Escape of HIV-1-Infected Dendritic Cells from TRAIL-Mediated NK Cell Cytotoxicity during NK-DC Cross-Talk—A Pivotal Role of HMGB1

Early stages of Human Immunodeficiency Virus-1 (HIV-1) infection are associated with local recruitment and activation of important effectors of innate immunity, i.e. natural killer (NK) cells and dendritic cells (DCs). Immature DCs (iDCs) capture HIV-1 through specific receptors and can disseminate the infection to lymphoid tissues following their migration, which is associated to a maturation process. This process is dependent on NK cells, whose role is to keep in check the quality and the quantity of DCs undergoing maturation. If DC maturation is inappropriate, NK cells will kill them (“editing process”) at sites of tissue inflammation, thus optimizing the adaptive immunity. In the context of a viral infection, NK-dependent killing of infected-DCs is a crucial event required for early elimination of infected target cells. Here, we report that NK-mediated editing of iDCs is impaired if DCs are infected with HIV-1. We first addressed the question of the mechanisms involved in iDC editing, and we show that cognate NK-iDC interaction triggers apoptosis via the TNF-related apoptosis-inducing ligand (TRAIL)-Death Receptor 4 (DR4) pathway and not via the perforin pathway. Nevertheless, once infected with HIV-1, DC_HIV become resistant to NK-induced TRAIL-mediated apoptosis. This resistance occurs despite normal amounts of TRAIL released by NK cells and comparable DR4 expression on DC_HIV. The escape of DC_HIV from NK killing is due to the upregulation of two anti-apoptotic molecules, the cellular-Flice like inhibitory protein (c-FLIP) and the cellular inhibitor of apoptosis 2 (c-IAP2), induced by NK-DC_HIV cognate interaction. High-mobility group box 1 (HMGB1), an alarmin and a key mediator of NK-DC cross-talk, was found to play a pivotal role in NK-dependent upregulation of c-FLIP and c-IAP2 in DC_HIV. Finally, we demonstrate that restoration of DC_HIV susceptibility to NK-induced TRAIL killing can be obtained either by silencing c-FLIP and c-IAP2 by specific siRNA, or by inhibiting HMGB1 with blocking antibodies or glycyrrhizin, arguing for a key role of HMGB1 in TRAIL resistance and DC_HIV survival. These findings provide evidence for a new strategy developed by HIV to escape immune attack, they challenge the question of the involvement of HMGB1 in the establishment of viral reservoirs in DCs, and they identify potential therapeutic targets to eliminate infected DCs.     

On the use of 2,1,3-benzothiadiazole derivatives as selective live cell fluorescence imaging probes

Newly designed 2,1,3-benzothiadiazole-containing fluorescent probes with four excited state intramolecular proton transfer (ESIPT) sites were successfully tested in live cell-imaging assays using a confluent monolayer of human stem-cells (tissue). All tested dyes were compared with the commercially available DAPI and gave far better results.     

Bloch Equations-Based Reconstruction of Myocardium T1 Maps from Modified Look-Locker Inversion Recovery Sequence

Modified Look-Locker Inversion recovery (MOLLI) sequence is increasingly performed for myocardial T1 mapping but is known to underestimate T1 values. The aim of the study was to quantitatively analyze several sources of errors when T1 maps are derived using standard post-processing of the sequence and to propose a reconstruction approach that takes into account inversion efficacy (η), T2 relaxation during balanced steady-state free-precession readouts and B1+ inhomogeneities. Contributions of the different sources of error were analyzed using Bloch equations simulations of MOLLI sequence. Bloch simulations were then combined with the acquisition of fast B1+ and T2 maps to derive more accurate T1 maps. This novel approach was evaluated on phantoms and on five healthy volunteers. Simulations show that T2 variations, B1+ heterogeneities and inversion efficiency represent major confounders for T1 mapping when MOLLI is processed with standard 3-parameters fitting. In vitro data indicate that T1 values are accurately derived with the simulation approach and in vivo data suggest that myocardium T1 are 15% underestimated when processed with the standard 3-parameters fitting. At the cost of additional acquisitions, this method might be suitable in clinical research protocols for precise tissue characterization as it decorrelates T1 and T2 effects on parametric maps provided by MOLLI sequence and avoids inaccuracies when B1+ is not homogenous throughout the myocardium.     

Head-to-head bis-hairpin polyamide minor groove binders and their conjugates with triplex-forming oligonucleotides: studies of interaction with target double-stranded DNA

Two hairpin hexa(N-methylpyrrole)carboxamide DNA minor groove binders (MGB) were linked together via their N-termini in head-to-head orientation. Complex formation between these bis-MGB conjugates and target DNA has been studied using DNase I footprinting, circular dichroism, thermal dissociation, and molecular modeling. DNase I footprint revealed binding of these conjugates to all the sites of 492 b.p. DNA fragment containing (A/T)(n)X(m)(A/T)(p) sequences, where n>3, p>3; m=1,2; X = A,T,G, or C. Binding affinity depended on the sequence context of the target. CD experiments and molecular modeling showed that oligo(N-methylpyrrole)carboxamide moieties in the complex form two short antiparallel hairpins rather than a long parallel head-to-head hairpin. Binding of bis-MGB also stabilized a target duplex thermodynamically. Sequence specificity of bis-MGB/DNA binding was validated using bis-conjugates of sequence-specific hairpin (N-methylpyrrole)/(N-methylimidazole) carboxamides. In order to increase the size of recognition sequence, the conjugates of bis-MGB with triplex-forming oligonucleotides (TFO) were synthesized and compared to TFO conjugated with single MGB hairpin unit. Bis-MGB-oligonucleotide conjugates also bind to two blocks of three and more A.T/T.A pairs similarly to bis-MGB alone, independently of the oligonucleotide moiety, but with lower affinity. However, the role of TFO in DNA recognition was demonstrated for mono-MGB-TFO conjugate where the binding was detected mainly in the area of the target sequence consisting of both MGB and TFO recognition sites. Basing on the molecular modeling, three-dimensional models of both target DNA/bis-MGB and target DNA/TFO-bis-MGB complexes were built, where bis-MGB forms two antiparallel hairpins. According to the second model, one MGB hairpin is in the minor groove of 5'-adjacent A/T sequence next to the triplex-forming region, whereas the other one occupies the minor groove of the TFO binding polypurine tract. All these data together give a key information for the construction of MGB-MGB and MGB-oligonucleotide conjugates possessing high specificity and affinity for the target double-stranded DNA.     

Assembly,plasticity and selective vulnerability to disease of mouse neuromuscular junctions

Although physiological differences among neuromuscular junctions (NMJs) have long been known, NMJs have usually been considered as one type of synapse, restricting their potential value as model systems to investigate mechanisms controlling synapse assembly and plasticity. Here we discuss recent evidence that skeletal muscles in the mouse can be subdivided into two previously unrecognized subtypes, designated FaSyn and DeSyn muscles. These muscles differ in the pattern of neuromuscular synaptogenesis during embryonic development. Differences between classes are intrinsic to the muscles, and manifest in the absence of innervation or agrin. The distinct rates of synaptogenesis in the periphery may influence processes of circuit maturation through retrograde signals. While NMJs on FaSyn and DeSyn muscles exhibit a comparable anatomical organization in postnatal mice, treatments that challenge synaptic stability result in nerve sprouting, NMJ remodeling, and ectopic synaptogenesis selectively on DeSyn muscles. This anatomical plasticity of NMJs diminishes greatly between 2 and 6 months postnatally. NMJs lacking this plasticity are lost selectively and very early on in mouse models of motoneuron disease, suggesting that disease-associated motoneuron dysfunction may fail to initiate maintenance processes at “non-plastic” NMJs. Transgenic mice overexpressing growth-promoting proteins in motoneurons exhibit greatly enhanced stimulus-induced sprouting restricted to DeSyn muscles, supporting the notion that anatomical plasticity at the NMJ is primarily controlled by processes in the postsynaptic muscle. The discovery that entire muscles in the mouse differ substantially in the anatomical plasticity of their synapses establishes NMJs as a uniquely advantageous experimental system to investigate mechanisms controlling synaptic rearrangements at defined synapses in vivo.     

Effect of Forest Fragmentation on Tick Infestations of Birds and Tick Infection Rates by Rickettsia in the Atlantic Forest of Brazil

Habitat loss and modifications affect biodiversity, potentially contributing to outbreaks of infectious diseases. We evaluated if the patch sizeinfragmented areas of Atlantic Forest in southeastern Brazil influences the diversity of forest birds and consequently the prevalence of ticks on birds and the rickettsial infection of these ticks. During 2 years, we collected ticks from birds in 12 sites: four small forest patches (80–140 ha), four large ones (480–1,850 ha), and four forest control areas within the much larger Morro do Diabo State Park (~36,000 ha). A total of 1,725 birds were captured (81 species, 24 families), from which 223 birds were infested by 2,339 ticks of the genus Amblyomma, mostly by the species A. nodosum. Bird diversity and richness were higher in larger than smaller forest fragments. The prevalence of ticks on birds was inversely correlated with bird diversity and richness. Among 174 A. nodosum tested for rickettsial infection by polymerase chain reaction, 51 were found to be infected by Rickettsia bellii or Rickettsia parkeri. However, tick infection rates by Rickettsia spp. were not statistically different between forest patch sizes. The higher prevalence of ticks on birds in degraded patches might be caused by a dominance of a few generalist bird species in small patches, allowing an easier transmission of parasites among individuals. It could also be related to more favorable microclimatic conditions for the free-living stages of A. nodosum in smaller forest fragments.The higher burden of ticks on birds in smaller forest fragments is an important secondary effect of habitat fragmentation, possibly increasing the likelihood of Rickettsia contagion.     

Translation initiator EIF4G1 mutations in familial Parkinson disease

Genome-wide analysis of a multi-incident family with autosomal-dominant parkinsonism has implicated a locus on chromosomal region 3q26-q28. Linkage and disease segregation is explained by a missense mutation c.3614G>A (p.Arg1205His) in eukaryotic translation initiation factor 4-gamma (EIF4G1). Subsequent sequence and genotype analysis identified EIF4G1 c.1505C>T (p.Ala502Val), c.2056G>T (p.Gly686Cys), c.3490A>C (p.Ser1164Arg), c.3589C>T (p.Arg1197Trp) and c.3614G>A (p.Arg1205His) substitutions in affected subjects with familial parkinsonism and idiopathic Lewy body disease but not in control subjects. Despite different countries of origin, persons with EIF4G1 c.1505C>T (p.Ala502Val) or c.3614G>A (p.Arg1205His) mutations appear to share haplotypes consistent with ancestral founders. eIF4G1 p.Ala502Val and p.Arg1205His disrupt eIF4E or eIF3e binding, although the wild-type protein does not, and render mutant cells more vulnerable to reactive oxidative species. EIF4G1 mutations implicate mRNA translation initiation in familial parkinsonism and highlight a convergent pathway for monogenic, toxin and perhaps virally-induced Parkinson disease.     

Hematological responses of the Neotropical teleost matrinxã (Brycon cephalus) to environmental nitrite

Environmental increase in nitrite impairs the function of several aquatic species, including fishes. Nitrite reacts with hemoglobin yielding the non-functional methemoglobin (metHb), and many physiological disturbances can arise. The physiological mechanisms to cope with nitrite are still unclear in fish. Hematological parameters, the role of NADH-methemoglobin reductase system and the electrolytic balance were studied in the freshwater teleost Brycon cephalus (matrinx?) exposed to 0.2, 0.4 and 0.6 mg/L of nitrite N-NO(2) for 24 and 96 h. Hematocrit, total hemoglobin and the red blood cell (RBC) number decreased. Methemoglobin content increased from 1% to 69% for 24 h of exposure and drastically from 5-6% to 90% for 96 h. The activity of NADH-methemoglobin reductase system displayed a tendency of increase in response to nitrite concentration or time of exposure. In the plasma, nitrite was accumulated to values 30-fold higher than the environmental concentration. The plasma K(+) concentration increased only in fish exposed to NO(2) for 24 h. No changes in plasma protein and Na(+) were observed during nitrite exposure but Cl-presented a punctual increase at 0.2 mg/L N-NO(2)-96 h. The hematological data suggest that nitrite caused functional and hemolytic anemia. Furthermore, the electrolytic balance was relatively undisturbed, and the nitrite clearance in matrinx? is likely depending on other factors than NADH-methemoglobin reductase system.     

Germ-line DNA copy number variation frequencies in a large North American population

Genomic copy number variation (CNV) is a recently identified form of global genetic variation in the human genome. The Affymetrix GeneChip 100 and 500 K SNP genotyping platforms were used to perform a large-scale population-based study of CNV frequency. We constructed a genomic map of 578 CNV regions, covering approximately 220 Mb (7.3%) of the human genome, identifying 183 previously unknown intervals. Copy number changes were observed to occur infrequently (<1%) in the majority (>93%) of these genomic regions, but encompass hundreds of genes and disease loci. This North American population-based map will be a useful resource for future genetic studies. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Has the kouprey (Bos sauveli Urbain, 1937) been domesticated in Cambodia?

The kouprey (Bos sauveli Urbain, 1937) is a very rare bovid species of Cambodia, which may be extinct in the wild, as no living specimen has been observed for a long time. Here, we describe a complete taxidermy mount, which presents astonishing morphological similarities with the kouprey. The animal was mounted in 1871 at the National Museum of Natural History in Paris, where it was referenced as No. 1871-576. It was deposited at the Natural History Museum of Bourges, France, in 1931, where it is still conserved today. To clarify the taxonomic status of the specimen of Bourges, DNA was extracted from a piece of bone taken on the mandible, and two different fragments of the mitochondrial cytochrome b gene were independently amplified and sequenced. The phylogenetic analyses show that the specimen of Bourges is robustly associated with the holotype of the kouprey, and that both are related to other wild species of Bos found in Indochina, i.e., banteng (B. javanicus) and gaur (B. frontalis). Because of doubts for sexing the animal, we applied a molecular test based on the PCR amplification of a DNA fragment specific to the Y chromosome. The results indicate that the specimen of Bourges is a male. The comparisons with male kouprey previously described in the literature reveal important differences concerning the body size, general coloration and horns. As these differences involve phenotypic traits that are strongly selected in case of domestication, we suggest that the specimen of Bourges was a domestic ox. This implies therefore that the kouprey may have been domesticated in Cambodia, and that several extant local races may be directly related to the kouprey.