On the Three-Finger Protein Domain Fold and CD59-Like Proteins in Schistosoma mansoni

Background

It is believed that schistosomes evade complement-mediated killing by expressing regulatory proteins on their surface. Recently, six homologues of human CD59, an important inhibitor of the complement system membrane attack complex, were identified in the schistosome genome. Therefore, it is important to investigate whether these molecules could act as CD59-like complement inhibitors in schistosomes as part of an immune evasion strategy.

Methodology/Principal Findings

Herein, we describe the molecular characterization of seven putative SmCD59-like genes and attempt to address the putative biological function of two isoforms. Superimposition analysis of the 3D structure of hCD59 and schistosome sequences revealed that they contain the three-fingered protein domain (TFPD). However, the conserved amino acid residues involved in complement recognition in mammals could not be identified. Real-time RT-PCR and Western blot analysis determined that most of these genes are up-regulated in the transition from free-living cercaria to adult worm stage. Immunolocalization experiments and tegument preparations confirm that at least some of the SmCD59-like proteins are surface-localized; however, significant expression was also detected in internal tissues of adult worms. Finally, the involvement of two SmCD59 proteins in complement inhibition was evaluated by three different approaches: (i) a hemolytic assay using recombinant soluble forms expressed in Pichia pastoris and E. coli; (ii) complement-resistance of CHO cells expressing the respective membrane-anchored proteins; and (iii) the complement killing of schistosomula after gene suppression by RNAi. Our data indicated that these proteins are not involved in the regulation of complement activation.

Conclusions

Our results suggest that this group of proteins belongs to the TFPD superfamily. Their expression is associated to intra-host stages, present in the tegument surface, and also in intra-parasite tissues. Three distinct approaches using SmCD59 proteins to inhibit complement strongly suggested that these proteins are not complement inhibitors and their function in schistosomes remains to be determined.     

Impact of Perfusion Map Analysis on Early Survival Prediction Accuracy in Glioma Patients

Studies investigating dynamic susceptibility contrast magnetic resonance imaging-determined relative cerebral blood volume (rCBV) maps as a metric of treatment response assessment have generated conflicting results. We evaluated the potential of various analytical techniques to predict survival of patients with glioma treated with chemoradiation. rCBV maps were acquired in patients with high-grade gliomas at 0, 1, and 3 weeks into chemoradiation therapy. Various analytical techniques were applied to the same cohort of serial rCBV data for early assessment of survival. Three different methodologies were investigated: 1) percentage change of whole tumor statistics (i.e., mean, median, and percentiles), 2) physiological segmentation (low rCBV, medium rCBV, or high rCBV), and 3) a voxel-based approach, parametric response mapping (PRM). All analyses were performed using the same tumor contours, which were determined using contrast-enhanced T1-weighted and fluid attenuated inversion recovery images. The predictive potential of each response metric was assessed at 1-year and overall survival. PRM was the only analytical approach found to generate a response metric significantly predictive of patient 1-year survival. Time of acquisition and contour volume were not found to alter the sensitivity of the PRM approach for predicting overall survival. We have demonstrated the importance of the analytical approach in early response assessment using serial rCBV maps. The PRM analysis shows promise as a unified early and robust imaging biomarker of treatment response in patients diagnosed with high-grade gliomas.     

Linkage Maps of Lowland and Upland Tetraploid Switchgrass Ecotypes

Switchgrass (Panicum virgatum L.) is a native perennial warm season (C₄) grass that has been identified as a promising species for bioenergy research and production. Consequently, biomass yield and feedstock quality improvements are high priorities for switchgrass research. The objective of this study was to develop a switchgrass genetic linkage map using a full-sib pseudo-testcross mapping population derived from a cross between two heterozygous genotypes selected from the lowland cultivar ‘Alamo’ (AP13) and the upland cultivar ‘Summer’ (VS16). The female parent (AP13) map consists of 515 loci in 18 linkage groups (LGs) and spans 1,733 cM. The male parent (VS16) map arranges 363 loci in 17 LGs and spans 1,508 cM. No obvious cause for the lack of one LG in VS16 could be identified. Comparative analyses between the AP13 and VS16 maps showed that the two major ecotypic classes of switchgrass have highly colinear maps with similar recombination rates, suggesting that chromosomal exchange between the two ecotypes should be able to occur freely. The AP13 and VS16 maps are also highly similar with respect to marker orders and recombination levels to previously published switchgrass maps. The genetic maps will be used to identify quantitative trait loci associated with biomass and quality traits. The AP13 genotype was used for the whole genome-sequencing project and the map will thus also provide a tool for the anchoring of the switchgrass genome assembly.     

Vagal nerve stimulation protects cardiac injury by attenuating mitochondrial dysfunction in a murine burn injury model

Mitochondria play a central role in the integration and execution of a wide variety of apoptotic signals. In the present study, we examined the deleterious effects of burn injury on heart tissue. We explored the effects of vagal nerve stimulation (VNS) on cardiac injury in a murine burn injury model, with a focus on the protective effect of VNS on mitochondrial dysfunction in heart tissue. Mice were subjected to a 30% total body surface area, full‐thickness steam burn followed by right cervical VNS for 10 min. and compared to burn alone. A separate group of mice were treated with the M₃‐muscarinic acetylcholine receptor (M₃‐AchR) antagonist 4‐DAMP or phosphatidylinositol 3 Kinase (PI3K) inhibitor LY294002 prior to burn and VNS. Heart tissue samples were collected at 6 and 24 hrs after injury to measure changes in apoptotic signalling pathways. Burn injury caused significant cardiac pathological changes, cardiomyocyte apoptosis, mitochondrial swelling and decrease in myocardial ATP content at 6 and 24 hrs after injury. These changes were significantly attenuated by VNS. VNS inhibited release of pro‐apoptotic protein cytochrome C and apoptosis‐inducing factor from mitochondria to cytosol by increasing the expression of Bcl‐2, and the phosphorylation level of Bad (pBad¹³⁶) and Akt (pAkt³⁰⁸). These protective changes were blocked by 4‐DAMP or LY294002. We demonstrated that VNS protected against burn injury–induced cardiac injury by attenuating mitochondria dysfunction, likely through the M₃‐AchR and the PI3K/Akt signalling pathways.     

BEHAVIOUR OF A RADIO-TAGGED TILAPIA RENDALLI BOULENGER IN LAKE NGEZI,ZIMBABWE

SUMMARY

The behaviour of a single specimen of Tilapia rendalli Boulenger was observed June through October 1986 in Lake Ngezi, Zimbabwe, by the use of a surgically-implanted radiotransmitter. The individual displayed a distinct homing response on two occasions, returning to the cove of capture (1) 1.5 km away within 48 hours of release (95Z of the distance was covered in 24 hours), and (2) after a lengthy migration some 6.0 km up the lake with other tilapia. During the only observed migration in late June, the fish travelled a conservative distance of 6.7 km over a 5-day period, sometimes at rates > 150 m/hr. The fish was also characterized by strong territoriality (i.e., preference to the cove of capture) and avoidance to the approach of the researcher by boat.     

Spatial Pattern Dynamics of 3D Stem Cell Loss of Pluripotency via Rules-Based Computational Modeling

Pluripotent embryonic stem cells (ESCs) have the unique ability to differentiate into cells from all germ lineages, making them a potentially robust cell source for regenerative medicine therapies, but difficulties in predicting and controlling ESC differentiation currently limit the development of therapies and applications from such cells. A common approach to induce the differentiation of ESCs in vitro is via the formation of multicellular aggregates known as embryoid bodies (EBs), yet cell fate specification within EBs is generally considered an ill-defined and poorly controlled process. Thus, the objective of this study was to use rules-based cellular modeling to provide insight into which processes influence initial cell fate transitions in 3-dimensional microenvironments. Mouse embryonic stem cells (D3 cell line) were differentiated to examine the temporal and spatial patterns associated with loss of pluripotency as measured through Oct4 expression. Global properties of the multicellular aggregates were accurately recapitulated by a physics-based aggregation simulation when compared to experimentally measured physical parameters of EBs. Oct4 expression patterns were analyzed by confocal microscopy over time and compared to simulated trajectories of EB patterns. The simulations demonstrated that loss of Oct4 can be modeled as a binary process, and that associated patterns can be explained by a set of simple rules that combine baseline stochasticity with intercellular communication. Competing influences between Oct4+ and Oct4− neighbors result in the observed patterns of pluripotency loss within EBs, establishing the utility of rules-based modeling for hypothesis generation of underlying ESC differentiation processes. Importantly, the results indicate that the rules dominate the emergence of patterns independent of EB structure, size, or cell division. In combination with strategies to engineer cellular microenvironments, this type of modeling approach is a powerful tool to predict stem cell behavior under a number of culture conditions that emulate characteristics of 3D stem cell niches.     

Early Apoptosis of Macrophages Modulated by Injection of Yersinia pestis YopK Promotes Progression of Primary Pneumonic Plague

Yersinia pestis causes pneumonic plague, a disease characterized by inflammation, necrosis and rapid bacterial growth which together cause acute lung congestion and lethality. The bacterial type III secretion system (T3SS) injects 7 effector proteins into host cells and their combined activities are necessary to establish infection. Y. pestis infection of the lungs proceeds as a biphasic inflammatory response believed to be regulated through the control of apoptosis and pyroptosis by a single, well-conserved T3SS effector protein YopJ. Recently, YopJ-mediated pyroptosis, which proceeds via the NLRP3-inflammasome, was shown to be regulated by a second T3SS effector protein YopK in the related strain Y. pseudotuberculosis. In this work, we show that for Y. pestis, YopK appears to regulate YopJ-mediated apoptosis, rather than pyroptosis, of macrophages. Inhibition of caspase-8 blocked YopK-dependent apoptosis, suggesting the involvement of the extrinsic pathway, and appeared cell-type specific. However, in contrast to yopJ, deletion of yopK caused a large decrease in virulence in a mouse pneumonic plague model. YopK-dependent modulation of macrophage apoptosis was observed at 6 and 24 hours post-infection (HPI). When YopK was absent, decreased populations of macrophages and dendritic cells were seen in the lungs at 24 HPI and correlated with resolution rather than progression of inflammation. Together the data suggest that Y. pestis YopK may coordinate the inflammatory response during pneumonic plague through the regulation of apoptosis of immune cells.