Ultrastructural and cytochemical examination of the nuclear crystalloid inclusions of Olea europaea L. mesophyll cells

Summary The nuclei of mesophyll cells of olive trees contain numerous sizeable crystalloid inclusions. Cytochemical examination using epoxy resin-embedded, semithin-sectioned tissue indicated the presence of proteins and oligoor polysaccharides in these inclusions. Their electron microscopical analysis revealed a crystalline substructure consisting of intersected subunits of high order. The spacing of the lattice fibrils and the angles of intersection were determined and used to establish a model of the unit cell of crystallization. It is suggested that the nuclear crystalloids of olive trees consist of glycoprotein molecules. They differ from the intranuclear crystalloids observed in other species predominantly in the high density of their subunit arrangement.     

Carbon oxidation and incorporation patterns in batch cultures ofMethylomonas L3

Summary A recently developed methodology of directly measuring the oxidation and incorporation patterns of carbon substrate in continuous cultures of RuMP-type methylotrophs is extended to batch cultures of the obligate methylotrophMethylomonas L3. The ratio of cyclic to total substrate oxidation varies with the initial methanol concentration from 0 to 68%. Formaldehyde, as a methanol cosubstrate, enhances the net substrate oxidation. The substrate oxidation and incorporation pattern is also affected by the state of the culture inoculum.     

Intracellular reaction rates,enzyme activities and biomass yields in Methylomonas L3: growth rate and substrate composition effects

Summary The effects of specific growth rate and medium feed composition on the metabolic reactions of methanol incorporation and oxidation have been studied in carbon-limited, chemostatic cultures of Methylomonas L3. An in situ radioisotopic tracer technique was employed. The in vivo rates of substrate-carbon flow and the corresponding steady-state levels of several key RuMP-type methylotrophic enzymes were determined over a range of dilution rates from 0.19 to 0.41 h^-1 on methanol and methanol/formaldehyde substrates. It was determined that an absolute correlation does not exist between the in vivo specific carbon flux and the in vitro specific activity of any of the key enzymes studied. Oxidation of substrate-carbon via 6-phosphogluconate dehydrogenase is not stringently regulated in this methylotroph and the extent of its operation may be dependent on kinetic factors which make immediate cellular detoxification of formaldehyde imperative. As such, the cyclic oxidation mechanism in this methylotroph does not appear to be coupled to efficient energy utilization, since it was observed that high levels of the cyclic oxidation flux are commensurate with depressed biomass yields.     

The effects of normalization method on antagonistic activity patterns during eccentric and concentric isokinetic knee extension and flexion

The purpose of this study was to compare different normalization methods of electromyographic (EMG) activity of antagonists during isokinetic eccentric and concentric knee movements. Twelve women performed three maximum knee extensions and flexions isometrically and at isokinetic concentric and eccentric angular velocities of 30 °·s⁻¹, 90 °·s⁻¹, 120 °·s⁻¹ and 150 °·s⁻¹. The EMG activity of the vastus lateralis, rectus femoris, vastus medialis and hamstrings was recorded. The antagonist integrated IEMG values were normalized relative to the EMG of the same muscle during an isometric maximal action (static method). The values were also expressed as a percentage of the EMG activity of the same muscle, at the same angle, angular velocity and muscle action (dynamic method) when the muscle was acting as an agonist. Three-way analysis of variance (ANOVA) designs indicated significantly greater IEMG normalized with the dynamic method compared to the EMG derived using the static method (P < 0.05). These differences were more evident at concentric angular velocities and at the first and last 20 ° of the movement. The present findings demonstrate that the method of normalization significantly influences the conclusions on antagonistic activity during isokinetic maximum voluntary efforts. The dynamic method of normalization is more appropriate because it considers the effects of muscle action, muscle length and angular velocity on antagonist IEMG.     

Fluoride effects on leaf cell ultrastructure of olive trees growing in the vicinity of the Aluminium Factory of Greece

Summary Leaves of olive trees growing in the vicinity of the Aluminium Factory of Greece were ultrastructurally investigated in order to determine any malformations caused by environmental air pollutants, especially hydrogen fluoride, in comparison with control samples and normal seasonal senescence. Estimation of some elements accumulated by these leaves showed that they contained high amounts of F and Al attributable to the operation of the nearby factory. The most seriously effected cell components were found to be the mesophyll chloroplasts that show a dilation of the intrathylakoid space, increase of the number of plastoglobuli, discoloration of plastoglobuli, accumulation of large starch grains and an overall disorganized appearance of the organelle. The nuclear crystalloid inclusions have unusual shapes, while the vacuoles contain a fibrillar/granular material that increases their electron density. It is concluded that the ultrastructural malformations are caused by a combination of environmental stresses and air pollutants.This paper is dedicated to Professor Dr. Eberhard Schnepf on the occasion of his 60^th birthday.     

Anti-MRSA agent discovery using Caenorhabditis elegans -based high-throughput screening

Staphylococcus aureus is a leading cause of hospital- and community-acquired infections. Despite current advances in antimicrobial chemotherapy, the infections caused by S. aureus remain challenging due to their ability to readily develop resistance. Indeed, antibiotic resistance, exemplified by methicillin-resistant S. aureus (MRSA) is a top threat to global health security. Furthermore, the current rate of antibiotic discovery is much slower than the rate of antibiotic-resistance development. It seems evident that the conventional in vitro bacterial growth-based screening strategies can no longer effectively supply new antibiotics at the rate needed to combat bacterial antibiotic-resistance. To overcome this antibiotic resistance crisis, screening assays based on host–pathogen interactions have been developed. In particular, the free-living nematode Caenorhabditis elegans has been used for drug screening against MRSA. In this review, we will discuss the general principles of the C. elegans-based screening platform and will highlight its unique strengths by comparing it with conventional antibiotic screening platforms. We will outline major hits from high-throughput screens of more than 100,000 small molecules using the C. elegans–MRSA infection assay and will review the mode-of-action of the identified hit compounds. Lastly, we will discuss the potential of a C. elegans-based screening strategy as a paradigm shift screening platform.     

Effects of bisphenol A on the microtubule arrays in root meristematic cells of Pisum sativum L.

Bisphenol A (BPA), a widely used chemical in the plastics industry that displays weak oestrogenic properties, is an emerging environmental pollutant, potentially harmful to living organisms. The presumed cytotoxicity of BPA to plant cells has been poorly studied. To understand how BPA might influence plant cell division and affect the underlying cytoskeleton, the effects of BPA on the microtubule (MT) arrays of meristematic root-tip cells of Pisum sativum L. were investigated. Root tips of young seedlings were exposed to 20, 50 and 100 mg/L BPA for 1, 3, 6, 12 and 24 h. The effects of each treatment were determined by means of confocal laser scanning microscopy after immunolabelling of tubulin and counterstaining of DNA, and by use of light and transmission electron microscopy. It was found that BPA affected normal chromosome segregation, hampered the completion of cytokinesis and deranged interphase and mitotic MT arrays. BPA effects were dependent on the stage of each cell at the time of BPA entrance. Moreover, BPA induced the formation of macrotubules with a mean diameter of 32 ± 0.14 nm, compared with 23 ± 0.70 nm for the MT arrays in untreated cells. Finally, all MT arrays and macrotubules were depolymerised upon longer treatment. Taken together, the data suggest that BPA exerts acute anti-mitotic effects on meristematic root-tip cells of P. sativum, MT arrays constitute a primary sub-cellular target of BPA toxicity, and the manifested chromosomal abnormalities could be attributed to the disruption of the MT cytoskeleton.     

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.