An “Inclusion-Body-Like” Configuration of Some Cell Nuclei in Moose

At routine post-mortem of a moose an inclusion-body-like configuration of the nuclei of the cells in the granule layer of the cerebellum was found. This was also observed to a lesser extent in glial cells of the cerebrum, epithelial cells of the collecting tubules of kidney and in the hepatocytes.     

Effects in vitro of alloxan on the glucose metabolism of mouse pancreatic B-cells

To facilitate detailed studies of the B-cytotoxic action of alloxan we developed a model using isolated pancreatic islets of normal mice. An essential feature of this model is the low temperature employed during exposure to alloxan, which minimizes the degradation of the drug. The islets were incubated with alloxan for 30min at 4 degrees C and subsequently various aspects of their metabolism were studied. The O(2) consumption was measured by the Cartesian-diver technique. Islets exposed to 2mm-alloxan and control islets had the same endogenous respiration, whereas the O(2) uptake of the alloxan-treated islets was inhibited and that of the control islets stimulated when they were incubated with 28mm-glucose as an exogenous substrate. The islet glucose oxidation was estimated by measurement of the formation of (14)CO(2) from [U-(14)C]glucose at 37 degrees C. Compared with the controls, alloxan-treated islets showed a decrease in the glucose-oxidation rate in a dose-dependent manner. Pretreatment of the islets with 28mm-glucose for 30min at 37 degrees C completely protected against this effect, whereas preincubations at glucose concentrations below 16.7mm failed to exert any protective effect. The glucose utilization was estimated as the formation of (3)H(2)O from [5-(3)H]glucose. Alloxan (2mm) failed to affect islet glucoseutilization rate in the presence of either 2.8 or 28mm-glucose. In contrast, islets exposed to 5 or 10mm-alloxan exhibited lowered glucose utilization. It is concluded that in vitro alloxan has an acute inhibitory effect on the islet glucose metabolism, and that this action can be prevented by previous exposure to a high glucose concentration. The results are consistent with the idea that the B-cytotoxicity of alloxan reflects an interaction with intracellular sites involved in the oxidative metabolism of the B-cell.     

Comparison of Light and Electron Microscopic Determinations of the Number of Bacteria and Algae in Lake Water

Determinations of the number of microorganisms in lake water samples with the bright-field light microscope were performed using conventional counting chambers. Determinations with the fluorescence microscope were carried out after staining the organisms with acridine orange and filtering them onto Nuclepore filters. For transmission electron microscopy, a water sample was concentrated by centrifugation. The pellet was solidifed in agar, fixed, dehydrated, embedded in Epon, and cut into thin sections. The number and area of organism profiles per unit area of the sections were determined. The number of organisms per unit volume of the pellet was then calculated using stereological formulae. The corresponding number in the lake water was obtained from the ratio of volume of solidified pellet/volume of water sample. Control experiments with pure cultures of bacteria and algae showed good agreement between light and electron microscopic counts. This was also true for most lake water samples, but the electron microscopic preparations from some samples contained small vibrio-like bodies and ill-defined structures that made a precise comparison more difficult. Bacteria and small blue-green and green algae could not always be differentiated with the light microscope, but this was easily done by electron microscopy. Our results show that transmission electron microscopy can be used for checking light microscopic counts of microorganisms in lake water.     

SOMA: A Single Oligonucleotide Mutagenesis and Cloning Approach

Modern biology research requires simple techniques for efficient and restriction site-independent modification of genetic material. Classical cloning and mutagenesis strategies are limited by their dependency on restriction sites and the use of complementary primer pairs. Here, we describe the Single Oligonucleotide Mutagenesis and Cloning Approach (SOMA) that is independent of restriction sites and only requires a single mutagenic oligonucleotide to modify a plasmid. We demonstrate the broad application spectrum of SOMA with three examples. First, we present a novel plasmid that in a standardized and rapid fashion can be used as a template for SOMA to generate GFP-reporters. We successfully use such a reporter to assess the in vivo knock-down quality of morpholinos in Xenopus laevis embryos. In a second example, we show how to use a SOMA-based protocol for restriction-site independent cloning to generate chimeric proteins by domain swapping between the two human hRMD5a and hRMD5b isoforms. Last, we show that SOMA simplifies the generation of randomized single-site mutagenized gene libraries. As an example we random-mutagenize a single codon affecting the catalytic activity of the yeast Ssy5 endoprotease and identify a spectrum of tolerated and non-tolerated substitutions. Thus, SOMA represents a highly efficient alternative to classical cloning and mutagenesis strategies.     

Local temperatures inferred from plant communities suggest strong spatial buffering of climate warming across Northern Europe

Recent studies from mountainous areas of small spatial extent (<2500 km²) suggest that fine‐grained thermal variability over tens or hundreds of metres exceeds much of the climate warming expected for the coming decades. Such variability in temperature provides buffering to mitigate climate‐change impacts. Is this local spatial buffering restricted to topographically complex terrains? To answer this, we here study fine‐grained thermal variability across a 2500‐km wide latitudinal gradient in Northern Europe encompassing a large array of topographic complexities. We first combined plant community data, Ellenberg temperature indicator values, locally measured temperatures (LmT) and globally interpolated temperatures (GiT) in a modelling framework to infer biologically relevant temperature conditions from plant assemblages within <1000‐m² units (community‐inferred temperatures: CiT). We then assessed: (1) CiT range (thermal variability) within 1‐km² units; (2) the relationship between CiT range and topographically and geographically derived predictors at 1‐km resolution; and (3) whether spatial turnover in CiT is greater than spatial turnover in GiT within 100‐km² units. Ellenberg temperature indicator values in combination with plant assemblages explained 46–72% of variation in LmT and 92–96% of variation in GiT during the growing season (June, July, August). Growing‐season CiT range within 1‐km² units peaked at 60–65°N and increased with terrain roughness, averaging 1.97 °C (SD = 0.84 °C) and 2.68 °C (SD = 1.26 °C) within the flattest and roughest units respectively. Complex interactions between topography‐related variables and latitude explained 35% of variation in growing‐season CiT range when accounting for sampling effort and residual spatial autocorrelation. Spatial turnover in growing‐season CiT within 100‐km² units was, on average, 1.8 times greater (0.32 °C km^?1) than spatial turnover in growing‐season GiT (0.18 °C km^?1). We conclude that thermal variability within 1‐km² units strongly increases local spatial buffering of future climate warming across Northern Europe, even in the flattest terrains.     

A Novel Implantable Glaucoma Valve Using Ferrofluid

Purpose

To present a novel design of an implantable glaucoma valve based on ferrofluidic nanoparticles and to compare it with a well-established FDA approved valve.

Setting

Massachusetts Eye & Ear Infirmary, Boston, USA.

Methods

A glaucoma valve was designed using soft lithography techniques utilizing a water-immiscible magnetic fluid (ferrofluid) as a pressure-sensitive barrier to aqueous flow. Two rare earth micro magnets were used to calibrate the opening and closing pressure. In-vitro flow measurements were performed to characterize the valve and to compare it to Ahmed™ glaucoma valve. The reliability and predictability of the new valve was verified by pressure/flow measurements over a period of three months and X-ray diffraction (XRD) analysis over a period of eight weeks. In vivo assessment was performed in three rabbits.

Results

In the in vitro experiments, the opening and closing pressures of the valve were 10 and 7 mmHg, respectively. The measured flow/pressure response was linearly proportional and reproducible over a period of three months (1.8 µl/min at 12 mmHg; 4.3 µl/min at 16 mmHg; 7.6 µl/min at 21 mmHg). X-ray diffraction analysis did not show oxidization of the ferrofluid when exposed to water or air. Preliminary in vivo results suggest that the valve is biocompatible and can control the intraocular pressure in rabbits.

Conclusions

The proposed valve utilizes ferrofluid as passive, tunable constriction element to provide highly predictable opening and closing pressures while maintaining ocular tone. The ferrofluid maintained its magnetic properties in the aqueous environment and provided linear flow to pressure response. Our in-vitro tests showed reliable and reproducible results over a study period of three months. Preliminary in-vivo results were very promising and currently more thorough investigation of this device is underway.     

Oxygen-depleted surfaces: a new antifouling technology

A novel, non-toxic strategy to combat marine biofouling is presented. The technology is paint with additions of up to 43% of industrial protein. Through microbial degradation of the protein component, an oxygen-depleted layer rapidly forms in a 0.2 mm layer close to the paint surface. With the present paint formulations, a stable, O₂-depleted layer can persist for 16 weeks. Barnacle larvae (cyprids) did not settle on panels where oxygen saturation was <20%, and cyprids were killed when exposed to O₂-free water for more than 1 h. It is also shown that the O₂-depleted layer will rapidly reform (within 15 min) after exposure to turbulent flow. Field exposure of panels for 16 weeks showed that paint with protein reduced fouling by barnacles and bryozoans by 80% and close to 100%, respectively. The results suggest that this novel technology may be developed into a non-toxic alternative to copper-based antifouling paints, especially for pleasure boats in sensitive environments. There is clearly potential for further development of the paint formulation, and a full-scale test on a boat-hull suggested that service-life under realistic operations needs to be improved.