Inhibition of Na+/H+ exchanger enhances low pH-induced L-selectin shedding and beta2-integrin surface expression in human neutrophils

Ischemia-reperfusion injury is a common pathological occurrence causing tissue damage in heart attack and stroke. Entrapment of neutrophils in the vasculature during ischemic events has been implicated in this process. In this study, we examine the effects that lactacidosis and consequent reductions in intracellular pH (pH(i)) have on surface expression of adhesion molecules on neutrophils. When human neutrophils were exposed to pH 6 lactate, there was a marked decrease in surface L-selectin (CD62L) levels, and the decrease was significantly enhanced by inclusion of Na(+)/H(+) exchanger (NHE) inhibitor 5-(N,N-hexamethylene)amiloride (HMA). Similar effects were observed when pH(i) was reduced while maintaining normal extracellular pH, by using an NH(4)Cl prepulse followed by washes and incubation in pH 7.4 buffer containing NHE inhibitors [HMA, cariporide, or 5-(N,N-dimethyl)amiloride (DMA)]. The amount of L-selectin shedding induced by different concentrations of NH(4)Cl in the prepulse correlated with the level of intracellular acidification with an apparent pK of 6.3. In contrast, beta(2)-integrin (CD11b and CD18) was only slightly upregulated in the low-pH(i) condition and was enhanced by NHE inhibition to a much lesser extent. L-selectin shedding was prevented by treating human neutrophils with inhibitors of extracellular metalloproteases (RO-31-9790 and KD-IX-73-4) or with inhibitors of intracellular signaling via p38 MAP kinase (SB-203580 and SB-239063), implying a transmembrane effect of pH(i). Taken together, these data suggest that the ability of NHE inhibitors such as HMA to reduce ischemia-reperfusion injury may be related to the nearly complete removal of L-selectin from the neutrophil surface.     

Factors influencing the geographical distribution of Dendroctonus rhizophagus (Coleoptera: Curculionidae: Scolytinae) in the Sierra Madre Occidental, México

The bark beetle, Dendroctonus rhizophagus Thomas & Bright, is endemic to the Sierra Madre Occidental (SMOC) in México. This bark beetle is a major pest of the seedlings and young saplings of several pine species that are of prime importance to the nation's forest industry. Despite the significance of this bark beetle as a pest, its biology, ecology, and distribution are poorly known. Three predictive modeling approaches were used as a first approximation to identify bioclimatic variables related to the presence of D. rhizophagus in the SMOC and to obtain maps of its potential distribution within the SMOC, which is a morphotectonic province. Our results suggest that the bark beetle could have an almost continuous distribution throughout the major mountain ranges of the SMOC. This beetle has a relatively narrow ecological niche with respect to some temperature and precipitation variables and inhabits areas with climatic conditions that are unique from those usually prevalent in the SMOC. However, the bark beetle has a broad ecological niche with respect to the number of hosts that it attacks. At the macro-scale level, the D. rhizophagus distribution occurs within the wider distribution of its main hosts. The limit of the geographical distribution of this bark beetle coincides with the maximum temperature isotherms. Our results imply a preference for temperate habitats, which leads to the hypothesis that even minor changes in climate may have significant effects on its distribution and abundance.     

Role of alpha adrenoceptors and nitric oxide on cardiovascular responses in acute and chronic hypertension

The contribution of α-adrenoceptors and nitric oxide (NO) on the alterations of sympathetically mediated cardiovascular responses after acute (AcH) and chronic (ChH) hypertension was evaluated in pithed aortic coarcted hypertensive rats. Pressor and tachycardia response produced by electrical stimulation of preganglionic sympathetic fibers or exogenous noradrenaline (NA) were recorded in the absence and presence of prazosin (α₁-antagonist), rauwolscine (α₂-antagonist), or N ^G-nitro-l-arginine methyl ester (l-NAME; an inhibitor of NO synthase). Compared with age-matched sham-operated rats (Nt), the pressor response produced by electrical stimulation or NA was smaller in AcH rats and larger in ChH rats. Prazosin caused a decrease of pressor response elicited by electrical stimulation or NA in all groups. However, this effect was higher in ChH. Rauwolscine produced a similar increase of sympathetically mediated pressor response in Nt and AcH rats. Nevertheless, this antagonist did not affect the sympathetically mediated pressor response in ChH rats. In addition, rauwolscine did not affect the NA-induced pressor response in all groups. The pressor response elicited by l-NAME was larger in all groups compared without l-NAME and in presence of l-arginine. Moreover, l-NAME in the presence of NA increased sympathetically mediated pressor response is in all groups, compared without it or in the presence of l-arginine. Compared with Nt, basally produced NO in aortic rings was increased in AcH but decreased in ChH. Collectively, our data suggest that decreased cardiovascular reactivity in AcH is due to an increase in basally produced NO. In ChH, enhanced cardiovascular response appears to be associated with a decrease in produced NO and an increase in released NA from sympathetic nerves.     

The impact of livestock grazing on the spatial pattern of vegetation in north-eastern Patagonia,Argentina

Background: The occurrence of shrub patches, alternating with either bare soil or low herbaceous cover, is a common feature in arid and semi-arid shrublands throughout the world. This patchy pattern of vegetation may result from water limitation, modulated by plant interactions; grazing (offtake and tramping) by livestock may cause further patchiness vegetation structure.

Aims: We hypothesised that vegetation patchiness in the semi-arid shrublands of north-eastern Patagonia would be increased by livestock grazing, but not by positive interactions between adult plants of shrubs and grasses.

Methods: We compared vegetation cover and pattern at three grazing intensities (exclosure, light and heavy grazing) and measured the growth of a representative shrub and grass in the presence and absence of the other to quantify the role of plant-to-plant interactions and its interaction with grazing for vegetation structure.

Results: In the grazing exclosure and in moderately grazed areas, vegetation cover among shrub patches was larger, whereas the top cover of shrubs was lower than in the heavily grazed areas. We did not find any evidence of positive interactions between shrub and grass life forms.

Conclusions: Our results were consistent with the hypothesis that livestock grazing increased the formation of patchy vegetation cover in arid and semi-arid shrublands.     

Leishmania infantum Amastigotes Trigger a Subpopulation of Human B Cells with an Immunoregulatory Phenotype

Visceral leishmaniasis is caused by the protozoan parasites Leishmania infantum and Leishmania donovani. This infection is characterized by an uncontrolled parasitization of internal organs which, when left untreated, leads to death. Disease progression is linked with the type of immune response generated and a strong correlation was found between disease progression and serum levels of the immunosuppressive cytokine IL-10. Other studies have suggested a role for B cells in the pathology of this parasitic infection and the recent identification of a B-cell population in humans with regulatory functions, which secretes large amounts of IL-10 following activation, have sparked our interest in the context of visceral leishmaniasis. We report here that incubation of human B cells with Leishmania infantum amastigotes resulted in upregulation of multiple cell surface activation markers and a dose-dependent secretion of IL-10. Conditioned media from B cells incubated with Leishmania infantum amastigotes were shown to strongly inhibit CD4⁺ T-cell activation, proliferation and function (i.e. as monitored by TNF and IFNγ secretion). Blockade of IL-10 activity using a soluble IL-10 receptor restored only partially TNF and IFNγ production to control levels. The parasite-mediated IL-10 secretion was shown to rely on the activity of Syk, phosphatidylinositol-3 kinase and p38, as well as to require intracellular calcium mobilization. Cell sorting experiments allowed us to identify the IL-10-secreting B-cell subset (i.e. CD19⁺CD24⁺CD27^-). In summary, exposure of human B cells to Leishmania infantum amastigotes triggers B cells with regulatory activities mediated in part by IL-10, which could favor parasite dissemination in the organism.     

S-layer proteins of Lactobacillus acidophilus inhibits JUNV infection

We investigated the regulation of Hsp27 phosphorylation by protein kinase C δ (PKCδ) during etoposide-induced apoptosis. The phosphorylation of Hsp27 at Ser78 was temporally correlated with the proteolytic activation of PKCδ during apoptosis. Hsp27 phosphorylation was dependent on the activity of PKCδ since treatment with rottlerin, a chemical inhibitor of PKCδ, or overexpression of a PKCδ dominant negative mutant abolished the phosphorylation. In addition, recombinant PKCδ phosphorylated Hsp27 at Ser78 in vitro. Moreover, caspase-3 was specifically activated following Hsp27 phosphorylation at Ser78. Pull-down assays using a phosphomimetic Hsp27 mutant revealed that binding between Hsp27 and cytochrome c was abolished by the phosphorylation. These results suggest that Hsp27 dissociates from cytochrome c following PKCδ-mediated phosphorylation at Ser78, which allows formation of the apoptosome and stimulates apoptotic progression.     

Recent Rapid Rise of a Permethrin Knock Down Resistance Allele in Aedes aegypti in México

Background

Aedes aegypti, the ‘yellow fever mosquito’, is the primary vector to humans of dengue and yellow fever flaviviruses (DENV, YFV), and is a known vector of the chikungunya alphavirus (CV). Because vaccines are not yet available for DENV or CV or are inadequately distributed in developing countries (YFV), management of Ae. aegypti remains the primary option to prevent and control outbreaks of the diseases caused by these arboviruses. Permethrin is one of the most widely used active ingredients in insecticides for suppression of adult Ae. aegypti. In 2007, we documented a replacement mutation in codon 1,016 of the voltage-gated sodium channel gene (para) of Ae. aegypti that encodes an isoleucine rather than a valine and confers resistance to permethrin. Ile1,016 segregates as a recessive allele conferring knockdown resistance to homozygous mosquitoes at 5–10 µg of permethrin in bottle bioassays.

Methods and Findings

A total of 81 field collections containing 3,951 Ae. aegypti were made throughout México from 1996 to 2009. These mosquitoes were analyzed for the frequency of the Ile1,016 mutation using a melting-curve PCR assay. Dramatic increases in frequencies of Ile1,016 were recorded from the late 1990''s to 2006–2009 in several states including Nuevo León in the north, Veracruz on the central Atlantic coast, and Yucatán, Quintana Roo and Chiapas in the south. From 1996 to 2000, the overall frequency of Ile1,016 was 0.04% (95% confidence interval (CI95) = 0.12%; n = 1,359 mosquitoes examined). The earliest detection of Ile1,016 was in Nuevo Laredo on the U.S. border in 1997. By 2003–2004 the overall frequency of Ile1,016 had increased ∼100-fold to 2.7% (±0.80% CI95; n = 808). When checked again in 2006, the frequency had increased slightly to 3.9% (±1.15% CI95; n = 473). This was followed in 2007–2009 by a sudden jump in Ile1,016 frequency to 33.2% (±1.99% CI95; n = 1,074 mosquitoes). There was spatial heterogeneity in Ile1,016 frequencies among 2007–2008 collections, which ranged from 45.7% (±2.00% CI95) in the state of Veracruz to 51.2% (±4.36% CI95) in the Yucatán peninsula and 14.5% (±2.23% CI95) in and around Tapachula in the state of Chiapas. Spatial heterogeneity was also evident at smaller geographic scales. For example within the city of Chetumal, Quintana Roo, Ile1,016 frequencies varied from 38.3%–88.3%. A linear regression analysis based on seven collections from 2007 revealed that the frequency of Ile1,016 homozygotes accurately predicted knockdown rate for mosquitoes exposed to permethrin in a bioassay (R² = 0.98).

Conclusions

We have recorded a dramatic increase in the frequency of the Ile1,016 mutation in the voltage-gated sodium channel gene of Ae. aegypti in México from 1996 to 2009. This may be related to heavy use of permethrin-based insecticides in mosquito control programs. Spatial heterogeneity in Ile1,016 frequencies in 2007 and 2008 collections may reflect differences in selection pressure or in the initial frequency of Ile1,016. The rapid recent increase in Ile1,016 is predicted by a simple model of positive directional selection on a recessive allele. Unfortunately this model also predicts rapid fixation of Ile1,016 unless there is negative fitness associated with Ile1,016 in the absence of permethrin. If so, then spatial refugia of susceptible Ae. aegypti or rotational schedules of different classes of adulticides could be established to slow or prevent fixation of Ile1,016.     

p38gamma regulates the localisation of SAP97 in the cytoskeleton by modulating its interaction with GKAP

Activation of the p38 MAP kinase pathways is crucial for the adaptation of mammalian cells to changes in the osmolarity of the environment. Here we identify SAP97/hDlg, the mammalian homologue of the Drosophila tumour suppressor Dlg, as a physiological substrate for the p38gamma MAP kinase (SAPK3/p38gamma) isoform. SAP97/hDlg is a scaffold protein that forms multiprotein complexes with a variety of proteins and is targeted to the cytoskeleton by its association with the protein guanylate kinase-associated protein (GKAP). The SAPK3/p38gamma-catalysed phosphorylation of SAP97/hDlg triggers its dissociation from GKAP and therefore releases it from the cytoskeleton. This is likely to regulate the integrity of intercellular-junctional complexes, and cell shape and volume in response to osmotic stress.