Analysis of second messenger pathways stimulated by different chemokines acting at the chemokine receptor

The chemokine receptor, CCR5, responds to several chemokines leading to changes in activity in several signalling pathways. Here, we investigated the ability of different chemokines to provide differential activation of pathways. The effects of five CC chemokines acting at CCR5 were investigated for their ability to inhibit forskolin- stimulated 3'-5'-cyclic adenosine monophosphate (cAMP) accumulation and to stimulate Ca2+ mobilisation. in Chinese hamster ovary (CHO) cells expressing CCR5. Macrophage inflammatory protein 1 alpha (D26A) (MIP-1 alpha (D26A), CCL3 (D26A)), regulated on activation, normal T-cell expressed and secreted (RANTES, CCLS), MIP-1 beta (CCL4) and monocyte chemoattractant protein 2 (MCP-2, CCL8) were able to inhibit forskolin -stimulated CAMP accumulation, whilst MCP-4 (CCL13) could not elicit a response. CCL3 (D26A), CCL4, CCLS, CCL8 and CCL13 were able to stimulate Ca2+ mobilisation. through CCRS, although CCL3 (D26A) and CCL5 exhibited biphasic concentration-response curves. The Ca2+ responses induced by CCL4, CCL5, CCL8 and CCL13 were abolished by pertussis toxin, whereas the response to CCL3 (D26A) was only partially inhibited by pertussis toxin, indicating G(i/o)-independent signalling induced by this chemokine. Although the rank order of potency of chemokines was similar between the two assays, certain chemokines displayed different pharmacological profiles in cAMP inhibition and Ca2+ mobilisation assays. For instance, whilst CCL13 could not inhibit forskolin-stimulated cAMP accumulation, this chemokine was able to induce Ca2+ mobilisation via CCR5. It is concluded that different chemokines acting at CCR5 can induce different pharmacological responses, which may account for the broad spectrum of chemokines that can act at CCRS. (C) 2007 Elsevier Inc. All rights reserved.

IsdA protects Staphylococcus aureus against the bactericidal protease activity of apolactoferrin

An important facet of the Staphylococcus aureus host-pathogen interaction is the ability of the invading bacterium to evade host innate defenses, particularly the cocktail of host antimicrobial peptides. In this work, we showed that IsdA, a surface protein of S. aureus which is required for nasal colonization, binds to lactoferrin, the most abundant antistaphylococcal polypeptide in human nasal secretions. The presence of IsdA on the surface of S. aureus confers resistance to killing by lactoferrin. In addition, the bactericidal activity of lactoferrin was inhibited by addition of phenylmethylsulfonyl fluoride, implicating the serine protease activity of lactoferrin in the killing of S. aureus. Recombinant IsdA was a competitive inhibitor of lactoferrin protease activity. Reciprocally, antibody reactive to IsdA enhanced killing of S. aureus. Thus, IsdA can protect S. aureus against lactoferrin and acts as a protease inhibitor.

Chaos of Wolbachia Sequences Inside the Compact Fig Syconia of Ficus benjamina (Ficus: Moraceae)

Figs and fig wasps form a peculiar closed community in which the Ficus tree provides a compact syconium (inflorescence) habitat for the lives of a complex assemblage of Chalcidoid insects. These diverse fig wasp species have intimate ecological relationships within the closed world of the fig syconia. Previous surveys of Wolbachia, maternally inherited endosymbiotic bacteria that infect vast numbers of arthropod hosts, showed that fig wasps have some of the highest known incidences of Wolbachia amongst all insects. We ask whether the evolutionary patterns of Wolbachia sequences in this closed syconium community are different from those in the outside world. In the present study, we sampled all 17 fig wasp species living on Ficus benjamina, covering 4 families, 6 subfamilies, and 8 genera of wasps. We made a thorough survey of Wolbachia infection patterns and studied evolutionary patterns in wsp (Wolbachia Surface Protein) sequences. We find evidence for high infection incidences, frequent recombination between Wolbachia strains, and considerable horizontal transfer, suggesting rapid evolution of Wolbachia sequences within the syconium community. Though the fig wasps have relatively limited contact with outside world, Wolbachia may be introduced to the syconium community via horizontal transmission by fig wasps species that have winged males and visit the syconia earlier.

The impacts of European and Asian anthropogenic sulphur dioxide emissions on Sahel rainfall

In this study, the atmospheric component of a state-of-the-art climate model (HadGEM2-ES) has been used to investigate the impacts of regional anthropogenic sulphur dioxide emissions on boreal summer Sahel rainfall. The study focuses on the transient response of the West African monsoon (WAM) to a sudden change in regional anthropogenic sulphur dioxide emissions, including land surface feedbacks, but without sea surface temperature (SST) feedbacks. The response occurs in two distinct phases: 1) fast adjustment of the atmosphere on a time scale of days to weeks (up to 3 weeks) through aerosol-radiation and aerosol-cloud interactions with weak hydrological cycle changes and surface feedbacks. 2) adjustment of the atmosphere and land surface with significant local hydrological cycle changes and changes in atmospheric circulation (beyond 3 weeks). European emissions lead to an increase in shortwave (SW) scattering by increased sulphate burden, leading to a decrease in surface downward SW radiation which causes surface cooling over North Africa, a weakening of the Saharan heat low and WAM, and a decrease in Sahel precipitation. In contrast, Asian emissions lead to very little change in sulphate burden over North Africa, but they induce an adjustment of the Walker Circulation which leads again to a weakening of the WAM and a decrease in Sahel precipitation. The responses to European and Asian emissions during the second phase exhibit similar large scale patterns of anomalous atmospheric circulation and hydrological variables, suggesting a preferred response. The results support the idea that sulphate aerosol emissions contributed to the observed decline in Sahel precipitation in the second half of the twentieth century.

Effects of land management on different forms of soil carbon in olive groves in Mediterranean areas

This study analyses soil organic carbon (SOC) and hot-water extractable carbon (HWC), both measures of soil quality, under different land management: (1) conventional tillage (CT); (2) CT plus the addition of oil mill waste alperujo (A); (3) CT plus the addition of oil mill waste olive leaves (L); (4) no tillage with chipped pruned branches (NT1); and (5) no tillage with chipped pruned branches and weeds (NT2); in a typical Mediterranean agricultural area; the olive groves of Andalucía, southern Spain. SOC values in CT, A, NT1 and NT2 decreased with depth, but in NT2 the surface horizon (0-5 cm) had higher values than the other treatments, 47% more than the average values in the other three soils. In L, SOC also decreased with depth, although there was an increase of 88.5% from the first (0-10 cm) to the second horizon (10-16 cm). Total SOC stock values were very similar under A (101.9 Mg ha−1), CT (101.7 Mg ha−1), NT1 (105.8 Mg ha−1) and NT2 (111.3 Mg ha−1, if we consider the same depth of the others). However, SOC under L was significantly higher (p < 0.05) at 250.2 Mg ha−1. HWC decreased with depth in A, CT and NT1. NT2 and L followed the same pattern as the other management types but with a higher value in the surface horizon (2.3 and 4.9 mg g−1 respectively). Overall, our results indicate that application of oil mill waste olive leaves under CT (L) is a good management practice to improve SOC and reduce waste.

Cluster analysis of downscaled and explicitly simulated North Atlantic tropical cyclone tracks

A realistic representation of the North Atlantic tropical cyclone tracks is crucial as it allows, for example, explaining potential changes in US landfalling systems. Here we present a tentative study, which examines the ability of recent climate models to represent North Atlantic tropical cyclone tracks. Tracks from two types of climate models are evaluated: explicit tracks are obtained from tropical cyclones simulated in regional or global climate models with moderate to high horizontal resolution (1° to 0.25°), and downscaled tracks are obtained using a downscaling technique with large-scale environmental fields from a subset of these models. For both configurations, tracks are objectively separated into four groups using a cluster technique, leading to a zonal and a meridional separation of the tracks. The meridional separation largely captures the separation between deep tropical and sub-tropical, hybrid or baroclinic cyclones, while the zonal separation segregates Gulf of Mexico and Cape Verde storms. The properties of the tracks’ seasonality, intensity and power dissipation index in each cluster are documented for both configurations. Our results show that except for the seasonality, the downscaled tracks better capture the observed characteristics of the clusters. We also use three different idealized scenarios to examine the possible future changes of tropical cyclone tracks under 1) warming sea surface temperature, 2) increasing carbon dioxide, and 3) a combination of the two. The response to each scenario is highly variable depending on the simulation considered. Finally, we examine the role of each cluster in these future changes and find no preponderant contribution of any single cluster over the others.

Regulation of actin assembly by SCAR/WAVE proteins.

Actin reorganization is a tightly regulated process that co-ordinates complex cellular events, such as cell migration, chemotaxis, phagocytosis and adhesion, but the molecular mechanisms that underlie these processes are not well understood. SCAR (suppressor of cAMP receptor)/WAVE [WASP (Wiskott-Aldrich syndrome protein)-family verprolin homology protein] proteins are members of the conserved WASP family of cytoskeletal regulators, which play a critical role in actin dynamics by triggering Arp2/3 (actin-related protein 2/3)-dependent actin nucleation. SCAR/WAVEs are thought to be regulated by a pentameric complex which also contains Abi (Abl-interactor), Nap (Nck-associated protein), PIR121 (p53-inducible mRNA 121) and HSPC300 (haematopoietic stem progenitor cell 300), but the structural organization of the complex and the contribution of its individual components to the regulation of SCAR/WAVE function remain unclear. Additional features of SCAR/WAVE regulation are highlighted by the discovery of other interactors and distinct complexes. It is likely that the combinatorial assembly of different components of SCAR/WAVE complexes will prove to be vital for their roles at the centre of dynamic actin reorganization.

The impact of a thermodynamic sea-ice module in the COSMO numerical weather prediction model on simulations for the Laptev Sea, Siberian Arctic

Previous versions of the Consortium for Small-scale Modelling (COSMO) numerical weather prediction model have used a constant sea-ice surface temperature, but observations show a high degree of variability on sub-daily timescales. To account for this, we have implemented a thermodynamic sea-ice module in COSMO and performed simulations at a resolution of 15 km and 5 km for the Laptev Sea area in April 2008. Temporal and spatial variability of surface and 2-m air temperature are verified by four automatic weather stations deployed along the edge of the western New Siberian polynya during the Transdrift XIII-2 expedition and by surface temperature charts derived from Moderate Resolution Imaging Spectroradiometer (MODIS) satellite data. A remarkable agreement between the new model results and these observations demonstrates that the implemented sea-ice module can be applied for short-range simulations. Prescribing the polynya areas daily, our COSMO simulations provide a high-resolution and high-quality atmospheric data set for the Laptev Sea for the period 14-30 April 2008. Based on this data set, we derive a mean total sea-ice production rate of 0.53 km3/day for all Laptev Sea polynyas under the assumption that the polynyas are ice-free and a rate of 0.30 km3/day if a 10-cm-thin ice layer is assumed. Our results indicate that ice production in Laptev Sea polynyas has been overestimated in previous studies.

Effect of meloxicam and diclofenac sodium on peri-implant bone healing in rats

Background: This study evaluated the effects of diclofenac sodium and meloxicam on peri-implant bone healing. Methods: Thirty male rats were divided into three groups: the control group (CG) received no drug; the diclofenac sodium group (DSG) received 1.07 mg/kg twice a day for 5 days; and the meloxicam group (MG) received 0.2 mg/kg daily for 5 days. A screw-shaped titanium implant was placed in the tibia. Fluorochromes, oxytetracycline (OxT), calcein (CA), and alizarin (AL), were injected at 7, 14, and 21 days, respectively, after implantation, and the animals were sacrificed 28 days after implant placement. The percentages of OxT-, CA-, and AL-labeled bone as well as the percentages of bone-to-implant contact (BIC), cortical bone area (CBA), and trabecular bone area (TBA) within the implant threads were evaluated. Results: Bone healing was delayed in the DSG during the first 14 days after implant placement (OxT-labeled bone: DSG: 5.3% +/- 7.3% versus CG: 13.2% +/- 9.8%, P= 0.002, and versus MG: 14.4% +/- 13.1%, P = 0.05). The percentages of BIC (DSG: 49.6% +/- 21.9%; MG: 67.1% +/- 22.8%; and CG: 68.1% +/- 22.8%) and CBA (DSG: 63.7% +/- 21.2%; MG: 82.7% +/- 12.4%; CG: 84.9% +/- 10.6%) were lower in the DSG compared to the MG and CG (P<0.001). The percentage of TBA was significantly greater in the DSG compared to the MG and CG (DSG: 36.3% +/- 21.2% versus MG: 17.3% +/- 12.7% and versus CG: 15.1% +/- 10.6%; P<0.001). Conclusion: Diclofenac sodium seemed to delay peri-implant bone healing and to decrease BIC, whereas meloxicam had no negative effect on peri-implant bone healing.

Long-Term Culture of Mouse Embryonic Stem Cell-Derived Adherent Neurospheres and Functional Neurons

Innumerous protocols, using the mouse embryonic stem (ES) cells as model for in vitro study of neurons functional properties and features, have been developed. Most of these protocols are short lasting, which, therefore, does not allow a careful analysis of the neurons maturation, aging, and death processes. We describe here a novel and efficient long-lasting protocol for in vitro ES cells differentiation into neuronal cells. It consists of obtaining embryoid bodies, followed by induction of neuronal differentiation with retinoic acid of nonadherent embryoid bodies (three-dimensional model), which further allows their adherence and formation of adherent neurospheres (AN, bi-dimensional model). The AN can be maintained for at least 12 weeks in culture under repetitive mechanical splitting, providing a constant microenvironment (in vitro niche) for the neuronal progenitor cells avoiding mechanical dissociation of AN. The expression of neuron-specific proteins, such as nestin, sox1, beta III-tubulin, microtubule-associated protein 2, neurofilament medium protein, Tau, neuronal nuclei marker, gamma-aminobutyric acid, and 5-hydroxytryptamine, were confirmed in these cells maintained during 3 months under several splitting. Additionally, expression pattern of microtubule-associated proteins, such as lissencephaly (Lis1) and nuclear distribution element-like (Ndel1), which were shown to be essential for differentiation and migration of neurons during embryogenesis, was also studied. As expected, both proteins were expressed in undifferentiated ES cells, AN, and nonrosette neurons, although presenting different spatial distribution in AN. In contrast to previous studies, using cultured neuronal cells derived from embryonic and adult tissues, only Ndel1 expression was observed in the centrosome region of early neuroblasts from AN. Mature neurons, obtained from ES cells in this work, display ionic channels and oscillations of membrane electrical potential typical of electrically excitable cells, which is a characteristic feature of the functional central nervous system (CNS) neurons. Taken together, our study demonstrated that AN are a long-term culture of neuronal cells that can be used to analyze the process of neuronal differentiation dynamics. Thus, the protocol described here provides a new experimental model for studying neurological diseases associated with neuronal differentiation during early development, as well as it represents a novel source of functional cells that can be used as tools for testing the effects of toxins and/or drugs on neuronal cells.

REACTIVE OXYGEN SPECIES AND THE STRUCTURAL REMODELING OF THE VISUAL SYSTEM AFTER OCULAR ENUCLEATION

Redox processes associated with controlled generation of reactive oxygen species (ROS) by NADPH oxidase (Nox) add an essential level of regulation to signaling pathways underlying physiological processes. We evaluated the ROS generation in the main visual relays of the mammalian brain, namely the superior colliculus (SC) and the dorsal lateral geniculate nucleus (DLG), after ocular enucleation in adult rats. Dihydroethidium (DHE) oxidation revealed increased ROS generation in SC and DLG between 1 and 30 days postlesion. ROS generation was decreased by the Nox inhibitors diphenyleneiodonium chloride (DPI) and apocynin. Real-time PCR results revealed that Nox 2 was upregulated in both retinorecipient structures after deafferentation, whereas Nox 1 and Nox 4 were upregulated only in the SC. To evaluate the role of ROS in structural remodeling after the lesions, apocynin was given to enucleated rats and immunohistochemistry was conducted for markers of neuronal remodeling into SC and DLG. Immunohistochemical data showed that ocular enucleation produces an increase of neurofilament and microtubule-associated protein-2 immunostaining in both SC and DLG, which was markedly attenuated by apocynin treatment. Taken together, the findings of the present study suggest a novel role for Nox-induced ROS signaling in mediating neuronal remodeling in visual areas after ocular enucleation. (C) 2010 IBRO. Published by Elsevier Ltd. All rights reserved.

In vitro analysis of human tooth pulp chamber temperature after low-intensity laser therapy at different power outputs

In vitro studies have provided conflicting evidence of temperature changes in the tooth pulp chamber after low-level laser irradiation of the tooth surface. The present study was an in vitro evaluation of temperature increases in the human tooth pulp chamber after diode laser irradiation (GaAlAs, lambda = 808 nm) using different power densities. Twelve human teeth (three incisors, three canines, three premolars and three molars) were sectioned in the cervical third of the root and enlarged for the introduction of a thermocouple into the pulp chamber. The teeth were irradiated with 417 mW, 207 mW and 78 mW power outputs for 30 s on the vestibular surface approximately 2 mm from the cervical line of the crown. The highest average increase in temperature (5.6A degrees C) was observed in incisors irradiated with 417 mW. None of the teeth (incisors, canines, premolars or molars) irradiated with 207 mW showed temperature increases higher than 5.5A degrees C that could potentially be harmful to pulp tissue. Teeth irradiated with 78 mW showed lower temperature increases. The study showed that diode laser irradiation with a wavelength of 808 nm at 417 mW power output increased the pulp chamber temperature of certain groups of teeth, especially incisors and premolars, to critical threshold values for the dental pulp (5.5A degrees C). Thus, this study serves as a warning to clinicians that ""more"" is not necessarily ""better"".

Worldwide sequence conservation of transmission-blocking vaccine candidate Pvs230 in Plasmodium vivax

Pfs230, surface protein of gametocyte/gamete of the human malaria parasite, Plasmodium falciparum, is a prime candidate of malaria transmission-blocking vaccine. Plasmodium vivax has an ortholog of Pfs230 (Pvs230), however, there has been no study in any aspects on Pvs230 to date. To investigate whether Pvs230 can be a vivax malaria transmission-blocking vaccine, we performed evolutionary and population genetic analysis of the Pvs230 gene (pvs230: PVX_003905). Our analysis of Pvs230 and its orthologs in eight Plasmodium species revealed two distinctive parts: an interspecies variable part (IVP) containing species-specific oligopeptide repeats at the N-terminus and a 7.5 kb interspecies conserved part (ICP) containing 14 cysteine-rich domains. Pvs230 was closely related to its orthologs, Pks230 and Pcys230, in monkey malaria parasites. Analysis of 113 pvs230 sequences obtained from worldwide, showed that nucleotide diversity is remarkably low in the non-repeat 8-kb region of pvs230 (theta pi = 0.00118) with 77 polymorphic nucleotide sites, 40 of which results in amino acid replacements. A signature of purifying selection but not of balancing selection was seen on pvs230. Functional and/or structural constraints may limit the level of polymorphism in pvs230. The observed limited polymorphism in pvs230 should ground for utilization of Pvs230 as an effective transmission-blocking vaccine. (C) 2011 Elsevier Ltd. All rights reserved.

Extensive microsatellite diversity in the human malaria parasite Plasmodium vivax

The population structure of Plasmodium vivax remains elusive. The markers of choice for large-scale population genetic studies of eukaryotes, short tandem repeats known as microsatellites, have been recently reported to be less polymorphic in R vivax. Here we investigate the microsatellite diversity and geographic structure in P vivax, at both local and global levels, using 14 new markers consisting of tri- or tetranucleotide repeats. The local-level analysis, which involved 50 field isolates from Sri Lanka, revealed unexpectedly high diversity (average virtual heterozygosity [H-E], 0.807) and significant multilocus linkage disequilibrium in this region of low malaria endemicity. Multiple-clone infections occurred in 60% of isolates sampled in 2005. The global-level analysis of field isolates or monkey-adapted strains identified 150 unique haplotypes among 164 parasites from four continents. Individual P. vivax isolates could not be unambiguously assigned to geographic populations. For example, we found relatively low divergence among parasites from Central America, Africa, Southeast Asia and Oceania, but substantial differentiation between parasites from the same continent (South Asia and Southeast Asia) or even from the same country (Brazil). Parasite relapses, which may extend the duration of P. vivax carriage in humans, are suggested to facilitate the spread of strains across continents, breaking down any pre-existing geographic structure. (C) 2008 Elsevier B.V. All rights reserved.

Inflammation of the Hypothalamus Leads to Defective Pancreatic Islet Function

Type 2 diabetes mellitus results from the complex association of insulin resistance and pancreatic beta-cell failure. Obesity is the main risk factor for type 2 diabetes mellitus, and recent studies have shown that, in diet-induced obesity, the hypothalamus becomes inflamed and dysfunctional, resulting in the loss of the perfect coupling between caloric intake and energy expenditure. Because pancreatic beta-cell function is, in part, under the control of the autonomic nervous system, we evaluated the role of hypothalamic inflammation in pancreatic islet function. In diet-induced obesity, the earliest markers of hypothalamic inflammation are present at 8 weeks after the beginning of the high fat diet; similarly, the loss of the first phase of insulin secretion is detected at the same time point and is restored following sympathectomy. Intracerebroventricular injection of a low dose of tumor necrosis factor a leads to a dysfunctional increase in insulin secretion and activates the expression of a number of markers of apoptosis in pancreatic islets. In addition, the injection of stearic acid intracerebroventricularly, which leads to hypothalamic inflammation through the activation of tau-like receptor-4 and endoplasmic reticulum stress, produces an impairment of insulin secretion, accompanied by increased expression of markers of apoptosis. The defective insulin secretion, in this case, is partially dependent on sympathetic signal-induced peroxisome proliferator receptor-gamma coactivator Delta a and uncoupling protein-2 expression and is restored after sympathectomy or following PGC1 alpha expression inhibition by an antisense oligonucleotide. Thus, the autonomic signals generated in concert with hypothalamic inflammation can impair pancreatic islet function, a phenomenon that may explain the early link between obesity and defective insulin secretion.