The NOX toolbox: validating the role of NADPH oxidases in physiology and disease

Reactive oxygen species (ROS) are cellular signals but also disease triggers; their relative excess (oxidative stress) or shortage (reductive stress) compared to reducing equivalents are potentially deleterious. This may explain why antioxidants fail to combat diseases that correlate with oxidative stress. Instead, targeting of disease-relevant enzymatic ROS sources that leaves physiological ROS signaling unaffected may be more beneficial. NADPH oxidases are the only known enzyme family with the sole function to produce ROS. Of the catalytic NADPH oxidase subunits (NOX), NOX4 is the most widely distributed isoform. We provide here a critical review of the currently available experimental tools to assess the role of NOX and especially NOX4, i.e. knock-out mice, siRNAs, antibodies, and pharmacological inhibitors. We then focus on the characterization of the small molecule NADPH oxidase inhibitor, VAS2870, in vitro and in vivo, its specificity, selectivity, and possible mechanism of action. Finally, we discuss the validation of NOX4 as a potential therapeutic target for indications including stroke, heart failure, and fibrosis

Investigating the microstructure of plant leaves in 3D with lab-based X-ray Computed Tomography

Background Leaf cellular architecture plays an important role in setting limits for carbon assimilation and, thus, photosynthetic performance. However, the low density, fine structure, and sensitivity to desiccation of plant tissue has presented challenges to its quantification. Classical methods of tissue fixation and embedding prior to 2D microscopy of sections is both laborious and susceptible to artefacts that can skew the values obtained. Here we report an image analysis pipeline that provides quantitative descriptors of plant leaf intercellular airspace using lab-based X-ray Computed Tomography (microCT). We demonstrate successful visualisation and quantification of differences in leaf intercellular airspace in 3D for a range of species (including both dicots and monocots) and provide a comparison with a standard 2D analysis of leaf sections. Results We used the microCT image pipeline to obtain estimates of leaf porosity and mesophyll exposed surface area (Smes) for three dicot species (Arabidopsis, tomato and pea) and three monocot grasses (barley, oat and rice). The imaging pipeline consisted of (1) a masking operation to remove the background airspace surrounding the leaf, (2) segmentation by an automated threshold in ImageJ and then (3) quantification of the extracted pores using the ImageJ ‘Analyze Particles’ tool. Arabidopsis had the highest porosity and lowest Smes for the dicot species whereas barley had the highest porosity and the highest Smes for the grass species. Comparison of porosity and Smes estimates from 3D microCT analysis and 2D analysis of sections indicates that both methods provide a comparable estimate of porosity but the 2D method may underestimate Smes by almost 50%. A deeper study of porosity revealed similarities and differences in the asymmetric distribution of airspace between the species analysed. Conclusions Our results demonstrate the utility of high resolution imaging of leaf intercellular airspace networks by lab-based microCT and provide quantitative data on descriptors of leaf cellular architecture. They indicate there is a range of porosity and Smes values in different species and that there is not a simple relationship between these parameters, suggesting the importance of cell size, shape and packing in the determination of cellular parameters proposed to influence leaf photosynthetic performance

Thermal effect on the life-cycle parameters of the medically important freshwater snail species lymnaea (radix) luteola (lamarck)

The snails Lymnaea (Radix) luteola exhibited marked variations in growth, longevity, and attaining sexual maturity at different temperatures and diets. At 10 C, irrespective of foods, pH and salinity of water, the snails had minimum life span, maximum death rate and lowest growth rate. At 15 C, the growth rate was comparatively higher and the snails survived for a few more days. But at these temperatures they failed to attain sexual maturity. Snails exposed to pH 5 and 9 at 20 , 25 , 30 , 35 C and room temperatures (19.6 -29.6 C); to 0.5, 1.5 and 2.5 NaCl at 20 and 35 C; to 2.5 NaCl at 25 C and room temperatures failed to attain sexual maturity. The snails exposed to pH 7 and different salinity grades at 20 , 25 , 30 , 35 C and room temperatures became sexually mature between 25-93 days depending upon the type of foods used in the culture