Energy transduction in transmembrane ion pumps

Recent crystallographic structures of three different ion pumps provide a first view of the mechanisms by which these molecular machines transfer ions across cell membranes against an electrochemical gradient. Each of the structures reinforces the concept that several buried counter ions have central roles in substrate recruitment, substrate binding and energy transduction during ion pumping. The spatial organization of the counter ions suggests that, initially, one or more counter ions lowers the Born energy cost of binding a substrate ion in the low-dielectric interior of the membrane. Subsequently, a ligand-induced conformational change seems to close a charged access gate to prevent backflow from a subsequent, low-affinity state of the pump. A final role of the buried counter ions might be to couple the input of external energy to a small charge separation between the substrate ion and the buried counter ions, thereby decreasing the binding affinity for the substrate ion in preparation for its release on the high-energy side of the membrane.     

In Vitro Silencing of Brugia malayi Trehalose-6-Phosphate Phosphatase Impairs Embryogenesis and In Vivo Development of Infective Larvae in Jirds

Background

The trehalose metabolic enzymes have been considered as potential targets for drug or vaccine in several organisms such as Mycobacterium, plant nematodes, insects and fungi due to crucial role of sugar trehalose in embryogenesis, glucose uptake and protection from stress. Trehalose-6-phosphate phosphatase (TPP) is one of the enzymes of trehalose biosynthesis that has not been reported in mammals. Silencing of tpp gene in Caenorhabditis elegans revealed an indispensable functional role of TPP in nematodes.

Methodology and Principal Findings

In the present study, functional role of B. malayi tpp gene was investigated by siRNA mediated silencing which further validated this enzyme to be a putative antifilarial drug target. The silencing of tpp gene in adult female B. malayi brought about severe phenotypic deformities in the intrauterine stages such as distortion and embryonic development arrest. The motility of the parasites was significantly reduced and the microfilarial production as well as their in vitro release from the female worms was also drastically abridged. A majority of the microfilariae released in to the culture medium were found dead. B. malayi infective larvae which underwent tpp gene silencing showed 84.9% reduced adult worm establishment after inoculation into the peritoneal cavity of naïve jirds.

Conclusions/Significance

The present findings suggest that B. malayi TPP plays an important role in the female worm embryogenesis, infectivity of the larvae and parasite viability. TPP enzyme of B. malayi therefore has the potential to be exploited as an antifilarial drug target.     

Adipose tissue-derived stem cell response to the differently processed 316L stainless steel substrates

Stainless steel (SS) is one of the most applicable materials in fabrication of cardiac implants. The aim of this study is to investigate the effect of atomic structure of polycrystalline stainless steel on the response of adipose tissue-derived stem cells (ADSCs). Samples are prepared from differently processed extruded rod and rolled sheet of 316L SS having different crystallographic structure. X-ray diffraction analysis indicated (2 0 0) and (1 1 1) orientations with distinct volume fractions in the specimens. Morphology and ADSCs behavior including adhesion, proliferation and differentiation are assessed. The expression of cardiac specific protein (cardiac troponin I) and genes of differentiating cardiomyocytes is analyzed by immunofluorescence and RT-PCR. The number of attached and grown cells on the rod sample is higher than the sheet sample also the scanning electron microscopy (SEM) analysis of ADSCs grown on the samples demonstrates higher cell density and spreading pattern on the surface of rod sample. In differentiated ADSCs on the rod sample the expression of all genes except ANF are detectable, while on the sheet sample only the MEF2C and β-MHC are expressed. This study shows that the cellular response is influenced by the crystal structure of the substrate therefore; the skill to alter the structure of substrate may lend itself to engineer a biomaterial which could be suitable for differentiation of stem cells into a definite lineage.     

Relationship between vitamin B12, folate and homocysteine levels and H. pylori infection in patients with functional dyspepsia: A cross-section study

ABSTRACT: BACKGROUND: H. pylori infection has been associated with many micronutrient deficiencies. There is a dearth of data from communities with nutritional deficiencies and high prevalence of H. pylori infection. The aim of this study was to determine the impact of H. pylori infection on serum levels of vitamin B12, folate and homocysteine in patients with functional dyspepsia (FD). METHODS: One hundred and thirty-two patients with FD undergoing gastroscopy were enrolled. The serum was analyzed for B12, folate and homocysteine levels before gastroscopy. H. pylori infection was diagnosed by histopathological examination of gastric biopsies and urea breath test. An independent sample t-test and the Mann-Whitney test were used to compare mean serum concentrations of biomarkers between H. pylori-positive and H. pylori-negative groups of patients. A Chi-square test was performed to assess the differences among proportions, while Spearman's rho was used for correlation analysis between levels of B12 and homocysteine. RESULTS: The mean age of the group was 40.3 +/- 11.5 (19-72) years. Folate deficiency was seen in 43 (34.6%), B12 deficiency in 30 (23.1%) and hyperhomocysteinemia in 60 (46.2%) patients. H. pylori was present in 80 (61.5%) patients with FD while it was absent in 50 (38.5%). Mean serum levels of B12, folate and homocysteine in the H. pylori-positive group of patients were not significantly different from the levels in the H. pylori-negative group (357 +/- 170 vs. 313 +/- 136 pg/mL; p = 0.13), (4.35 +/- 1.89 vs. 4.42 +/- 1.93 ng/mL; p = 0.84); (15.88 +/- 8.97 vs. 16.62 +/- 7.82 mumol/L; p = 0.24); respectively. B12 deficiency ([less than or equal to]200 pg/mL) was 23.8% in the H. pylori-positive patients versus 22.0% in the H. pylori-negative patients. Folate deficiency ([less than or equal to]3.5 ng/mL) was 33.8% in the H. pylori-positive group versus 36% in the H. pylori-negative group. Hyperhomocysteinemia (>15 mumol/L) was present in 46.2% of H. pylori-positive patients compared to 44% in the H. pylori-negative group. Correlation analysis indicated that serum B12 levels were inversely associated with serum levels of homocysteine in patients with FD (rho = 0.192; p = 0.028). CONCLUSIONS: This study demonstrated an inverse relationship between serum levels of B12 and homocysteine in patients with FD. Moreover, no impact of the presence of H. pylori was found on B12, folate and homocysteine levels in such patients.     

The effects of MicroRNA transfections on global patterns of gene expression in ovarian cancer cells are functionally coordinated

ABSTRACT: BACKGROUND: MicroRNAs (miRNAs) are a class of small RNAs that have been linked to a number of diseases including cancer. The potential application of miRNAs in the diagnostics and therapeutics of ovarian and other cancers is an area of intense interest. A current challenge is the inability to accurately predict the functional consequences of exogenous modulations in the levels of potentially therapeutic miRNAs. In an initial effort to systematically address this issue, we conducted miRNA transfection experiments using two miRNAs (miR-7, miR-128) and characterized global changes in levels of gene expression. RESULTS: While ~20% of the changes in expression patterns of hundreds to thousands of genes could be attributed to direct miRNA-mRNA interactions, the majority of the changes are indirect, involving the downstream consequences of miRNA-mediated changes in regulatory gene expression. We find that the changes in gene expression induced by individual miRNAs are functionally coordinated but distinct between the two miRNAs. MiR-7 transfection into ovarian cancer cells induces changes in cell adhesion and other developmental networks previously associated with epithelial-mesenchymal transitions (EMT) and other processes linked with metastasis. In contrast, miR-128 transfection induces changes in cell cycle control and other processes commonly linked with cellular replication. CONCLUSION: The functionally coordinated patterns of gene expression displayed by different families of miRNAs have the potential to provide clinicians with a strategy to treat cancers from a systems rather than a single gene perspective.     

Effects of foliar application of BAP on source and sink strength in four six-rowed barley (Hordeum vulgare L.) cultivars

A field experiment was conducted to investigate the effects of foliar application of a synthetic cytokinin (BAP) on source and sink strength of four different six-rowed barley (Hordeum vulgare L.) cultivars. Different spraying treatments consisting of spraying on whole plant, spraying only on leaves and spraying only on ears started at anthesis and continued for 7 days. One additional spraying was carried out on late period of grain filling. Results showed that spraying only on leaves did not affect ear weight, grain yield and 1,000-grain weight, while the two other treatments increased all above mentioned traits. Neither of treatments affected stem weight, biological yield and contribution of stem reserves in grain filling. Exogenous cytokinin did not increase photosynthetic rate and chlorophyll content in treated leaves until late period of grain filling, although there was no significant increase in final grain weight due to late application of BAP. Our results suggested that effects of foliar application of BAP were mostly due to increased sink size soon after anthesis and increased sink demand probably met by current photosynthesis of organs other than leaves, like ear green tissues. An erratum to this article is available at .     

Sparse representation and Bayesian detection of genome copy number alterations from microarray data

MOTIVATION: Genomic instability in cancer leads to abnormal genome copy number alterations (CNA) that are associated with the development and behavior of tumors. Advances in microarray technology have allowed for greater resolution in detection of DNA copy number changes (amplifications or deletions) across the genome. However, the increase in number of measured signals and accompanying noise from the array probes present a challenge in accurate and fast identification of breakpoints that define CNA. This article proposes a novel detection technique that exploits the use of piece wise constant (PWC) vectors to represent genome copy number and sparse Bayesian learning (SBL) to detect CNA breakpoints. METHODS: First, a compact linear algebra representation for the genome copy number is developed from normalized probe intensities. Second, SBL is applied and optimized to infer locations where copy number changes occur. Third, a backward elimination (BE) procedure is used to rank the inferred breakpoints; and a cut-off point can be efficiently adjusted in this procedure to control for the false discovery rate (FDR). RESULTS: The performance of our algorithm is evaluated using simulated and real genome datasets and compared to other existing techniques. Our approach achieves the highest accuracy and lowest FDR while improving computational speed by several orders of magnitude. The proposed algorithm has been developed into a free standing software application (GADA, Genome Alteration Detection Algorithm). AVAILABILITY: http://biron.usc.edu/~piquereg/GADA     

Activation of the p38 mitogen-activated protein kinase mediates the suppressive effects of type I interferons and transforming growth factor-beta on normal hematopoiesis.

Type I interferons (IFNs) are potent regulators of normal hematopoiesis in vitro and in vivo, but the mechanisms by which they suppress hematopoietic progenitor cell growth and differentiation are not known. In the present study we provide evidence that IFN alpha and IFN beta induce phosphorylation of the p38 mitogen-activated protein (Map) kinase in CD34+-derived primitive human hematopoietic progenitors. Such type I IFN-inducible phosphorylation of p38 results in activation of the catalytic domain of the kinase and sequential activation of the MAPK-activated protein kinase-2 (MapKapK-2 kinase), indicating the existence of a signaling cascade, activated downstream of p38 in hematopoietic progenitors. Our data indicate that activation of this signaling cascade by the type I IFN receptor is essential for the generation of the suppressive effects of type I IFNs on normal hematopoiesis. This is shown by studies demonstrating that pharmacological inhibitors of p38 reverse the growth inhibitory effects of IFN alpha and IFN beta on myeloid (colony-forming granulocytic-macrophage) and erythroid (burst-forming unit-erythroid) progenitor colony formation. In a similar manner, transforming growth factor beta, which also exhibits inhibitory effects on normal hematopoiesis, activates p38 and MapKapK-2 in human hematopoietic progenitors, whereas pharmacological inhibitors of p38 reverse its suppressive activities on both myeloid and erythroid colony formation. In further studies, we demonstrate that the primary mechanism by which the p38 Map kinase pathway mediates hematopoietic suppression is regulation of cell cycle progression and is unrelated to induction of apoptosis. Altogether, these findings establish that the p38 Map kinase pathway is a common effector for type I IFN and transforming growth factor beta signaling in human hematopoietic progenitors and plays a critical role in the induction of the suppressive effects of these cytokines on normal hematopoiesis.     

A model of anterior cruciate ligament reconstructive surgery: A validation construct and computational insights

This study sought to develop a computational framework that emulates the anterior cruciate ligament reconstruction surgery using transtibial portal technique. The proposed model included the tibia–femoral and patella–femoral joints, articular cartilage and menisci. Key surgical parameters were incorporated including bone-patellar-tendon-bone graft excision and pre-tensioning, tunnel morphology, bone plugs, and bone plug fixation. Several simulation steps were parameterized to reflect the clinically reported surgical procedure. Our focus was to explore the intra-operative effects of variations in tunnel directions on the selected metrics of joint mechanics during Lachman and Anterior Drawer tests. A mathematical construct capable of transforming the limited and heterogeneous experimental and surgical data to evidence-based validation was adopted to ensure the viability of the finite element models. We found that the proposed models, subject to a variation in tunnel directions, resulted in simulation outputs that favor the reported experimental data of Lachman and Anterior Drawer tests under uncertainty. Simulation results for a population of three-dimensional tunnel orientations provided insights into the graft–tunnel contact mechanics and the spatial stress distribution in the graft, insights that have been anecdotally observed in prior experimental studies. The intraarticular graft tension was found to be higher than the estimated in tunnel graft force, and larger differences were found for the least inclined tunnels exhibiting higher contact pressures, transverse bending and twisting of the graft and Von-Mises stress at the graft–femoral tunnel interface. Conversely, tunnels with high inclination angles exhibited higher intraarticular graft tension and Von-Mises stress at the graft–tibial bone plug interface.