Third generation of matrix metalloprotease inhibitors: Gain in selectivity by targeting the depth of the S1′ cavity

Following the disappointment of clinical trials with early broad-spectrum synthetic inhibitors of matrix metalloproteases (MMPs), the field is now resurging with a new focus on the development of more selective inhibitors. Compounds able to fully discriminate between different members of the MMP family are sorely needed for therapeutic applications. Chemical efforts over the past years have led to very few selective inhibitors of MMPs. The over-exploitation of the hydroxamate function, or other strong zinc-binding groups, might be responsible for this failure. By resorting to weaker zinc-chelating groups, like phosphoryl or carboxylic groups, inhibitors with improved selectivity profiles have been developed. However, the most encouraging results have been obtained with compounds that avoid targeting the zinc but gain their affinity from plunging deeper into the MMP S₁′ cavity. Analyses of the crystal structures of MMP-13 and MMP-8 complexes with such compounds provide novel insights for the design of more selective inhibitors for other members of the MMP family.     

A solid-phase assay for the quantitative analysis of hyaluronic acid at the nanogram level

A sensitive and accurate solid-phase assay for the quantitative determination of hyaluronic acid (HA) is described. The wells of the polystyrene microplates used were coated with glutaraldehyde followed, via a Schiff's base bond, with spermine to introduce amino groups. HA was added to the activated microwells in the presence of carbodiimide and left to bind via a peptide bond to the amino groups. Then aggrecan solution was added to the wells of the microtiter plates to interact with its G1 domain with hyaluronic acid, and the amounts of aggrecan bound were measured immunochemically. The inhibition of the binding between aggrecan and immobilized HA due to soluble HA present in reference solutions showed linearity in the range of concentrations 0.1 to 0.7 microg/ml. The reaction is specific and rapid and can be widely used for the calculation of HA in body fluids directly and in tissue samples after a brief digestion with a proteolytic enzyme.     

Predicting the consequences of species loss using size‐structured biodiversity approaches

Understanding the consequences of species loss in complex ecological communities is one of the great challenges in current biodiversity research. For a long time, this topic has been addressed by traditional biodiversity experiments. Most of these approaches treat species as trait‐free, taxonomic units characterizing communities only by species number without accounting for species traits. However, extinctions do not occur at random as there is a clear correlation between extinction risk and species traits. In this review, we assume that large species will be most threatened by extinction and use novel allometric and size‐spectrum concepts that include body mass as a primary species trait at the levels of populations and individuals, respectively, to re‐assess three classic debates on the relationships between biodiversity and (i) food‐web structural complexity, (ii) community dynamic stability, and (iii) ecosystem functioning. Contrasting current expectations, size‐structured approaches suggest that the loss of large species, that typically exploit most resource species, may lead to future food webs that are less interwoven and more structured by chains of interactions and compartments. The disruption of natural body‐mass distributions maintaining food‐web stability may trigger avalanches of secondary extinctions and strong trophic cascades with expected knock‐on effects on the functionality of the ecosystems. Therefore, we argue that it is crucial to take into account body size as a species trait when analysing the consequences of biodiversity loss for natural ecosystems. Applying size‐structured approaches provides an integrative ecological concept that enables a better understanding of each species' unique role across communities and the causes and consequences of biodiversity loss.     

High content selection of cells

Conventional methods for clonal analysis are mostly based on limiting dilution or FACS sorting. Purity of clones can only be verified retrospectively. This process is tedious and time-consuming. The Elektra™ improves and expedites the cell cloning process using IACS™ (Image Activated Cell Selection) for isolation of clonal cells providing high-content single cell information. A micro fluidic Sorter Chip within the Elektra™ uses CellProcessor technology, i. e. negative dielectrophoretic (nDEP) force to guide, cage and sort cells (about 500 to 1000 cells per run). The system is equipped with a highly sensitive CCD camera and a 40× lens. Cells are analyzed on-line and selected according to phase contrast, fluorescence and size while passing the micro device. Target cells are trapped and an image series can be acquired. Post process evaluation of the images enables quantitative analysis of e. g. subcellular fluorescence distribution and co-localization. Elektra™ produces micro titer plates containing single viable and 100% pure clones without the need for multiple iterations. This is demonstrated for different cell lines (e. g. CHO, U2-OS, hybridoma cells). The technology is shown to be compatible to label free detection techniques like impedance, electrorotation and dielectrophoretic cell deformation.     

Reductions in the incidence of nitrofen-induced diaphragmatic hernia by vitamin A and retinoic acid

Congenital diaphragmatic hernia (CDH) is a serious medical condition in which the developing diaphragm forms incompletely, leaving a hole through which the abdominal contents can enter the thoracic space and interfere with lung growth. A perturbation of the retinoid system has been linked to the etiology of CDH. This includes findings that nitrofen, which induces CDH in rodents, inhibits the key enzyme for retinoic acid (RA) production, retinaldehyde dehydrogenase-2 (RALDH2) in vitro. Published studies indicate that antenatal vitamin A administration on gestational day (D) 12 in the nitrofen model of CDH reduced the severity and incidence of right-sided defects and lung hypoplasia. In this study, we administered nitrofen on D8, to include the induction of clinically more prevalent left-sided defects, and examined the efficacy of several vitamin A administration paradigms to gain insights into the developmental stage of susceptibility. Furthermore, we tested the hypothesis that administration of RA, the product of RALDH2 activity, is more potent than administering the substrate, vitamin A, in reducing the incidence of CDH. The incidence of CDH was reduced from approximately 54% (nitrofen alone) to approximately 32% with vitamin A treatment. The efficacy of RA treatment was very marked, with a reduction in the incidence of CDH to approximately 15%. Administration of vitamin A or RA on approximately D10 was most effective. These data lend further support for the potential involvement of retinoid signaling pathways and the etiology of CDH and support data from in vitro studies demonstrating a nitrofen-induced suppression of RALDH2.     

Rapid alterations of serum fatty acids with the intravenous administration of an arachidonate solution

Polyunsaturated fatty acids (PUFAs) have been shown to possess a considerable anti-tumor and anti-bacterial effect in vitro. In an attempt to achieve serum concentrations of these acids similar to those applied in vitro, a solution of ethyl ester of arachidonic acid (AA) was administered intravenously at 25 mg/kg within 10 min in six male rabbits. Blood samples were collected before and 60 min after infusion from catheters inserted in the hepatic veins and in the carotid artery. Analysis of serum fatty acids was performed by gas chromatography mass spectrometry. Elevated concentrations of elongated fatty acids were detected in the hepatic veins after infusion. Mean concentrations of arachidonate in the hepatic veins and the carotid arteries after infusion of AA were 2.77 and 3.73 microM, respectively. It is concluded that the intravenous administration of a solution of AA might result in increased hepatic biosynthesis of serum saturated and unsaturated fatty acids of elongated carbon chains. The increasing interest for the application of PUFAs in therapeutics renders further study mandatory to clarify the significance of these findings.     

Crucial role of the peroxyketal function for antimalarial activity in the G-factor series

New endoperoxides, related to the natural phytohormones known as G factors (G1, G2, G3), were modified on the side chain and the ketalic position. An unexpected rearrangement, specific to one diastereoisomer was observed in the deprotection step of O-silylated compounds and attributed to a hexacoordinated fluorosilicon intermediate. The reduction potential of these new peroxides was determined. They exhibited good to moderate antimalarial activity, greatly related to the presence of peroxyketal function.     

Test of pangamy by genetic analysis of Schistosoma mansoni pairs within its natural murine host in Guadeloupe

Mating system plays a determinant role in the maintenance and distribution of genetic variations. It can be assessed indirectly by analyzing the distribution of the genetic variability within populations or directly by considering how mating pairs are formed. In the present study, 71 pairs of adult Schistosoma mansoni worms sampled from naturally infected rats were genotyped to investigate how male and female schistosomes paired according to their genetic relatedness. Among all samples, pangamy, the random association between males and females, could not be rejected. Whereas the schistosome mating system has been intensively studied under experimental conditions, to the best of our knowledge, our study is the first to attempt to understand the way in which males and females pair in natural conditions.     

Upregulation of DNA repair genes in active cirrhosis associated with hepatocellular carcinoma

Phenotypic changes in injured livers involve complex network of genes whose interplays may lead to fibrosis and cirrhosis, a major risk of hepatocellular carcinoma. Gene expression profiles in fibrotic livers were analyzed by using cDNA microarray, hierarchical clustering and gene ontology. Analyses of a major cluster of upregulated genes in cirrhosis identified a new set of genes involved in DNA repair and damage. The upregulation of DNA repair genes was confirmed by real-time quantitative polymerase chain reaction and associated with necroinflammatory activity (P<0.001). Increased DNA repair activity in cirrhosis with inflammatory activity may reflect increased DNA damages as a consequence of chronic liver injury.