Atómica , Leche/química , Nanoestructuras/química , Tamaño de la Partícula , Dispersión del Ángulo Pequeño , Soluciones/química , Temperatura , Difracción de Rayos XRESUMEN
The operon containing the genes encoding the subunits of the binary crystal toxin of Bacillus sphaericus strain LP1-G, BinA and BinB (41.9 kDa and 51.4 kDa, respectively), was cloned and sequenced. Purified crystals were not toxic to Culex pipiens larvae. Comparison of the amino-acid sequences of this strain (Bin4) with those of the three other known toxin types (Bin1, Bin2 and Bin3) revealed mutations at six positions, including a serine at position 93 of BinA4, whereas all other types of BinA toxin from B. sphaericus had a leucine at this position. Reciprocal site-directed mutagenesis was performed to replace this serine in BinA4 from LP1-G with a leucine and the leucine in the BinA2 protein from strain 1593 with a serine. Native and mutated genes were cloned and overexpressed. Inclusion bodies were tested on C. pipiens larvae. Unlike the native Bin4 toxin, the mutated protein was toxic, and the reciprocal mutation in Bin2 led to a significant loss of toxicity. In vitro receptor-binding studies showed similar binding behaviour for native and mutated toxins. In the absence of any experimental data on the 3D structure of these proteins, sequence analysis and secondary-structure predictions were performed. Amino acid 93 of the BinA polypeptide probably belongs to an alpha helix that is sensitive to amino-acid modifications. Position 93 may be a key element in the formation of the BinA-BinB complex responsible for the toxicity and stability of B. sphaericus Bin toxins.
Asunto(s)
Bacillus/química , Toxinas Bacterianas/química , Secuencia de Aminoácidos , Animales , Bacillus/genética , Toxinas Bacterianas/genética , Toxinas Bacterianas/toxicidad , Secuencia de Bases , Unión Competitiva , Culex/efectos de los fármacos , Culex/metabolismo , Cartilla de ADN/genética , Sistema Digestivo/metabolismo , Larva/efectos de los fármacos , Larva/metabolismo , Datos de Secuencia Molecular , Estructura Molecular , Mutagénesis Sitio-Dirigida , Control Biológico de Vectores , Estructura Secundaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/toxicidad , Homología de Secuencia de AminoácidoRESUMEN
Similar total doses of Ara-C given as a single injection or given in a fractionated protocol have reverse effects on CFU-S differentiation pathways. Whereas a single dose of 20 mg channels CFU-S towards erythropoiesis, 5 X 5 mg given at 8 or 24 hour intervals channels CFU-S to granulopoiesis and megakaryocytopoiesis. It therefore seems possible to manipulate CFU-S differentiation not only by varying the inducing agents but also by varying the protocols using the same agent. Hypotheses to explain the mechanisms of CFU-S regulation are presented.
Asunto(s)
Citarabina/farmacología , Células Madre Hematopoyéticas/efectos de los fármacos , Animales , Diferenciación Celular/efectos de los fármacos , Citarabina/administración & dosificación , Eritropoyesis/efectos de los fármacos , Ratones , Ratones Endogámicos CBA , Organismos Libres de Patógenos EspecíficosRESUMEN
Regulation of pluripotent stem cell (CFU-S) proliferation kinetics by humoral factors is now well documented. However, the mechanism of choice of CFU-S differentiation pathways is still a controversial issue. We suggest that long-range humoral factors (pluripoietins) are capable of preferentially channelling CFU-S towards one of the cell lineages after various perturbations such as irradiation and drug treatment. The differentiation pathway depends on the treatment protocol. We present data concerning one of the protocols: injection of 20 mg of cytosine arabinoside (Ara-C). When conditioned medium from bone marrow of treated mice is incubated with normal marrow, CFU-S of the latter generate spleen colonies with an E/G ratio above normal values. Clonal analyses of spleen colonies demonstrate that they are generated by pluripotent stem cells. Therefore, any modification in the E/G ratio is due to modifications of CFU-S channelling and not to the variations of committed cells. It was of interest to determine the mechanism of pluripoietin activity. This was studied at three levels: membrane receptors, gene activation and protein synthesis. These studies suggest that CFU-S have receptors for pluripoietins which activate genes responsible for specific mRNA synthesis. De novo synthesis of proteins is a necessary prerequisite for pluripoietin activity to be expressed. Hypotheses for these mechanisms are presented.
Asunto(s)
Hematopoyesis/efectos de los fármacos , Animales , Diferenciación Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Ensayo de Unidades Formadoras de Colonias , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Ratones , Modelos Biológicos , Receptores de Superficie Celular/fisiología , Activación TranscripcionalRESUMEN
A comparison was made, by autoradiographic studies, of the nucleic acids synthesis in the target organs of the male hormone, in normal mice, castrate and castrated mice with an implantation of a testosterone pellet. The results show that the response of each of these organs, to a physiological supply of hormone or to a supraphysiological supply is quite different, both in intensity and in evolution. The differences are still apparent in castrated animals. The transfer of RNA from the nucleus to the cytoplasm seems to be affected by the castration. The signification of these results is discussed.
Asunto(s)
ADN/biosíntesis , Genitales Masculinos/metabolismo , ARN/biosíntesis , Testosterona/farmacología , Animales , Autorradiografía , Castración , Citidina/metabolismo , Preparaciones de Acción Retardada , Epidídimo/metabolismo , Genitales Masculinos/efectos de los fármacos , Masculino , Ratones , Próstata/metabolismo , Vesículas Seminales/metabolismo , Testosterona/administración & dosificación , Timidina/metabolismo , Conducto Deferente/metabolismoRESUMEN
The advantages and drawbacks of urethrocytogram in the course of pregnancy, as compared with vaginal smear to determine how near the term is, have been studied by the authors. They have come to the conclusion that urethrocytogram was an easy to perform examination requiring no highly special equipment, easy to read and capable of giving an immediate answer when the vaginal smear cannot be interpreted because If the existing inflammation, which occurs in one third of cases. Modifications of the urethrocytogram show a slight delay as compared with those shown by the vaginal smear.
Asunto(s)
Trimestres del Embarazo , Uretra/citología , Adulto , Femenino , Humanos , Embarazo , Sensibilidad y Especificidad , Frotis VaginalRESUMEN
A case of feminizing testicle in a family with 9 children, of which girls, has been reported by the authors. A familial study showed 4 out of the 7 girls to be affected by the same disease. The mother, aged 81, presented with no anomaly, save the absence of pubic hair.