Placentation in the Mexican lizard Sceloporus mucronatus (Squamata: Phrynosomatidae)

We used light microscopy to study placental structure of the lizard Sceloporus mucronatus throughout 6 months of embryonic development. Three stages of placental development could be assigned to embryos based on the arrangement of the extraembryonic membranes. A highly vascular choriovitelline placenta was present in the embryonic hemisphere and a nonvascular bilaminar omphalopleure covered most of the abembryonic hemisphere of the egg during embryonic Stages 10-28. A chorioallantoic placenta replaced the choriovitelline placenta by embryonic Stage 29 and an omphaloplacenta covered the abembryonic hemisphere at this stage. The combination of these two placental types occurred in Stage 29-36 embryos. The final stage of placentation, embryonic Stages 37-40, was characterized by an omphalallantoic placenta in the abembryonic hemisphere and a chorioallantoic placenta in the embryonic hemisphere of the egg. The choriovitelline and chorioallantoic placentae are well vascularized, with closely apposed maternal and embryonic blood vessels. These structures are the most likely sites of respiratory exchange. In contrast, the omphaloplacenta and omphalallantoic placentae contain cuboidal or columnar epithelia and these structures may function in histotrophic exchange. Placentation of S. mucronatus is similar to that of predominantly lecithotrophic species in other squamate lineages suggesting that the evolution of this placental morphology is a response to similar factors and is independent of phylogeny.     

Effect of ion-binding and chemical phospholipid structure on the nanomechanics of lipid bilayers studied by force spectroscopy

The nanomechanical response of supported lipid bilayers has been studied by force spectroscopy with atomic force microscopy. We have experimentally proved that the amount of ions present in the measuring system has a strong effect on the force needed to puncture a 1,2-dimyristoyl-sn-glycero-3-phosphocholine bilayer with an atomic force microscope tip, thus highlighting the role that monovalent cations (so far underestimated, e.g., Na(+)) play upon membrane stability. The increase in the yield threshold force has been related to the increase in lateral interactions (higher phospholipid-phospholipid interaction, decrease in area per lipid) promoted by ions bound into the membrane. The same tendency has also been observed for other phosphatidylcholine bilayers, namely, 2-dilauroyl-sn-glycero-3-phosphocholine, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, and 1,2-dioleoyl-sn-3-phosphocholine, and also for phosphatidylethanolamine bilayers such as 1-palmitoyl-2-oleoyl-sn-3-phosphoethanolamine. Finally, this effect has been also tested on a natural lipid bilayer (Escherichia coli lipid extract), showing the same overall tendency. The kinetics of the process has also been studied, together with the role of water upon membrane stability and its effect on membrane nanomechanics. Finally, the effect of the chemical structure of the phospholipid molecule on the nanomechanical response of the membrane has also been discussed.     

Comparative cytogenetic analysis of eleven species of subfamilies Neoplecostominae and Hypostominae (Siluriformes: Loricariidae)

The family Loricariidae with about 690 species divided into six subfamilies, is one of the world’s largest fish families. Recent studies have shown the existence of several problems in the definition of natural groups in the family, which has made the characterization of the subfamilies and even of some genera quite difficult. With the main objective of contributing for a better understanding of the relationships between loricariids, cytogenetic analysis were conducted with two species of Neoplecostominae and nine species of Hypostominae that, according to morphological and molecular data, may belong to a new monophyletic unit. The results obtained showed a marked chromosomal conservation with the presence of 2n = 54 chromosomes and single interstitial Ag-NORs in all species analyzed. Considering that Neoplecostominae is the primitive sister-group of all other loricariids, with exception of Lithogeneinae, this karyotypic structure may represent the primitive condition for the family Loricariidae. The cytogenetic characteristics partaken by the species of Neoplecostominae and Hypostominae analyzed in the present study reinforce the hypothesis that the species of both these subfamilies might belong to a natural group.     

Probing the interaction between vesicular stomatitis virus and phosphatidylserine

The entry of enveloped animal viruses into their host cells always depends on membrane fusion triggered by conformational changes in viral envelope glycoproteins. Vesicular stomatitis virus (VSV) infection is mediated by virus spike glycoprotein G, which induces membrane fusion between the viral envelope and the endosomal membrane at the acidic environment of this compartment. In this work, we evaluated VSV interactions with membranes of different phospholipid compositions, at neutral and acidic pH, using atomic force microscopy (AFM) operating in the force spectroscopy mode, isothermal calorimetry (ITC) and molecular dynamics simulation. We found that the binding forces differed dramatically depending on the membrane phospholipid composition, revealing a high specificity of G protein binding to membranes containing phosphatidylserine (PS). In a previous work, we showed that the sequence corresponding amino acid 164 of VSV G protein was as efficient as the virus in catalyzing membrane fusion at pH 6.0. Here, we used this sequence to explore VSV–PS interaction using ITC. We found that peptide binding to membranes was exothermic, suggesting the participation of electrostatic interactions. Peptide–membrane interaction at pH 7.5 was shown to be specific to PS and dependent on the presence of His residues in the fusion peptide. The application of the simplified continuum Gouy–Chapman theory to our system predicted a pH of 5.0 at membrane surface, suggesting that the His residues should be protonated when located close to the membrane. Molecular dynamics simulations suggested that the peptide interacts with the lipid bilayer through its N-terminal residues, especially Val¹⁴⁵ and His¹⁴⁸. Fabiana A.Carneiro and Pedro A. Lapido-Loureiro contributed equally to this work An erratum to this article can be found at     

Comparative spermatology in Plecoptera (Insecta): an ultrastructural investigation on four species

The ultrastructure of spermatozoa of four Plecoptera species, Leuctra fusca Linnaeus, Brachyptera risi (Morton), Taeniopteryx stankovitchi Ikonomov, T. kuehtreiberi (Aubert), was investigated by light microscopy, scanning and transmission electron microscopy, and immunofluorescence microscopy. The spermatozoa of all the species have a complex acrosome, are filiform and flagellate, with a '9+9+2' axoneme flanked by two mitochondrial derivatives. However, the structure shows a certain interspecific heterogeneity. L. fusca has a short conical nucleus in the apical position and an axoneme flanked by two accessory bodies. In the sperm of the Taeniopterygidae, the nucleus is a long cylindrical body which lies between the two mitochondrial derivatives flanking the axoneme for most of its length. Our results suggest a close phylogenetic affinity between Plecoptera and the other orders of Polyneoptera, although the proposed sister-group relationship between Plecoptera and Embioptera cannot be confirmed.     

Sorafenib sensitizes hepatocellular carcinoma cells to physiological apoptotic stimuli

Sorafenib increases survival rate of patients with advanced hepatocellular carcinoma (HCC). The mechanism underlying this effect is not completely understood. In this work we have analyzed the effects of sorafenib on autocrine proliferation and survival of different human HCC cell lines. Our results indicate that sorafenib in vitro counteracts autocrine growth of different tumor cells (Hep3B, HepG2, PLC-PRF-5, SK-Hep1). Arrest in S/G2/M cell cycle phases were observed coincident with cyclin D1 down-regulation. However, sorafenib's main anti-tumor activity seems to occur through cell death induction which correlated with caspase activation, increase in the percentage of hypodiploid cells, activation of BAX and BAK and cytochrome c release from mitochondria to cytosol. In addition, we observed a rise in mRNA and protein levels of the pro-apoptotic "BH3-domain only" PUMA and BIM, as well as decreased protein levels of the anti-apoptotic MCL1 and survivin. PUMA targeting knock-down, by using specific siRNAs, inhibited sorafenib-induced apoptotic features. Moreover, we obtained evidence suggesting that sorafenib also sensitizes HCC cells to the apoptotic activity of transforming growth factor-β (TGF-β) through the intrinsic pathway and to tumor necrosis factor-α (TNF) through the extrinsic pathway. Interestingly, sensitization to sorafenib-induced apoptosis is characteristic of liver tumor cells, since untransformed hepatocytes did not respond to sorafenib inducing apoptosis, either alone or in combination with TGF-β or TNF. Indeed, sorafenib effectiveness in delaying HCC late progression might be partly related to a selectively sensitization of HCC cells to apoptosis by disrupting autocrine signals that protect them from adverse conditions and pro-apoptotic physiological cytokines.     

Complete Genome Sequence of a Novel Human Enterovirus C (HEV-C117) Identified in a Child with Community-Acquired Pneumonia

The new enterovirus C-117 strain belongs to the human enterovirus C species in the Picornaviridae family. We describe the characterization of the complete genome of this strain identified in a respiratory specimen of a child enrolled in the Community-Acquired Pneumonia Pediatric Research Initiative (CAP-PRI) study evaluating the etiology of community-acquired pneumonia (CAP).     

Plasma protein changes in horse after prolonged physical exercise: a proteomic study

Physical exercise induces various stress responses and metabolic adaptations that have not yet been completely elucidated. Novel biomarkers are needed in sport veterinary medicine to monitor training levels and to detect subclinical conditions that can develop into exercise-related diseases. In this study, protein modifications in horse plasma induced by prolonged, aerobic physical exercise were investigated by using a proteomic approach based on 2-DE and combined mass spectrometry procedures. Thirty-eight protein spots, associated with expression products of 13 genes, showed significant quantitative changes; spots identified as membrane Cu amine oxidase, α-1 antitrypsin, α-1 antitrypsin-related protein, caeruloplasmin, α-2 macroglobulin and complement factor C4 were augmented in relative abundance after the race, while haptoglobin β chain, apolipoprotein A-I, transthyretin, retinol binding protein 4, fibrinogen γ chain, complement factor B and albumin fragments were reduced. These results indicate that prolonged physical exercise affects plasma proteins involved in pathways related to inflammation, coagulation, immune modulation, oxidant/antioxidant activity and cellular and vascular damage, with consequent effects on whole horse metabolism.     

Characterization of PbPga1, an Antigenic GPI-Protein in the Pathogenic Fungus Paracoccidioides brasiliensis

Paracoccidioides brasiliensis is the etiologic agent of paracoccidioidomycosis (PCM), one of the most prevalent mycosis in Latin America. P. brasiliensis cell wall components interact with host cells and influence the pathogenesis of PCM. Cell wall components, such as glycosylphosphatidylinositol (GPI)-proteins play a critical role in cell adhesion and host tissue invasion. Although the importance of GPI-proteins in the pathogenesis of other medically important fungi is recognized, little is known about their function in P. brasiliensis cells and PCM pathogenesis. We cloned the PbPga1 gene that codifies for a predicted GPI-anchored glycoprotein from the dimorphic pathogenic fungus P. brasiliensis. PbPga1 is conserved in Eurotiomycetes fungi and encodes for a protein with potential glycosylation sites in a serine/threonine-rich region, a signal peptide and a putative glycosylphosphatidylinositol attachment signal sequence. Specific chicken anti-rPbPga1 antibody localized PbPga1 on the yeast cell surface at the septum between the mother cell and the bud with stronger staining of the bud. The exposure of murine peritoneal macrophages to rPbPga1 induces TNF-α release and nitric oxide (NO) production by macrophages. Furthermore, the presence of O-glycosylation sites was demonstrated by β-elimination under ammonium hydroxide treatment of rPbPga1. Finally, sera from PCM patients recognized rPbPga1 by Western blotting indicating the presence of specific antibodies against rPbPga1. In conclusion, our findings suggest that the PbPga1gene codifies for a cell surface glycoprotein, probably attached to a GPI-anchor, which may play a role in P. brasiliensis cell wall morphogenesis and infection. The induction of inflammatory mediators released by rPbPga1 and the reactivity of PCM patient sera toward rPbPga1 imply that the protein favors the innate mechanisms of defense and induces humoral immunity during P. brasiliensis infection.