Preparation of protoplasts from the green alga Enteromorpha intestinalis (L.) Link

Protoplasts have been obtained from vegetative thallus of the green seaweed Enteromorpha following enzymic digestion with driselase and pectinase. The viability of purified protoplast fractions was assessed by staining and measurements of O₂ uptake and evolution.Abbreviations MES 2-(N-morpholino) ethanesulphonic acid - TES N-tris(hydoxymethyl) methyl-2 aminoethanesulphonic acid     

The role of monocyte/macrophages as vehicles of dissemination of Simkania negevensis: an in vitro simulation model

Exposure to Simkania negevensis (Sn), an intracellular microorganism that has been associated with respiratory tract infections in infants and adults, is prevalent. Sn can multiply within free-living amoebae and has been detected in domestic water supplies, which may constitute a source of infection with the organism. Its path of transport from its portal of entry to the body to its target organs is unknown. In this study, the possibility that monocytes/macrophages may serve as vehicles of transmission was examined. In vitro cocultivation of Sn-infected Acanthamoeba polyphaga with the monocyte/macrophage cell line U937 resulted in the death of the amoebae and infection of the U937 cells. Sn entered and multiplied in U937 cells within short periods of time, and the microorganism could be transferred from U937 cells to cell cultures of various origins. Uninfected monocyte/macrophages could become infected when in contact with either actively or persistently Sn-infected cell cultures. Persistently infected cultures in contact with uninfected U937 cells became actively infected. The results of this study provide a basis for determination of the molecular mechanisms of monocyte/macrophage-cell interactions in transfer of infection and may contribute to a better understanding of the pathogenesis of Sn infections in vivo.     

A centralized model for creating shared,standardized, microsatellite data that simplifies inter-laboratory collaboration

We demonstrate an efficient model for standardizing microsatellite DNA data among laboratories studying Oncorhynchus mykiss. Eight laboratories standardized 13 microsatellite loci following allele nomenclature of a central laboratory (average inter-laboratory genotyping concordance >98%). Following this central model, we have currently standardized 298 alleles from throughout the species native range. Although we focus here on O. mykiss, our experiences and recommendation apply equally to other broadly distributed species that may benefit from multi-laboratory collaborative data collection.     

The contribution of naturally occurring polymorphisms in altering the biochemical and structural characteristics of HIV-1 subtype C protease

Fourteen subtype B and C protease variants have been engineered in an effort to study whether the preexistent baseline polymorphisms, by themselves or in combination with drug resistance mutations, differentially alter the biochemical and structural features of the subtype C protease when compared with those of subtype B protease. The kinetic studies performed in this work showed that the preexistent polymorphisms in subtype C protease, by themselves, do not provide for a greater level of resistance. Inhibition analysis with eight clinically used protease inhibitors revealed that the natural polymorphisms found in subtype C protease, in combination with drug resistance mutations, can influence enzymatic catalytic efficiency and inhibitor resistance. Structural analyses of the subtype C protease bound to nelfinavir and indinavir showed that these inhibitors form similar interactions with the residues in the active site of subtype B and C proteases. It also revealed that the naturally occurring polymorphisms could alter the position of the outer loops of the subtype C protease, especially the 60's loop.     

Life-cycle assessment and techno-economic analysis of biochar produced from forest residues using portable systems

The International Journal of Life Cycle Assessment - Producing biochar from forest residues can help resolve environmental issues by reducing forest fires and mitigating climate change. However,...     

Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

The mouse is now the primary animal used to model a variety of lung diseases. To study the mechanisms that underlie such pathologies, phenotypic methods are needed that can quantify the pathologic changes. Furthermore, to provide translational relevance to the mouse models, such measurements should be tests that can easily be done in both humans and mice. Unfortunately, in the present literature few phenotypic measurements of lung function have direct application to humans. One exception is the diffusing capacity for carbon monoxide, which is a measurement that is routinely done in humans. In the present report, we describe a means to quickly and simply measure this diffusing capacity in mice. The procedure involves brief lung inflation with tracer gases in an anesthetized mouse, followed by a 1 min gas analysis time. We have tested the ability of this method to detect several lung pathologies, including emphysema, fibrosis, acute lung injury, and influenza and fungal lung infections, as well as monitoring lung maturation in young pups. Results show significant decreases in all the lung pathologies, as well as an increase in the diffusing capacity with lung maturation. This measurement of lung diffusing capacity thus provides a pulmonary function test that has broad application with its ability to detect phenotypic structural changes with most of the existing pathologic lung models.     

Design and synthesis of MMP inhibitors with appended fluorescent tags for imaging and visualization of matrix metalloproteinase enzymes

We describe the synthesis, MMP-2 and 9 potency, and in vitro evaluation of a series of α-sulfone hydroxmate MMP inhibitors conjugated to a series of dyes with different absorption/emission lamina maxima’s that can be used to visualize tumors.