Optimization of protein crystallization:the OptiCryst project

Protein crystallization has gained a new strategic and commercial relevance in the postgenomic era due to its pivotal role in structural genomics. Producing high quality crystals has always been a bottleneck to efficient structure determination, and this problem is becoming increasingly acute. This is especially true for challenging, therapeutically important proteins that typically do not form suitable crystals. The OptiCryst consortium has focused on relieving this bottleneck by making a concerted effort to improve the crystallization techniques usually employed, designing new crystallization tools, and applying such developments to the optimization of target protein crystals. In particular, the focus has been on the novel application of dual polarization interferometry (DPI) to detect suitable nucleation; the application of in situ dynamic light scattering (DLS) to monitor and analyze the process of crystallization; the use of UV-fluorescence to differentiate protein crystals from salt; the design of novel nucleants and seeding technologies; and the development of kits for capillary counterdiffusion and crystal growth in gels. The consortium collectively handled 60 new target proteins that had not been crystallized previously. From these, we generated 39 crystals with improved diffraction properties. Fourteen of these 39 were only obtainable using OptiCryst methods. For the remaining 25, OptiCryst methods were used in combination with standard crystallization techniques. Eighteen structures have already been solved (30% success rate), with several more in the pipeline.

The use of sodium carboxymethylcellulose in the preparation of spray-dried proteins for pulmonary drug delivery

The use of sodium carboxymethylcellulose (NaCMC) as a spray-drying excipient in the preparation of inhalable formulations of proteins was investigated, using alkaline phosphatase as a model functional protein. Two spray-dried powders were investigated: a control powder comprising 100% (w/w) alkaline phosphatase and a test powder comprising 67% (w/w) NaCMC and 33% (w/w) alkaline phosphatase. Following physicochemical characterisation, the powders were prepared as both dry powder inhaler (DPI) and pressurised metered dose inhaler (pMDI) formulations. The aerosolisation performance of the formulations was assessed using a Multi-Stage Liquid Impinger, both immediately after preparation and over a 16-week storage period. Formulating the control powder as a DPI resulted in a poor fine particle fraction (FPF: 10%), whereas the FPF of the NaCMC-modified DPI formulation was significantly greater (47%). When the powders were formulated as pMDI systems, the control and NaCMC-modified powders demonstrated FPFs of 52% and 55%, respectively. Following storage, reduced FPF was observed for all formulations except the NaCMC-modified pMDI system; the performance of this formulation following storage was statistically equivalent to that immediately following preparation. Co-spray-drying proteins and peptides with NaCMC may therefore offer an alternative method for the preparation of stable and respirable pMDI formulations for pulmonary delivery. © 2010 Elsevier B.V.

Optimized square Fresnel zone plates for microoptics applications

Polygonal Fresnel zone plates with a low number of sides have deserved attention in micro and nanoptics, because they can be straightforwardly integrated in photonic devices, and, at the same time, they represent a balance between the high-focusing performance of a circular zone plate and the easiness of fabrication at micro and nano-scales of polygons. Among them, the most representative family are Square Fresnel Zone Plates (SFZP). In this work, we propose two different customized designs of SFZP for optical wavelengths. Both designs are based on the optimization of a SFZP to perform as close as possible as a usual Fresnel Zone Plate. In the first case, the criterion followed to compute it is the minimization of the difference between the area covered by the angular sector of the zone of the corresponding circular plate and the one covered by the polygon traced on the former. Such a requirement leads to a customized polygon-like Fresnel zone. The simplest one is a square zone with a pattern of phases repeating each five zones. On the other hand, an alternative SFZP can be designed guided by the same criterion but with a new restriction. In this case, the distance between the borders of different zones remains unaltered. A comparison between the two lenses is carried out. The irradiance at focus is computed for both and suitable merit figures are defined to account for the difference between them.

Far field of binary phase gratings with errors in the height of the strips

Diffraction gratings are not always ideal but, due to the fabrication process, several errors can be produced. In this work we show that when the strips of a binary phase diffraction grating present certain randomness in their height, the intensity of the diffraction orders varies with respect to that obtained with a perfect grating. To show this, we perform an analysis of the mutual coherence function and then, the intensity distribution at the far field is obtained. In addition to the far field diffraction orders, a "halo" that surrounds the diffraction order is found, which is due to the randomness of the strips height.

Binary gratings with random heights

We analyze the far-field intensity distribution of binary phase gratings whose strips present certain randomness in their height. A statistical analysis based on the mutual coherence function is done in the plane just after the grating. Then, the mutual coherence function is propagated to the far field and the intensity distribution is obtained. Generally, the intensity of the diffraction orders decreases in comparison to that of the ideal perfect grating. Several important limit cases, such as low- and high-randomness perturbed gratings, are analyzed. In the high-randomness limit, the phase grating is equivalent to an amplitude grating plus a “halo.” Although these structures are not purely periodic, they behave approximately as a diffraction grating.

Isoellipticines: novel compounds for leukaemia treatment

Ellipticine, an anticancer agent, has had limited clinical success due to low solubility and toxic side effects. To overcome these limitations, a panel of novel ellipticine isomers were designed and synthesised with the aim of evaluating their anti-cancer effects on selected cancer cell lines. A preliminary NCI 60-cell screen demonstrated that these isoellipticines displayed promising anti-tumour activity across a number of different cell types, particularly leukaemia cell lines. We consequently examined the effect of these derivatives in detail on the Acute Myeloid Leukaemia (AML) cell line, MV4-11. Cell cycle analyses revealed that the compounds had a range of distinctive cell cycle effects on MV4-11 cells. 7-Hydroxyisoellipticine showed the most promise with respect to cytostatic activity. We demonstrated that this compound inhibited proliferation of leukaemia cells by preventing cells from progressing from G2 phase. Our research suggests that this is mediated by an induction of reactive oxygen species (ROS), which in turn activates the DNA damage response pathway. More extensive research on the source of ROS generated by the most potent derivative, 7-formyl-10-methylisoellipticine showed that this compounds cytotoxicity is partially mediated by an induction of mitochondrial derived reactive oxygen species (ROS). We showed that 7-formyl-10-methylisoellipticine has synergistic effects when used in combination with the clinically used AML drug, daunorubicin, as well as DPI, a Nox inhibitor. Additionally, combination experiments with other drugs served to give us a deeper insight into 7- formyl-10-methylisoellipticine mechanism of action. 7-Formyl-10-methylisoellipticine also displayed promising in vivo results. Treatment resulted in a lack of toxicity, as measured by body weight changes and liver enzyme analyses. Most importantly, 7-formyl-10-methylisoellipticine demonstrated potent anti-tumour activity in the in vivo xenograft mouse model, implying the potential of isoellipticines as novel chemotherapeutic agents in the treatment of leukaemia. In summary, this study provides for the first time detailed cellular information on the potential use of isoellipticines as chemotherapeutic agents. Our study documents for the first time, the therapeutic potential of an isoellipticine compound in a subcutaneous AML cell-derived xenograft (CDX) model. By probing the mechanism of action of this novel compound class we have uncovered a potential clinical application in the field of adjuvant therapy. We anticipate that the recent research on ellipticine derivatives, such as this study, will lead the development of an ellipticine analogue that may be employed clinically.

Effects of silicon treatment and inoculation with Fusarium oxysporum f. sp. vasinfectum on cellular defences in root tissues of two cotton cultivars

BACKGROUND AND AIMS: Silicon has been shown to enhance the resistance of plants to fungal and bacterial pathogens. Here, the effect of potassium silicate was assessed on two cotton (Gossypium hirsutum) cultivars subsequently inoculated with Fusarium oxysporum f. sp. vasinfectum (Fov). Sicot 189 is moderately resistant whilst Sicot F-1 is the second most resistant commercial cultivar presently available in Australia. METHODS: Transmission and light microscopy were used to compare cellular modifications in root cells after these different treatments. The accumulation of phenolic compounds and lignin was measured. KEY RESULTS: Cellular alterations including the deposition of electron-dense material, degradation of fungal hyphae and occlusion of endodermal cells were more rapidly induced and more intense in endodermal and vascular regions of Sicot F-1 plants supplied with potassium silicate followed by inoculation with Fov than in similarly treated Sicot 189 plants or in silicate-treated plants of either cultivar not inoculated with Fov. Significantly more phenolic compounds were present at 7 d post-infection (dpi) in root extracts of Sicot F-1 plants treated with potassium silicate followed by inoculation with Fov compared with plants from all other treatments. The lignin concentration at 3 dpi in root material from Sicot F-1 treated with potassium silicate and inoculated with Fov was significantly higher than that from water-treated and inoculated plants. CONCLUSIONS: This study demonstrates that silicon treatment can affect cellular defence responses in cotton roots subsequently inoculated with Fov, particularly in Sicot F-1, a cultivar with greater inherent resistance to this pathogen. This suggests that silicon may interact with or initiate defence pathways faster in this cultivar than in the less resistant cultivar.

Cumplimiento de metas de hipertensión arterial en pacientes con enfermedad renal crónica terminal en el Instituto Salvadoreño del Seguro Social

La enfermedad renal crónica es un problema de salud al alza, que para el año 2010 se clasificó como la 18ª causa de mortalidad a nivel mundial, con una tasa de mortalidad de 16.3 por 100,000 habitantes. La diabetes mellitus e hipertensión arterial son las causas principales de la enfermedad renal crónica a nivel mundial tanto en países desarrollados como subdesarrollados. En las últimas revisiones de las guías de manejo de la Sociedad Europea de Hipertensión y la Sociedad Europea de Cardiología 2013, como las guías estadounidenses del grupo de expertos citados al Comité Nacional Colectiva (por sus siglas en inglés JNC8), establecen la meta de presión arterial por debajo de 130/90 mmHg para el paciente con enfermedad renal crónica. El objetivo de este estudio fue conocer el cumplimiento de metas de presión arterial en pacientes con enfermedad renal crónica en terapia de sustitución renal en servicios de Medicina Interna (MI) y Diálisis Peritoneal Intermitente DPI) del Hospital Médico Quirúrgico (HMQ). El estudio se estructuró para ser de tipo observacional descriptivo de corte transversal. La población de estudio comprendió pacientes con enfermedad renal crónica en terapia de sustitución renal en los servicios de Medicina Interna y Diálisis Peritoneal Intermitente del Hospital Médico y Quirúrgico del Instituto Salvadoreño del Seguro Social. Con un universo de pacientes que totalizó 346. La muestra para la realización del estudio se calculó en 182 pacientes que cumplieron con un nivel de confianza del 95% y un porcentaje máximo de error del 5%. Con este grupo se realizó un muestreo aleatorizado simple con lo que se seleccionaron los pacientes para participar de la muestra.

Thief Stealing Wallet From Woman's Purse

3435 x 5150 px | 29,08 x 43,60 cm | 300 dpi

Two men sitting in court

6.271 x 4.180 px | 53,09 x 35,39 cm | 300 dpi

La conservación de recursos genéticos vegetales y los derechos de propiedad intelectual

Introducción El presente artículo trata sobre los derechos de propiedad intelectual (DPI) y la conservación in situ. En primer término describe la conservación y uso de los recursos fitogenéticos como un “sistema” en el cual distintos agentes desempeñan papeles diferentes, Se tipifica la creación de conocimiento por parte le comunidades tradicionales e indígenas y se la compara con la producción de conocimiento que ocurre dentro del sistema de la “ciencia” y la  “tecnología”. Los DPI actualmente rigen para las actividades comerciales, mientras que el conocimiento generado por dichas comunidades se considera de dominio público, pese a su valor económico, Se analiza brevemente el papel de los derechos de propiedad intelectual en cada subsistema…

El papel de la FIFA en la soberanía del Estado. Una aproximación a la relación entre actores y sujetos del derecho internacional en la copa del mundo de fútbol Sudáfrica 2010

Este estudio de caso se realiza con el ánimo de analizar las relaciones de la actividad de la FIFA con la soberanía del Estado, teniendo como objeto de estudio principal la Copa del Mundo Sudáfrica 2010. Asimismo, se busca analizar el papel y categorización de la FIFA, tanto en las relaciones internacionales, como en el derecho internacional. Para tal fin, a lo largo del trabajo se procederá a exponer los principales aspectos en las relaciones entre actores no tradicionales y los Estados soberanos, se identificará la conexión existente entre la FIFA y el Soft Law. Por último, se describirán las acciones concretas que realizó la FIFA que de alguna manera afectaron la soberanía sudafricana.