Light reception and circadian behavior in 'blind' and 'clock-less' mutants of Neurospora crassa

The filamentous fungus Neurospora crassa is a model organism for the genetic dissection of blue light photoreception and circadian rhythms. WHITE COLLAR-1 (WC-1) and WC-2 are considered necessary for all light responses, while FREQUENCY (FRQ) is required for light-regulated asexual development (conidia formation); without any of the three, self-sustained (circadian) rhythmicity in constant conditions fails. Here we show that light-regulated and self-sustained development occur in the individual or mutant white collar strains. These strains resemble wild type in their organization of the daily bout of light-regulated conidiation. Molecular profiles of light- induced genes indicate that the individual white collar-1 and white collar-2 mutants utilize distinct pathways, despite their similar appearance in all aspects. Titration of fluence rate also demonstrates different light sensitivities between the two strains. The data require the existence of an as-yet-unidentified photoreceptor. Furthermore, the extant circadian clock machinery in these mutant strains supports the notion that the circadian system in Neurospora involves components outside the WC-FRQ loop.     

Ordination and classification of vegetation along an altitudinal gradient in the Venezuelan páramos

By means of ordination and classification techniques, the relationships between climate, soils, human activities and vegetation along an altitudinal gradient of the Venezuelan páramos are analyzed and interpreted. The altitudinal gradient chosen is characterized by decrease of temperature, precipitation, soil fertility, soil water-holding capacity, and plant cover as altitude increases. The ordination results suggest vegetation changes to be primarily related to environmental changes occurring with altitude, and secondly to disturbances caused mainly by grazing. Some results point toward a disjunction in the vegetational gradient occurring at ca. 3 500 m.a.s.l. and separating low and high páramo. This disjunction might have been caused by the glacial history of the páramos and the occurrence of frequent night-frosts.The soil samples were kindly analyzed by the Laboratorio de Edafologia, Centro Nacional de Investigaciones Agropecuarias. Help in plant identification was generously obtained from the specialists of Instituto Botánico, Instituto Nacional de Parques, Caracas.Nomenclature follows Vareschi (1970).Acknowledgements: This work was supported by the Decanato de Investigaciones, Universidad Simón Bolivar. I wish to thank A. Pacheco for help in the field sampling. Dr O. Arenas help was invaluable in the mathematical treatment of the data. Drs A. Vivas and J. M. B. Smith provided useful criticism to an earlier version of this work.     

Patterns of energy content in plants from the venezuelan paramos

Summary Determinations of energy and ash content were made on the organs of some common plant species from an elevational gradient in the Venezuelan páramos. The results indicate differences in energy content among the life forms sampled. Giant rosette plants show the highest values in the roots while woody plants have the highest energy content in the leaves. The reproductive organs show a higher energy content than the vegetative organs. There is a tendency to increase energy content as elevation and environmental stress increase. The adaptive significance of these patterns is discussed. The high average energy value for páramo plants, 20.18 KJ/g ash free dry weight, is similar to values reported for alpine tundra plants and conifer forests.     

Regulation of Hsp70 function by HspBP1: structural analysis reveals an alternate mechanism for Hsp70 nucleotide exchange

HspBP1 belongs to a family of eukaryotic proteins recently identified as nucleotide exchange factors for Hsp70. We show that the S. cerevisiae ortholog of HspBP1, Fes1p, is required for efficient protein folding in the cytosol at 37 degrees C. The crystal structure of HspBP1, alone and complexed with part of the Hsp70 ATPase domain, reveals a mechanism for its function distinct from that of BAG-1 or GrpE, previously characterized nucleotide exchange factors of Hsp70. HspBP1 has a curved, all alpha-helical fold containing four armadillo-like repeats unlike the other nucleotide exchange factors. The concave face of HspBP1 embraces lobe II of the ATPase domain, and a steric conflict displaces lobe I, reducing the affinity for nucleotide. In contrast, BAG-1 and GrpE trigger a conserved conformational change in lobe II of the ATPase domain. Thus, nucleotide exchange on eukaryotic Hsp70 occurs through two distinct mechanisms.     

Optimization of cultivation conditions in spin tubes for Chinese hamster ovary cells producing erythropoietin and the comparison of glycosylation patterns in different cultivation vessels

This article describes the optimization of cultivation factor settings, that is the shaking rate and working volume in 50 mL spin tubes for a Chinese hamster ovary cell line expressing recombinant human α‐erythropoietin, using a response D‐optimal surface method. The main objectives of the research were, firstly, to determine a setting in which the product titer and product quality attributes in spin tubes are equivalent to those in 250 mL shake flasks in a seven day batch and, secondly, to find a setting in which the product titer is maximal. The model for product titer prediction as a function of shaking rate and working volume in the defined design space was successfully applied to the optimization of cultivation conditions in spin tubes for the tested cell line. Subsequently, validation experiments were carried out simultaneously in spin tubes, shake flasks and bench scale bioreactors to compare cell culture performance parameters such as growth, productivity and product quality attributes in the form of isoform profiles and glycan antennarity structures. The results of the experiments showed that similar cell culture performance and product quality could be achieved in spin tubes when compared to shake flasks. Additionally, bioreactor titers could be reproduced in spin tubes at high shaking rates and low working volumes, but with differing product quality. Cultivation at lower shaking rates in spin tubes and shake flasks produced a glycoprotein with a product quality slightly comparable to that from bioreactors, but with titers being only two thirds. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

Improved Visualization of Intracranial Vessels with Intraoperative Coregistration of Rotational Digital Subtraction Angiography and Intraoperative 3D Ultrasound

Introduction

Ultrasound can visualize and update the vessel status in real time during cerebral vascular surgery. We studied the depiction of parent vessels and aneurysms with a high-resolution 3D intraoperative ultrasound imaging system during aneurysm clipping using rotational digital subtraction angiography as a reference.

Methods

We analyzed 3D intraoperative ultrasound in 39 patients with cerebral aneurysms to visualize the aneurysm intraoperatively and the nearby vascular tree before and after clipping. Simultaneous coregistration of preoperative subtraction angiography data with 3D intraoperative ultrasound was performed to verify the anatomical assignment.

Results

Intraoperative ultrasound detected 35 of 43 aneurysms (81%) in 39 patients. Thirty-nine intraoperative ultrasound measurements were matched with rotational digital subtraction angiography and were successfully reconstructed during the procedure. In 7 patients, the aneurysm was partially visualized by 3D-ioUS or was not in field of view. Post-clipping intraoperative ultrasound was obtained in 26 and successfully reconstructed in 18 patients (69%) despite clip related artefacts. The overlap between 3D-ioUS aneurysm volume and preoperative rDSA aneurysm volume resulted in a mean accuracy of 0.71 (Dice coefficient).

Conclusions

Intraoperative coregistration of 3D intraoperative ultrasound data with preoperative rotational digital subtraction angiography is possible with high accuracy. It allows the immediate visualization of vessels beyond the microscopic field, as well as parallel assessment of blood velocity, aneurysm and vascular tree configuration. Although spatial resolution is lower than for standard angiography, the method provides an excellent vascular overview, advantageous interpretation of 3D-ioUS and immediate intraoperative feedback of the vascular status. A prerequisite for understanding vascular intraoperative ultrasound is image quality and a successful match with preoperative rotational digital subtraction angiography.     

Hemiarthroplasty is an effective surgical method to manage unstable trochanteric fractures in elderly people

Trochanteric femoral fractures are a major problem in the elderly because of higher bone fragility due to osteoporosis. Numerous chronic illnesses, which usually affect the elderly, aggravate and complicate their surgical treatment. Trochanteric femoral fractures results in high morbidity and mortality in elderly patients. The aim of our study is to evaluate the effectiveness of hemiarthroplasty in the treatment of unstable trochanteric femoral fractures in elderly patients. Between 2000 and 2005, 50 patients with unstable trochanteric femoral fractures (41 women) aged 75 to 92 years (mean 86 years) underwent cemented hemiarthroplasty. The surgical procedure was performed within first 48 hours after the fracture (out of which 14 in the first 12 hours, 27 in the first 24 hours and 9 in the first 48 hours), with minimal blood loss. Hemiarthroplasty was indicated in patients where stability was important to allow early mobilization. In forty patients (80%) early ambulation with full weight bearing was achieved during the short period of hospitalization (9-14 days). Given that the affected population is predominantly the elderly, who are less mobile and demanding and thus put less strain on the endoprosthesis, we believe that this kind of treatment is the treatment of choice for unstable trochanteric femoral fractures in these patients. This assumption is corroborated by the fact that we did not have any endoprosthesis luxation, apparent acetabular protrusion or instability during the mean follow up period of 15 months (range 12-18 months).     

Formation of adhesions at surgical meshes in a rat experimental model

Abdominal wall hernias are surgical problem that are easily solved with laparoscopic surgery. The determining factor for the success of the operation is the right choice and use of surgical mesh as the support material. The most common complication of surgical mesh placement is the formation of adhesions. Aim of this paper is to determine whether there is a statistic difference in formation of adhesions between different surgical meshes in lab environment. Wistar rats were used as the experimental model. After the anaesthesia a 1x1 cm defect of the abdominal wall was made, but the skin was left intact. The mesh was placed directly on the internal organs. The experiment considered four different mesh types. After set time periods of one, two or four weeks the animals were sacrificed and the amount of formed adhesions were evaluated based on the modified Diamond scale. Immediately after the first week we found a statistically significant difference in the adhesion occurrence rate between compared materials. The smallest amount of adhesions was caused by polypropylen + polydoksanon mesh, and the most by polypropilen mesh. Polypropylen + polyglactin mesh showed significant reduction of adhesion formation between the tested weeks. We can conclude that polypropylen + polydoxanon meshes are superior for ventral hernia operation, because those defects are in close contact with the internal organs and it is very important to have the smallest amount of adhesions.     

Novel series of pyrrolotriazine analogs as highly potent pan-Aurora kinase inhibitors

The synthesis and SAR for a novel series of pyrrolotriazines as pan-Aurora kinase inhibitors are described. Optimization of the cyclopropane carboxamide terminus of lead compound 1 resulted in analogs with high cellular activity and improved rat PK profiles. Notably, compound 17l demonstrated tumor growth inhibition in a mouse xenograft model.