Cloning of Choriocarcinoma Cells Shows That Invasion Correlates with Expression and Activation of Gelatinase A

Implantation and placental development are dependent upon trophoblast invasion of the endometrium. While the villous trophoblast does not display invasive behavior, the extravillous cytotrophoblast is highly invasive. By cloning BeWo choriocarcinoma cells, we have isolated two distinct clones that share similarities with villous and extravillous cytotrophoblasts. When cultured at the surface of a type I collagen gel, BeWo MC-1 cells were not invasive, whereas BeWo MC-2 cells rapidly invaded this matrix. When injected subcutaneously in nude mice, BeWo MC-1 cells developed a localized tumor and BeWo MC-2 cells developed larger tumors with micrometastases. Gelatinase A expression and minute amounts of gelatinase B were detected in the parental cell line and in both clones. However, the parental and the BeWo MC-2 cells secreted 5- to 10-fold more gelatinase A than the BeWo MC-1 cells. Laminin and matrigel stimulated the production of gelatinase A in BeWo MC-2 cells. Type I collagen promoted the conversion of the 72-kDa progelatinase A in an active enzyme only in parental BeWo and in BeWo MC-2 cells. These clones provide an interesting model for studying the complex mechanisms regulating implantation as well as the controlled invasiveness during implantation compared to tumor invasion.     

Rosuvastatin 20 mg restores normal HDL-apoA-I kinetics in type 2 diabetes

Catabolism of HDL particles is accelerated in type 2 diabetes, leading to a reduction in plasma residence time, which may be detrimental. Rosuvastatin is the most powerful statin to reduce LDL-cholesterol, but its effects on HDL metabolism in type 2 diabetes remain unknown. We performed a randomized double-blind cross-over trial of 6-week treatment period with placebo or rosuvastatin 20 mg in eight patients with type 2 diabetes. An in vivo kinetic study of HDL-apolipoprotein A-I (apoA-I) with ¹³C leucine was performed at the end of each treatment period. Moreover, a similar kinetic study was carried out in eight nondiabetic normolipidemic controls. Rosuvastatin significantly reduced plasma LDL-cholesterol (−51%), triglycerides (TGs) (−38%), and HDL-TG (−23%). HDL-apoA-I fractional catabolic rate (FCR) was decreased by rosuvastatin (0.25 ± 0.06 vs. 0.32 ± 0.07 pool/day, P = 0.011), leading to an increase in plasma HDL-apoA-I residence time (4.21 ± 1.02 vs. 3.30 ± 0.73 day, P = 0.011). Treatment with rosuvastatin was associated with a concomitant reduction of HDL-apoA-I production rate. The decrease in HDL-apoA-I FCR, induced by rosuvastatin, was correlated with the reduction of plasma TGs and HDL-TG. HDL apoA-I FCR and production rate values in diabetic patients on rosuvastatin were not different from those found in controls. Rosuvastatin is responsible for a 22% reduction of HDL-apoA-I FCR and restores to normal the increased HDL turnover observed in type 2 diabetes. These kinetic modifications may have beneficial effects by increasing HDL plasma residence time.     

The nucleoside diphosphate kinase from mimivirus: a peculiar affinity for deoxypyrimidine nucleotides

The first viral Nucleoside Diphosphate Kinase was recently identified in the giant double-stranded DNA virus Acanthamoeba polyphag a Mimivirus (ApM). Here we report its expression and detailed biochemical characterization. NDK_apm exhibits unique features such as a shorter Kpn-loop, a structural motif previously reported to be part of the active site and involved in oligomer formation. Enzymatic activity measurements on the recombinant NDK_apm revealed its preferential affinity for deoxypyrimidine nucleotides. This property might represent an adaptation of NDK_apm to the production of the limiting TTP deoxynucleotide required for the replication of the large A+T rich (72%) viral genome. The NDK_apm might also assume a role in dUTP detoxification to compensate for the surprising absence of Mimivirus dUTPase (deoxyuridine triphosphate pyrophosphatase) an important enzyme conserved in most viruses. Although the phylogenetic analysis of NDK sequences sampled through organisms from the three domains of life is only partially informative, it favors an ancestral origin for NDK_apm over a recent acquisition from a eukaryotic organism by horizontal gene transfer.     

Peptide translocation across MOMP,the major outer membrane channel from Campylobacter jejuni

Here we report on translocation of short poly-arginines across the MOMP porin, the major outer membrane protein in the cell wall of Campylobacter jejuni. MOMP was purified to homogeneity from a pathogenic strain of C. jejuni. Its reconstitution in lipid membranes and measuring the ion-current revealed two main distinct populations of protein channels which we interpreted as mono and trimers. Addition of poly-arginines causes concentration and voltage dependent ion-current fluctuations. Increasing the transmembrane potential decreases the residence time of the peptide inside the channel indicating successful translocation. We conclude that poly-arginines can cross the outer membrane of Campylobacter through the MOMP channel.     

Purification of the blue-green pigment “marennine” from the marine tychopelagic diatom Haslea ostrearia (Gaillon/Bory) Simonsen

The diatom Haslea ostrearia that lives in oyster ponds has the distinctive feature of synthesizing “marennine”, a blue-green pigment of which the chemical nature still remains unknown. This pigment is responsible for the greening of oyster gills. Here, we report a new method for extraction and purification of intracellular (accumulated in the apex of the cell) and extracellular (released into the external medium) forms of the pigment. Intracellular marennine is obtained by extraction from blue algal pellets with a carbonate buffer. The extract is then centrifuged and filtered. Extracellular marennine is obtained by clarification of blue-coloured culture medium. Both extracts are then purified by a semi-preparative process, using ultrafiltration through membranes and anion-exchange chromatography. This procedure allows us to produce native pigment displaying the degree of purity required to enter upon the molecular characterisation of marennine. By this process, about 35% of the initial amount of pigment can be recovered. If necessary, this method could be easily scaled up to a larger production system to accommodate potential industrial applications.     

Parasitoid polydnaviruses: evolution,pathology and applications

One of the more unusual groups of insect pathogens consists of members of the family Polydnaviridae, insect DNA viruses that live in mutual symbioses with their associated parasitoid wasp (Hymentoptera) carriers until they are injected into specific lepidopteran hosts. Once inside this secondary host, polydnaviruses cause a wide variety of negative effects that ultimately ensure the survival of the parasitoid larvae. Because of their unusual life strategy and genetic features, it had been difficult to fully characterise polydnaviruses in terms of evolutionary history, replication cycle and functions in the host that might normally be well characterised for more conventional viruses. Recently, our understanding of polydnavirus evolutionary origins, gene content, genome organisation and functions in parasitism has greatly increased. Key findings are summarised in this review with emphasis on evolution of polydnavirus genes and genomes, their functional roles in insect pathology and their potential applications in insect biological control and biotechnology.     

Active aggregation among sexes in bean flower thrips (Megalurothrips sjostedti) on cowpea (Vigna unguiculata)

Male sexual aggregations are a common territorial, mating‐related or resource‐based, behaviour observed in diverse organisms, including insects such as thrips. The influence of factors such as plant substrate, time of day, and geographic location on aggregation of thrips is uncertain, therefore we monitored the dispersion of male and female bean flower thrips (BFT), Megalurothrips sjostedti (Trybom) (Thysanoptera: Thripidae), on cowpea, Vigna unguiculata (L.) Walp. (Fabaceae), over three cowpea growth stages and across three cowpea‐growing areas of Kenya. Our results indicated that for all the crop growth stages, the density of BFTs varied over the time of day, with higher densities at 10:00, 13:00, and 16:00 hours than at 07:00 hours. Thrips densities did not differ among blocks at the budding stage, but they did at peak flowering and podding stages. Dispersion indices suggested that both male and female BFTs were aggregated. Active male aggregation occurred only on green plant parts and it varied across blocks, crop stages, and locations. Similarly, active female aggregation was observed in peak flowering and podding stages. Such active aggregation indicates a semiochemical or behaviour‐mediated aggregation. Identification of such a semiochemical may offer new opportunities for refining monitoring and management strategies for BFT on cowpea, the most important grain legume in sub‐Saharan Africa.     

Domain 3 of NS5A protein from the hepatitis C virus has intrinsic alpha-helical propensity and is a substrate of cyclophilin A

Nonstructural protein 5A (NS5A) is essential for hepatitis C virus (HCV) replication and constitutes an attractive target for antiviral drug development. Although structural data for its in-plane membrane anchor and domain D1 are available, the structure of domains 2 (D2) and 3 (D3) remain poorly defined. We report here a comparative molecular characterization of the NS5A-D3 domains of the HCV JFH-1 (genotype 2a) and Con1 (genotype 1b) strains. Combining gel filtration, CD, and NMR spectroscopy analyses, we show that NS5A-D3 is natively unfolded. However, NS5A-D3 domains from both JFH-1 and Con1 strains exhibit a propensity to partially fold into an α-helix. NMR analysis identifies two putative α-helices, for which a molecular model could be obtained. The amphipathic nature of the first helix and its conservation in all genotypes suggest that it might correspond to a molecular recognition element and, as such, promote the interaction with relevant biological partner(s). Because mutations conferring resistance to cyclophilin inhibitors have been mapped into NS5A-D3, we also investigated the functional interaction between NS5A-D3 and cyclophilin A (CypA). CypA indeed interacts with NS5A-D3, and this interaction is completely abolished by cyclosporin A. NMR heteronuclear exchange experiments demonstrate that CypA has in vitro peptidyl-prolyl cis/trans-isomerase activity toward some, but not all, of the peptidyl-prolyl bonds in NS5A-D3. These studies lead to novel insights into the structural features of NS5A-D3 and its relationships with CypA.     

Protective effect of ε-viniferin on β-amyloid peptide aggregation investigated by electrospray ionization mass spectrometry

Abnormal β-amyloid peptide accumulation and aggregation is considered to be responsible for the formation and cerebral deposition of senile plaques in the brains of patients with Alzheimer's disease (AD). Inhibition of the formation of β-amyloid (Aβ) fibrils would be an attractive therapeutic target for the treatment of AD. Resveratrol and its derivatives exhibit a broad range of pharmacological properties such as protection against cardiovascular diseases and cancers, as well as promoting antiaging effects. We reported previously that ε-viniferin glucoside (VG), a resveratrol-derived dimer, strongly inhibits Aβ (25-35) fibril formation in vitro. In this study, we investigated the effects of VG on the aggregation of the full-length peptides (Aβ (1-40) and Aβ (1-42)) and on the β-amyloid-induced toxicity in PC12 cells. VG inhibited Aβ cytotoxicity and the non-covalent complex between VG and Aβ was observed by electrospray ionization mass spectrometry.     

Metagenomic exploration of antibiotic resistance in soil

The ongoing development of metagenomic approaches is providing the means to explore antibiotic resistance in nature and address questions that could not be answered previously with conventional culture-based strategies. The number of available environmental metagenomic sequence datasets is rapidly expanding and henceforth offer the ability to gain a more comprehensive understanding of antibiotic resistance at the global scale. Although there is now evidence that the environment constitutes a vast reservoir of antibiotic resistance gene determinants (ARGDs) and that the majority of ARGDs acquired by human pathogens may have an environmental origin, a better understanding of their diversity, prevalence and ecological significance may help predict the emergence and spreading of newly acquired resistances. Recent applications of metagenomic approaches to the study of ARGDs in natural environments such as soil should help overcome challenges concerning expanding antibiotic resistances.