Celecoxib exerts protective effects in the vascular endothelium via COX-2-independent activation of AMPK-CREB-Nrf2 signalling

Although concern remains about the athero-thrombotic risk posed by cyclo-oxygenase (COX)-2-selective inhibitors, recent data implicates rofecoxib, while celecoxib appears equivalent to NSAIDs naproxen and ibuprofen. We investigated the hypothesis that celecoxib activates AMP kinase (AMPK) signalling to enhance vascular endothelial protection. In human arterial and venous endothelial cells (EC), and in contrast to ibuprofen and naproxen, celecoxib induced the protective protein heme oxygenase-1 (HO-1). Celecoxib derivative 2,5-dimethyl-celecoxib (DMC) which lacks COX-2 inhibition also upregulated HO-1, implicating a COX-2-independent mechanism. Celecoxib activated AMPKα(Thr172) and CREB-1(Ser133) phosphorylation leading to Nrf2 nuclear translocation. Importantly, these responses were not reproduced by ibuprofen or naproxen, while AMPKα silencing abrogated celecoxib-mediated CREB and Nrf2 activation. Moreover, celecoxib induced H-ferritin via the same pathway, and increased HO-1 and H-ferritin in the aortic endothelium of mice fed celecoxib (1000 ppm) or control chow. Functionally, celecoxib inhibited TNF-α-induced NF-κB p65(Ser536) phosphorylation by activating AMPK. This attenuated VCAM-1 upregulation via induction of HO-1, a response reproduced by DMC but not ibuprofen or naproxen. Similarly, celecoxib prevented IL-1β-mediated induction of IL-6. Celecoxib enhances vascular protection via AMPK-CREB-Nrf2 signalling, a mechanism which may mitigate cardiovascular risk in patients prescribed celecoxib. Understanding NSAID heterogeneity and COX-2-independent signalling will ultimately lead to safer anti-inflammatory drugs

Sequence Variants of the Phytophthora sojae RXLR Effector Avr3a/5 Are Differentially Recognized by Rps3a and Rps5 in Soybean

The perception of Phytophthora sojae avirulence (Avr) gene products by corresponding soybean resistance (Rps) gene products causes effector triggered immunity. Past studies have shown that the Avr3a and Avr5 genes of P. sojae are genetically linked, and the Avr3a gene encoding a secreted RXLR effector protein was recently identified. We now provide evidence that Avr3a and Avr5 are allelic. Genetic mapping data from F2 progeny indicates that Avr3a and Avr5 co-segregate, and haplotype analysis of P. sojae strain collections reveal sequence and transcriptional polymorphisms that are consistent with a single genetic locus encoding Avr3a/5. Transformation of P. sojae and transient expression in soybean were performed to test how Avr3a/5 alleles interact with soybean Rps3a and Rps5. Over-expression of Avr3a/5 in a P. sojae strain that is normally virulent on Rps3a and Rps5 results in avirulence to Rps3a and Rps5; whereas silencing of Avr3a/5 causes gain of virulence in a P. sojae strain that is normally avirulent on Rps3a and Rps5 soybean lines. Transient expression and co-bombardment with a reporter gene confirms that Avr3a/5 triggers cell death in Rps5 soybean leaves in an appropriate allele-specific manner. Sequence analysis of the Avr3a/5 gene identifies crucial residues in the effector domain that distinguish recognition by Rps3a and Rps5

A Rapid Assessment of the Availability and Use of Obstetric Care in Nigerian Healthcare Facilities

Background: As part of efforts to reduce maternal deaths in Nigeria, pregnant women are being encouraged to give birth in healthcare facilities. However, little is known about whether or not available healthcare facilities can cope with an increasing demand for obstetric care. We thus carried out this survey as a rapid and tactical assessment of facility quality. We visited 121 healthcare facilities, and used the opportunity to interview over 700 women seeking care at these facilities. Findings Most of the primary healthcare facilities we visited were unable to provide all basic Emergency Obstetric Care (bEmOC) services. In general, they lack clinical staff needed to dispense maternal and neonatal care services, ambulances and uninterrupted electricity supply whenever there were obstetric emergencies. Secondary healthcare facilities fared better, but, like their primary counterparts, lack neonatal care infrastructure. Among patients, most lived within 30 minutes of the visited facilities and still reported some difficulty getting there. Of those who had had two or more childbirths, the conditional probability of a delivery occurring in a healthcare facility was 0.91 if the previous delivery occurred in a healthcare facility, and 0.24 if it occurred at home. The crude risk of an adverse neonatal outcome did not significantly vary by delivery site or birth attendant, and the occurrence of such an outcome during an in-facility delivery may influence the mother to have her next delivery outside. Such an outcome during a home delivery may not prompt a subsequent in-facility delivery. Conclusions: In conclusion, reducing maternal deaths in Nigeria will require attention to both increasing the number of facilities with high-quality EmOC capability and also assuring Nigerian women have access to these facilities regardless of where they live

Efficiency in a forced contribution threshold public good game

We contrast and compare three ways of predicting efficiency in a forced contribution threshold public good game. The three alternatives are based on ordinal potential, quantal response and impulse balance theory. We report an experiment designed to test the respective predictions and find that impulse balance gives the best predictions. A simple expression detailing when enforced contributions result in high or low efficiency is provided

Pharmacological Inhibition of Caspase and Calpain Proteases: A Novel Strategy to Enhance the Homing Responses of Cord Blood HSPCs during Expansion

Background: Expansion of hematopoietic stem/progenitor cells (HSPCs) is a well-known strategy employed to facilitate the transplantation outcome. We have previously shown that the prevention of apoptosis by the inhibition of cysteine proteases, caspase and calpain played an important role in the expansion and engraftment of cord blood (CB) derived HSPCs. We hypothesize that these protease inhibitors might have maneuvered the adhesive and migratory properties of the cells rendering them to be retained in the bone marrow for sustained engraftment. The current study was aimed to investigate the mechanism of the homing responses of CB cells during expansion. Methodology/Principal Findings: CB derived CD34 + cells were expanded using a combination of growth factors with and without Caspase inhibitor-zVADfmk or Calpain 1 inhibitor- zLLYfmk. The cells were analyzed for the expression of homingrelated molecules. In vitro adhesive/migratory interactions and actin polymerization dynamics of HSPCs were assessed. In vivo homing assays were carried out in NOD/SCID mice to corroborate these observations. We observed that the presence of zVADfmk or zLLYfmk (inhibitors) caused the functional up regulation of CXCR4, integrins, and adhesion molecules, reflecting in a higher migration and adhesive interactions in vitro. The enhanced actin polymerization and the RhoGTPase protein expression complemented these observations. Furthermore, in vivo experiments showed a significantly enhanced homing to the bone marrow of NOD/SCID mice

Computational Prediction and Molecular Characterization of an Oomycete Effector and the Cognate Arabidopsis Resistance Gene

Hyaloperonospora arabidopsidis (Hpa) is an obligate biotroph oomycete pathogen of the model plant Arabidopsis thaliana and contains a large set of effector proteins that are translocated to the host to exert virulence functions or trigger immune responses. These effectors are characterized by conserved amino-terminal translocation sequences and highly divergent carboxyl-terminal functional domains. The availability of the Hpa genome sequence allowed the computational prediction of effectors and the development of effector delivery systems enabled validation of the predicted effectors in Arabidopsis. In this study, we identified a novel effector ATR39-1 by computational methods, which was found to trigger a resistance response in the Arabidopsis ecotype Weiningen (Wei-0). The allelic variant of this effector, ATR39-2, is not recognized, and two amino acid residues were identified and shown to be critical for this loss of recognition. The resistance protein responsible for recognition of the ATR39-1 effector in Arabidopsis is RPP39 and was identified by map-based cloning. RPP39 is a member of the CC-NBS-LRR family of resistance proteins and requires the signaling gene NDR1 for full activity. Recognition of ATR39-1 in Wei-0 does not inhibit growth of Hpa strains expressing the effector, suggesting complex mechanisms of pathogen evasion of recognition, and is similar to what has been shown in several other cases of plant-oomycete interactions. Identification of this resistance gene/effector pair adds to our knowledge of plant resistance mechanisms and provides the basis for further functional analyses

Intestinal Epithelial Cell-Specific Deletion of PLD2 Alleviates DSS-Induced Colitis by Regulating Occludin

Ulcerative colitis is a multi-factorial disease involving a dysregulated immune response. Disruptions to the intestinal epithelial barrier and translocation of bacteria, resulting in inflammation, are common in colitis. The mechanisms underlying epithelial barrier dysfunction or regulation of tight junction proteins during disease progression of colitis have not been clearly elucidated. Increase in phospholipase D (PLD) activity is associated with disease severity in colitis animal models. However, the role of PLD2 in the maintenance of intestinal barrier integrity remains elusive. We have generated intestinal specific Pld2 knockout mice (Pld2 IEC-KO) to investigate the mechanism of intestinal epithelial PLD2 in colitis. We show that the knockout of Pld2 confers protection against dextran sodium sulphate (DSS)-induced colitis in mice. Treatment with DSS induced the expression of PLD2 and downregulated occludin in colon epithelial cells. PLD2 was shown to mediate phosphorylation of occludin and induce its proteasomal degradation in a c-Src kinase-dependent pathway. Additionally, we have shown that treatment with an inhibitor of PLD2 can rescue mice from DSS-induced colitis. To our knowledge, this is the first report showing that PLD2 is pivotal in the regulation of the integrity of epithelial tight junctions and occludin turn over, thereby implicating it in the pathogenesis of colitis

Activation of Akt by the Bacterial Inositol Phosphatase, SopB, is Wortmannin Insensitive

Salmonella enterica uses effector proteins translocated by a Type III Secretion System to invade epithelial cells. One of the invasion-associated effectors, SopB, is an inositol phosphatase that mediates sustained activation of the pro-survival kinase Akt in infected cells. Canonical activation of Akt involves membrane translocation and phosphorylation and is dependent on phosphatidyl inositide 3 kinase (PI3K). Here we have investigated these two distinct processes in Salmonella infected HeLa cells. Firstly, we found that SopB-dependent membrane translocation and phosphorylation of Akt are insensitive to the PI3K inhibitor wortmannin. Similarly, depletion of the PI3K regulatory subunits p85α and p85ß by RNAi had no inhibitory effect on SopB-dependent Akt phosphorylation. Nevertheless, SopB-dependent phosphorylation does depend on the Akt kinases, PDK1 and rictor-mTOR. Membrane translocation assays revealed a dependence on SopB for Akt recruitment to Salmonella ruffles and suggest that this is mediated by phosphoinositide (3,4) P2 rather than phosphoinositide (3,4,5) P3. Altogether these data demonstrate that Salmonella activates Akt via a wortmannin insensitive mechanism that is likely a class I PI3K-independent process that incorporates some essential elements of the canonical pathway